Forest Health Technology 

Enterprise Team 

Technology 

Transfer Biological

Control
 

ImplementatIon and StatuS
 

of BIologIcal control
 

of the hemlock Woolly adelgId
 

United states forest health forest fhTeT-2011-04 

Department Technology service December 2011 

of agriculture enterprise Team 



 

 

 

Forest Health Technology 

Enterprise Team 

Technology 

Transfer Biological

Control
 

ImplementatIon and StatuS
 

of BIologIcal control
 

of the hemlock Woolly adelgId
 

United states forest health forest fhTeT-2011-04 

Department Technology service December 2011 

of agriculture enterprise Team 

IMPLEMENTATION AND STATUS
 

OF BIOLOGICAL CONTROL
 

OF THE HEMLOCK WOOLLY ADELGID
 


 

 

 

The Forest Health Technology Enterprise Team (FHTET) was created in 

1995 by the Deputy Chief for State and Private Forestry, Forest Service, U.S. 

Department of Agriculture, to develop and deliver technologies to protect 

and improve the health of American forests. This book was published by 

FHTET as part of the technology transfer series. 

http://www.fs.fed.us/foresthealth/technology/ 

On the cover: 

Center photo: Hemlock woolly adelgid white woolly masses on a hemlock branch (USDA Forest Service, Karen Felton) 

Top right: Sasajiscymnus tsugae predatory beetle (USDA Forest Service, Lynn Jones); 

Middle right: Collecting and checking hemlock branch samples for Laricobius nigrinus predatory beetle larvae 

(USDA Forest Service, Brad Onken); 

Bottom center: Collecting Laricobius nigrinus predatory beetles in Idaho (USDA Forest Service, Brad Onken); 

Middle left: Releasing Laricobius nigrinus predatory beetles (USDA Forest Service, Brad Onken); 

Top left: Laricobius nigrinus predatory beetle (USDA Forest Service, Lynn Jones) 

For additional copies of this publication, contact: 

Brad Onken Richard Reardon 

U.S. Forest Service U.S. Forest Service
 

180 Canfi eld Street 180 Canfi eld Street
 

Morgantown, WV 26505 Morgantown, WV 26505
 

(304) 285-1546 (304) 285-1566
 

bonken@fs.fed.us rreardon@fs.fed.us
 

The entire publication is available online at 

http://www.fs.fed.us/na/morgantown/fhp/hwa 

The U.S. Department of Agriculture (USDA) prohibits discrimination in all its programs and 

activities on the basis of race, color, national origin, sex, religion, age, disability, political 

beliefs, sexual orientation, and marital or family status. (Not all prohibited bases apply to 

all programs.) Persons with disabilities who require alternative means for communication 

of program information (Braille, large print, audiotape, etc.) should contact USDA’s TARGET 

Center at 202-720-2600 (voice and TDD). 

To file a complaint of discrimination, write USDA, Director, Office of Civil Rights, Room 326-W, 

Whitten Building, 1400 Independence Avenue SW, Washington, DC 20250-9410 or call 

202-720-5964 (voice and TDD). USDA is an equal opportunity provider and employer. 

The use of trade, firm, or corporation names in this publication is for the information 

and convenience of the reader. Such use does not constitute an offi cial endorsement 

or approval by the U.S. Department of Agriculture or the Forest Service of any product 

or service to the exclusion of others that may be suitable. 

Federal Recycling Program
 

Printed on recycled paper
 
IMPLEMENTATION AND STATUS
 

OF BIOLOGICAL CONTROL
 

OF THE HEMLOCK WOOLLY ADELGID
 

Technical Coordinators 

Brad Onken
 

Forest Health Protection, U.S. Forest Service
 

180 Canfield Street, Morgantown, WV 26505
 

Richard Reardon
 

Forest Health Technology Enterprise Team, U.S. Forest Service
 

180 Canfield Street, Morgantown, WV 26505
 

U.S. Forest Service
 

Publication FHTET-2011-04
 
ACKNOWLEDGMENTS
 

We thank the authors of the individual chapters 

for their expertise in reviewing and summarizing 

the literature and providing current information 

on biological control of hemlock woolly adelgid. 

Thanks to Wendy Harding for design and layout 

of this publication. Special thanks to the many 

graduate students, technicians, and summer fi eld 

and laboratory assistants, too numerous to list 

here, for their hard work and dedication to the 

development of biological control for hemlock 

woolly adelgid. 

We would also like to thank the U.S. Department 

of Agriculture, Forest Service, Forest Health 

Protection, Northeastern Area, Region 8, and Forest 

Health Technology Enterprise Team, for providing 

funding for the preparation and printing of this 

publication. 
PREFACE
 

Hemlock woolly adelgid (HWA), Adelges tsugae 

(Annand), remains the single greatest threat to the 

health and sustainability of hemlock as a forest 

resource in the eastern United States. It is an exotic 

pest native to Asia and western North America. 

First discovered in the eastern United States in 1951 

near Richmond, VA, the pathway and source of the 

introduction are believed to have originated from 

southern Japan on nursery stock. In the late 1980s, 

HWA began to spread rapidly from its point of 

introduction. It has since spread to 17 eastern states 

threatening two species of hemlock: the eastern 

hemlock, Tsuga canadensis (L.) Carr., and Carolina 

hemlock, Tsuga caroliniana Engelm. On these 

hemlocks, HWA has two generations per year and 

is parthogenic; that is, all individuals are female, 

capable of reproducing. 

The Hemlock Woolly Adelgid Initiative began in 

2003 with the development of a 5-year program 

(2003-2007) in cooperation with the U.S. 

Department of Agriculture Forest Service and the 

Animal Plant Health Inspection Service (APHIS), 

the National Association of State Foresters, and 

the National Plant Board for the development and 

implementation of management options to reduce 

the spread and impact of HWA. A second 5-year 

program (2008-2012) was funded to continue 

and to accelerate the development of research and 

technology, management, and information transfer 

program components. 

Although the earliest investigations into HWA 

biological control date back to the early 1990s, the 

biological control effort has been greatly expanded 

with the onset of the Hemlock Woolly Adelgid 

Initiative and has involved 28 federal and state 

agencies, 24 universities, 7 institutions in China and 

Japan, and numerous private industries. Biological 

control offers a potential long-term solution to 

suppression of HWA and needs to become the focal 

component of an integrated management program. 

The purpose of this book is to provide a reference 

guide for field workers and land managers on the 

historical and current status of the biological control 

of hemlock woolly adelgid. This book is a substantial 

revision of FHTET-2004-04, Biological Control of 

Hemlock Woolly Adelgid. 
TABLE OF CONTENTS
 

SECTION I INTRODUCTION 

Chapter 1: Hemlock Woolly Adelgid and its Hemlock Hosts: A Global Perspective . . . . . . . . . . . . . .3
 

Nathan Havill, Michael E. Montgomery, and Melody A. Keena 

Chapter 2: Simulations of Population Dynamics of Hemlock Woolly Adelgid 


 and Potential Impact of Biological Control Agents . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .15
 

Joseph S. Elkinton, Robert T. Trotter, and Ann F. Paradis 

Chapter 3: Understanding Federal Regulations as Guidelines 


 for Classical Biological Control Programs. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .25
 

Michael E. Montgomery 

SECTION II AGENTS FOR BIOLOGICAL CONTROL 

Chapter 4: Sasajiscymnus (=Pseudoscymnus) tsugae, a ladybeetle from Japan . . . . . . . . . . . . . . . . . .43
 

Carole Cheah 

Chapter 5: Scymnus (Neopullus) Lady Beetles from China . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .53
 

Michael E. Montgomery and Melody A. Keena 

Chapter 6: Laricobius nigrinus Fender (Coleoptera: Derodontidae) . . . . . . . . . . . . . . . . . . . . . . . . . . .77
 

D.L. Mausel, G.A. Davis, A.S. Lamb, G.M.G. Zilahi-Balogh, 

L.T. Kok, and S.M. Salom 

Chapter 7: Laricobius osakensis, a Hemlock Woolly Adelgid Predator from Japan . . . . . . . . . . . . . . .90
 

Ashley Lamb, Michael E. Montgomery, Ligia Cota Viera, 

Shigehiko Shiyake, and Scott Salom 

Chapter 8: Chamaemyiid Predators of the Hemlock Woolly Adelgid 


 from the Pacific Northwest . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .97
 

Darrell W. Ross, Stephen D. Gaimari, Glenn R. Kohler, 

Kimberly F. Wallin, and Sarah M. Grubin 

Chapter 9: Insect-Killing Fungi for HWA Management: Current Status . . . . . . . . . . . . . . . . . . . . . . 107
 

Scott D. Costa 

Chapter 10: Other Species Considered . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 116
 

Michael E. Montgomery, Thomas J. McAvoy, and Scott M. Salom 

SECTION III LABORATORY REARING FOR FIELD RELEASE 

Chapter 11: Rearing labs and distribution of predators for release. . . . . . . . . . . . . . . . . . . . . . . . . . 125
 

Carrie S. Jubb 

Chapter 12: Microsporidian Disease in Predatory Beetles . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 133
 

Leellen Solter, Wei-Fone Huang, and Brad Onken 

Chapter 13: Defining PC/QC Standards for Mass-Rearing HWA Predators. . . . . . . . . . . . . . . . . . . 139
 

Allen C. Cohen, Carole Cheah, Kathleen Kidd, and Thom Hodgson 

Chapter 14: Development of Artifi cial Diets 


 for Predators of Hemlock Woolly Adelgids. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 148
 

Allen C. Cohen and Carole Cheah 
 

 

 

 

SECTION IV ESTABLISHMENT AND MONITORING 

Chapter 15: Whole Tree Enclosures: A Tool to Assess Introduced Predators 


 of Hemlock Woolly Adelgid, Adelges tsugae. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 161
 

Jerome F. Grant, Rusty Rhea, Gregory J. Wiggins, 

Abdul Hakeem, and Paris L. Lambdin
 

Chapter 16: A Case Study of a Release of the Predator Laricobius nigrinus Fender 


 against Hemlock Woolly Adelgid, Adelges tsugae, Annand, 


 at the Urban Community Forest Interface: Hemlock Hill, 


 Lees-McRae College, Banner Elk, North Carolina . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 168
 

Richard McDonald, David Mausel, Scott Salom, and Loke T. Kok 

Chapter 17: The HWA Predator Release and Recovery Database . . . . . . . . . . . . . . . . . . . . . . . . . . 176
 

Andy Roberts, Ashley Lamb, Brad Onken, and Scott Salom 

SECTION V ADDITIONAL TOPICS 

Chapter 18: Field Insectary: Concept for Future Predator Production . . . . . . . . . . . . . . . . . . . . . . . 195
 

Scott Salom, Loke T. Kok, Tom McAvoy, and Richard McDonald 

Chapter 19: Integrating Chemical and Biological Control. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 199
 

Scott Salom, Albert Mayfi eld, and Tom McAvoy 

Chapter 20: Integrating the Early Steps of Host Selection Behavior 


 into Biological Control of HWA . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 202
 

Kimberly F. Wallin 

Chapter 21: The Introduction of Laricobius nigrinus as a Biological Control Agent 


 for the Hemlock Woolly Adelgid: Is there a Threat


 to the Native Congener, L. rubidus? . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 212
 

Nathan Havill, Gina Davis, Melissa Fischer, Scott Salom, 

Dave Mausel, and Brad Onken 

Chapter 22: An Overview and Outlook for Biological Control 


 of Hemlock Woolly Adelgid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 222
 

Brad P. Onken and Richard C. Reardon 
SECTION I INTRODUCTION
 
CHAPTER 1: HEMLOCK WOOLLY ADELGID AND ITS HEMLOCK HOSTS: 


A GLOBAL PERSPECTIVE 

Nathan Havill, Michael Montgomery, and Melody Keena 

U.S. Forest Service, Northern Research Station, Hamden, CT 

INTRODUCTION 

The hemlock woolly adelgid (HWA), Adelges 

tsugae Annand (Hempitera: Adelgidae), threatens 

the health and sustainability of the native eastern 

North American hemlocks, Tsuga canadensis (L.) 

Carrière and T. caroliniana Engelman. Th e lineage 

of HWA that was introduced to the eastern United 

States came from Japan sometime prior to 1951, 

and did not co-evolve with eastern North American 

forest ecosystems (Havill et al. 2006). As a result, 

eastern hemlock species cannot adequately resist 

or tolerate the impacts of adelgid feeding, and the 

native community of natural enemies in eastern 

North America is not capable of maintaining HWA 

populations below damaging levels. This results in 

widespread death of hemlock trees, which is having 

serious consequences for biodiversity, ecosystem 

functions, and ornamental and urban resources. 

There are quite a few studies that have examined 

the effects of HWA on hemlock ecosystems. 

Hemlocks are the most shade tolerant of conifers 

(Farjon 1990) and hemlock dominated forests 

provide a uniquely cool and densely shaded microenvironment. Loss of hemlock because of HWA is 

changing forest composition and structure (Orwig 

and Foster 1998, Spaulding and Rieske 2010), 

nutrient cycling (Kizlinski et al. 2002, Stadler et 

al. 2006; Nuckolls et al. 2009, Albani et al. 2010, 

Cobb 2010), and the composition of wildlife 

communities (Becker et al. 2008, Allen et al. 2009). 

Since the late 1980’s, there has been a concerted 

effort to understand HWA biology, evolutionary 

history, host effects, ecological impacts, and natural 

enemies, with the goal of finding ways to control 

this devastating pest. Although progress has been 

made on all of these fronts, trees continue to die at 

an alarming rate. Individual trees can be protected 

with repeated application of insecticidal soap, 

horticultural oil, or systemic insecticides (Ward et al. 

2004), and silvicultural thinning is being evaluated 

as a way to prolong the health of hemlock stands 

(Fajvan and Wood 2008), but these interventions are 

expensive and not sustainable at the landscape scale. 

Manipulating hemlock resistance to HWA is 

another approach with potential to control 

adelgid populations. Researchers are searching 

for naturally resistant trees (Ingwell and Preisser 

2011), developing resistant crosses between North 

American and Asian species (Montgomery et al. 

2009), and establishing protected plantings to 

conserve hemlock genetic diversity (Jetton et al. 

2011). Unfortunately, restoring forest and urban 

ecosystems with these trees would take many 

decades and there is no guarantee that this can 

be completed in time to safeguard hemlock’s 

unique role in eastern forests. As a consequence, 

the establishment of effective biological control 

agents is a critical component of efforts to maintain 

hemlock resources in eastern North America. 

Predicting the safety and success of biological 

control is challenging because natural enemies 

function within a complex system of multispecies, multi-trophic interactions. In this chapter, 

we summarize the evolutionary history of the 

interaction among hemlocks and adelgids. At the 

end of the chapter, we discuss how this information 

can help in the selection and establishment 

3 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

of biological control agents to maximize their 

potential to control HWA populations and 

minimize undesirable non-target eff ects. 

DIVERSITY AND DISTRIBUTION 

OF HEMLOCK 

There are nine species of hemlock currently 

accepted (Farjon 1990), five are found in 

Asia and four in North America (Fig. 1). 

Hemlock trees grow naturally in cool, humid 

areas from sea-level to the subalpine zone, 

depending on the species and region. All species 

have a strict requirement of adequate soilmoisture throughout the growing season. 

Both species of hemlock native to eastern North 

American are susceptible to HWA. Eastern 

hemlock, T. canadensis, has a broad distribution, 

spanning from New Brunswick in the north, to 

Alabama in the south, and west to Minnesota, 

with isolated disjunct populations to the south 

and west of its main range. Eastern hemlock is 

also highly valued as an ornamental, thus HWA 

is impacting property values throughout the 

eastern United States (Holmes et al. 2010). Th e 

other hemlock species in eastern North America, 

Carolina hemlock, Tsuga caroliniana, has a very 

limited distribution in the southern United States, 

and is not commonly planted as an ornamental. It 

is typically found in small, isolated populations in 

the southern Appalachians on exposed ridges and 

rocky outcroppings where it can escape competition 

from hardwoods (Jetton et al. 2008). Because of 

its restricted range, Carolina hemlock is at even 

higher risk from HWA than eastern hemlock. 

A recent molecular phylogeny of Tsuga provided 

new information about the diversity, evolutionary 

relationships, and historical biogeography of 

hemlock (Havill et al. 2008). The results suggest 

that in addition to the nine species typically 

recognized, there are two endemic island species 

of hemlock that should probably be given species 

status. Hemlock from Taiwan is often treated 

as a variety of T. chinensis (Franchet) Pritzel in 

Diels (e.g. Farjon 1990); however, phylogenetic 

analyses show that this variety is not closely related 

to hemlocks from mainland China. It appears 

that Taiwanese hemlock was correctly described 

as a separate species, T. formosana, by Hayata in 

1908. Hemlocks on Ullung Island, Korea, a small 

volcanic island in the Sea of Japan, were previously 

thought to be T. sieboldii Carrière, the southern 

Japanese hemlock. The Ullung Island hemlocks 

are actually more closely related to, but distinct 

from, T. diversifolia (Maximowicz) Masters, the 

northern Japanese hemlock. Work is underway to 

determine whether the Ullung Island hemlock is 

distinct enough to be considered a new species. 

Figure 1. Map showing the distribution of hemlock species worldwide (reprinted with permission from Havill et 

al. 2008). 

4 
Chapter 1: Hemlock Woolly Adelgid and its Hemlock Hosts: A Global Perspective 

Havill et al. (2008) also showed that the two 

eastern North American hemlocks are not close 

relatives. Tsuga canadensis diverged from all the 

other hemlocks very early in the evolutionary 

history of the genus, but T. caroliniana diverged 

more recently and is more closely related to the 

Asian species. The observation that T. caroliniana is 

closely related to the Asian hemlocks is consistent 

with its ability to successfully hybridize with the 

HWA resistant Asian species, whereas attempts to 

cross T. canadensis with other species have failed 

(Bentz et al. 2002). Hybrids between T. caroliniana 

and T. chinensis are resistant to HWA and could 

be good replacements for T. canadensis in the 

urban environment (Montgomery et al. 2009). 

Tsuga chinensis by itself is also highly resistant to 

HWA and grows well in the northeast (Del Tredici 

and Kitajima 2004; Evans 2008; Weston and 

Harper 2009). Although the two western North 

American species appear to be resistant or tolerant 

to HWA, they do not survive well in the east. 

Tsuga canadensis has low genetic variation compared 

to other hemlock species and other eastern North 

American conifers (Zabinsky 1992; Potter et al. 

2007). This could have implications for HWA 

control if this pattern translates into less natural 

variation in resistance to HWA. According to 

the pollen record, there were two periods of 

rapid decline in eastern hemlock that occurred 

approximately 9,800 and 5,300 years ago (Zhao 

et al. 2010). The more recent decline has been 

attributed to insect feeding (e.g. Bhiry and Filion 

1996), but it is more likely that both periods 

of decline were due to increased variation in 

temperature and drought that occurred during the 

early- to mid-Holocene (Foster et al. 2006, Shuman 

et al. 2009, Zhao et al. 2010). Among conifers, 

hemlocks are the most susceptible to drought 

(Farjon 1990), which may explain why the decline 

of eastern hemlock was more severe than other 

tree species experiencing the same environmental 

changes. The pattern of genetic diversity in 

T. canadensis in the southern part of its range 

suggests that when its range contracted, 

there was a refuge southeast of the Appalachians 

out of which the species eventually spread to 

re-occupy its current distribution (Potter et al. 

2007). 

Carolina hemlock was found to have moderate 

levels of genetic diversity and the genetic signature 

of a similar glacial refuge southeast of the 

Appalachians (Potter et al. 2011). Interestingly, 

Havill et al. (2008) hypothesized that T. caroliniana 

was closely related to European hemlocks based 

on an analysis that took into account hemlock 

phylogeny, molecular dating, the fossil record, 

and the timing of ancient connections among 

the continents. Hemlock eventually recovered in 

eastern North America, but this was not the case 

in Europe. Hemlock was common throughout 

Europe until approximately 750,000 years ago 

when it went extinct due to drier climate and 

repeated glaciations (LePage 2003; Follieri 2010). 

The combination of eastern hemlock’s low genetic 

diversity and relatively narrow site requirements may 

make the search for resistant trees diffi cult. With this 

in mind, Camcore (International Tree Conservation 

and Domestication, N.C. State University) is 

collecting seeds from eastern and Carolina hemlocks 

throughout their ranges, placing them in long-term 

storage, and growing them in protected plantations 

to conserve their genetic diversity for future 

restoration (Jetton et al. 2011). If biological control 

of HWA is successful, these trees could be used 

as a source to restore hemlock to eastern forests. 

DIVERSITY AND BIOGEOGRAPHY 

OF HEMLOCK ADELGIDS 

HWA has been documented on all hemlock species 

including those present on Taiwan and Ullung 

Island (Annand 1924, Takahashi 1937, Inouye 

1953, Ghosh 1975, Montgomery et al. 2000). 

The earliest reports of HWA in North America are 

from the west coast. The earliest North American 

specimens were collected in 1907 from South Bend, 

Washington (U.S. National Collection of Insects, 

Beltsville, Maryland). Other early records from the 

west include a report of damage to western hemlocks 

in Vancouver, British Columbia (Chrystal 1916), 

and specimens collected in Oregon and California 

used to formally describe A. tsugae as a new species 

(Annand 1924). In eastern North America, the 

earliest specimens were collected decades later in 

5 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Richmond, Virginia in 1951, and damage on eastern 

hemlocks was reported in Pennsylvania starting 

in 1969 (Gouger 1971). Widespread mortality 

of eastern hemlocks began in the Mid-Atlantic 

States then spread to southern New England in 

the mid 1980’s. HWA is currently established in 

more than half of the range of eastern hemlock, 

occupying 18 states from Maine to Georgia (Fig. 2) 

This sequence of records led to the incorrect 

assumption that HWA was first introduced into 

western North America in the early 20th century, 

and then brought to eastern North America from 

the west some time after that. This is understandable 

since HWA collected from different regions do not 

show any obvious morphological differences. It was 

only after a series of genetic analyses that A. tsugae 

was recognized as a diverse group of related insect 

lineages with a complex evolutionary history 

(Havill et al. 2006, Havill et al. 2007, Havill 

et al. 2009). Molecular dating methods estimated 

that the diversification of hemlock adelgids 

began approximately 30 million years ago, which 

corresponds to when much of the genus Tsuga 

was also diversifying. There are at least six distinct 

lineages of hemlock adelgids endemic to diff erent 

parts of the world: one each in China, Taiwan, and 

western North America, and two in Japan. HWA 

is also found in India and Nepal, but it is not yet 

known how these populations relate to the others. 

We now know that HWA was introduced to the 

eastern United States directly from Japan, and that 

the lineage in western North America is native. 

Figure 2. Native range of hemlock in the eastern United States (green) and range of hemlock woolly adelgid 

(brown) in 2010. 

6 
Chapter 1: Hemlock Woolly Adelgid and its Hemlock Hosts: A Global Perspective 

In Japan, there are two lineages of HWA that 

specialize on each of the Japanese hemlock species, 

T. sieboldii and T. diversifolia. Tsuga sieboldii grows 

at lower elevations and further south while T. 

diversifolia grows to the north at higher elevations. 

The geographic ranges of the two species overlap 

in central Honshu, but it is rare for them to grow 

naturally in the same stands. For example, they 

could be found on the same mountain, but T. 

sieboldii will grow at the base, and T. diversifolia will 

grow at the top. Extensive sampling throughout 

Japan has confirmed that the two Japanese HWA 

lineages are not exchanging genes despite the 

existence of areas where the two species grow in 

close proximity (Havill, unpublished data). 

The adelgids that were introduced to the eastern 

United States are from the lineage that lives on 

T. sieboldii, the southern Japanese hemlock. We 

know this because DNA sequences from adelgids 

in the eastern United States are an exact match to 

adelgids living on T. sieboldii in Japan. In the eastern 

United States, we observe only a fraction of the 

genetic variation found naturally in Japan (Havill 

et al. 2009). This is characteristic of a recently 

introduced species with a single introduction. 

In contrast, HWA in western North America is 

much more genetically diverse than in the east 

and their DNA does not match any of the Asian 

lineages (Havill et al. 2009). Hemlock adelgids 

from China and Taiwan are conspicuously diff erent 

from those in Japan and North America; perhaps 

enough to consider them separate species. 

ADELGID BIOLOGY 

Adelgids have multi-generation, complex lifecycles with many different morphological forms 

within a single species (Havill and Foottit 

2007). The typical adelgid life-cycle involves 

alternation between spruce (Picea) primary 

hosts where they form galls and where there is a 

sexual generation, and other conifer secondary 

hosts where reproduction is strictly asexual. 

The lineage of HWA that was introduced to the 

eastern United States alternates between T. sieboldii 

and tigertail spruce, Picea torano (K. Koch) Koehne, 

in Japan. The HWA gall is morphologically 

different than the typical “pineapple” adelgid gall. 

It is nearly spherical and can be quite large, up 

to 4 cm in diameter (Fig. 3). Tigertail spruce is a 

protected species in Japan where it is uncommon 

and patchily distributed on the Japanese landscape. 

Like many other adelgid species, HWA can 

maintain continuous asexual generations on 

its secondary hosts in areas where there are no 

suitable spruce primary hosts. In Japan, this has 

resulted in a patchwork of sexual and asexual 

populations of HWA, depending the proximity 

and availability of primary and secondary host 

trees. If both host species are present, a proportion 

of the HWA population migrates from hemlock 

to spruce where there is a sexual generation. 

When tigertail spruce is absent, winged migrants 

do not survive to reproduce and the population 

is limited to asexual generations on hemlock. In 

southwestern China, HWA alternates between T. 

chinensis and Picea likiangensis (Franchet) Pritzel, 

and P. brachytyla (Franchet) Pritzel (Montgomery 

and Havill, unpublished data). In western North 

America, HWA feeds on both T. heterophylla and 

T. mertensiana but winged migrants have not been 

observed and it does not alternate to spruce. 

Figure 3. Hemlock woolly adelgid gall on tigertail 

spruce in Japan. 

7 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

The ability of HWA to continue reproducing 

asexually on hemlock probably contributed to its 

successful invasion of eastern North America where 

there are no suitable spruce species to support the 

sexual generation (McClure 1989). HWA has two 

generations per year on hemlock in eastern North 

America. One generation consists of wingless sistens 

individuals that hatch in late spring to early summer, 

and quickly enter a diapause which continues 

through late summer and into the fall. Sistentes then 

overwinter as nymphs, becoming adults in early 

spring when they lay a large clutch of eggs. Th e next 

generation consists of both wingless progrediens 

and winged sexuparae. Most individuals develop as 

wingless progredientes in early spring, progress very 

quickly to the adult stage, and lay eggs in late spring 

and early summer. The remaining individuals of this 

generation develop into the winged sexuparae that 

would give rise to the sexual generation, though 

these individuals do not reproduce because suitable 

spruce species are not available. The phenology of 

the life cycle is somewhat more accelerated in the 

southern versus the northern areas of the introduced 

range because of the warmer climate (Mausel et al. 

2008), and overwintering mortality is much higher 

in the north than in the south (Trotter and Shields 

2009). The overwintering sistentes are generally 

more fecund than the progredientes (Fig. 4). 

Population genetic analyses indicated that HWA 

has very little genetic variation in the eastern 

United States, and since it only reproduces 

asexually, new genotypes can only arise from 

mutation, not from recombination. However, its 

extremely high population sizes are likely to harbor 

enough mutations to allow for adaptation to local 

environmental conditions as it spreads. For example, 

there is evidence that it is evolving increased cold 

tolerance as it moves north (Butin et al. 2005). 

Figure 4. Hemlock woolly adelgid life cycle. In Japan, the adelgid alternates between hemlock and tigertail 

spruce. Tigertail spruce supports a sexual generation and gall formation. In the eastern United States 

there are only two generations on hemlock, because winged migrants do not find suitable spruce 

species on which to complete the entire life cycle. (Vince D’Amico and Nathan Havill created the 

artwork for this fi gure.) 

N2 

N3 N3 

N4 

egg 

egg 

winter spring summer autumn

winged females 

to spruce 

winged females

to hemlock 

Spruce 

Hemlock 

adult adult wingless females

remain on hemlock 

N1 

N2 

N3 

adult N4

N2 

N3 

N4 

adult 

N1 

adult 

egg 

N2 

N3 

N1 

egg 

N2 

N3 

N1 

egg 

N1 N1

winged females

from spruce 

N2 

within gall 

8 
Chapter 1: Hemlock Woolly Adelgid and its Hemlock Hosts: A Global Perspective 

HOST IMPACTS 

HWA settles at the base of hemlock needles, 

primarily on the underside of branches. HWA has 

piercing-sucking mouth parts that extend through 

the leaf cushion into the ray parenchyma cells 

where it extracts stored plant nutrients (Young 

et al. 1995). Feeding causes the loss of hemlock 

needles, and the mortality of buds and branch tips. 

Loss of foliage and dieback can become apparent 2 

to 4 years after infestation in many locations, and 

trees can die within a few years, or can survive in a 

weakened state for many years (Paradis et al. 2008). 

Recent work suggests the damage caused by HWA 

may be more complicated than simple depletion of 

nutrients. We know that adelgids feeding on spruce 

induce dramatic changes in the host, as evidenced 

by the formation of galls. Adelgids settled at the 

base of spruce buds cause the developing needles 

to be stunted, to expand laterally, and merge 

together into a gall rather than form a normal 

shoot. Gall tissue is high in lipids and starch and 

low in phenolic compounds making them more 

suitable for adelgid feeding (reviewed in Havill and 

Foottit 2007). Secondary hosts such as hemlock 

do not respond to adelgid feeding by producing 

galls, but they may have similarly complex reactions 

to adelgid feeding. Another introduced adelgid 

species, Adelges piceae (Ratzeburg) for example, 

induces abnormal growth of bark and wood tissue 

in susceptible Abies species called “gout disease” or 

“rotholtz” (Balch 1952). HWA feeding appears to 

induce similar responses in hemlock including: 1) 

formation of abnormal xylem that limits the tree’s 

ability to transport water (Rivera et al. 2010); 2) 

increased foliar nitrogen (Stadler et al. 2005); 3) 

dramatic changes in amino acid concentration 

and composition (Gómez et al. 2011); and 4) 

the maintenance of high levels of starch at the 

feeding site (Schwartzburg and Montgomery 

2011). Thus, HWA may be able to induce localized 

changes in both its primary and secondary 

hosts that favor its survival and reproduction. 

In eastern North America, where hemlocks 

did not co-evolve with adelgids, tree response 

to feeding appears to be hypersensitive, which 

coupled with the lack of population suppression 

by natural enemies, can result in tree death. 

The terpenoid chemistry of the two eastern 

North American hemlock species is an intriguing 

example of herbivore/host co-evolution. 

Although T. canadensis and T. caroliniana are 

not phylogenetically closely related, both species 

have relatively high levels of isobornyl acetate 

(about 40%), which is twice the percentage of 

total terpenoids found in other hemlock species, 

and low levels of alpha-humulene (2-4%), which 

is less than half the percentage found in all other 

species, except T. mertensiana (Lagalante et al. 

2003). In terms of the chemical signature of all 40 

terpenoids detected, the terpenoid composition 

of T. caroliniana is more similar to the Asian 

species than the North American species. Since 

eastern North American hemlocks have evolved 

with more chewing insects such as the hemlock 

looper, Lambdina fi scellaria (Guenée), and fewer 

piercing-sucking insects, their defenses may 

not be effective against sucking insects such as 

HWA (Lagalante et al. 2007, Montgomery and 

Lagalante 2008). HWA infestation is also known 

to increase the release rate of volatile monoterpenes 

in eastern hemlock branches (Broeckerling and 

Salom 2003), but the role of terpenes in defense 

against HWA is not known, nor is their role in 

attracting natural enemies to HWA infested trees. 

IMPLICATIONS FOR 

BIOLOGICAL CONTROL 

The evolutionary history of the interaction among 

hemlocks, adelgids, and their natural enemies 

should be considered when developing a successful 

biological control program for HWA. We know 

of eleven distinct hemlock taxa that support 

six different HWA lineages. Each population 

represents a multitrophic community that is 

a potential source of HWA biological control 

agents. These communities share fundamental 

traits that can be traced to shared ancestry within 

each trophic level (i.e. within Tsuga, Adelges 

tsugae, Laricobius, Chamaemyiidae, etc.). Each 

community of predators, herbivores, and hosts has 

unique adaptations to local climatic conditions 

and to the community of specific species present. 

For example, each endemic adelgid lineage coevolved with different host species, has adapted 

9 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

different variations on the life cycle to fi t local 

ecological conditions, and contends with a diff erent 

community of natural enemies. Knowledge of these 

systems can inform and guide the development 

of biological control agents. For example, because 

HWA was introduced from southern Japan, this 

region could yield biological control agents that 

are well adapted to feed on this specifi c lineage 

of HWA. Laricobius osakensis Montgomery 

and Shiyake, a beetle recently described from 

populations of HWA in Japan is therefore a 

promising biological control of HWA (Montgomery 

et al. 2011). In western North America, there is an 

additional assemblage of adelgid-specifi c natural 

enemies (Kohler et al. 2008) on a diff erent lineage 

of HWA which, like the one in the eastern United 

States, does not alternate hosts. This lineage is also 

closely related to the one in southern Japan, and so 

predators of the western North American lineage 

of HWA may also be effective biological controls 

in the eastern United States. One such western 

predator, the beetle Laricobius nigrinus Fender, 

collected from western hemlock has been widely 

established as a biological control of HWA in the 

east (Mausel et al. 2010). Predaceous flies in the 

family Chamaemyiidae from the west also show 

potential as biological controls. In southwest China, 

there is a remarkably diverse assemblage of adelgid 

predators, especially in the lady beetle Scymnus 

(Montgomery et al. 2000). This diverse community 

of natural enemies could yield eff ective biological 

controls, especially if the native and non-native 

climates are similar, but careful attention should 

be paid to the prominent diff erences between 

the Chinese and Japanese lineage of HWA. 

Predators of specific herbivores often use 

characteristic volatile chemicals released by specifi c 

plants as cues to locate their prey. Th e interaction 

between HWA and different host species could 

influence the ability of predators to control HWA. 

To examine this, Wallin et al. (2011) tested 

whether L. nigrinus was attracted to conifer hosts 

of different adelgid species. Laricobius nigrinus 

collected from western hemlock infested with 

western HWA was more attracted to western 

hemlock volatiles than to Ponderosa pine, Douglas 

fir, white spruce, or eastern hemlock volatiles. 

They were also attracted to western white pine. 

Beetles that were lab reared on eastern hemlock 

infested with Japanese HWA did not respond 

well to volatiles from either hemlock species. Put 

in the context of what we know about HWA in 

western North America, this lends further evidence 

that L. nigrinus is adapted to locating HWA in 

the west, and raises questions about the roles of 

learning and pre-conditioning in prey location. 

Tsuga sieboldii, the host species with which the 

introduced HWA lineage co-evolved, is genetically 

and chemically different than the eastern North 

American hemlock species. Th e importance 

of a shared evolutionary history is highlighted 

when we recognize that Tsuga canadensis and T. 

caroliniana are not closely related to each other, 

yet both have independently evolved similar 

chemical signatures, perhaps because of the 

absence of pressure from sucking insects. For 

HWA control to be effective, it may be necessary 

to combine biological control with more resistant 

hemlocks. This may be diffi cult because T. 

canadensis has low genetic variation and does not 

readily hybridize with other hemlock species. 

These studies are examples of why it is useful to 

consider adelgid biology and host interactions 

in the context of its evolutionary history when 

evaluating biological control agents. Diff erent 

natural enemy species will behave diff erently in 

the context of the complex interaction between 

HWA and its hosts in different environments. An 

understanding of the diversity of hemlock species 

and HWA lineages in different parts of the world, 

and careful consideration of how natural enemies 

perform in their native and introduced ranges can 

be used to optimize the impact of biological control. 

10 
Chapter 1: Hemlock Woolly Adelgid and its Hemlock Hosts: A Global Perspective 

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14 
 

CHAPTER 2: SIMULATIONS OF POPULATION DYNAMICS
 

OF HEMLOCK WOOLLY ADELGID AND POTENTIAL IMPACT
 

OF BIOLOGICAL CONTROL AGENTS
 

Joseph S. Elkinton1, Robert T. Trotter2, and Ann F. Paradis1 

1

Department of Plant, Soil and Insect Sciences, University of Massachusetts-Amherst, MA 

2

U.S. Forest Service, Northern Research Station, Hamden CT 

ABSTRACT 

The hemlock woolly adelgid ( Adelges tsugae) is a 

small invasive Hemipteran herbivore that threatens 

the continued presence and abundance of hemlock 

in eastern North America. Efforts to control 

the adelgid have focused on the introduction of 

classical biological control agents. Th ese biological 

controls include six different species of predatory 

beetles, two of which have established. Of these 

predatory beetle species, Laricobius nigrinus has the 

most potential to alter the dynamics of hemlock 

woolly adelgid, because it has been recovered in 

large numbers at multiple release sites in eastern 

North America. However, establishment does not 

guarantee the predators will maintain the density 

of the adelgid at levels sufficiently low to prevent 

hemlock damage and death. Here we present results 

of a simulation model in which the potential impact 

of the introduced predator L. nigrinus is explored in 

the context of what is known about the dynamics of 

hemlock woolly adelgid in the absence of predators. 

INTRODUCTION 

The hemlock woolly adelgid ( Adelges tsugae Annand) 

was first documented in eastern North America 

near Richmond, Virginia in 1951 (Gouger 1971). 

Since then, the adelgid has moved through natural 

stands of hemlock and now infests at least 17 states 

from Georgia to Maine (http://na.fs.fed.us/fhp/ 

hwa/maps/distribution.shtm). The rapid growth 

and expansion of adelgid populations, and the 

resulting decline in hemlock health and abundance 

has prompted research into the biology, ecology, and 

management of this forest pest over the last 30 years. 

Research on the biology of the hemlock woolly 

adelgid in North America has shown the annual lifecycle of hemlock woolly adelgid (hereafter HWA) to 

consist of two generations, both of which reproduce 

asexually (McClure 1989) on members of the genus 

Tsuga. The overwintering sistentes generation begins 

development in the fall after a summer aestivation 

period. Development continues through the winter, 

with adults maturing and laying eggs in early spring. 

These eggs hatch after a short development period 

to produce progredientes crawlers. Th ese crawlers 

then settle on the previous year’s growth and mature 

in June. Some individuals in this generation become 

sexuparae that disperse and settle on spruce (Picea 

spp.). Sexuparae in the native range of China and 

Japan initiate a sexually reproducing generation 

on spruce, but McClure (1989) showed that none 

of the North American species of spruce trees are 

suitable hosts. The progredientes which do not 

develop into sexuparae continue to develop into 

adults, and lay eggs that hatch in June. Th e crawlers 

of the new sistentes generation settle primarily on 

new (current year) hemlock growth and enter a 

summer aestivation phase that lasts until October, 

whereupon they resume development and feeding. 

Development is completed the following March. 

15 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

In eastern North America, HWA has very few 

known natural enemies, and those that have been 

observed are not believed to have any signifi cant 

impact on HWA populations (Montgomery and 

Lyon 1996; Wallace and Hain 2000). Th ere are 

no known parasitoids that attack any species in 

the family Adelgidae. Little is known about the 

potential or actual impact of pathogens, and the 

dynamics of the invaded system are largely driven 

by the interaction of HWA and its hemlock host as 

shown by McClure (1991) who provided the fi rst 

comprehensive account of the population dynamics 

of HWA. Using artificially established populations 

on uninfested plantation trees and recently infested 

forest trees in Connecticut in 1986-1989, McClure 

showed that the HWA populations built to densities 

as high as 25 per cm of hemlock twigs in as little 

as one year after the initial infestation (Fig. 1a). 

As a result, trees produced little or no new growth 

the following year. This reduction in new growth 

forced the crawlers to colonize 1-2 year old growth 

where survival of nymphs was much lower relative 

to new growth. Additionally, the proportion 

of nymphs which became sexuparae instead of 

progredientes nymphs increased. Because sexuparae 

are unable to reproduce in North America, 

population densities declined dramatically in the 

second year (Fig. 1). The lower HWA densities 

allowed the trees to partially recover, leading in 

turn to an increase in HWA densities. Th is caused 

a second precipitous decline in hemlock growth 

that ended in the death of all the trees and HWA. 

a 

b 

2004 2005 2006 2007 2008

Year

Figure 1. 	 (a) Fluctuations in density of HWA at three naturally infested forest sites studied by McClure (1991) 

in Connecticut from 1986-1989 (figure redrawn from McClure 1991); (b) Six sites studied by Paradis 

(2011) in Massachusetts and Connecticut 2004-2008 using methods designed to be comparable with 

McClure’s original studies. In 2008 and 2009 only the April counts of adult sistentes were made. 

16 
Chapter 2: Simulations of Population Dynamics of Hemlock Woolly Adelgid 

and Potential Impact of Biological Conrol Agents 

The pattern of hemlock death four years (or even 

two years) after infestation has been described by 

many observers in the southern Appalachians, an 

area which has recently been invaded by HWA. In 

northern New England, however, hemlock trees 

have been known to host infestations of HWA 

for ten years and continue to survive (Orwig and 

Foster 1998, Orwig et al. 2002, Paradis 2011). 

Several investigators have shown that overwintering 

mortality is much higher in northern compared 

to southern states (e.g. Trotter and Shields 2009) 

due to colder winter temperatures (Paradis 2008). 

Paradis (2011) (Fig. 1b) conducted a study 

of naturally established HWA populations in 

Massachusetts and Connecticut between 2004 and 

2008 that was designed to be comparable with 

previous work by McClure (Fig. 1a). Her purpose 

was to gain insight as to why very few hemlock 

trees were dying from HWA at these northern 

sites. Like McClure, Paradis (2011) (Fig. 1b) 

recorded declines in the production of new growth 

by infested hemlocks at high HWA densities. In 

contrast to McClure’s results, however, some new 

growth was produced even on highly infested trees, 

and none of the 60 trees in her study were dead 

after six years. Paradis also investigated various 

factors that might explain the apparent stability 

of HWA densities in her study sites, and found 

that higher sistentes densities increased mortality 

and decreased fecundity in the progredientes 

generation (Fig. 2). Overwintering mortality 

varied from year to year depending primarily on 

winter temperature, and was not consistently 

density dependent (Paradis et al. 2008). 

y = -0.8647x - 0.1817

R2 = 0.7986

-3

-2.5

-2

-1.5

-1

-0.5

0

1.5 1.7 1.9 2.1 2.3 2.5 2.7

Log 10 progrediens survival

Log 10 progrediens crawler density

y = -13.898x + 22.128

R2 = 0.5445

0

5

10

15

20

25

30

35

-0.6 -0.4 -0.2 0 0.2 0.4 0.6

Progrediensfecundity

Log10 sistens density

a 

b 

c 

d 

Figure 2. 	 Density dependent a) production of sexuparae; b) mortality in the progredientes stage from McClure 

1991; c) density dependent survival of progredientes; and d) progredientes fecundity recorded by 

Paradis (2011). Figure 2b plots mortality versus density of progredientes whereas Figure 2c plots 

survival versus density. They presumably represent the same process in the two different studies. 

17 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

It is clear from the yearly variation in HWA density 

(Fig. 1) that some factor(s) limit the growth of 

HWA populations; this limitation is presumably 

caused by interaction of HWA with its hemlock 

host (McClure 1991). Both McClure (1991) and 

Paradis (2011) found density-dependent mortality 

among immature progredientes (Fig. 2). McClure 

attributed at least part of this mortality to densitydependent production of sexuparae (Fig. 2a), which 

disperse to settle on spruce. However, because 

none of the North American spruce species are 

suitable for HWA, none of these adelgids survive. 

Paradis (2011) also showed that sistentes density 

is negatively correlated with the fecundity of the 

following progredientes generation (Fig. 2d). 

Paradis (2011) found no evidence for densitydependent mortality in any other life stage, or for 

density-dependent changes in sistentes fecundity. 

In the absence of native predators and parasites, 

the natural enemies of HWA in the invaded range 

appear to be the two predatory beetle species 

Sasajiscymnus tsugae and Laricobius nigrinus 

Fender. Both have been successfully introduced in 

the eastern United States as part of the ongoing 

biological control effort against HWA. Th e longterm impact of these releases is not yet known, 

though it appears unlikely that S. tsugae has a 

measurable impact on HWA population densities. 

Although the beetle was released in the mid 1990s 

(Cheah et al. 2004) at three of the sites that 

Paradis (2011) studied, intensive sampling for the 

beetles by Paradis (2011) did not recover any S. 

tsugae at these sites. Although S. tsugae has been 

recovered at sites many years after release, in the 

northeastern states it is rarely if ever abundant 

enough to signifi cantly affect HWA densities.

 In contrast, the predatory beetle Laricobius nigrinus 

(Derodontidae) was imported from the Pacifi c 

Northwest and has been recovered in large numbers 

at many release sites throughout the eastern United 

States (Mausel 2007, Mausel et al. 2008). It is by 

far the most promising biological control agent 

released against HWA so far, yet we still do not 

know how much of an impact it is having on 

HWA densities or dynamics. For this reason we 

concentrate our simulation efforts on this species. 

SIMULATION 

Data collected by Paradis (2011) (Fig. 1b), was used 

to parameterize simulations of HWA population 

densities. We began by constructing a simulation 

of the system in the absence of any predators based 

on generalized (averaged) values of sistentes and 

progridientes survival and fecundity. Th e data 

consist of information collected from eight locations, 

with 3-5 years of data for each location (Table 1). 

These data provided the basic values needed to 

generate an empirically-based simulation of HWA 

dynamics. We constructed our model using two 

parallel approaches. In the first, a fl ow model 

was built to describe the population dynamics 

of the adelgid over the course of multiple lifestages and generations using Stella (V9.1 IE 

systems).For verification, a similar model was 

constructed in MS Excel®. In Stella we built a 

flow model using the following basic structure: 

SA = (PA )*SS (t+1) (t)*PF(t) (t+1) 

PA = (SA )*PS (t+2) (t+1)*SF(t+1) (t+2) 

in which SA = Density of Sistentes Adults, 

PA = Density of Progredientes Adults, 

Table 1. Average values of density, mortality and fecundity of HWA life stages recorded by Paradis 

(2011) (Fig. 1b). 

Density: HWA per cm 

OW Progred Immature 

sistens adults sistentes 

Mortality: proportion dying 

OW Sistens 

sistens Progredientes crawlers 

Prewinter 

sistentes 

Fecundity

Sistentes Progredientes 

Mean 

Std. dev 

2.21 

0.90 

1.33 

0.43 

10.60 

5.41 

0.52 

0.27 

0.99 

0.01 

0.64 

0.13 

0.75 

0.15 

146.97 

29.77 

22.54 

4.85 

18 
Chapter 2: Simulations of Population Dynamics of Hemlock Woolly Adelgid 

and Potential Impact of Biological Conrol Agents 

SF = Sistentes Fecundity, PF = Progredientes 

Fecundity, SS = Sistentes Survival Rate, and 

PS = Progredientes Survival Rate. Based on this set 

of relationships, and using the above parameters, our 

simulated HWA populations grow exponentially. 

This behavior was expected, since any population 

with constant rates of mortality and fecundity 

will either grow or decline exponentially, and no 

doubt this behavior describes the growth of HWA 

populations when they first colonize a tree or stand. 

Such populations soon reach the carrying capacity 

represented by the available hemlock foliage and 

the densities are subsequently governed by the 

density-dependent processes described above. We 

thus incorporated density-dependent progredientes 

survival (Fig. 2c) and fecundity (Fig. 2d) into our 

simulation. The result is a population system that 

stabilizes at around two adult sistentes per cm 

of hemlock twig, but that exhibits the expected 

variation in density of different life stages evident 

in Figs. 1a and 1b. If we add a modest amount 

of annual variation in overwintering mortality, 

drawn at random from a normal distribution 

with a mean and standard deviation equal to that 

recorded by Paradis (2011) (Table 1), we generate 

fluctuations in simulated densities (Fig. 3a) similar 

to those observed by McClure (1991) (Fig. 2a) 

and Paradis (2011) (Fig. 1b). These results give 

us confidence that our simulation has captured 

the essence of HWA dynamics in the absence of 

predators, at least in the northeastern US where 

McClure and Paradis conducted their studies. 

Year

HWA per cm

a 

b 

Figure 3. 	 Our simulated time series of HWA density incorporating mean values of fecundity and survival of HWA 

life stages recorded by Paradis (2011) (Table 1) and density-dependence in progredientes survival 

(Fig. 2c) and fecundity (Fig. 2d). In Figure 3a we plot HWA densities at 3 times per year as in Figure 1 

and we add variation in overwintering mortality recorded by Paradis (2011). In Figure 3b, we plot only 

one life stage per year (the overwintering adult sistentes) and we remove the variation in overwintering 

mortality. Otherwise they are the same simulation. 

19 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Although the population shown in Figure 3a 

demonstrates the fundamental stability evident 

in this system caused by the density dependence 

in mortality and fecundity discussed above, the 

stability is partially obscured by the inclusion of 

variable HWA overwintering mortality. Th is is 

highlighted if we remove this variation and plot 

only one life stage per year (overwintering sistentes, 

Fig. 3b) rather than the three life stages plotted in 

Figures 1 and 3a. As Figure 3b shows, the system 

stabilizes at an equilibrium value of approximately 

2.1 sistentes per cm. Although instructive with 

regards to evaluating the fundamental parameters 

responsible for long-term population dynamics, 

it is important to remember that this simulation 

represents an idealized system. In natural 

settings, annual fluctuations still occur, and 

sometimes result in tree death (McClure 1991). 

With these results in hand, we simulated the 

impact of predation by L. nigrinus on the eggs 

produced in early spring by the overwintering 

sistentes generation. This was accomplished by 

reducing the survival of HWA progredientes 

eggs (eggs laid by sistentes). We were surprised 

to learn that the removal of even 80 or 90 

percent of these eggs had negligible eff ects on 

the subsequent density of the next generation 

in our simulation (Fig. 4a). This result occurred 

because the density-dependent mortality in the 

progredientes crawler stage (Fig. 2b,c) decreased as 

egg predation increased, offsetting the population 

decreases that would otherwise have occurred. 

On further reflection, we realized that this result 

depends on the actual cause and timing of the 

mortality evident in this stage. If most or all of it is 

due to density dependent production of sexuparae 

(Fig. 2a) as McClure (1991) suggests, then the 

density dependence presumably occurs when the 

eggs are laid, rather than after the progredientes 

hatch. If that is true, then egg predation occurs 

after sexuparae production. If we incorporate 

this into our simulation, then egg predation has 

a profound effect on the equilibrium density 

(Fig. 4b), because it is now preceded instead of 

followed by the compensating density-dependent 

Mature sistens per cm

a 

b 

Figure 4. Simulated effect of predation on HWA eggs 

produced by overwintering sistentes a) after 

or b) before density dependent mortality 

occurring in the progrediens generation as 

in Figure 2a,b,c. 

mortality. Alternatively, it seems likely that the 

response of the hemlock host is involved in some 

way that is not well understood in causing the 

density dependent mortality evident in Figure 2, 

as McClure (1991) suggested. Recent work by 

Radville et al. (2011) shows that HWA attack 

induces a systemic hypersensitive response in 

hemlocks. If the host response causes the mortality 

documented in Figure 2 and it occurs in response 

to HWA density during the overwintering sistentes 

stage, then the mortality may occur whether or 

not the egg densities are reduced by predation. 

The resulting effect would be like Figure 4b. If, on 

the other hand, the host response occurs when the 

immature progredientes settle, then the eff ect of 

egg predation would be compensated as in Figure 

4a. The experiments by Radville et al. involved 

inoculating hemlock foliage with progredientes 

crawlers and their results thus suggest the latter 

20 
 

Chapter 2: Simulations of Population Dynamics of Hemlock Woolly Adelgid 

and Potential Impact of Biological Conrol Agents 

conclusion (Fig. 4a). These considerations make 

it clear that understanding the exact cause and 

timing of the mortality evident in Figure 2 should 

be a high research priority. We have initiated 

experiments designed to accomplish this need. 

Thus far we have only considered the eff ects of 

constant rates of egg predation on HWA densities. 

In nature, egg predation rates presumably vary with 

density of L. nigrinus and we know from Mausel 

et al. (2008) that density of L. nigrinus increases 

with the density of HWA (Fig. 5). Furthermore, 

we know that adult L. nigrinus feed on late-instar 

sistentes in autumn. Incorporating these eff ects into 

our simulation also has profound effects on the 

equilibrium density, including the version of the 

simulation where egg predation precedes (Fig. 4a) 

rather than follows (Fig. 4b) progredientes mortality. 

In this version of our simulation, we make 

assumptions about the number of eggs and late 

instar sistentes consumed per beetle and allow 

beetle density to vary according to Figure 5. 

In Figure 6, we show the effects of varying the 

number of late-instar sistentes eaten per beetle. Th e 

resulting dynamics are complex. As the numbers of 

sistentes eaten per beetle increases, the mean HWA 

density declines, but the amplitude of the density 

fluctuations increases. As the number of HWA eaten 

per beetle increases, the HWA density progresses 

from equilibrium (10 sistentes per beetle), to 

damped oscillations, to evident two- and four-year 

cycles, and eventually extinction, which occurs when 

the fluctuating densities overlap zero. Th is behavior 

is reminiscent of the progression from equilibrium, 

to damped oscillations, to cycles with periods of 2n, 

and finally to chaos first explored by May (1974) 

with the discrete logistic model. This behavior was 

demonstrated subsequently in many other models. 

Whether our system is chaotic under particular 

parameter values is difficult to answer and of little 

practical importance. We present these results, not 

because we believe it to be an accurate description 

of the impact of L. nigrinus on HWA dynamics, 

but because it illustrates the complex dynamics 

that even simple models such as ours can exhibit. 

No. HWA ovisacs/cm shoot

No. immature predators/cmshoot

Figure 5. Increases in density of L. nigrinus as a function of HWA density recorded by Mausel et al. (2008). 

21 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Figure 6. Variation in simulated HWA density as a function of numbers of late-instar HWA consumed by each 

individual adult L. nigrinus in the autumn. 

0

0.5

1

1.5

2

2.5

3

3.5

4

0 adults eaten/beetle

10 adults eaten/beelte

15 adults eaten/beetle

0

0.5

1

1.5

2

2.5

0 2 4 6 8 10 12 14 16 18

20 adults eaten/beetle

23 adults eaten/beetle

Overwintering sistens

25 adults eaten /beetle

per cm

Year

a 

b 

22 
Chapter 2: Simulations of Population Dynamics of Hemlock Woolly Adelgid 

and Potential Impact of Biological Conrol Agents 

DISCUSSION AND CONCLUSION 

We are tempted to add more and more details to 

our model, incorporating various other factors 

that we already know affect HWA population 

dynamics. We might include host tree eff ects and 

factors influencing survival and fecundity of other 

life stages. We might also include the impact of 

elongate hemlock scale, Fiorinia externa, another 

invasive species co-infesting hemlock in the eastern 

US that Preisser and Elkinton (2008) have shown 

has a significant impact on HWA. We are mindful 

however, of the rather sorry history of complex 

simulations in forest entomology. The Gypsy Moth 

Life System Model (Sharov and Colbert 1994) for 

example, was constructed in the 1980s and 1990s at 

great expense and incorporated much of what was 

known about gypsy moth population dynamics. It 

was one of several such models, all of which were 

soon abandoned, because they were too complex 

to understand and they yielded little insight into 

the dynamics of the system (Liebhold 1994, Sharov 

1996). Rather, models that incorporate minimal 

to intermediate complexity (Liebhold 1994, Logan 

1994, Sharov 1996) have been shown to yield the 

most insight. The art of building such models is to 

know what to put in and what to leave out. It is 

our belief that any useful model of HWA dynamics 

must include the strong density dependent mortality 

in the absence of predators (Fig. 2) documented by 

McClure (1991) and Paradis (2011). Th ese eff ects 

involve the interaction of HWA with its hemlock 

host, as McClure (1991) described, but they 

continue to govern the system even in the absence 

of tree mortality, as Paradis (2011) has shown. 

Our simulation suggests that the impact of 

biological control agents on HWA can be counterintuitive and can only be understood in the context 

of the rest of the population dynamics of the system. 

For example, our simulations revealed that very 

high rates of predation can have almost no eff ect on 

mean HWA densities, if they are followed by other 

compensatory density-dependent mortalities. Th ese 

findings underscore the importance of introducing 

biocontrol agents that prey on multiple life stages, 

and the importance of the timing of predation. 

Finally, we believe our simulation shows the 

importance of understanding the various factors that 

affect mortality and fecundity of different life stages 

of a pest population system undergoing biological 

control and can suggest critical experiments that 

need to be done to understand these eff ects. As 

additional information on the biology of the 

controls becomes available, our ability to estimate 

their potential impact should continue to improve. 

LITERATURE CITED 

Cheah, C.A.S.-J.; Montgomery, M.E.; Salom, S.M.; 

Parker, B.L.; Costa, S.; Skinner, M. 2004. 

Biological Control of Hemlock Woolly Adelgid. 

Reardon, R.; Onken, B., tech. coords. FHTET2004-04. Morgantown, WV: U.S. Department 

of Agriculture, Forest Service: 22 p. 

Gouger, R.J. 1971. Control of Adelges tsugae on 

hemlock in Pennsylvania. Scientific Tree Topics 

3: 9. 

Liebhold, A.M. 1994. Use and abuse of insect and 

disease models in forest pest management: 

past, present, and future. In: Covington, 

W.W.; DeBano, L.F., eds. Sustainable Ecological 

Systems: Implementing an Ecological Approach to 

Land Management. Tech. Rep. RM-247. U.S. 

Department of Agriculture, Forest Service: 

204-210. 

Logan, J.A. 1994. In defense of big ugly models. 

American Entomologist 40: 202-207. 

Mausel, D.L. 2007. Release and Monitoring of 

Laricobius nigrinus (Coleoptera: Derodontidae) for 

Biological Control of the Hemlock Woolly Adelgid 

in the Eastern US. Virginia Polytechnic Institute 

and State University, Blacksburg, VA: 156 p. 

Ph.D. dissertation. 

Mausel, D.L.; Salom, S.M.; Kok, L.T.; Fidgen, 

F.G. 2008. Propagation, synchrony, and impact 

of introduced and native Laricobius spp. 

(Coleoptera: Derodontidae) on hemlock woolly 

adelgid in Virginia. Environmental Entomology 

37: 1498-1507. 

23 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

McClure, M.S. 1989. Evidence of a polymorphic 

life-cycle in the hemlock woolly adelgid, Adelgestsugae (Homoptera: Adelgidae). Annals of the 

Entomological Society of America 82: 50-54. 

McClure, M.S. 1991. Density-dependent feedback 

and population-cycles in Adelges-tsugae 

(Homoptera: Adelgidae) on Tsuga-canadensis. 

Environmental Entomology 20: 258-264. 

May, R.M. 1974. Biological populations with 

nonoverlapping generations: stable points, stable 

cycles, and chaos. Science 186: 645-647. 

Montgomery, M.E.; Lyon, S.M. 1996. Natural 

enemies of adelgids in North America: their 

prospect for biological control of Adelges 

tsugae (Homoptera: Adelgidae). In: Salom, S.; 

Tigner, T.; Reardon, R., eds. Proceedings: Th e 

First Hemlock Woolly Adelgid Review; 1995 

October 12; Charlottesville, VA. FHTET96-10. Morgantown, WV: U.S. Department 

of Agriculture, Forest Service, Forest Health 

Technology Enterprise Team: 89-102. 

Orwig, D.A.; Foster, D.R. 1998. Forest response 

to the introduced hemlock woolly adelgid in 

southern New England, USA. Journal of the 

Torrey Botanical Society 125: 60-73. 

Orwig, D.A.; Foster, D.R.; Mausel, D.L. 2002. 

Landscape patterns of hemlock decline in New 

England due to the introduced hemlock woolly 

adelgid. Journal of Biogeography 29: 1475-1487. 

Paradis, A.; Elkinton, J.; Hayhoe, K.; Buonaccorsi, 

J. 2008. Role of winter temperature and 

climate change on the survival and future 

range expansion of the hemlock woolly adelgid 

(Adelges tsugae) in eastern North America. 

Mitigation and Adaptation Strategies for Global 

Change 13: 541-554. 

Paradis, A. 2011. Population Dynamics of Hemlock 

Woolly Adelgid ( Adelges tsugae) in New England. 

University of Massachusetts, Amherst. 

114 pages. Ph.D. dissertation. 

Preisser, E.L.; Elkinton, J.S. 2008. Exploitative 

competition between invasive herbivores 

benefits a native host plant. Ecology 89: 

2671-2677. 

Radville, L.; Chaves, A.; Preisser, E.L. 2011. 

Variation in plant defense against invasive 

herbivores: evidence for a hypersensitive 

response in eastern hemlocks. Journal of 

Chemical Ecology 37: 592-597. 

Sharov, A.A. 1996. Modeling insect dynamics. In: 

Korpilahti, E.; Mukkela, H.; Salonen, T., eds. 

Caring for the Forest: Research in a Changing 

World. Congress Report, Vol. II., IUFRO XX 

World Congress, 6-12 August 1995, Tampere, 

Finland. Jyvaskyla (Finland): Gummerus 

Printing: 293-303. 

Sharov, A.A.; Colbert, J.J. 1994. Gypsy Moth Life 

System Model. Integration of Knowledge and a 

User’s Guide. Virginia Polytechnic Institute and 

State University, Blacksburg, VA: 232 p. 

Trotter, R.T.; Shields, K.S. 2009. Variation in winter 

survival of the invasive hemlock woolly adelgid 

(Hemiptera: Adelgidae) across the eastern 

United States. Environmental Entomology 38: 

577-587. 

Wallace, M.S.; Hain, F.P. 2000. Field surveys and 

evaluation of native and established predators 

of the hemlock woolly adelgid (Homoptera: 

Adelgidae) in the Southeastern United States. 

Environmental Entomology 29(3): 638-644. 

24 
 

 

CHAPTER 3: UNDERSTANDING FEDERAL REGULATIONS
 

AS GUIDELINES FOR CLASSICAL BIOLOGICAL CONTROL PROGRAMS
 

Michael E. Montgomery 

U.S. Forest Service, Northern Research Station, Hamden, CT (retired)1 

INTRODUCTION 

This chapter reviews the legislation and rules that 

provide the foundation for federal regulation of 

the introduction of natural enemies of insects as 

biological control agents. It also outlines the steps 

for complying with regulatory requirements, using 

biological control of Adelges tsugae Annand, the 

hemlock woolly adelgid (HWA), as an example. 

The program to establish biological control agents 

for HWA in eastern North America dates from 

1993 to present and involves importation from 

other continents, from other countries in the 

North American continent, as well as the interstate 

movement of biological control agents. Th us, 

biological control of HWA provides examples of 

regulation under old and new federal regulations 

and rules for foreign importation and interstate 

movement. With these regulations in mind, the 

several steps involved in putting a biological control 

program into practice will be reviewed—fi nding and 

importing agents, rearing and studying the agents in 

a containment facility (aka quarantine), acquiring 

data to show that the agent will be effective and safe, 

and release into a new environment. Monitoring 

the biological control agent’s establishment 

and efficacy is also part of a biological control 

program, but this is not discussed here. 

1

Disclaimer: Views and statements are those of the author 

and should not be interpreted as official policy of any 

federal agency. Anyone considering releases of biological 

control organisms must follow offi cial regulations and 

should not rely only on the information in this chapter. 

In its broadest meaning, biological control is 

the action of an organism that maintains the 

population of another organism at a lower average 

density than would occur if it was absent. From 

an applied viewpoint, biological control is the 

use of natural enemies (predators, parasites, 

pathogens) to reduce a pest population and thus 

the damage it causes. Classical biological control 

is the introduction and permanent establishment 

of natural enemies in order to reduce populations 

of a non-indigenous pest. It is the movement 

(importation from a foreign country, release from 

containment, and interstate transport) of biological 

control organisms that is federally regulated. 

The federal government recognizes that biological 

control is often a desirable, low-risk means to 

reduce pests of crops and other plants; however, 

the unregulated movement of certain biological 

control organisms may present an unacceptable 

risk. Thus, the government has a dual role to 

facilitate biological control and also to assess the 

risks and benefits of releasing specific organisms for 

biological control of pests. Regulations regarding 

the movement of entomophagous (insect-eating) 

biological control organisms have not been 

promulgated, although federal legislation was passed 

in 2000 authorizing regulation of all biological 

control organisms. Because regulatory policies now 

in place are not transparent and widely available, 

many practitioners may not be adequately informed. 

This review of the underlying legal framework and 

the steps in importing and releasing biological 

control organisms will hopefully fill some of 

the information gaps practitioners may have. 

25 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

DEFINITIONS 

The terminology associated with biological control 

varies with disciplines and can have uncertain and 

conflicting meanings. Even familiar terms when 

used by government agencies can have meanings 

that are narrower or broader than when used 

scientifi cally. Therefore, the following defi nitions 

are provided for terms used in this chapter. 

Act means a public law passed by the United States 

Congress. These acts are listed sequentially 

by Congressional session (e.g., Public Law 

106-224 is the Agricultural Risk Protection 

Act of 2000). These are organized by topic 

in the United States Code (USC). 

Biological Control has various meanings, but the 

definition of DeBach (1964) is appropriate for 

this chapter—“the action of parasites, predators, 

or pathogens (disease-causing organisms) in 

maintaining another organism’s population 

density at a lower average than would occur 

in its absence.” DeBach recognized three 

approaches to achieve biological control: 

classical biological control is the purposeful 

introduction and permanent establishment 

of natural enemies to suppress populations of 

a pest; augmentation is the supplemental or 

inundative release of natural enemies in areas 

where they are missing, too scarce to provide 

adequate control, or arrive too late in the season 

to be eff ective; and conservation of natural 

enemies is management to enhance the survival 

and impact of established natural enemies. 

Biological Control Organism, as defi ned by 

public law, means “any enemy, antagonist, 

or competitor used to control a plant 

pest or noxious weed” (7 USC 7702). 

Indirect Damage is when an organism adversely 

affects another organism that is benefi cial 

to plants, and those adverse eff ects cause 

losses in yields of crops or forage plants 

or a reduction in the viability or vigor of 

ornamental or native plants (cf. defi nition 

proposed but not adopted (USDA 2001)). 

Movement, Move, and Moving includes release 

into the environment as well as the transport 

or facilitating the transport, by any means, 

into the country or between states. 

Nonindigenous refers to a plant or animal 

that is not native to a place. Introduced, 

adventive, alien, exotic, nonnative, and 

nonindigenous (non-indigenous) species are 

used rather interchangeably to indicate a 

species living outside its native distributional 

range and that arrived there by human 

activity, either accidental or deliberate. 

Regulations are general and permanent rules 

developed by executive branch agencies to 

administer and enforce the Acts passed by 

the U.S. Congress. These rules (also called 

administrative laws) are published in the 

Federal Register and codified under 50 

titles in the Code of Federal Regulations 

(CFR), which is updated annually. 

LEGAL FRAMEWORK 

Legislation 

In the United States, there are many acts that may 

pertain to biological control organisms, but three 

are especially important. The most signifi cant is 

the Plant Protection Act (PPA) passed by Congress 

on June 20, 2000 (U.S. Congress 2000). Th is Act 

consolidated all or part of ten existing plant health 

laws into one law that gives the Secretary of the 

United States Department of Agriculture (USDA) 

broad authority to regulate movement of any 

plant, plant product, biological control organism, 

noxious weed, and plant pest. This authority has 

been delegated to the Department’s Animal and 

Plant Health Inspection Service (APHIS) and 

the regulation of biological control organisms has 

been assigned to APHIS’s Plant Protection and 

Quarantine (PPQ) program. Most of the provisions 

of the Federal Plant Pest Act (FPPA) of 1957, which 

regulated the importation and interstate movement 

of plant pests, were retained with the new act 

providing additional authority for regulation of 

noxious weeds and biological control organisms. 

26 
 

Chapter 3: Understanding Federal Regulations as Guidelines for Classical Biological Control Programs 

The PPA recognizes the need to prevent the 

dissemination of plant pests and to facilitate the 

use of biological control to protect plants from 

their pests, including noxious weeds. Excerpts from 

the Act (Table 1) illustrate how regulation of the 

movement of plant pests (Section 411) is simple 

and direct—movement of plant pests is prohibited 

without a permit; whereas Section 412, regulating 

the movement of biological control organisms, is 

much more complex. For example, Section 412 

(a) states that any biological control organism 

would be regulated only if it is determined that 

this is necessary to prevent the introduction or 

dissemination of a plant pest; and Section 412 (l) 

indicates that certain biological control organisms 

may be listed as exempt from restrictions on 

movement in interstate commerce. Th e following 

section on regulations illustrates the diffi culty of 

developing administrative rules to both promote and 

ensure the safety of biological control organisms. 

The second important piece of legislation aff ecting 

biological control programs in the United States is 

the National Environmental Policy Act (NEPA). 

This law, effective January 1, 1970, applies to 

all federal activities, including projects receiving 

federal funds and permits issued by federal agencies. 

Non-federal applicants are not responsible for 

compliance; rather, the federal agency that proposes 

the action, provides the funds, manages the land 

where the activity will occur, or issues the permit 

is responsible for compliance. To fulfi ll NEPA 

provisions, the agency first determines which of 

three levels of analysis is required. Th e simplest 

level of analysis is for an activity that has been 

predetermined to be Categorically Excluded (CE). 

These are specific activities listed by the agency 

that it has determined have limited environmental 

effects. Examples of CE activities listed by APHIS 

include (a) interstate movement of nonindigenous 

species between containment facilities; (b) 

importation of nonindigenous species into 

containment facilities; and (c) releases into a State’s 

environment of pure cultures of organisms that are 

either native or are established introductions (see 7 

CFR Ch. III 372.5). If the activity is characterized 

Table 1.—Excerpts from the Plant Protection Act. 

Section 411. REGULATION OF MOVEMENT OF PLANT 

PESTS. (7 USC 7711) 

(a) PROHIBITION OF UNAUTHORIZED MOVEMENT 

OF PLANT PESTS.—No person shall import, enter, export, 

or move in interstate commerce any plant pest, unless the 

importation, entry, exportation, or movement is authorized 

under general or specifi c permit. 

Section 412. REGULATION OF MOVEMENT OF 

PLANTS, PLANT PRODUCTS, BIOLOGICAL CONTROL 

ORGANISMS, NOXIOUS WEEDS, ARTICLES, AND 

MEANS OF CONVEYANCE. (7 USC 7712) 

(a) IN GENERAL—The Secretary may prohibit or restrict 

the importation, entry, exportation, or movement in interstate 

commerce of any plant, plant product, biological control 

organism, noxious weed, article, or means of conveyance, if 

the Secretary determines that the prohibition or restriction is 

necessary to prevent the introduction into the United States or 

the dissemination of a plant pest or noxious weed within the 

United States. 

(b) POLICY.—The Secretary shall ensure that processes 

used in developing regulations under this section governing 

consideration of import requests are based on sound science 

and are transparent and accessible. 

(c) REGULATIONS.—The Secretary may issue regulations to 

implement subsection (a), including regulations requiring that 

any plant, plant product, biological control organism, noxious 

weed, article, or means of conveyance imported, entered, to be 

exported, or moved in interstate commerce. 

(e) STUDY AND REPORT ON SYSTEMS APPROACH 

[for plant pathogens] 

(e) NOXIOUS WEEDS.— 

(g) BIOLOGICAL CONTROL ORGANISMS.— 

(1) REGULATIONS.—In the case of biological control 

organisms, the Secretary may publish, by regulation, a list 

of organisms whose movement in interstate commerce 

is not prohibited or restricted. Any listing may take into 

account distinctions between organisms such as indigenous, 

nonindigenous, newly introduced, or commercially raised. 

(2) PETITION TO ADD OR REMOVE BIOLOGICAL 

CONTROL ORGANISMS FROM THE REGULATIONS.— 

Any person may petition the Secretary to add a biological 

control organism to, or remove a biological control organism 

from, the regulations issued by the Secretary under this 

subsection. 

(3) DUTIES OF THE SECRETARY.—In the case of a petition 

submitted under paragraph (2), the Secretary shall act on the 

petition within a reasonable time and notify the petitioner of the 

final action the Secretary takes on the petition. The Secretary’s 

determination on the petition shall be based on sound science. 

27 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

as being limited in scope to specific sites, specifi c 

species, or activities that potentially would impact 

few environmental values or systems, then the 

agency would prepare an Environmental Assessment 

(EA). The purpose of the EA is to determine the 

significance of the environmental effects and to 

examine alternative means to achieve the objective. 

The EA includes a brief discussion of: (1) the need 

for the proposal, (2) alternative courses of action, 

(3) the environmental impacts of the proposed 

action and alternatives, and (4) a listing of agencies, 

institutions, and persons consulted. Th e applicant, 

the public, and other agencies may be involved in 

preparing or commenting on the draft EA. Th e 

process concludes with either a Finding of No 

Significant Impact (FONSI), or the application 

is denied. The third NEPA category, the 

Environmental Impact Statement (EIS), is 

used for changes in policy, such as changes 

in agency regulations, and for activities 

that do not qualify for a FONSI. 

The third important piece of federal legislation 

is The Endangered Species Act of 1973. Th is act 

requires federal agencies, in consultation with the 

U.S. Fish and Wildlife Service (F&WS) or the 

NOAA Fisheries Service, to ensure that any action 

that a federal agency authorizes, funds, or carries out 

is unlikely to jeopardize the continued existence of a 

listed species, or result in the destruction or adverse 

modification of designated critical habitat of such 

species. Proposals to release a nonindigenous species 

into a new environment would be reviewed by the 

F&WS for potential impacts on endangered species. 

The federal agency, such as APHIS, conducting 

an EA usually asks the F&WS for its opinion 

regarding threats to federally-listed endangered or 

threatened species. The F&WS is also responsible 

for monitoring the movement of wildlife, which 

includes insects (50 CFR 14—Importation, 

Exportation and Transportation of Wildlife). Th e 

F&WS is particularly concerned about species 

that are listed according to the Convention on 

International Trade in Endangered Species of Wild 

Fauna and Flora (CITES). Import permits from 

F&WS are required for species that are on the 

CITES list. Currently there are no entomophagous 

insects on the list; however, shipments of all 

biological control organisms should be accompanied 

by a Declaration for Importation or Exportation 

of Fish or Wildlife (USFWS Form 3-177). 

Regulations 

Implementation of the Acts of Congress is done 

usually by agencies in the Executive Branch by a 

process of “rule-making.” The proposed regulations 

are published in the Federal Register for public 

comment and as final regulations. These are then 

compiled in the Code of Federal Regulations 

(CFR), which is available on the internet. 

Regulations for movement of biological control 

organisms are provided in 7 CFR Part 330–Federal 

Plant Pest Regulations; General; Plant Pests; Soil, 

Stone, and Quarry Products; Garbage. In response 

to passage of the PPA of 2000, Section 330.102 was 

revised in April, 2001 to include biological control 

organisms among the items that “the Secretary of 

Agriculture may prohibit or restrict the importation, 

entry, exportation, or movement in interstate 

commerce, . . . if the Secretary determines that the 

prohibition or restriction is necessary to prevent 

the introduction into or the dissemination within 

the United States of a plant pest or noxious weed.” 

Although the CFR was updated to refl ect USDA’s 

current authority to regulate the importation and 

movement of entomophagous biological control 

organisms, new rules to exercise this authority 

have not been incorporated into the Regulations. 

Extensive revisions were proposed in the Federal 

Register of October 9, 2011, but public concerns 

about the proposed rules and reorganization of 

security following events of September 11, 2001 

sidetracked their adoption. Nonetheless, these 

proposed rules (USDA 2001) provide insight into 

the complexity of developing detailed rules for 

the regulations for the many aspects of biological 

control of arthropods and noxious weeds. 

28 
Chapter 3: Understanding Federal Regulations as Guidelines for Classical Biological Control Programs 

In 2009, APHIS-PPQ asked for public comment 

to help it determine which alternative it should 

examine in preparing an Environmental Impact 

Statement for Movement of Plant Pests, Biological 

Control Organisms, and Associated Articles 

(USDA 2009). The alternatives proposed were: 

(1) Take no action—leave current rules unchanged 

(2) Revise requirements for movement of plant pests 

to cover biological control organisms consistent 

with the scope of the PPA (preferred alternative) 

(3) Implement a comprehensive risk reduction 

program (more expansive regulations 

to address specifi c risk categories) 

The statement “Establishment of clear, coherent, 

and streamlined regulations at the national level 

will be important to ensuring objective assessment 

of the risks and benefits of biological control in 

the U.S.” (Mason et al. 2005) still applies today. 

Although new regulations of biological control 

organisms have not yet been issued, APHIS has 

new policies and procedures for regulation of 

entomophagous biological control organisms that 

are largely unpublished or available on web pages; 

thus, it is important to check with the agency 

regarding compliance if you wish to import, 

release, or move any biological control organism. 

HISTORICAL REGULATION OF 

BIOLOGICAL CONTROLS FOR HWA 

Regulation of the importation of entomophagous 

biological control agents from foreign countries to 

approved containment facilities has changed little in 

the last 50 years. What has changed is regulation of 

the first-time release of an entomophagous biological 

control agents into the environment. Th e hemlock 

woolly adelgid (HWA) provides a good example of 

these changes (Table 2), which fall into three groups: 

Table 2.—Imported arthropods released from containment for biological control of the 

hemlock woolly adelgid and procedures used to assess potential risk of release. 

Species, author, year described Origin 

Released from 

containment 

Evaluation process 

used by APHIS 

NEPA 

compliance5 

Diapterobates humeralis (Hermann) 1804 Japan1 1992 OPRA3

, 

Limited Review ? 

Sasajiscymnus tsugae (Sasaji & McClure) 1997 Japan 1995 OPRA, 

Limited Review 

APHIS, FS, 

NPS 

Scymnus sinuanodulus Yu & Yao 1997 China 1998 First-tier Risks 

(Not Regulated) FS, NPS 

Laricobius nigrinus Fender 1945 Canada2 2000 First-tier Risks 

(Not Regulated) FS, NPS 

Scymnus ningshanensis Yu & Yao 2000 China 2000 First-tier Risks 

(Not Regulated) FS 

Scymnus camptodromus Yu & Liu 1997 China 2000 First-tier Risks 

(Not Regulated) —6 

Laricobius osakensis Montgomery & Shiyake 2011 Japan 2010 

NAPPO4, 

Independent Review, 

Public Comment 

APHIS 

1

Widespread, including North America 

2

Endemic to western North America; permits not issued for subsequent movement from western States to eastern States 

3

OPRA=Organism Permitting and Risk Analysis conducted by the Biological Assessment and Support Team (BATS) of APHIS-PPQ 

4

Standards to release entomological biological control agents adopted by the North American Plant Protection Organization 

5

Compliance with the National Environmental Policy Act by Federal Agencies, Animal & Plant Health Inspection Service, Forest Service 

or National Park Service 

6

Has not been released into the environment 

29 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

(1) Following the passage of NEPA in 1970, 

APHIS prepared an EA for the fi rst-time 

release of all biological control organisms into 

the environment. The process for obtaining 

a release permit was not much diff erent from 

the process currently used, except that it was 

less rigorous in the information required 

and the scope of the review. Th e permit 

granted, however, was usually restricted to 

a single State and of limited duration. 

(2) Sometime between 1995 and 1997, Federal 

lawyers interpreted that the FPPA of 1957 did 

not apply to the release of entomophagous 

insects, but only to their importation and 

holding in containment. When an application 

to release an entomophagous insect from 

containment was received, APHIS would 

decline jurisdiction if it determined that the 

organism met First-Tier Risk criteria (Table 3). 

Because a permit was not issued, the need for 

an EA was not triggered. After APHIS declined 

jurisdiction, biological control practitioners 

could move entomophagous organisms to the 

laboratory for further research or mass rearing; 

however, if they were federal employees, received 

federal funds for the project, or the organisms 

would be released on federal lands, then NEPA 

applied and an EA was needed prior to a 

release into the environment. Biological control 

agents for weeds (e.g., herbivorous insects) still 

required an EA for release from containment 

and into the environment. For these “weedeaters”, APHIS has had in place, for more 

than fifty-years, published guidelines and a 

Technical Advisory Group (TAG) to evaluate 

applications for release (http://www.aphis.usda. 

gov/plant_health/permits/tag/index.shtml). 

(3) In early 2006, under the authority of the PPA 

of 2000, APHIS resumed issuing permits 

for the release of entomophagous organisms 

from containment and preparing EAs for 

their first-time release into the environment. 

The current procedure is similar to that used 

before 1995, except that the EA provides a 

more thorough analysis of risks and benefi ts, 

and public comment on the EA is solicited. 

Although it can take a year or more for fi nal 

approval, the permit is usually comprehensive, 

with few restrictions, and is often a key to 

quickly obtaining subsequent approvals from 

States and other federal agencies, if needed. 

Although APHIS’s policies and procedures for 

the regulation of entomophagous biological 

control organisms are not formally established, 

my understanding of these is incorporated in 

the following procedures. It is strongly advised 

to first check with APHIS before attempting the 

first-time introduction of any biological control 

organism in any State, regardless of whether the 

source of the organism is domestic or foreign. 

Table 3.—First-tier risk assessment of nonindigenous invertebrates and micro-organisms proposed 

as candidates for release from containment. 

1) This organism has been identified to species /strain /biotype by a recognized authority.
 

2) All reasonable efforts have been made to exclude undesirable plant pests and other contaminants.
 

3) This organism does not feed on or infect living plant tissues.
 

4) This organism does not feed on, infect or contaminate plant products.
 

5) This organism does not transmit plant pathogens.
 

6) No life stage or sex of this organism develops as a parasite or pathogen of a primary parasite.
 

7) Release of this organism is not expected to cause significant losses in yields of crop or forage plants by causing major, 

population-level damage to commercially important pollinator or important natural enemies of plant pests or weeds. 

8) This organism is not expected to feed upon, attack, infect or otherwise adversely impact endangered or threatened 

plants or animals in the United States. 

30 
Chapter 3: Understanding Federal Regulations as Guidelines for Classical Biological Control Programs 

THE PROCESS FROM EXPLORATION 

TO RELEASE 

The process for introduction of biological control 

agents is often presented as a series of sequential 

steps; however, the path usually is neither linear 

nor uniform. In reality an outline of procedures is 

analogous to a roadmap that shows a fairly straight 

mountain road, which upon travelling is found to 

have ups and downs, switch-backs, and wrong turns, 

but also the excitement of new discoveries. Table 4 

is an example of a “roadmap” intended to help with 

planning. Chapters in other books give a broader, 

more general explanation for each step of a program 

to introduce new natural enemies (Van Driesche 

and Bellows 1996, Bellows and Fisher 1999). Th e 

emphasis in this chapter is on compliance with 

federal regulations and aspects of the process that 

usually are not provided in generalized descriptions. 

Initial Surveys 

The literature on the target pest and its natural 

enemies should be compiled for both where the 

pest is indigenous and where the natural enemies 

will be introduced. This literature survey should 

also include relatives of the target pest and their 

natural enemies. Besides taxonomic information, 

this compilation should include the distribution 

and host records of the natural enemies, if available. 

This information not only forms the basis to defi ne 

suitable areas to explore and what groups of natural 

enemies to search for in these areas, but also what 

taxonomic expertise may be needed. The ability to 

identify natural enemies is critical for biological 

control programs as the potential candidates often 

are undescribed species—five of the seven natural 

enemies imported for control of HWA (Table 2) 

were species previously unknown to science. 

Table 4.—Synopsis of steps and activities for obtaining and introducing biological control organisms. 

Step Partial List of Activities 

Survey potential release areas For potential release areas, define the climate and existing natural enemies 

attacking the target pest; start monitoring target population and potential 

non-target hosts. 

Initial planning/preparation 

(Where, What, When) 

Define search area and targets; review rules for export from search area; 

establish support team and funding; obtain use of approved containment 

facility; obtain import permit (PPQ-526) 

Exploration/collection Have suitable equipment for collecting, studying and keeping agents 

alive; do preliminary host range study; make arrangements for additional 

collections and study of natural enemies in collection area(s) 

Shipment Have proper forms for export (if needed) and import (PPQ-526); have 

suitable shipping materials; alert APHIS and your quarantine offi cer of 

shipment 

Rearing, and evaluation in containment Free agents of contaminants, obtain positive identification of candidate 

agents, develop rearing methods, study biology and potential host range in 

target release area(s) 

Biology in indigenous habitat Obtain information on biology and feeding range of selected natural 

enemies in indigenous area 

Release from containment Prepare release petition using NAPPO guidelines; apply for release permit 

(PPQ-526); APHIS-PPQ prepares an EA and solicits comments 

Field release/establishment Mass rear, as needed; decide where, when, how many/location for 

releases; obtain State and local permission for release; initial assessment of 

establishment and efficacy 

31 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

A field survey for natural enemies of the target 

pest, and its relatives, in the areas where it has 

become established should be made prior to 

making plans to collect and import natural enemies 

of the target pest. What native natural enemies 

may interact with the prospective natural enemy 

(e.g., its parasites and competitors) should also be 

identifi ed. This background information about 

existing fauna in potential release areas helps to 

define the missing components (e.g., natural 

enemies) and to identify potential nontarget or 

alternative hosts of the prospective natural enemy. 

Potential nontarget or alternate hosts and possible 

interactions with native natural enemies are very 

important considerations in risk analysis. 

Another purpose of the pre-introduction surveys 

is to establish a reference collection of positively 

identified natural enemies in the prospective 

introduction areas. The biological control 

program for the balsam woolly adelgid, Adelges 

piceae (Ratzeburg), failed to recognize native 

congeners of some of the introduced species 

resulting in false reports of the establishment of 

introduced species (Montgomery et al. 2011). 

Pre-introduction surveys of HWA natural enemies 

made in Connecticut and North Carolina 

(Montgomery and Lyon 1995, Wallace and Hain 

2000) provided some background information 

for the HWA biological control program. 

Where, What, When 

Although the literature on the distribution of the 

target pest may indicate where to search for its 

natural enemies, its biology and natural enemies 

may not have been reported, since introduced 

insects are often not pests in their indigenous 

regions. For HWA, we know where it originated— 

Japan (Havill et al. 2006)—and that HWA is also 

indigenous in western North America and in China, 

but these populations differ genetically from the 

population in the eastern United States (Havill et 

al. 2007). Other areas where HWA is indigenous 

include Taiwan, Nepal, Vietnam, and India, but 

these areas have climates that are less similar to 

target release areas in the eastern United States than 

areas already explored. Because of concerns about 

climate matching, the populations of Laricobius 

nigrinus, from moderate, coastal climates that 

were released for biological control of HWA in the 

eastern U.S. were supplemented with populations 

from cold, mountainous areas (Mausel et al. 2011). 

Unlike most insects, adelgids have no parasites 

and no specific pathogens; thus, the search for 

natural enemies is limited to predators. Past 

introductions of biological controls for HWA 

have been limited to predatory beetles—lady 

beetles and derodontids (Cheah et al. 2004). 

Surveys in western North America have identifi ed 

Leucopis flies (Diptera: Chamaemyiidae) that are 

part of the predator complex that feeds on HWA 

(Kohler et al. 2008), but their introduction is 

hampered by their abundant parasites, difficulty 

in rearing them, and taxonomic problems. 

Ideally, what to import would be based on the 

knowledge that the natural enemy actually regulates 

indigenous HWA populations. Life table analysis is 

a robust method, introduced by Varley et al. (1973), 

for describing the sources of and the quantifying 

mortality of a population in order to provide 

insight into the regulation of insect populations. 

Unfortunately, life tables are diffi cult to construct 

for field populations, especially when predators, 

rather than parasites, are the source of mortality. 

Additional information on construction of a life 

table can be found in Morris (1957), Royama 

(1981), Buonaccorsi and Elkinton (1990), and 

Bellows and Van Driesche (1999). A study of white 

fly mortality is a good example of application of a 

life table (Naranjo and Ellsworth 2005). McClure’s 

(1995, 1997) analysis of HWA mortality in Japan is 

one of the few efforts to collect mortality caused by 

prospective biological controls in their indigenous 

environment. He concluded that both densitydependent negative feedback (host resistance) and 

natural enemies played important roles in keeping 

populations of HWA low in Japan (McClure 

1997), but it is unclear what natural enemies were 

responsible. For example, one article points to an 

orbatid mite, which dislodged HWA eggs (McClure 

1995), another to four insect predators (McClure 

1997), and another to a lady beetle (Sasaji and 

32 
 

Chapter 3: Understanding Federal Regulations as Guidelines for Classical Biological Control Programs 

McClure 1997). He also noted that HWA mortality 

was high (>99%) in forests, where HWA density 

was low, but that mortality from predators was 

very low (<9%) on cultivated hemlocks with high 

HWA populations. He concluded that “the best 

biological control agents of introduced pests may 

not be those that help maintain pest populations at 

non-outbreak levels in natural habitats, but rather 

those that are most responsive to pest outbreaks in 

cultivated and disturbed habitats” (McClure 1997). 

In considering if natural enemies of relatives of 

the target species should be introduced, the life 

history and habitat requirements of the relatives and 

targets should be considered. The natural enemies 

released to control balsam woolly adelgid in North 

America were not successful, partly because they 

were collected from other adelgids and were not 

adapted to the climate (Schooley et al. 1984). It 

seems unlikely that reintroduction of these species 

for biological control of HWA would be successful. 

The best time to observe natural enemies in their 

indigenous habitat may not be the best time to 

collect them to establish colonies in containment. 

The greatest diversity and abundance of natural 

enemies of HWA seems to be in the spring, 

when the eggs of the overwintering and spring 

generations of HWA are present. Most discovery 

and first-time importations of natural enemies 

of HWA were made in the spring, but successful 

establishment of breeding colonies was done 

with fall collections. This is because most HWA 

predators are univoltine and lay eggs in the spring; 

thus spring imports may have already produced 

most of their eggs. Predators that feed on HWA 

during the fall can be collected then and stored 

in the containment facility until spring, enabling 

oviposition by the predator to be synchronized 

with the life history of HWA in the target region. 

The Containment Facility 

Organisms imported for classical biological control 

are usually brought first into a containment facility 

(aka quarantine). Thus, well before a permit is 

requested, the applicant must have access to a 

containment facility that has been inspected and 

approved by APHIS-PPQ. The process of certifying 

a facility for containment of arthropods takes 1-4 

months and APHIS should be consulted prior 

to its construction. APHIS consults with state 

officials about the construction of the containment 

facility and before issuing permits to import 

or release organisms into the state. It is good 

protocol to inform the state offi cial about your 

program in advance. An approved containment 

facility must have a Standard Operating Procedure 

(SOP) and this should be reviewed in advance of 

importations to make sure that your activity fi ts 

within the SOP. The Quarantine Offi cer should 

also be provided with an outline of your proposed 

activities and a copy of the approved permit 

so that he or she can ensure all protocols and 

restrictions are followed. Additional information 

regarding containment can be found at http://www. 

aphis.usda.gov/plant_health/permits/organism/ 

containment_facility_inspections.shtml. 

Import Permits 

The form PPQ-526, titled “Application for Permit 

Move Live Plant Pests or Noxious Weeds,” is used 

to obtain a permit for importing biological control 

agents. (As the title suggests, this form is also used 

for movement of plant pests and noxious weeds.) 

It has existed for more than 25 years, although it 

has been modified several times and adapted for 

electronic filing. On the form, the word “pest” 

means the organism(s) for which you are seeking a 

permit, thus organisms intended for use as biological 

control agents are to be listed on the form as “pests 

to be moved.” Use scientific names, but species 

group names—genus, family, order—may be 

acceptable since APHIS recognizes that little may 

be known about the natural enemy complex of the 

target organism in its indigenous habitat. If hosts 

of the natural enemy or foliage will be included in 

shipments, the scientific names of these are listed 

in a separate category. The countries where the 

collections will be made must be listed. Methods 

of containment and final disposition are required; 

thus, an approved containment facility and standard 

operating procedure usually is a prerequisite 

for importing biological control candidates. 

Application for permits should be made by the 

33 
 

 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

research scientist or other leader of the project rather 

than the quarantine officer. A minimum of eight 

weeks should be allowed to receive the permit. 

The application (PPQ-526) can be fi led 

electronically or using a paper form (APHIS 

recommends the former). Electronic fi ling requires 

that the applicant receive a USDA eAuthentication 

Account with Level 2 Access. Obtaining an 

eAuthentication account involves filling out a 

simple form online and then going to the nearest 

USDA Service Center to show a driver’s license 

or other government-issued photo ID. Th ere are 

several advantages in using eAuthentication to 

apply for permits: (1) there is helpful guidance in 

filling out the permit, such as pull down menus for 

countries and organism names, (2) processing is 

much faster—initial review takes one week whereas 

a paper application takes one month, (3) progress 

of the approval can be tracked, (4) your template is 

saved for renewal or application for another permit, 

(5) tasks, such as ordering shipping labels and fi ling 

annual reports, can be done by email, and (6) you 

will receive advance notice of permit expiration. 

Shipping 

After receiving the permit, the permit holder will 

need to request the PPQ Form 599 Red/White 

labels to enable foreign shipments to enter the 

United States (the labels are not issued or used 

for domestic, interstate shipments). Each of 

these distinctive labels has an individual number 

and an address of a USDA Plant Inspection 

Station (PIS). The red and white labels are not 

Table 5.—Outline of invoice letter. 

reusable or transferable and records of each use 

are tracked electronically using a barcode on 

each label. When shipping natural enemies, a red 

and white label is affixed to the outside of the 

package and supplemental information is placed 

inside (minimum is the permittee name, permit 

number and label number). Legally, the package 

should include the USFWS Form 3-177 on the 

outside (see http://www.fws.gov/le/ImpExp/faqs. 

htm). To expedite the shipment, I make an invoice 

(Table 5) with all the information that may be 

needed to clear the package and place this invoice 

in a clear pouch on the outside of the container 

addressed to Inspectors with a copy between the 

inner and outer layer of the shipment packaging. 

The permit includes detailed information on 

packaging the shipment, conditions regarding 

what may be shipped, as well as detailed step by 

step instructions regarding clearance for U.S. 

Customs and Border Protection (CBP) and APHIS 

PPQ Agriculture Plant Inspection Station (PIS). 

Shipments brought to the U.S. via commercial 

bonded-carriers or hand carried go first to CBP for 

clearance, with PIS and F&WS helping as needed. 

After clearance, the shipment may be transported to 

the containment facility by the same bonded carrier 

or it may be reshipped to the containment facility by 

APHIS-PPQ using the permittee’s designated carrier, 

billed to the permittee’s account. My experience is 

that the inspectors from these three Departments 

work closely, shipments are cleared very quickly, and 

delays can be traced to necessary information not 

being provided with the shipment, or to the carrier. 

1) Permit number and label number, permittee name.
 

2) The species (genus or family, if species unknown), both plants and animals, in the shipment
 

3) Statement that the shipment does not include CITES species
 

4) Statement that no venomous animals are in the shipment.
 

5) Statement that the contents have no commercial value
 

6) Address to forward the shipment to its fi nal destination
 

7) Your carrier and billing account number for reshipment (APHIS will not pay shipping costs) 

8) Name and phone number of a contact (quarantine officer) at the fi nal destination.
 

34 
 

 

 

Chapter 3: Understanding Federal Regulations as Guidelines for Classical Biological Control Programs 

Although hand-carrying live natural enemies from a 

foreign country directly to the containment facility 

can be arranged, this may not be safer or more 

expedient than shipping by bonded carrier. Th is 

privilege must be requested when applying for the 

permit and can be done only by the permittee or 

others designated on the permit. At least 20 days 

before the entry, each hand-carry event must be 

submitted and pre-authorized by the PPQ Permit 

Compliance Officer, who will notify CBP and 

provide you with a red and white label specifi cally 

prepared for the hand-carry event. Th e request 

should include details about who, when and where 

(i.e., the person who will carry, specific date, fl ight, 

and scheduled arrival), as well as details about 

what the package will contain, including foliage 

or other host material. Any deviations from what 

was pre-authorized, or changes in the airline or the 

travel date, will create the risk that the CPB officer 

will seize the package and send it to the nearest 

PIS or have it destroyed. Flight delays typical of 

airline travel should not create problems. After the 

package is released by CBP, it must be taken directly 

to the containment facility, and the quarantine 

officer (not the person who carried the package) 

must notify the PPQ Compliance Officer of the 

organisms received within 24 hours of their arrival. 

Biology and Host Specifi city Research 

Information on the agent’s biology and host 

specificity should be conducted in its indigenous 

habitat as well as in the containment facility. Th is 

is listed as two steps in Table 4, but one does 

not necessarily precede the other and they may 

occur simultaneously. Frequently, it is not until 

the species is imported and in the containment 

facility that its identity and potential for biological 

control is recognized. It often is necessary to return 

to where the species was collected to examine 

its biology and host range more thoroughly. 

The biological information for a petition to release 

an entomophagous biological control agent not only 

includes its identity but also methods to distinguish 

it from its relatives. Additional information should 

be provided about closely related species so that 

potential interactions with these can be assessed. 

For example, Laricobius nigrinus Fender, which 

was introduced in the eastern United States, can 

hybridize in nature with the indigenous L. rubidus 

LeConte (Davis et al. 2011). Th e likelihood that 

the agent may compete for food resources with 

native predators and be attacked by predators, 

parasites, and pathogens currently established in 

the proposed release area should be discussed. 

Information should also be provided on the 

agent’s dispersal capability and potential to thrive 

in the climate of the proposed release area. 

Predictions of host range should be based on 

observations in the putative agent’s indigenous 

environment as well as host specifi city testing 

conducted in the laboratory. When a potential 

biological control is first discovered, the collectors 

should also search for relatives of the target pest 

and determine if the putative agent also attacks 

it. This can be done using simple tests—for 

parasitoids, potential hosts can be recovered and 

reared to see if the putative agent emerges, and 

with predators simple feeding tests in small dishes 

may be done overnight. Notes should be made 

of the flora and fauna in the putative agent’s 

indigenous environment. Once the identity of 

the putative agent is confirmed, then literature 

searches may reveal other potential hosts. 

Laboratory evaluation of host specifi city begins 

with compiling a list of potential non-target species 

for testing that includes species with phylogenetic 

and ecological similarities, and species that may 

be endangered or of special ecological signifi cance 

(Kuhlmann et al. 2005). Often a hierarchical 

framework is used that starts with no-choice tests 

done in small arenas followed by choice tests, 

to determine prey that are attacked and prey 

preferences, and then rearing trials to determine 

prey suitability for development (van Lenteren 

et al. 2006). A good example of hierarchical 

testing is the evaluation of L. nigrinus (ZilahiBalogh 2005a,b), although Simberloff (2011) has 

questioned the adequacy of this. Natural enemies 

in the laboratory may utilize hosts that they 

would not utilize in nature; thus it is important 

to validate host specificity testing with knowledge 

35 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

of the host range in the candidate’s native areas. 

Anomalies in host specificity and biology in the 

laboratory can often be clarified with information 

about the natural enemy in its native range. 

Movement from Containment 

and Environmental Release 

In terms of federal regulation, the removal of 

entomophagous agents from containment facilities 

is regarded as a release into the environment. Th e 

same extensive information and thorough review 

as done for a “full” release into the environment 

would be needed for a “partial” release to rearing 

laboratory or for caged field studies. Th e process 

to remove a biological control agent from a 

containment facility begins with the same form 

used to import the organism into containment— 

Form PPQ-526. However, this time the permit 

application should include a separate report with 

the information (Table 6) requested in the North 

American Plant Protection Organization (NAPPO) 

“Guidelines for Petition for First Release of Nonindigenous Entomophagous Biological Control 

Agents” (NAPPO, 2008). The United States does 

not have a committee to review these petitions, as 

it has for biological controls for weeds; therefore, 

APHIS asks the Biological Control Review 

Committee, which has members from Mexico, 

the United States, and Canada, and is coordinated 

by Agriculture & Agri-Food Canada, to review 

petitions for release of entomophagous agents. 

Preparing a draft environmental assessment (EA) 

is the next step if the above review is favorable and 

APHIS concurs. This draft is prepared by APHIS 

based on information supplied by the applicant in 

the petition, and other resources the agency may 

have. Native Tribes and states in affected areas are 

contacted for comments on the draft. Then, a notice 

is published in the Federal Register of the availability 

of the draft EA and that anyone can comment 

on it for a 30-day period. Public involvement is 

required by NEPA and APHIS implementing 

regulations (7 CFR 372.5). If warranted, a fi nding 

of no significant impacts to the environment 

(FONSI) is issued along with a final EA. Only 

then may the permit to release the organism 

from containment be issued. Although not up to 

date, Hunt et al. (2008) provides a review of the 

procedure in the United States and other countries. 

The EA for the field release of L. osakensis in the 

continental United States is an example of this 

aspect of the regulatory process (APHIS 2009). 

The petition for its release was submitted October 

30, 2008 and on April 9, 2009, the Biological 

Table 6.—NAAPO guidelines for petitions for first release of nonindigenous entomophagous agents.* 

(1) Proposed action, with the purpose, need, and reasons for the release as well as specific location, timing, and 

method for the initial release. 

(2) Target pest information, including its taxonomy, economic impact, life history, and distribution as well as knowledge 

of other natural enemies (native and introduced) that attack the pest, and potential non-target species related 

phylogenetically or ecologically to the target pest. 

(3) Biological control agent information, consisting of its taxonomy and recognition characters; depository of voucher 

specimens (some must be deposited in the U. S. National Collection); other closely related species or genera in 

North America; its current and potential geographic, habitat, and climatic range; source of the agent; its life history; its 

known host range; and its natural enemies and that it will be free of these when released. 

(4) Environmental and economic impacts of the proposed release, based on known impact on vertebrates, direct 

impact on target and non-target species, indirect effects including competition with resident natural enemies, and any 

potential effects on threatened and endangered species. 

(5) Post-release monitoring, including its establishment and spread and affect on target population densities, and, 

when sufficient data are available, the economic and environmental impact of the program. 

*Abbreviated from North American Plant Protection Organization, Regional Standards for Phytosanitary Measures, No. 12 

(NAPPO 2008) 

36 
 

Chapter 3: Understanding Federal Regulations as Guidelines for Classical Biological Control Programs 

Control Review Committee issued a favorable 

evaluation. The notice of the availability of the EA 

was published a year later on 20 May in Federal 

Register (USDA 2010), and the FONSI was 

issued June 22, 2010. The process from receipt 

of the petition to issue of the permit took 18 

months, which is within published timelines. 

Movement within the 

Continental United States 

The PPA uses the level of a State in defining the area 

where an organism is considered to be established 

or native. Although this definition may not have 

scientific basis, it reflects the importance of States 

in federal laws. The procedure to obtain a federal 

permit to move an entomophagous biological 

control organism from a State where it is native 

to another State where it is not established is the 

same as for the release of an organism imported 

from a foreign country, except that a petition 

following NAPPO guidelines is not prepared since 

NAPPO addresses only species not established 

in the North American continent. The policy of 

regulating movement of entomophagous insects 

between States began sometime after 2006—a new 

edition of PPQ Form 526 (Dec 2011) has deleted 

the statement on its reverse side that it does not 

apply to interstate shipment of entomophagous 

insects (http://www.aphis.usda.gov/plant_health/ 

permits/downloads/forms/ppqform526.pdf). 

Many States also regulate the importation of 

biological control organisms into their State from 

other States. These states generally “piggy-back” 

on the federal permit and the purpose of the State 

permit is usually to provide a notification of specifi c 

release information, such as release date, place, and 

number to be released. Since new federal regulations 

regarding interstate movement of entomophagous 

biological control organisms have not been 

issued, it is best to consult with APHIS-PPQ and 

affected states prior to interstate movement of 

any biological control organism. It is anticipated 

that the pending proposal of new regulations 

will nominate more than 150 phytophagous and 

entomophagous biological control organisms for 

interstate movement without a permit and have 

procedures for nominating additional species. 

This will provide for dissemination of biological 

controls still expanding their range and not 

established in all states where the target pest occurs. 

IMPLICATIONS FOR 

BIOLOGICAL CONTROL 

Federal regulations are often viewed as an 

inconvenience or obstacle to biological control 

programs; however, understanding the regulations 

can not only facilitate compliance but also guide 

the development of biological control programs. 

The information required for a permit aligns 

with the information a conscientious scientist 

would obtain prior to releasing a new biological 

control organism. The regulations reflect the need 

to assure the safety of biological controls and 

to facilitate their dissemination. Th e NAPPO 

guidelines not only provide a framework for 

scientifically based risk assessment by regulatory 

agencies, but also can serve as guidelines in 

planning a biological control program. 

The hemlock woolly adelgid illustrates the need 

to regulate the movement of biological controls 

between ecological regions, whether the biological 

controls are native or imported from another 

country. This adelgid has regional populations in 

the United States, one that is indigenous to western 

North America, and another that is nonindigenous 

to eastern U.S., which originated in Japan (Havill 

et al. 2006, Havill et al. 2007). The permit for the 

release of Laricobius osakensis, a HWA predator 

from Japan, is valid for the continental United 

States. While it seems unlikely that someone would 

deliberately introduce L. osakensis to the western 

U.S., the consequences of its establishment there 

were not considered by the EA. Laricobius nigrinus, 

from the western U.S. and western Canada, was 

released in the eastern U.S. at a time when APHIS 

did not regulate the environmental release of 

entomophagous insects. It has since been found 

that L. nigrinus hybridizes with Laricobius rubidus, 

a related beetle native to eastern U.S. (Havill et 

al. 2010). It is only through the use of molecular 

37 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

genetics that this problem was identifi ed, and 

this issue highlights the need for both classical 

morphological identification, and information on 

the phylogeny of pest populations and their natural 

enemies in their native and introduced habitats. 

In summary, under existing regulatory authority 

and current policies, APHIS requires a permit for 

all importations and for any movement that crosses 

the “border” of the containment facility, or the 

border of a state. Federal regulatory authority is 

necessarily grounded in political (State) boundaries, 

and often fails to incorporate concepts such as 

ecological zones. While the example of biological 

control of HWA illustrates the need to regulate the 

movement of entomophagous insects, it remains 

unclear how to do this in a manner that facilitates 

biological control programs while protecting 

the environment from adverse impacts. Public 

response to new, proposed regulations will likely 

reflect the complex interactions of ecological and 

political boundaries, and variation between the 

intended use and the behavior of an organism. 

LITERATURE CITED 

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Beetle for Biological Control of Hemlock Woolly 

Adelgid ( Adelges tsugae), in the Continental 

United States—Environmental Assessment, 

December 2009. APHIS, PPQ, Riverdale, MD: 

37 p. http://www.aphis.usda.gov/plant_health/ 

ea/downloads/Laricobius-osakensis2009.pdf 

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of Biological Control. London: Academic Press: 

1046 p. 

Bellows, T.S.; Van Driesche, R.G. 1999. Life 

table construction and analysis for evaluating 

biological control agents. In: Bellows, T.S.; 

Fisher, T.W., eds. Handbook of Biological Control. 

London: Academic Press: 199-224. 

Buonaccorsi, J.P.; Elkinton, J.S. 1990. Estimation of 

contemporaneous mortality factors. Researches 

on Population Ecology 32: 151-171. 

Cheah, C.A.S.-J.; Montgomery, M.E.; Salom, 

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Caccone, A.; Kok, L.T.; Salom, S.M. 2011. 

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(Coleoptera: Derodontidae) species in eastern 

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DeBach, P. 1964. The scope of biological control. 

In: DeBach, P., ed. Biological Control of Insect 

Pests and Weeds. New York: Reinhold: 3-20. 

Havill, N.P.; Foottit, R.G.; Von Dohlen, C.D. 2007. 

Evolution of host specialization in the Adelgidae 

(Insecta: Hemiptera) inferred from molecular 

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Evolution 44: 357-370. 

Havill, N.P.; Montgomery, M.E.; Yu, G.; Shiyake, 

S.; Caccone, A. 2006. Mitochondrial DNA 

from hemlock woolly adelgid (Hemiptera: 

Adelgidae) suggests cryptic speciation and 

pinpoints the source of the introduction 

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Entomological Society of America 99: 195-203. 

Havill, N.P.; Davis, G.A.; Klein, J.; Caccone, A.; 

Salom, S.M. 2010. Hemlock woolly adelgid 

biological control: molecular methods to 

distinguish Laricobius nigrinus, L. rubidus, and 

their hybrids. In: McManus, K.A.; Gottschalk, 

K.W., eds. Proceedings, 21st U.S. Department 

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Invasive Species 2010; 2010 January 12-15; 

Annapolis, MD. Gen. Tech. Rep. NRS-P-75. 

Newtown Square, PA: U.S. Department of 

Agriculture, Forest Service, Northern Research 

Station: 25-28. 

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Chapter 3: Understanding Federal Regulations as Guidelines for Classical Biological Control Programs 

Hunt, E.J.; Kuhlmann, U.; Sheppard, A.; Qin, 

T.K.; et al. 2008. Review of invertebrate 

biological control agent regulation in 

Australia, New Zealand, Canada and the USA: 

Recommendations for a harmonized European 

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89-123. 

Kohler, G.R.; Stiefel, V.L.; Wallin, K.F.; Ross, D.W. 

2008. Predators associated with the hemlock 

woolly adelgid (Hemiptera: Adelgidae) in the 

Pacifi c Northwest. Environmental Entomology 

37: 494-504. 

Kuhlmann, U.; Schaffner, U.; Mason, P.G. 2005. 

Selection of non-target species for host 

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control agents. In: Hoddle, M.S., comp. Second 

International Symposium on Biological Control 

of Arthropods; 2005 September 12-16; Davos, 

Switzerland. FHTET-2005-08. Morgantown, 

WV: U.S. Department of Agriculture, Forest 

Service, Forest Health Technology Enterprise 

Team: 566-583. 

Mason, P.G.; Flanders, R.G.; Arrendondo-Bernal, 

H.A. 2005. How can legislation facilitate the 

use of biological control of arthropods in North 

America? In: Hoddle, M.S., comp. Second 

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Switzerland. FHTET-2005-08. Morgantown, 

WV: U.S. Department of Agriculture, Forest 

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matching of coastal and inland Laricobius 

nigrinus (Coleoptera: Derodontidae), a 

biological control agent of hemlock woolly 

adelgid. Biological Control 58: 96-102. 

McClure, M.S. 1995. Diapterobates humeralis 

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and introduced habitats: Lessons learned from 

the sap-feeding guilds on hemlock and pine. 

In: Andow, D.A.; Ragsdale, D.W.; Nyvall, R.F., 

eds. Ecological Interactions and Biological Control. 

Boulder: Westview Press: 31-52. 

Montgomery, M.E.; Lyon, S.M. 1995. Natural 

enemies of adelgids in North America: their 

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Tigner, T.; Reardon, R., eds. Proceedings: 

The First Hemlock Woolly Adelgid Review; 

1995 October 12; Charlottesville, VA. 

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Leschen, R.A.B. 2011. A new species of 

Laricobius (Coleoptera: Derodontidae) from 

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104: 389-401. 

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Biological Control Agents. Ottowa, Ontario: 

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Protection Organization. http://www.nappo. 

org/Standards/Standards(all)/RSPM12-Rev20­

10-08-e.pdf. (Last accessed 23 March 2011). 

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116: 93-108. 

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39 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Sasaji, H.; McClure, M.S. 1997. Description and 

distribution of Pseudoscymnus tsugae sp. nov. 

(Coleoptera: Coccinellidae), an important 

predator of hemlock woolly adelgid in Japan. 

Annals of the Entomological Society of America 90: 

563-568. 

Schooley, H.O.; Harris, J.W.E.; Pendrel, B. 

1984. Adelges piceae (Ratz.), balsam woolly 

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1969-1980. Farnham Royal, Slough, England: 

Commonwealth Agricultural Bureax: 229-234. 

Simberloff, D. 2011. Risks of biological control 

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Agricultural Risk Protection Act, June 20, 

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hemlock woolly adelgid. Federal Register 75(97): 

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Van Driesche, R.; Bellows, T. 1996. Biological 

Control. New York: Chapman and Hall. 447 p. 

van Lenteren, J.C.; Bale, J.; Bigler, F.; Hokkanen, 

H.M.T.; Loomans, A.J.M. 2006. Assessing risks 

of releasing exotic biological control agents of 

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Approach. London: Blackwell Scientifi c 

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Wallace, M.S.; Hain, F.P. 2000. Field surveys and 

evaluation of native predators of the hemlock 

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southeastern United States. Environmental 

Entomology 29: 638-644. 

Zilahi-Balogh, G.M.G.; Kok, L.T.; Salom, S.M. 

2005a. A predator case history: Laricobius 

nigrinus, a derodontid beetle introduced 

against the hemlock woolly adelgid. In: 

Hoddle, M.S., comp. Second International 

Symposium on Biological Control of Arthropods; 

2005 September 12-16; Davos, Switzerland. 

FHTET-2005-08. Morgantown, WV: U.S. 

Department of Agriculture, Forest Service, 

Forest Health Technology Enterprise Team: 

651-664. 

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Science and Technology 15: 165. 

40 
SECTION II AGENTS FOR
 

BIOLOGICAL CONTROL
 
CHAPTER 4: SASAJISCYMNUS (=PSEUDOSCYMNUS) TSUGAE, 


A LADYBEETLE FROM JAPAN
 

Carole Cheah 


The Connecticut Agricultural Experiment Station, Valley Laboratory, Windsor, CT
 

ORIGIN, DESCRIPTION, AND HISTORY 

OF INTRODUCTION 

Th e first exploration for native natural enemies of 

hemlock woolly adelgid, Adelges tsugae (HWA) in 

Japan, conducted by Th e Connecticut Agricultural 

Experiment Station, began in 1992. Several 

predators were collected from adelgid-infested 

Japanese hemlocks, but it was a tiny ladybeetle 

which proved to have the most potential for 

biological control in subsequent laboratory and 

field evaluations. This ladybeetle was collected 

from adelgid-infested Japanese hemlocks, Tsuga 

sieboldii and Tsuga diversifolia, between mid-May 

and late June 1992, in 13 of 37 forests and at 11 of 

37 ornamental sites in 12 prefectures throughout 

Honshu, Japan from sea-level to 1,980 m elevation 

(Sasaji and McClure 1997). In Japan, McClure 

determined that S. tsugae was responsible for 86% 

adelgid mortality in landscaped sites and 99% 

adelgid mortality in forest sites. In 1997, Sasaji 

and McClure named this new cocinellid species 

Pseudoscymnus tsugae (Coleoptera: Coccinellidae). 

In 2004, the genus was renamed Sasajiscymnus as a 

replacement for a name already in use for a genus of 

shark, and P. tsugae is now known as Sasajiscymnus 

tsugae (Vandenberg 2004). North American colonies 

of S. tsugae originated from field collections made in 

1994 and 1995 from Takatsuki, Osaka prefecture, 

Japan (approximately 34° N), and were much later 

diversified with more recent collections from the 

Kansai district, central Honshu, Japan (Shiyake et 

al. 2008). Sasajiscymnus tsugae belongs to the Tribe 

Scymnini, a group of small coccinellids, less than 

3mm in length, which are specialist predators of 

aphids, scales, mealybugs, and adelgids. Th e adult 

is entirely jet black, on average 2 (1.5-2.5) mm in 

length, with dorsal pubescence and 9-segmented 

antennae (Sasaji & McClure 1997) (Fig. 1a). 

Amber-colored eggs are laid singly or in small 

clusters in concealed locales on hemlock foliage, 

buds, cones and stem crevices. Eggs, measuring 

0.48 mm on average (Fig. 1b), hatch in 6-10 days, 

and development from egg to adult takes 24 and 

40 days at 25 °C and 20 °C, respectively (Cheah 

and McClure 1998). There are four larval instars 

and a pupal stage, with the mature fourth instar 

(Fig. 1c) consuming > 70% of the total adelgid 

stages required for completion of development to 

adult (Cheah and McClure 1996). Th e mature 

fourth instar larva is on average 2.25-3.30 mm in 

length and dark grey or reddish brown in color 

while the pupa is a reddish brown (Fig. 1d) (Cheah 

and McClure 1998). Sasajiscymnus tsugae highly 

prefers adelgids to aphids and can also complete 

development on other adelgid species such as balsam 

woolly adelgid, Adelges piceae, pine bark adelgid, 

Pineus strobi, and Cooley spruce gall adelgid, Adelges 

cooleyi (McClure and Cheah 1998, Cheah and 

Donahue 2003). Host range tests showed that S. 

tsugae preferred A. tsugae to Pineus strobi on Pinus 

strobus, Adelges laricis on Larix deciduas, Adelges 

cooleyi on Pseudotsuga menziesii and the woolly 

alder aphid, Paraprociphilus tessellatus on Alnus 

serrulata (Butin et al. 2004). This species has no 

reproductive diapause and is, therefore, amenable 

43 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

a. Adult b. Egg 

c. Mature larva d. Pupa 

Figure 1. Stages of Sasajiscymnus tsugae (photos by C. Cheah). 

44 
Chapter 4: Sasajiscymnus (=Pseudoscymnus) tsugae, a Ladybeetle from Japan 

to laboratory mass rearing on field collections of 

A. tsugae infested foliage from fall to mid-summer. 

Mass rearing magnitude is primarily limited by 

healthy adelgid prey availability. A federal permit 

for the release of S. tsugae in Connecticut was 

issued in April 1995 (Hennesy 1995). Th e fi rst 

field release of S. tsugae was made in a town park 

in Windsor, Connecticut in May 1995. A starter 

colony of S. tsugae was transferred to the Philip 

Alampi Beneficial Insects Laboratory (PABIL), New 

Jersey Department of Agriculture in 1997, where 

mass rearing for the release in multiple states was 

initiated. Releases of S. tsugae were then expanded 

to other states in 1999, largely through the early 

mass rearing efforts of PABIL and EcoScientifi c 

Solutions LLC (Scranton, PA). More insectaries 

rearing S. tsugae and other HWA predators were 

later established in the southern states, greatly 

expanding releases in the southern range of HWA. 

Since 1995, over two million S. tsugae have been 

released in more than 400 sites on federal and nonfederal lands to combat hemlock woolly adelgid in 

16 eastern states from South Carolina to Maine. 

BIOLOGY AND SYNCHRONY 

OF LIFE CYCLE WITH PREY 

Scientists at The Connecticut Agricultural 

Experiment Station investigated the biology, 

life cycle and potential of S. tsugae for biological 

control of HWA. Sasajiscymnus tsugae has a high 

lifetime fecundity, and females lay an average of 

280 (64-513) eggs over 14 (5-30) weeks (Cheah 

and McClure 1998). The adult has a long life 

span (> 1 year with overwintering), and exhibits 

excellent field synchrony with the both sistens and 

progrediens generations of A. tsugae. This species is 

the only multivoltine introduced HWA predator, 

producing two generations in the northeast (Cheah 

and McClure 2000). One hypothesis examined 

the relative developmental time of the predator in 

relation to its prey (Kindlmann and Dixon 1999). 

Kindlmann and Dixon predicted that predators 

that have a longer developmental time than their 

prey are unlikely to be successful biological control 

agents. Generation time ratio is defined as the 

ratio of predator to prey developmental times 

(Kindlmann and Dixon 1999). Th ree year fi eld 

studies in Connecticut indicated that temperature 

regulated S. tsugae F1 generation time is about 5 

weeks in late spring and early summer and is similar 

for the F2 generation in mid to late summer (Cheah 

and McClure 2000). In contrast, generation time 

for the sistens A. tsugae is about 32 weeks, and 

10 weeks for the summer progrediens generation 

(McClure, 1987). Generation time ratios for the S. 

tsugae-A. tsugae relationship are very favorable and 

between 0.16 and 0.5, conferring an advantage on 

S. tsugae. Comparison of these relative development 

times indicates the effective predatory impact of 

S. tsugae. In addition, successive, overlapping F1 

cohorts of S. tsugae also span the second progrediens 

generation of A. tsugae and S. tsugae is the primary 

introduced predator with impact on this second 

adelgid generation. Furthermore, adults continue 

to feed and survive on aestivating first instar adelgid 

nymphs throughout the summer, augmenting the 

predation impact on its adelgid prey. Adults and 

larvae of S. tsugae are highly mobile and voraciously 

feed on all life stages of A. tsugae, from eggs and fi rst 

instars to adults. Each beetle larva consumes about 

500 adelgid eggs or 50 to 100 adelgid nymphs, 

depending upon their size, to complete development 

to adult. Adults can live for more than one year 

and may consume about 50 adelgid nymphs 

each week during times of peak reproductive and 

feeding activities (Cheah and McClure 1996). 

The synchrony between life cycles of S. tsugae and 

A. tsugae studied in Connecticut is shown in Fig. 2 

(Cheah and McClure 2000). Adult beetles emerge 

from overwintering sites in hemlock forests in 

March and April. Females generally mate before 

the onset of winter and begin oviposition on egg 

masses of the A. tsugae sistens generation in April, 

when daytime temperatures average 15 °C. Eggs of 

S. tsugae are laid throughout the spring into midsummer during periods of both adelgid generations. 

Incubation periods and larval developmental times 

are variable and dependent on seasonal spring 

temperatures. Th e fi rst field generation of adults 

generally emerges in June and July, but the timing 

is seasonally dependent on ambient temperatures. 

45 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Sasajiscymnus tsugae 

Figure 2. Seasonal synchrony between Sasajiscymnus tsugae, and its prey, the hemlock woolly adelgid, Adelges 

tsugae in Connecticut (adapted from Cheah and McClure 2000). 

46 
Chapter 4: Sasajiscymnus (=Pseudoscymnus) tsugae, a Ladybeetle from Japan 

A smaller second generation of S. tsugae is also 

produced on the progrediens generation of A. 

tsugae, with new adults emerging in mid-August 

into September. Adult S. tsugae can survive the 

late summer period of dormant settled adelgid 

nymphs and were found in hemlock forests during 

the late summer and early fall. Sampling at various 

different sites in Connecticut and Virginia over 

4 years showed that S. tsugae remained with its 

adelgid prey year round. In Connecticut, in the 

northern end of the adelgid distribution, S. tsugae 

adults were detected throughout the year during 

warm winter years, with larvae occurring from May 

to September in the field. Parallel sampling at two 

forest sites in Virginia, in the southern range of 

adelgid distribution, showed that S. tsugae adults 

were present from April to November (Cheah and 

McClure 2000). During milder winters in the 

northeast, adults overwinter on hemlock foliage 

(Cheah and McClure 2000). Sasajiscymnus tsugae 

has been documented in field cages in 2003 to 

survive minimum daily winter lows of -7 °F in 

northern Connecticut and -5.8 °F in 2002 in north 

central Maine during studies of adaptation to 

balsam woolly adelgid (Cheah & Donahue 2003). 

FIELD RECOVERIES 

Field studies and recoveries of the beetle have 

documented the ability of S. tsugae to reproduce 

after release, locally disperse, survive heat waves, 

survive mild and severe Connecticut winters, and 

establish in a variety of different hemlock habitats 

in Connecticut between 1995-2005 (Cheah 

and McClure 2000, 2002; Cheah et al. 2005). 

Recoveries of overwintered beetles (larvae and 

adults) were made in Connecticut release sites in 

years following severe Connecticut winters in 1996, 

2000, 2003, and 2004 (McClure et al. 1999, Cheah 

and McClure 2002, Cheah et al. 2005). All stages 

from larvae, pupae, to adults of S. tsugae could be 

found through intensive forest ground sampling 

after the year of release in 65% of Connecticut 

release sites from 1996-2001 (Cheah et al. 2005). 

However, the persistent challenge in predator 

field sampling efforts continues to be the use of 

the limiting beat sampling technique, which only 

samples accessible lower canopy foliage of hemlocks 

and results in the consequent misinterpretation of 

the establishment of S. tsugae. Bucket tree sampling 

of mature hemlocks in Connecticut and New Jersey 

has shown that the beetle is highly mobile and 

dispersed to upper canopies after release. Adults and 

larvae have been retrieved at 5-20 m heights on the 

outer hemlock branches in the forest canopy while 

concurrent ground sampling yielded no recoveries, 

indicating that the beetle was not evenly distributed 

in the hemlock canopy (Cheah et al. 2005). 

However, bucket truck sampling is expensive and 

not suitable for many release sites. In Japan, sweep 

net sampling up to 5 m in the canopy in a landscape 

setting in Osaka was effective in recovery of S. tsugae 

every month from March to December (Shiyake 

et al. 2008). Shiyake and colleagues recovered 

27 predacious species from adelgid-infested T. 

sieboldii during 2007 sampling and found that S. 

tsugae was observed on Japanese hemlocks for more 

months than any other predator; only absent in 

samples during January and February. Recoveries 

of S. tsugae have also been documented in western 

North Carolina, including in areas where S. tsugae 

was not known to have been released (McDonald 

et al. 2008). More recent detailed studies in the 

Great Smoky Mountains National Park using pole 

pruners and beat sampling techniques recovered 

adults and larvae of S. tsugae in 21.2% of sites 

sampled in 2008 and 2009 and these recoveries 

were significantly associated with older release sites 

(5-7 years after release) (Hakeem et al. 2010). 

FIELD IMPACT AND HEMLOCK RECOVERY 

IN CONNECTICUT 

In Connecticut, S. tsugae releases from 1995-2007 

have occurred in hemlock forests statewide over 

three climatic divisions and in a wide variety of 

soil types and habitats. This was also a period in 

which severe droughts occurred in 1995, 1999, 

and 2002. Initial hemlock mortality was recorded 

on marginal hemlock sites on ridge-tops, resulting 

from hemlock borer infestations and on the heels 

of devastating outbreaks of hemlock looper in the 

47 
 

 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

early 1990s. Concurrent with the expansion of 

HWA infestations in Connecticut in the 1990s, 

hemlocks were also stressed by the elongate hemlock 

scale, Fiorinia externa. Populations of this scale 

have significantly increased in density and range in 

Connecticut during the past 5 years, resulting in 

the decline of hemlock stands. Sasajiscymnus tsugae 

is the major introduced predator for biological 

control in Connecticut and efficacy of these releases 

on hemlock crown health has been monitored 

annually in a comparative approach since 2003. In 

the decade of the 1990s, severe winters punctuated 

the climate of Connecticut, and while the adelgid 

suffered major population reductions, populations 

tended to rebound to damaging levels in subsequent 

milder years. Ten years after the first release of S. 

tsugae, a period in which around 170,000 adult 

beetles were released in Connecticut for biological 

control of HWA, dramatic recovery of adelgidimpacted, declining hemlocks was recorded in many 

of the older established release sites, beginning in 

2005, after successive extreme winters in 2003 and 

2004 significantly reduced adelgid populations 

(Cheah 2006, Cheah 2011 in press). Recent 

winters in 2006-2008 and 2010 have been mild in 

comparison, but adelgid populations in the forests of 

Connecticut have not rebounded to original widespread damaging levels. Annual foliage transparency 

trends and other hemlock crown indicators such as 

new shoot production and tip dieback in S. tsugae 

release sites showed that even declining hemlocks 

recovered in one year when there was ample 

precipitation and a reduction in adelgid densities 

(Fig. 3). A comparative survey in Connecticut of 14 

non-release sites, which were matched to S. tsugae 

release sites climatically, topographically and in 

HWA infestation history, was performed in 2005 

to compare hemlock crown conditions in release 

AUGUST 2004 

Figure 3. Recovery of an adelgid-infested hemlock in one year in Connecticut where S. tsugae was released 

in 1999 (photos by C. Cheah). 

48 
 

Chapter 4: Sasajiscymnus (=Pseudoscymnus) tsugae, a Ladybeetle from Japan 

and non-release stands. Hemlock health assessments 

were also made in eight baseline hemlock sites which 

had no infestations of either adelgid or elongate 

hemlock scale. Crown health of hemlocks were rated 

(n = 15 trees/site) using standard U.S. Forest Service 

Forest Inventory Analysis (FIA) crown health 

assessment procedures, which has also been the 

method used to assess the conditions of hemlocks 

in release sites. A total of 287 hemlocks in release 

sites, 210 in non-release sites, and 90 hemlocks in 

baseline sites were compared statistically. In 2005, 

hemlock foliage transparency was signifi cantly 

lower in the 14 annually monitored S. tsugae sites, 

as compared to that in paired non-release matches 

(Cheah 2011 in press). Mean foliage transparency 

in 6-11 year release sites was also similar to that in 

the baseline sites, located at high elevation in the 

colder northwest corner of the state, indicating that 

recovery of hemlocks had approached that in non- Mean Foliage Transparency per Site Type

50

40

30

20

10

infested sites. An increase or stabilization of foliage 

transparency readings is interpreted to be due to 

the concurrent abundant new shoot production on 

previously infested hemlocks, an indication in itself 

that adelgid populations in the whole crown have 

been depressed. In addition, HWA crown levels 

in 2006 and 2007, measured in classes of <10%, 

11-50%, 51-75% and >75% showed that in the 

majority of release sites, average levels of adelgid in 

the site have been reduced from the initial prerelease levels. Annual hemlock crown health ratings 

from 2006-2010, show that this overall recovery 

has persisted to the present in S. tsugae release sites 

(Fig. 4; Cheah 2011 in press). This recovery has 

continued even in southern most sites, which have 

not had significant recent winter mortality of HWA 

until the winter of 2010-2011. With the reduction 

of adelgid populations during severe winters, 

abundant precipitation and cool growing seasons, 

0

2004 2005 2006 2007 2008 2009 2010

Year

Figure 4. Comparison between S. tsugae release , non-release , and uninfested baseline 

hemlock forest sites in Connecticut (Cheah 2011 in press). 

49 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

hemlock recovery in monitored release sites has been 

recorded in Connecticut in all types of soil types 

and sites, from rocky ridge tops to riparian, ravine 

to level habitats since 2005 to date (Fig. 5; Cheah 

unpublished). There has been negligible hemlock 

mortality in release sites since 2001. But in 2010, 

Connecticut sites which have had concurrently high 

infestations of elongate hemlock scale, had thinner 

crowns and higher foliage transparencies than sites 

with low or negligible scale infestations, indicating 

the deleterious impact of uncontrolled elongate 

hemlock scale populations (Cheah unpublished). 

Year

2002 2004 2006 2008 2010

0

20

40

60

80

100

a a

b b

b b

b

Mean Foliage Transparency

Figure 5. Mean annual foliage transparencies of previously adelgid-damaged hemlocks in 16 Connecticut 

S. tsugae release sites (9-15 years from year of first release), showing sustained crown recovery from 

2005-2009. Bars are followed by different letters showing significant differences at the p < 0.05 level 

(Cheah unpublished). 

50 
Chapter 4: Sasajiscymnus (=Pseudoscymnus) tsugae, a Ladybeetle from Japan 

LITERATURE CITED 

Butin, E.; Havill, N.P.; Elkinton, J.S.; Montgomery, 

M.E. 2004. Feeding preference of three lady 

beetle predators of the hemlock woolly adelgid 

(Homoptera: Adelgidae). Journal of Economic 

Entomology 97(5): 1635-1641. 

Cheah, C. 2006. Hope for hemlocks. Connecticut 

Woodlands 71(3): 13-15. 

Cheah, C. 2008. The case for Sasajiscymnus tsugae: 

Biological control has helped save Connecticut’s 

hemlocks. Abstract for a poster in: Onken, B.; 

Reardon, R., eds. Fourth Symposium on Hemlock 

Woolly Adelgid in the Eastern United States: 

Proceedings of the Meeting; 2008 February 12-14; 

Hartford, CT. FHTET-2008-01. Morgantown, 

WV: U.S. Department of Agriculture, Forest 

Service, Forest Health Technology Enterprise 

Team: 279-280. 

Cheah, C. 2011. Connecticut’s threatened 

landscape: natural enemies for biological control 

of invasive species. Frontiers of Plant Science. In 

press. 

Cheah, C.A.S.-J.; McClure, M.S. 1996. Exotic 

natural enemies of Adelges tsugae and their 

potential for biological control. In: Salom, S.; 

Tigner, T.; Reardon, R., eds. Proceedings: Th e 

First Hemlock Woolly Adelgid Review; 1995 

October 12; Charlottesville, VA. FHTET96-10. Morgantown, WV: U.S. Department 

of Agriculture, Forest Service, Forest Health 

Technology Enterprise Team: 103-112. 

Cheah, C.A.S-J.; McClure, M.S. 1998. Life history 

and development of Pseudoscymnus tsugae 

(Coleoptera: Coccinellidae), a new predator 

of the hemlock woolly adelgid, Adelges tsugae 

(Homoptera: Adelgidae). Environmental 

Entomology 27: 1531-1536. 

Cheah, C.A.S-J.; McClure, M.S. 2000. Seasonal 

synchrony of life cycles between the exotic 

predator, Pseudoscymnus tsugae (Coleoptera: 

Coccinellidae) and its prey, the hemlock woolly 

adelgid, Adelges tsugae (Homoptera: Adelgidae). 

Agricultural and Forest Entomology 2: 241-251. 

Cheah, C.A.S.-J.; McClure, M.S. 2002. 

Pseudoscymnus tsugae in Connecticut forests: Th e 

fi rst five years. In: Onken, B.; Reardon, R.C.; 

Lashcomb, J., eds. Proceedings: Hemlock Woolly 

Adelgid in the Eastern United States Symposium; 

2002 February 5-7; East Brunswick, NJ. New 

Brunswick, NJ: Rutgers, New Jersey Agricultural 

Experiment Station: 150-165. 

Cheah, C.; Donahue, C. 2003. Alternate 

adelgid prey species for Pseudocymnus tsugae 

establishment. Abstract for a presentation at 

the Northeast Forest Pest Council Meeting; 

2003 March 17-19; Grey Towers, PA. Online at 

http://cfs.nrcan.gc.ca/subsite/nefpc/abstracts/3. 

Cheah, C.A.S.-J.; Mayer, M.A.; Palmer, D.; 

Scudder, T.; Chianese, R. 2005. Assessments of 

biological control of hemlock woolly adelgid 

with Sasajiscymnus (=Pseudoscymnus) tsugae 

(Coleoptera: Coccinellidae) in Connecticut and 

New Jersey. In: Onken, B.; Reardon, R., comps. 

Proceedings of the Third Symposium on Hemlock 

Woolly Adelgid in the Eastern United States; 

2005 February 1-3; Asheville, NC. FHTET2005-01. Morgantown, WV: U.S. Department 

of Agriculture, Forest Service, Forest Health 

Technology Enterprise Team: 116-130. 

Hakeem, A.; Grant, J.F.; Lambdin, P.L.; Buckley, 

D.; Hale, F.A.; Rhea, J.R.; Widggins, G.; Taylor, 

G. 2010. Recovery of Sasajiscymnus tsugae, 

released against hemlock woolly adelgid, Adelges 

tsugae, in the southern Appalachians. Biocontrol, 

Science and Technology 20(10): 1069-1074. 

Hennessey, R. 1995. Field Release of a Nonindigenous 

Lady Beetle, Pseudoscymnus sp. (Coleoptera: 

Coccinellidae), for Biological Control of Hemlock 

Woolly Adelgid, Adelges tsugae (Homoptera: 

Adelgidae). Environmental Assessment. 

USDA-APHIS, Riverdale, MD. 

Kindlmann, P.; Dixon, A.F.G. 1999. Strategies of 

aphidophagous predators: lessons for modelling 

insect predator-prey dynamics. Journal of 

Applied Entomology 123: 397-399. 

51 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

McClure, M.S.; Cheah, C.A.S.-J. 1998. Released 

Japanese ladybugs are multiplying and killing 

hemlock woolly adelgids. Frontiers of Plant 

Science 50(2): 6-8. 

McClure, M.S.; Cheah, C.A.S.-J.; Tigner, T. 1999. 9999999

Is Pseudoscymnus tsugae the solution to the 

hemlock woolly adelgid problem? An early 

perspective. In: McManus, K.A.; Shields, K.S.; 

Souto, D.R., eds. Proceedings of a Symposium on 

Sustainable Management of Hemlock Ecosystems 

in Eastern North America; 1999 June 22-24; 

Durham, NH. Gen. Tech. Rep. NE-267. 

Newtown Square, PA: U.S. Department of 

Agriculture, Forest Service, Northeastern 

Research Station: 89-96. 

McDonald, R.; Mausel, D.; Salom, S.; Kok, L.; 

Montgomery, M.; Luker, G.; Steury, S.; Spears, 

G.; Skeate, S.; Graham, J.; Hamstead, B. 2008. 

Recovery of hemlock woolly adelgid predators 

in the high country of northwestern North 

Carolina 2004-2008. In: Onken, B.; Reardon, 

R., eds. Fourth Symposium on Hemlock Woolly 

Adelgid in the Eastern United States: Proceedings 

of the Meeting; 2008 February 12-14; Hartford, 

CT. FHTET-2008-01. Morgantown, WV: 

U.S. Department of Agriculture, Forest Service, 

Forest Health Technology Enterprise Team: 

53-60. 

Sasaji, H.; McClure, M.S. 1997. Description and 

distribution of Pseudoscymnus tsugae sp. nov. 

(Coleoptera: Coccinellidae), an important 

predator of hemlock woolly adelgid in Japan. 

Annals of the Entomological Society of America 90: 

563-568. 

Shiyake, S.; Miyatake, Y.; Montgomery, M.; Lamb, 

A. 2008. Hemlock woolly adelgid phenology 

and predacious beetle community on Japanese 

hemlocks. Abstract of a poster presented in: 

Onken, B.; Reardon, R., eds. Fourth Symposium 

on Hemlock Woolly Adelgid in the Eastern 

United States: Proceedings of the Meeting; 

2008 February 12-14; Hartford, CT. FHTET2008-01. Morgantown, WV: U.S. Department 

of Agriculture, Forest Service, Forest Health 

Technology Enterprise Team: 261-266. 

Vandenberg, N.J. 2004. Homonymy in the 

Coccinellidae (Coleoptera), or something fi shy 

about Pseudoscymnus Chapin. Proceedings of the 

Entomological Society of Washington 106: 

483-484. 

52 
 

CHAPTER 5: SCYMNUS (NEOPULLUS) LADY BEETLES FROM CHINA
 

Michael E. Montgomery and Melody A. Keena 

U.S. Forest Service, Northern Research Station, Hamden, CT 

INTRODUCTION 

In 1995, we found our fi rst Scymnus (Neopullus) lady 

beetle in China (Neopullus is a subgenus of Scymnus, 

the largest genus in the family Coccinellidae). At 

that time there were just a few known species in 

the subgenus and very little was known of their 

biology. By the end of the project, 14 years later, 

we had doubled the number of known Scymnus 

(Neopullus) species and thoroughly studied the 

biology of the three species that seemed the 

most promising candidates for biological control 

of Adelges tsugae Annand, the hemlock woolly 

adelgid (HWA) in the eastern United States. Th is 

discussion of the research on the three Scymnus 

(Neopullus) species is a good example of the 

process of classical biological control—(1) the 

discovery of potential agents and determination of 

their biology and host range in their indigenous 

environment; (2) importation and further study 

in quarantine of their biology and potential host 

range in the areas targeted for their release; (3) 

using sleeve cages to evaluate the response of 

natural enemies to their new environment and their 

impact on the pest; and (4) the free release of the 

biological control agents into the environment. 

The “we” refers to the several teams that participated 

in the project. The context of the research can be 

better understood by recognizing the principals 

of the China team, with a few remarks about how 

and where we worked. The project’s good luck 

was to have started with Yao Defu of the Chinese 

Academy of Forestry who was skilled in natural 

enemy research and forming partnerships. Yu 

Guoyue of the Beijing Academy of Agriculture and 

Forestry Science was indispensible in identifying 

or providing new descriptions for the more than 

70 species of lady beetles we found on hemlock in 

China. In Sichuan Province, Jianhua Zhou provided 

excellent leadership as well as hands-on help in the 

lab and field. In Yunnan, Li Li proved to be a most 

proficient collector and became so involved that he 

earned an engineering degree for his research on 

HWA natural enemies. Wenhua Lu of the University 

of Rhode Island did the initial studies in quarantine 

and acted as a liaison with the team in China. 

It is not possible to describe here the adventures 

and hardships this team experienced. Hemlock in 

China occurs in remote, rugged mountains; hence, 

access was a challenge (Havill and Montgomery 

2008). These forests have high diversity and 

both the Chinese and “Lao-wei” (foreigners) 

were excited about exploring these areas. Our 

working relationship was exemplified by the fun 

the Chinese had in speaking of “Lao-Mike” (“lao” 

literally means “old,” but depending on context it 

shows respect as an old friend or teacher or can be 

derogatory as a silly-old-fool) and the custom of 

sharing a meal and drink with local offi cials and 

forest workers who helped us with our research. 

OVERVIEW OF THE GENUS SCYMNUS 

Scymnus is the largest genus of lady beetles 

(Coccinellidae) with more than 600 described 

species. Adults are relatively small, less than 3 mm, 

compact, and pubescent. Larvae are characterized 

by a white waxy covering. Most of the species 

in the genus are aphidophagous, feeding on 

either aphids or adelgids. Th e genus Scymnus is 

divided into seven subgenera: Scymnus (Scymnus) 

Kugelann 1794, Scymnus (Pullus) Mulsant 1846, 

Scymnus (Didion) Casey 1899, Scymnus (Neopullus) 

53 
 

 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Sasaji 1971, Scymnus (Parapullus) Yang 1978, 

Scymnus (Mimopullus) Fursch 1987, and Scymnus 

(Orthoscymnus) Canepari 1997 (Kovar 2007). Th e 

subgenera Pullus and Scymnus are widespread, with 

the former having three-fourths of the species and 

the latter about 50 species. The other subgenera have 

less than 10 known species, except for the subgenus 

Neopullus, which has 22 known species (Kovar 

2007) that are Palaeartic, except for Parapullus 

and Didion, which also occur in North America. 

The subgenera can be separated by a combination 

of the number of antennal segments (10 or 

11), the length of the postcoxal line (complete 

or incomplete) and the presence or absence of 

distinct carinae on the intercoxal projection of 

the prosternum. Species in Scymnus (Neopullus) 

can be distinguished from the Scymnus species 

indigenous to North American by a combination 

of 10-antennal segments, a complete postcoxal 

line, and distinct carinae on the intercoxal process. 

Using a key to North American Coccinellidae 

(Gordon 1985), Scymnus (Neopullus) would key 

to Didion, except that the latter is distinguished 

by its feeble intercoxal carina. (Note: Gordon 

placed Didion in a separate genus rather than 

a subgenus of Scymnus.) Sasajiscymnus tsugae 

(Sasaji & McClure), imported from Japan 

to the United States for biological control of 

the hemlock woolly adelgid, has 9-segmented 

antennae and an incomplete postcoxal line. 

There have been 17 species of Scymnus lady beetles 

introduced to the United States, but only two are 

known to have established (Hagen et al. 1999). Both 

of the species that established are in the subgenus 

Pullus and attack adelgids (Gordon 1985). Scymnus 

(Pullus) impexus Mulsant, native to Europe, was 

introduced in the United States and Canada during 

1959-1963 to control the balsam woolly adelgid, 

Adelges piceae Ratzeburg. Although S. impexus was 

initially promising, it may have died out since its last 

recovery was 1978 in British Columbia (Harris and 

Dawsen 1979). Scymnus (Pullus) suturalis Th unberg, 

also native to Europe, was introduced to Michigan 

in 1961 to control adelgids on pine and is now 

established in several northeastern States (Gordon 

1985). It attacks Pineus strobi (Hartig), P. pini 

(Macquart) and A. tsugae in Connecticut (Lyon and 

Montgomery 1995). The abundance and seasonality 

of S. suturalis were examined on white pine and 

eastern hemlock, which had up to one larvae/branch 

(Montgomery and Lyon 1996). This lady beetle is 

now seldom collected in Connecticut (Montgomery, 

pers. obs.). There also is a Pullus lady beetle, 

Scymnus (Pullus) coniferarum Crotch, indigenous 

to the western United States, that feeds on both 

pine and hemlock adelgids (Whitehead 1967, 

Montgomery and McDonald 2010, chapter 10). 

Worldwide there are 22 described species in the 

subgenus Scymnus (Neopullus) and the hosts are 

known for only a few of these. Scymnus (Neopullus) 

hoff manii Weise is an important predator of 

aphids on crops in China and Japan (Yang and 

Zheng 1991, Kawauchi 1997). Seven Scymnus 

(Neopullus) species have been collected from 

hemlock in China (Yu et al. 2000) and three 

of these (Fig. 1) are the focus of this chapter: 

Scymnus (Neopullus) camptodromus Yu & Liu ( Sc), 

S. (N.) sinuanodulus Yu &Yao ( Ss) and S. (N.) 

ningshanensis Yu & Yao ( Sn). Although many species 

of Coccinellidae and other families of predators 

were collected, these three species seemed the 

most promising for biological control of HWA. 

DISTRIBUTION AND BIOLOGY 

IN NATIVE RANGE 

Distribution 

The search for natural enemies of the hemlock 

woolly adelgid in China focused on three 

provinces with extensive, widely distributed 

stands of hemlock—Yunnan and Sichuan in 

Southwestern China and Shaanxi in Central 

China (Fig. 2). These collection locations 

occur between 26.3°N and 33.3°N latitude 

and 1900 and 3200 meters elevation. 

Habitat 

The Sino-Himalayan region has three species of 

hemlock: Tsuga chinensis, which is widespread, 

occurring in 15 China provinces; T. dumosa, 

which occurs in a narrow zone across the southern 

Himalayan Range belt from Sichuan and Yunnan 

54 
Chapter 5: Scymnus (Neopullus) Lady Beetles from China 

Figure 1. Adults of three species of Scymnus (Neopullus): left to right, S. camptodromus (Sc), S. ningshanensis (Sn), 

and S. sinuanodulus (Ss). The beetles are about 2 mm in length. 

Figure 2. Map showing primary collecting area in China, with known distribution of three species of Scymnus 

(Neopullus) lady beetles outlined. 

55 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Provinces to Pakistan; and T. forrestii, which 

is limited to northwest Yunnan and southwest 

Sichuan (Fargon 1990). Like their eastern United 

States congeners, the hemlocks in China are shade 

tolerant and drought intolerant. Montgomery et al. 

(1999) provides additional information about the 

nomenclature and distribution of hemlock in China. 

In China, hemlock occurs only in mountainous 

regions, especially fog-belts at elevations between 

1,800 to 3,500 meters where moisture is plentiful 

during the growing season. Because the hemlock 

occurs in a limited elevation zone and the 

mountains are very steep and isolated, the hemlock 

often occurs in isolated “islands” (Wang 1961). 

This is a transitional area between broad-leaved 

deciduous forest and montane coniferous forest. 

These stands are extraordinarily diverse and have 

many of the same genera found in forests in the 

eastern United States. Besides Tsuga, there are 

species in the genera Abies, Aesculus, Carpinus, 

Cercis, Chamaecyparis, Clethra, Corylus, Ilex, 

Juglans, Juniperus, Lindera, Magnolia, Malus, 

Prunus, Picea, Pinus, Pseudotsuga, Sorbus, Taxus, 

Tilia, and Ulmus. There are also species in genera 

not present in eastern North America such as 

Castanopsis, Cercidiphyllum, Schima, Cunninghamia, 

Keteleeria, and Lithocarpus. This assemblage of a 

large number of tree species with the crown layer 

shared by several species is a characteristic of the 

mixed mesophytic forest type. This area of southwest 

China and the Southern Appalachian region of 

eastern North America are the only areas in the 

world where this forest type occurs (Wang 1961). 

In China, hemlock occurs only as a forest tree—it 

is not grown as an ornamental or landscape 

tree. Often it is the tallest tree in the forest with 

characteristic flat tops above the canopy. Although 

HWA and their lady beetle predators occur in the 

crowns of these trees, the crowns of big trees are 

not accessible. The distribution of HWA in the 

crown of T. dumosa is uniform vertically and by 

quadrant (Zhou et al. 2007, Li and Lu 2008). Most 

collecting is done in the lower crown, reachable 

from the ground, and on small trees growing near 

wet areas. The forests where collecting was done are 

protected and managed forest farms or preserves in 

which livestock are sometimes allowed to graze. 

Climate 

The areas where Scymnus (Neopullus) lady beetles 

were collected in China generally are more southerly 

in latitude and higher in altitude than the target 

areas for biological control in the eastern United 

States. Seasonal temperatures can be similar, 

however, because a 1000 m increase in altitude has 

roughly the same effect on air temperature as a 7.5 

degree increase in latitude (http://en.wikipedia. 

org/wiki/Alpine_climate). A comparison of average 

monthly temperatures for two of the collecting 

areas in China and two potential release areas in 

the United States provide examples of seasonal 

temperatures (Fig. 3). This shows that these four 

areas have similar winter temperatures, between 

-21 °C and -18 °C, but the areas in China have 

much lower summer temperatures. Th e lower 

summer temperatures are associated with the 

seasonal monsoon that occurs as warm tropical 

winds from the southeast collide with the rise of 

the Tibetan Plateau. Rainfall data, available for two 

of these areas (Fig. 4) shows the typical pattern of 

rainy summers and dry winters of the collecting 

areas in China and the more even pattern of rainfall 

in the Southern Appalachian area of the United 

States. Thus, the major climatic difference in the 

collecting area and the potential release area is the 

seasonal pattern of rainfall and cool summers. 

Host Associations in Endemic Area 

In addition to hemlock, other conifers and some 

angiosperms were sampled by beating limbs over 

an umbrella to determine the extent that alternate 

hosts may be used by the three Scymnus (Neopullus) 

species. Of the three species, only Ss was found 

on any host plant other than hemlock with a 

regularity that was not considered incidental 

(Table 1). During the first four years of collecting, 

the 5-needle white pine, Pinus armandii Franch, had 

heavy infestations of the adelgid Pineus armandicola 

Zhang, Zhong & Zhang, and the adelgid’s eggfilled ovisacs were on the pine’s needles in both 

spring and fall. In the spring and late fall, Ss was 

about one-fifth as abundant on the white pine as 

on hemlock, but Sc and Sn were not recovered from 

white pine. In Yunnan during September, when 

HWA was in diapause, Ss was more numerous on 

P. armandi than on hemlock. Numerous Scymnus 

56 
Chapter 5: Scymnus (Neopullus) Lady Beetles from China 

-5

0

5

10

15

20

25

Jan Feb Mar Apr May June Ju ly Aug Sep Oct Nov Dec

Nibago, Sichuan, PRC (37.36°N, 2750m) Mt. Lake, VA, USA (37.36°N, 1160m)

LJS, Yunnan, PRC (26.65°N, 2930m) Sugar Grove, NC, USA (36.25°N, 835m)

Figure 3. 	Average monthly temperatures (°C) of selected collecting locations in China and potential release sites 

in the United States. Latitude and meters altitude shown in brackets. Data collected by temperature 

recorders at the field sites for two or three year periods between 1999-2008. 

0

5

10

15

20

Mon th ly R a in fa ll

(% of Annual Tota l)

M t. Lake, V irgina N ibagou, C hina

Figure 4. Average monthly precipitation as a percentage of total annual precipitation. 

57 
 

 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

larvae collected from adelgid infested white pine 

at this time were identified as Scymnus (Pullus) 

yunshanpingensis Yu. Another Pullus species, S. 

(P.) geminus Yu and Montgomery, was abundant 

on white pine in the spring in Sichuan. Both 

of these Scymnus (Pullus) species will feed and 

oviposit on HWA, but their primary host seems to 

be pine adelgids. The other two species were not 

collected from white pine. It seems that the species 

in the subgenus Pullus may have pine adelgids 

as primary hosts whereas the three Neopullus 

species imported have HWA as the primary host, 

but may feed on pine adelgids to some extent. 

HWA Phenology 

Monthly sampling of HWA life stages and predators 

was done at three locations in Sichuan and Yunnan 

Provinces for one year. The number of ovisacs in 

each monthly sample was the most useful indicator 

of predator/HWA dynamics since the egg stage is 

targeted by most of its predators. As in the eastern 

United States, the onset of oviposition by HWA 

occurs earlier at more southerly latitudes. In Yunnan 

Province, the abundance of egg containing ovisacs 

peaks first in December and January and then 

in March whereas in colder Sichuan, the peak in 

abundance of ovisacs occurs in March and April 

(Fig. 5). In Sichuan, there is considerable overlap in 

eggs laid by the overwintering generation (sistens) 

and the spring generation (progrediens). In Sichuan, 

HWA ovisacs (and all nymphal stages) were found 

in low numbers throughout the summer and into 

early fall, especially in Nibagou, the coldest site. 

The cool summers of these regions may result in a 

longer period of survival and egg production by the 

progredientes. Other explanation for the presence 

of active stages throughout the summer include two 

progrediens generations or the progeny produced by 

migration of gallicolae from spruce. Th e diff erence 

in biology may also be related to the adelgids on 

hemlock in China being distinct genetically from 

HWA on other continents (Havill et al. 2006). 

Predator Abundance and Phenology 

Predators were collected by beating foliage over 

umbrellas. This technique is best suited for 

collecting adults. Usually 30 hemlock trees were 

beaten at each site every month. The closure of 

roads caused a switch to alternative sites in Sichuan; 

hence, the same three sites were not followed for 

Table 1. Occurrence of adult Scymnus sinuanodulus by beating foliage in Lijiang Prefecture, Yunnan, 

People’s Republic of China. 

Tree Species Occurrence Hemiptera present 

Tsuga dumosa Eichler Frequent Adelges tsugae, Diaspididae 

T. forrestii Downie Frequent A. tsugae 

Pinus armandii Franch. Moderate Pineus spp., Aphididae 

P. yunnanensis Franch. Rare Diaspididae 

Picea likiangensis Pritz. Rare Adelgid galls 

Abies delavayi Franch. Rare Adelges sp., Aphididae 

Larix potaninii Batalin None Adelges sp. 

Taxus yunnanensis Chang et L.K. Fu None Pseudococcidae 

Keteleeria evelyniana Mast. None Aphididae, Diaspididae 

Quercus pannosa Hand.-Mazz. None Coccidae 

Betula alnoidis Hamilt. None Aphididae 

Populus yunnanensis Dode None None 

Alnus ferdinandi-coburgii Schneider None None 

Rhododendron spp. None None 

58 
Chapter 5: Scymnus (Neopullus) Lady Beetles from China 

0

5

10

15

20

25

30

Jan Feb Mar Apr May June July Aug Sep Oct Nov Yunnan Sichuan Dec

Figure 5. Seasonal presence of HWA ovisacs in Sichuan and Yunnan Provinces; values are the average number 

on a twig for 60 twigs from each of three sites in each province. Data collected by Jianhua Zhou in 

Sichuan and Li Li in Yunnan. 

12 consecutive months. Overall 2,695 and 3,153 

specimens were collected in Sichuan and Yunnan, 

respectively. This is an average of 2-3 predators 

per umbrella sample with a range of zero for all 

samples taken in January and February in Sichuan 

to 8-9 per sample for May in Yunnan and October 

in Sichuan. The overall abundance of categories of 

natural enemies in the two Provinces was dissimilar 

(Table 2). Only seven specimens of Laricobius sp. 

were collected, all in Sichuan Province; thus, based 

on abundance, Laricobius spp. do not appear to 

be important regulators of HWA in China. In 

both provinces, the majority of predators collected 

are in the family Coccinellidae. In Yunnan, 62% 

of the Coccinellidae were the species Sc and Ss. 

In Sichuan, 48% of the Coccinellidae were the 

species Sc and Sn. Ss was not found in Sichuan and 

only three Sn individuals were found in Yunnan. 

The two most abundant predators were the large, 

colorful coccinellid Oenopia signatella (Mulsant) 

in Sichuan and the anthrocorid, Tetraphleps 

galchanoides Ghauri, in Yunnan. Both the literature 

and laboratory host range studies indicate that 

these last two predators are generalists feeding on 

aphids, adelgids, and other Heteroptera; thus, they 

appear to be non-specific, opportunistic species. 

59 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Table 2. Total number of predators recovered 

from umbrella samples taken monthly 

in two provinces. 

Species Sichuan Yunnan 

S. camptodromus 240 232 

S. ningshanensis 226 3 

S. sinuanodulus 0 269 

Oneopia spp. 549 23 

Other Coccinellidae 746 436 

Laricobius sp. 7 0 

Anthocoridae 22 305 

Other Predators 141 171 

The seasonal abundance of the three predators in 

each province that seem to have the greatest role in 

HWA population dynamics is presented in Figure 

6. Very few predators were collected during the 

coldest months of January and February in either 

province. There were two peaks in abundance of 

adult predators, one in the spring and another 

in the fall. In May and June, one site in Yunnan, 

which had the highest density of HWA, also had 

a high density of T. galchanoides nymphs and 

adults. Because this true bug pierces and sucks 

fluid from its prey and the HWA carcass remains 

attached to the stem, it was possible to assess, with 

the aid of a hand lens, that mortality of HWA 

exceeded 90 percent. The fall peak in predator 

abundance does not correspond well to HWA 

biology—HWA was still in diapause in Yunnan, but 

active stages of HWA were present in Sichuan—so 

its significance to HWA regulation is unclear. 

The abundance of the adults of the Scymnus 

(Neopullus) species peak in early spring, then 

increase in late spring and early summer (Fig. 6). 

0

20

40

60

80

100

120

140

160

Jan Feb Mar Apr May Jun Jul Aug Sep Oct Nov Dec

Scymnus camptodromus Scymnus sinuanodulus Tetraphleps galchanoides

YUNNAN

0

50

100

150

200

250

300

Jan Feb Mar Apr May Jun Jul Aug Sep Oct Nov Dec

Scymnus camptodromus Scymnus ningshanensis Oenopia signatella

SICHUAN 

Figure 6. Seasonal presence of selected predator species recovered from monthly foliage beating samples. 

60 
 

Chapter 5: Scymnus (Neopullus) Lady Beetles from China 

This decrease in mid-spring likely corresponds to 

mortality of the overwintered lady beetles after egg 

laying and is followed by the appearance of the 

newly eclosed adults about two months later. Th e 

adults are abundant on hemlock in the fall and this 

is the best time to collect them for export. Adults 

of Sc collected in the fall or spring will immediately 

lay eggs in quarantine, but Sn and Ss collected in 

the fall will not begin to lay eggs until after a few 

months in cold storage. Because Sn and Ss lay eggs 

in the spring, adults collected then for export may 

have laid most of their eggs and thus may produce 

few eggs when brought into the quarantine. 

Observations of immatures in the fi eld (Yao 

and Hongbin 1999; Montgomery, pers. obs.) 

indicate that larvae were present during April 

and May in both provinces. At the end of May, 

larvae with waxy plumes were seen walking on 

the trunk and branches, and pupae were found 

under the bark on the bole of the tree (Fig. 7). 

Mating of Ss adults was observed in early April 

and they oviposited from then until late May. 

In Sichuan, oviposition by Sc was observed only 

during April, but eggs were found in the fi eld 

in late summer. Based on the temperature data 

of the study locations (Fig. 3), development of 

the immature stages occurs when daily average 

temperatures are between 5 °C and 15 °C. 

Figure 7. 	Scymnus (Neopullus) larva feeding on HWA 

(left) and pupae in bark crevices on the bole 

of a hemlock tree in Yunnan, China (photos 

by Guoyue Yu). 

BIOLOGY STUDIES IN QUARANTINE 

Th e Scymnus (Neopullus) lady beetles imported 

from China exhibit two different life history 

schemes that synchronize their development with 

that of HWA, including the approximately three 

month aestivation period of HWA. Two species, 

Ss and Sn, both have an extended pre-oviposition 

period; new adults mate soon after emergence but 

need some exposure to cool temperatures before 

females will initiate oviposition. They lay eggs 

only in the spring that quickly hatch. Th e other 

species, Sc, can begin to oviposit one month after 

emergence if mated, but these eggs enter diapause 

and do not hatch until the next spring. Th e only 

other record of an egg diapause in Coccinellidae 

is for another Scymnus species, S. (Pullus) 

impexus (Mulsant), which feeds on Adelges piceae 

(Ratzeburg) in Europe (Delucchi 1954). Details 

on these two strategies and the data that supports 

them are provided in the following sections. 

Egg Deposition and Hatch 

The females of all three species select concealed, 

protected places, near HWA to lay their eggs, but 

in the laboratory will lay eggs in exposed areas and 

white gauze ( Ss and Sn only) if suitable oviposition 

sites are not available. Eggs are usually laid singly, 

but may be in groups if the adults are crowded. 

The eggs of all three species are fi rst yellow-orange, 

but become darker in 2-3 days. As they near hatch, 

the outer shell (chorion) becomes transparent and 

reddish eye spots are visible. Just before hatching, 

the egg surface becomes iridescent as the chorion 

separates from the embryo. Eggs of Sc are generally 

deposited in more concealed locations and have 

a more leather-like surface than the other two 

species (Fig. 8). This may provide added protection 

because the eggs of Sc will not hatch until 4-8 

months after being laid, whereas the eggs of Sn 

and Sc hatch in about two weeks after being laid. 

The eggs of Ss and Sn will hatch in an average 

of 10 and 8 days, respectively when held at 

18-20 °C. Egg hatch was about 90% for both these 

species. Storing eggs of these two species at 5 °C 

for two weeks does not affect hatch, but longer 

storage reduces percent egg hatch. Thus, the eggs 

of these two species are not able to overwinter. 

61 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Figure 8. Scymnus camptodromus egg inserted 

between bud scales of hemlock. 

At temperatures ≥ 15 °C, the eggs of Sc remain 

yellow-orange and show no signs of embryo 

development until exposed to temperatures < 15 °C 

for 1-3 months (Keena and Montgomery 2010). 

There is some variation among individual eggs in 

amount of chill required to break diapause, and 

diapause seems to be broken most quickly when 

eggs are held at temperatures near 5 °C. Once 

diapause has been broken, the Sc embryo will 

begin to develop, even at temperatures near 

0 °C, and the speed of development slowly 

increases with increasing temperatures up to 15 °C. 

At a constant 10 °C, the eggs will hatch after an 

average of 227 ± 32 days. Eggs will hatch after 

exposure to 5 °C and the percentage hatch increases 

with increasing time at 5 °C. Th e temperature 

regime for highest percentage hatch (90%) in the 

shortest time is 56 days at 5 °C followed by about 

2 months at 10 °C. The optimal temperature 

for Sc egg hatch is near 10 °C. Th us, Sc eggs will 

spend the summer in diapause, develop only 

after exposure to the cool temperatures of fall, 

and hatch after HWA has begun laying eggs. 

Larva and Pupa Development 

The larvae have four instars, are elongate, yellowish 

to reddish brown, densely setaceous on the head 

and plates, and the body has tubercles, but lacks 

prominent spines (Fig. 9) (Lu et al. 2002). Th e 

newly-hatched larvae are transparent with the 

color of the hemolymph and recently consumed 

adelgid visible (Fig 10). The larvae grasp HWA 

crawlers with their mandibles and suck the prey 

contents and then expel it back and forth into the 

dead prey’s exoskeleton to aid in digestion (Lu 

et al. 2002). By the last instar, the larvae produce a 

conspicuous waxy covering on the cuticle (Fig. 11). 

The pupae are naked, with the larval exuvium 

attached only to the last abdominal segment, but are 

covered with coarse setae with viscous droplets on 

the tips, which we believe are defensive (Fig. 12). 

Figure 9. 	Drawings of Scymnus sinuanodulus larvae, 

first instar (left) and fourth instar (from Lu 

et al. 2002). 

62 
 

Chapter 5: Scymnus (Neopullus) Lady Beetles from China 

Figure 10. Scymnus camptodromus first instar larva. 

Figure 11. Fourth instar Scymnus larva and HWA eggs. 

The total time for development at 20 °C for the 

larval and pupal stages is about 20 days and 10 days, 

respectively, for Ss and Sc. Sn has a shorter period in 

the larval stages (Table 3). The fourth larval instar 

of Sn is shorter than the other two species, which 

spend about half of the last larval stage wandering 

or inactive (Lu et al. 2002). This behavior has also 

been observed in another lady beetle that feeds 

on adelgids, S. (P.) impexus (Delucchi 1954). 

The larvae of Sc can develop at constant 

temperatures between 10 and 25 °C (Keena and 

Montgomery 2010). Larvae of Ss also complete 

development at temperatures between 15-25 °C, 

but rearing at lower temperatures was not attempted 

(Lu and Montgomery 2001). However, 10 °C 

and 25 °C may be sub-optimal, because survival 

to pupation for Sc larvae was only 50% at 10 °C, 

> 70% for 15-20 °C, and 25% for 25 °C (Keena, 

unpublished data). For Ss, survival from egg hatch 

to pupation was > 60% at 15-20 °C and only 5% 

at 25 °C (Lu and Montgomery 2001). Th e Sc larvae 

took an average of 90 ± 3 days and 20 ± 1 days to 

pupate, at 10 °C and 25 °C, respectively. In their 

indigenous environment, average temperatures 

range between 5 °C and 15 °C during the months 

the larvae and pupae are developing. The lab rearing 

indicates that these beetles should thrive at the 

warmer temperatures they would encounter in the 

Southern Appalachian Mountains (cf. Fig. 3). 

Figure 12. Droplets at the tips of the setae covering 

Scymnus pupa. 

Table 3. Development time (days) of larval and 

pupal stages of Scymnus sinuanodulus 

(Ss) and S. camptodromus (Sc) at 20 °C 

and S. ningshanensis (Sn) at 18-20 °C. 

Stage Ss1 Sn1 Sc2 Sc3 

L1 3.1 2.8 

L2 2.5 2.5 20 20 

L3 2.9 3.7 

L4 11.4 5.9 

Pupa 10.6 11.1 9.0 10 

Total 30.5 26.0 29.3 30 

Data sources: 1Montgomery et al. 2002; 2Keena and Montgomery 

2010; 3

Lu and Montgomery 2000. 

63 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Mating and Oviposition 

The adults of all three species are uniformly dull 

orange after emergence, with the elytral maculation 

that is distinctive for each species taking a few 

days to appear. After about three weeks, the adults 

of all three species become more active and are 

observed to mate and fly frequently (Fig. 13). 

Adult weights of the three species vary with larval 

food quality and rearing temperature, but Sc may 

be larger than Sn and Ss (Table 4). Th e greater 

weight of Sc may have been due in part to it being 

reared individually whereas Ss and Sn were reared 

in groups. Another factor may be that the weights 

of Ss and Sn were taken after they had been in lab 

culture for 12 and 11 generations, respectively, 

whereas the Sc colony was a mixture of populations 

established 5 to 4 generations ago. Measurements 

of the body size made on field collected specimens 

indicate that Sc and Ss are the same size, but Sn is 

a little smaller (Yu et al. 1997, Yu et al. 2000). 

Female Sc first mate when about 3 weeks old and 

begin oviposition about a week later if active stages 

of HWA are available. Only a few eggs are laid 

if HWA nymphs are the only food and total egg 

production is better if females are held at < 10 °C 

Figure 13. Scymnus sinuanodulus with wings spread 

open as in flight (photo by Nathan Havill). 

Table 4. Average adult weights (mg) of Scymnus 

sinuanodulus (Ss), S. camptodromus 

(Sc), and S.ningshanensis (Sn). 

Sex Ss Sn Sc 

Male 0.90 0.96 1.30 

Female 0.98 1.15 1.30 

until HWA eggs are available. When HWA eggs 

are available, oviposition will begin about 2 weeks 

after placement at > 10 °C. The females normally 

deposit eggs individually in concealed locations 

such as bud scales (especially ones that form a 

curl), but prefers the pollen cones of hemlock. 

When opened pollen cones are available, along 

with ample HWA eggs for food, they will average 

10-14 eggs/female/week at 20 °C and often lay 

multiple eggs in a cone. If the food supply is 

not optimal, only 1-2 eggs are laid in a week. 

Females of the other two species ( Ss and Sn) also 

mate at about 3 weeks post-eclosion, but have a 

prolonged pre-oviposition period that includes 

exposure to temperatures of 5-10 °C for at least 

two months (Lu and Montgomery 2001). After 

four months exposure to cool temperatures, they 

will lay eggs within 48 hours after being warmed 

to 20 °C and provided HWA eggs for food. Peak 

egg laying for Sn is 18 egg/female/week when held 

at 19 °C. The average number of Sn eggs produced 

per ovipositing female in the laboratory was 28 in 

China and 85 in the United States. Th is diff erence 

likely reflects the artificially extended laying 

period in quarantine in the United States and the 

availability of foliage with a higher density of ovisacs 

that had about 30% more eggs/sac. A reasonable 

expectation for oviposition in the laboratory is one 

to two eggs per day for a period of 5 to 10 weeks, 

if good quality and abundant food and oviposition 

sites are available. Both Ss and Sn oviposit single 

eggs in concealed locations such as bud scales and 

at the edge of ovisacs. If the ratio of oviposition 

sites to beetles is low or they are confined on the 

same foliage for several days, multiple eggs may be 

found in the same place and on the twigs, under 

dead needles on the bottom of the rearing container, 

and on substrates added to the cup such as gauze. 

Two experiments showed that hemlock woolly 

adelgid eggs must be present for Sn to lay eggs (see 

Montgomery et al. 2002). Adults removed from 

cold storage were given foliage either with adelgid 

eggs or with only adelgid nymphs (third instar). 

Beetles provided adelgid eggs laid an average of 

64 
Chapter 5: Scymnus (Neopullus) Lady Beetles from China 

2.3 ± 0.8 eggs/week (N = 15) over a three-week 

period whereas the 15 lady beetles given only 

nymphs laid a total of three eggs for the entire 

three weeks. These three eggs were laid in the fi rst 

week. When provided a diet simulating late winter 

conditions, with mostly adelgid nymphs and few 

hemlock woolly adelgid eggs, the lady beetle laid 

an average of only 6.42 ± 3.54 eggs during the 4 

weeks following removal from cold storage, whereas 

30.42 ± 8.98 eggs were laid by the beetles when they 

were provided adelgids at peak oviposition. Similar 

studies on Ss have shown that they will lay fewer 

eggs when provided only HWA nymphs than when 

provided HWA eggs (Lu and Montgomery 2001). 

During the summer months when HWA is in 

diapause, the adults of all three beetle species feed 

little and are inactive during the summer. Survival 

is best during the summer when the adults are held 

at 10-15 °C, which is equivalent to the summer 

temperatures where they are indiginous. During 

the winter months, adults survive well at 5 °C and 

will feed occasionally on artificial diet. Both Ss and 

Sn adults have been found to have super cooling 

points generally between -12 and -20 °C so are 

well adapted to survive in cold climates (Costa 

et al. 2008). Since Sc eggs overwinter, it would 

be beneficial to know if Sc eggs can withstand 

sub-freezing temperatures as well as the adults. 

Feeding Behavior on HWA 

Scymnus adults prey on adelgid eggs by chewing 

them, often leaving smeared egg contents or partially 

consumed eggs. The beetles feed first on eggs 

exposed outside the ovisac, then gradually crawl into 

the ovisac until little of their body is visible. Often 

the adult adelgid is dislodged and subsequently dies. 

When adult beetles attack adelgid adults or older 

nymphs, they take a single bite, which causes dark 

brown hemolymph to ooze from the adelgid that 

the beetles then drink; they sometimes chew on the 

adelgid’s body without entirely consuming it. Th e 

adult beetles feed little on the active crawlers, which 

easily escape when approached (Lu et al. 2002). 

In China, adult Ss and Sc ate, respectively, an 

average of 22 and 31 HWA eggs daily, taking 60-80 

seconds to consume an egg (Yao and Wang 1999). 

In quarantine at 20 °C, adult Sn, removed from cold 

storage, consumed 1.0 ± 0.29 nymphs, 0.8 ± 0.31 

adults, and 5.5 ± 0.23 eggs/day when given a mix 

of stages (Montgomery et al. 2002). Both Ss and 

Sn adults fed at 0 °C and consumed 7-10 and 9-17 

eggs in 20 hours when held at temperatures between 

2.5 °C and 10 °C, respectively (Costa et al. 2008). 

Scymnus larvae feed on all stages of adelgids, but 

mostly on eggs. When feeding on adelgid eggs, the 

larvae enter the adelgid ovisac and usually consume 

all of the eggs before leaving. Larvae suck the eggs, 

leaving the chorions, whereas adult beetles chew 

the eggs and do not leave the chorions. Although 

adelgid crawlers usually escape encounters with 

adult beetles, Ss and Sc larvae have been observed 

capturing and eating this active stage. Th e larvae 

appear to feed through a form of extraoral 

digestion on the crawlers. They bite and suck out 

the contents of the crawler, then regurgitate the 

contents back repeatedly into the crawler several 

times, before abandoning the empty corpse (Lu 

et al. 2002). When starved, 1st instar Ss larvae fed 

on crawlers of the hemlock scale Fiorinia externa 

Ferris, regurgitating in the same way (Lu et al. 

2002). The 1st instar Ss beetle larva have also 

been observed attacking the aestivating 1st instar 

adelgid nymph by turning the prey on its side, 

piercing the underside of the thorax near the stylet, 

and sucking out the hemolymph. Large beetle 

larvae, starved for 1-2 days, were cannibalistic. 

Larvae are voracious feeders. For example, Sn 

larvae in the third instar consumed 99.2 ± 11.7 

adelgid eggs/day. Chinese colleagues reported 

that consumption of HWA ovisacs by each larval 

instar (I-IV) was 1.8 ± 0.6, 3.9 ± 1.5, 5.3 ± 2.0, 

and 11.2 ± 3.1, respectively (Montgomery et al. 

2002). The adelgid ovisacs had an average of 31 

± 11 eggs/sac; thus, total consumption by a larva 

was 23 ovisacs, or 713 eggs. First instar larvae of all 

three Scymnus species do not survive if they do not 

have adelgid (or their own) eggs on which to feed. 

65 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

HOST EVALUATION IN QUARANTINE 

Methods for Host Preference Testing 

The methods and results are a compilation and 

summary of previously reported experiments 

(Montgomery and Lyon 1996, Montgomery et al. 

1997, Butin et al. 2002, Butin et al. 2004, Hoover 

et al. 2010), unpublished data of the authors, and 

personal communications from K. Hoover. Included 

in some of the tests for comparison purposes were 

two non-native coccinelid species established in the 

United States, Harmonia axyridis Pallas, a predator 

of arboreal aphids, and Scymnus (Pullus) suturalis 

Thunberg, a predator of pine adelgids. Feeding by 

adults was measured as it is the adults stage that 

make the choice of prey on which the larvae will 

feed. Tests were not conducted during the summer 

when the adults are normally inactive and feed little. 

A series of sequential tests were used. First, 

no-choice tests are made where adult beetles are 

confined with a single type of potential prey—this 

indicates what they will not eat even when in a 

starved condition. Where warranted, this is followed 

by tests where the beetles are offered a choice 

between two to four prey items—this indicates 

relative preference. Unless indicated otherwise, prey 

was presented on a small section of host material in 

a Petri dish with a filter paper on the bottom. Prey 

were carefully counted before placement in the dish. 

Beetles usually were tested individually, and starved, 

but given water, prior to testing. The number of 

prey remaining was counted after a period of 20 

hours or more, and the presence of fecal droplets 

on the filter paper was noted in no-choice tests. 

The prey evaluated included insects that could 

be encountered on hemlock foliage (e.g. scales, 

psocids, and predatory dipteran larvae), tree feeding 

aphids, and other adelgids present in eastern North 

America. There are only fi ve Adelges and seven 

Pineus species of adelgids reported from eastern 

North America; all of the Adelges are introduced 

species, while five of the Pineus species are native. 

One aphid tested, Paraprociphilius tesselatus 

(Fitch), is the primary prey of the only predaceous 

lepidopteran in the continental United States, 

Feniseca tarquinius F.; therefore, there was concern 

that the Scymnus beetles might attack this aphid. 

Prey Acceptance (No-choice Test Results) 

Four species of adelgids, five species of aphids, and 

representatives of non-aphidoid taxa were presented 

to the lady beetles (Table 5). Harmonia axyridis, a 

generalist predator, had a high acceptance rate for 

all the prey it was presented, except for Fiorinia 

elongate scale. It was the only predator that fed on 

the large aphid, Cinara pinea. This coccinellid is very 

active and capable of catching large, active prey. It 

also consumed the predaceous Diptera larvae that 

prey on aphids and adelgids. Th e Scymnus beetles 

were more restrictive in their prey acceptance. Th e 

aphids they were presented were all fi rst instar 

nymphs, most of which were larger than HWA 

adults; only Eucallipterus tiliae and Aphis gossypii 

are comparable in size to first instar HWA nymphs. 

Aphids were not appreciably consumed by these 

Scymnus beetles, except for the consumption of 

Aphis gossypii by Sc. Th e first instar aphids were 

presented in a small container off its host and 

without filter paper, and may have been unable to 

escape the beetles. Th e other Scymnus species tested 

will also consume small aphids off host. Th e only 

species that preyed on all the adult adelgids was Sc. 

The pine adelgid, Pineus strobi, which is smaller 

than HWA, was consumed at rate equal to HWA. 

Prey Preference (Choice Tests) 

Because prey preferences among the adelgids 

were not clear based on single prey tests, HWA 

and another adelgid on their respective host 

plant were offered together in a Petri dish. 

Extensive choice tests by Butin et al. (2004) 

indicated that Sn prefers HWA over Adelges 

laricis and A. cooleyi and any aphid tested, 

but HWA and P. strobi were eaten equally. 

When Sc females were presented with a choice 

of HWA and 2-3 other adelgids, they showed 

a strong preference for HWA. In the four-way 

choice tests, the relative preference was HWA > 

Pineus strobi >> Adelges cooleyi or Adelges laricis. 

The combination of other prey present, when 

only three were offered, resulted in signifi cant 

differences in the relative feeding preference. Sc 

females were 6.6-fold more likely to eat HWA 

eggs over larch adelgid, 4-fold more likely to eat 

eggs of the combined group of Adelges cooleyi 

66 
 

 

 

Chapter 5: Scymnus (Neopullus) Lady Beetles from China 

Table 5. No-choice feeding1

 by adult lady beetles during 24-72 hour confinement with prey. 

Harmonia Scymnus Scymnus Scymnus Scymnus 

Prey Item axyridis suturalis ningshanensis sinuanodulus camptodromus 

Adelgidae 

Adelges tsugae Annand

 egg +++ +++ +++ ++ +++

 crawler nt nt ++ + +

 aestivating nymph ++ + + + +

 nymph III & adult ++ +++ + ++ ++ 

Pineus strobi (Hartig) +++ +++ +++* ++ +++ 

Adelges laricis nt nt 0* nt +++ 

Adelges cooleyi (Gillete) nt nt ++* nt +++ 

Aphididiae 

Cinara pinea (Mordwilko) 

Eucallipterus tiliae (L.) nymph 

Prociphilus tessellatus (Fitch) 

Eriosoma lanigerum (Hausmann) 

Aphis gossypii Glover 

+++ 

+++ 

+++* 

nt 

0 

+ 

nt 

nt 

0 

+ 

+* 

nt 

0 

++ 

nt 

nt 

0 

nt 

0 

0 

+++ 

Diaspididae 

Fiorinia externa Ferris 0 0 0 0 0 

Pseudococcidae 

Pseudococcus sp. +++ 0 0 0 nt 

Psocoptera 

Pseudocaecilidae +++ 0 0 0 nt 

Diptera 

Syrphidae (larva) 

Chamaemyiidae (larva) 

++ 

++ 

0 

0 

0 

0 

0 

0 

nt 

nt 

1 Codes refer to the percentage of predators tested that fed on the prey: 0 = none; + = <33%; ++ = 33 to 67%; +++ = >67%; 

and “nt” = not tested. 

* Indicates score based on the proportion of alternative prey eaten in 2-way choice test between indicated species and HWA. 

and Pineus strobi over Adelges laricis, and 1.6­

fold more likely to eat HWA over the combined 

group of Adelges cooleyi and Pineus strobi (Hoover 

et al. 2011). Beetles only laid eggs on hemlock 

infested with HWA during these choice tests. 

Choice tests using only the host plant found 

that significantly more time was spent by S. 

suturalis on white pine than on hemlock, whereas 

hemlock foliage was preferred by Ss and Sn 

(Table 6). Thus, it appears that tree foliage itself 

may vary in attractiveness to Scymnus species. 

In summary, all three species of Scymnus imported 

from China are adelgid specialists, but when 

adelgids are not present, adults will minimally 

feed on slower moving aphids that are similar in 

size to HWA. Larvae seem to require HWA eggs 

to complete development, but will feed on other 

adelgids. They prefer HWA over other adelgids in 

choice tests and the adelgid’s host plays a role in 

host choice. They appear to more readily accept 

adelgids on hemlock than on pine, and on pine 

than on other conifers. Adelgid eggs are required 

for larvae to complete development and for optimal 

oviposition. Both adults and larvae will locate 

an HWA ovisac and generally feed on it until 

it is exhausted before moving to the next. Th ey 

will feed on crawlers, nymphs, and adults, often 

only injuring or partially consuming the later. 

67 
 

 

 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Table 6. Foliage preferences by adult lady beetles given a choice between pine and hemlock 

for 240 seconds. 

Predator species No. tests 

Time on foliage (seconds) 

Tsuga canadensis Pinus strobus Probability1 

S. suturalis

S. sinuanodulus

S. ningshanensis

 7 

13 

8 

24.1 

93.9 

80.7 

55.2 

41.6 

29.3 

<0.001 

<0.001 

<0.001 

1

Probability that means are equal, paired t-test 

Since these beetles are specialized on adelgids, of 

which seven of the 12 species in eastern North 

America are non-native, and they have a strong 

preference for HWA, they are not likely to have 

any appreciable impact on non-target prey. In 

addition, when Sc were presented with eggs of 

various adelgids without host or waxy coverings, 

they ate three times more eggs than when the 

same adelgid eggs were presented with the female 

parent on host material in a 48 hour time period. 

This indicates that prey acceptance is at least 

partially based on host characteristics. Sc adults 

also showed a significant preference for the eggs 

of the overwintering generation of the adelgids 

(both HWA and larch adelgids) when compared to 

the second generation, which may indicate some 

host quality differences between generations. 

ESTABLISHMENT 

Field Evaluation in Sleeve Cages 

Information on the efficacy of biological control 

agents can be difficult to obtain when they are 

released freely into the environment because 

they disperse and both the agent and its target 

are affected by external factors that cannot be 

controlled. Field cage studies are an eff ective method 

to evaluate efficacy of natural enemies in a controlled 

setting and are more realistic than the laboratory, 

although the cages may alter microclimate and 

interfere with some enemy/prey interactions (Luck 

et al. 1999). Between 1999 and 2008, a series of 

experiments were conducted using sleeve-cages 

(Fig. 14) to confi ne Sn, Ss, and Sasajiscymnus tsugae 

on HWA infested branches of eastern hemlock. 

Figure 14. Sleeve cages used to confine lady beetles 

on HWA infested hemlock branches. 

The recommended procedure, which was refi ned 

over several years of use, is as follows: sites should be 

selected several months before the cages and beetles 

will be placed on the trees. They should have healthy 

hemlocks that have branches reachable from the 

ground with a density of HWA ranging from 100­

300 per 0.5 meter of branch. The sleeve cages are 

made of light weight “No-see-um” knitted polyester 

fabric (nylon is more susceptible to UV rot) and are 

1 m long × 0.65 m wide. Each treatment should 

68 
Chapter 5: Scymnus (Neopullus) Lady Beetles from China 

have at least 30 replications, because variation 

within treatments is high. Just before the start of 

the test, branches are selected that appear to have 

between 150-350 ovisacs on the terminal 0.5 m. 

The number of ovisacs is estimated visually on each 

branch (it takes about 3 full days for two people to 

count 160 branches) and these are tagged where the 

open end of the bag will be closed over the branch. 

The branches are ranked by HWA density and the 

treatments assigned randomly within the sequential 

groups based on the number of treatments. (Initially, 

trees were considered blocks with complete sets of 

treatments assigned to each tree, but it was found 

that variation between trees was not a signifi cant 

component of random variation.) Ideally, mature 

beetles held over winter in cold storage are used 

in the experiments. They may be removed from 

the cold and held as a group overnight for mating; 

however, mating just prior to placement in the fi eld 

does not seem to increase fecundity. The beetles are 

sexed and placed individually in small containers 

that can easily be opened with one hand in the cage. 

Ideally, the sleeve cages are placed on the branches 

in April before HWA eggs have started to hatch and 

average daily temperatures are 5-7 °C, and removed 

in June, when the progredientes are just starting 

to lay eggs. The branch with the bag attached is 

cut from the tree and taken to the lab and placed 

in cold storage until the bags can be opened. Th e 

beetles and HWA are counted with the aid of a 

microscope. The statistic most useful in assessing 

impact on HWA is the per capita change in the 

0

1000

2000

3000

No Bag Empty Bag Fem ale (No

Pro g eny)

Fem ale (W ith

Pro g eny)

HWA/Caged Branch

Figure 15. Population change of HWA in cages for 

one month (sistens ovisacs to progredien 

nymphs). 

population (r=ln [Nt/No]) where No was the initial 

population (sistens ovisacs) and Nt was the fi nal 

population (progrediens ovisacs) in each sleeve cage. 

In the first experiment, the sleeve cages were 

installed at two sites in Connecticut on 20 April 

and most were removed one month later. During 

this month, the eggs in the sistens ovisacs hatched 

and progrediens nymphs were present when 

the cages were removed. The HWA population 

increased in all the cages during the month but 

those with a single female Ss had, on average, lower 

HWA populations than the control bags without a 

beetle (Fig. 15). Note that there was a cage eff ect. 

HWA numbers were higher on the branch in the 

cage without a beetle than on the branch without 

a cage—the effect of the beetle was compared to 

the caged branch without a beetle. Th e branches 

where beetles produced progeny reduced HWA 

populations more than uncaged branches. Some of Adelgids / Caged Branch

the cages in this experiment were allowed to remain 

on the branches until July 7. At this time, the HWA 

population in the cages with reproductive beetles 

was not signifi cantly different, on average, than 

in the cages without beetles (Fig. 16). By July, the 

cages contained aestivating neosistentes and most 

of the beetles were dead, perhaps from the lack 

of suitable food. In cages without the beetle, the 

HWA populations increased in the fi rst generation 

3000

2500

2000

1500

1000

500 Sisten

Adults

Progredien

Nymphs

Sisten

Nymphs

No Beetle

Beetle with Progeny

20 April 20 May 7 July

Figure 16. Population change of HWA in cages with 

or without beetles for two generations of 

HWA. 

69 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

and then crashed in the next generation whereas 

bags with the beetle stabilized the population, 

but at a level still high enough to damage trees. 

Stabilization of pest populations below damaging 

thresholds is the goal of biological control. 

In a similar caged study, the presence of Sn resulted 

in negative growth of HWA populations, whereas 

the population growth in the cages without the 

beetle was positive (Butin et al. 2003). Th is study 

also included S. tsugae as a treatment and this lady 

beetle also reduced HWA populations compared 

to the control, but not signifi cantly. Th e S. tsugae 

were new beetles that had not been overwintered 

in the laboratory. Th e S. tsugae did not reproduce 

in the bags, whereas one-third of the Sn did. Butin 

also reported that, in the laboratory, the fecundity 

of Sn increased with increasing densities of HWA 

ovisacs. With both Ss and Sn, the beetle cages with 

the most beetle progeny were those with the highest 

number of HWA at the end of the experiment 

and most of the progeny were produced within 

the first week after the beetles were placed in the 

bags. These results indicated that plentiful, fresh 

food is needed for good beetle reproduction. 

Other caged field trials, conducted from 2002 

to 2006, were less successful. In some, the HWA 

population crashed in the control cages and neither 

the presence of Ss nor Sn was able to drive the HWA 

populations significantly lower (although many 

cages with beetles contained no HWA at the end 

of the feeding period). The beetles did not produce 

progeny on these declining populations. Th ese trials 

showed the importance of “pre-conditioning” of the 

adult females placed in the cages. It was also found 

that degree day growth models could be used to 

predict when each stage of the lady beetle would be 

present in the cage and that the free released and 

caged beetles had the same development rates. 

In 2007, three species of lady beetles ( Sn, Ss, 

and S. tsugae) were evaluated in sleeve cages in 

northwestern North Carolina. All three species 

significantly reduced HWA compared to the cages 

without beetles (Table 7). However, there was 

not a signifi cant difference in reduction in HWA 

populations among beetle species. For this test, all 

three species had been held over winter at 7 °C and 

short daylength at the Forest Service laboratory in 

Hamden. This test demonstrated that placement 

of overwintered, reproductively active lady beetles 

in the field, when day-night temperatures ranged 

from -2 °C to 12 °C, results in good oviposition 

by all three species and full development of their 

progeny, including maturation feeding of the 

new adults, before HWA enters aestivation. 

In 2008, a sleeve cage evaluation focused on S. 

tsugae. For this trial, S. tsugae was obtained from the 

North Carolina Department of Agriculture rearing 

facility. These beetles are normally reared at 25 °C 

Table 7. Populations of HWA (average/caged branch) when the cages were placed on the branches 

April 15 and removed June 19-22. The change in HWA population (r) is (ln Final – ln Initial). 

The number of progeny the female beetle produced in each cage is based on number of all 

beetle stages recovered at the end of the test, minus the initial female. 

 Empty cage Scymnus Scymnus Sasajiscymnus 

Treatments (no beetle) ningshanensis sinuanodulus tsugae 

Sistentes (initial) 254.5 297.0 258.0 252.5 

Progredientes (final) 132.7 49.5 1.5 17.6 

HWA pop’n change (r) -0.77 -3.79 -6.02 -5.85 

HWA pop’n change (%) -41.0 -84.1 -99.0 -95.0 

Beetle progeny (avg) na 4.8 3.2 7.9 

Beetle progeny (max) na 24 29 31 

Replications (n)1

 30 34 31 24 

1

Initally 30 controls and 34 cages for each treatment, but some cages were damaged. 

70 
Chapter 5: Scymnus (Neopullus) Lady Beetles from China 

and stored at 18 °C prior to release. The adults are 

usually less than one-month-old when fi eld released. 

Thirty replicates of four different ages of S. tsugae 

were placed in cages on April 1. Complete sets of 

the four treatments were removed on a given day 

between July 5 and 16, and the progeny counted 

(Table 8). The two-week-old beetles did not produce 

any progeny and only one of the one-month-old 

beetles produced progeny. The beetles held for one 

month at 18 °C and then placed in cold storage 

for one month also did not produce progeny. Th e 

only beetles that reproduced well had been reared 

the previous year and had been producing progeny 

for field release prior to being placed in the bags. 

These beetles were noticeably smaller and much less 

active than the much younger beetles in the other 

treatments. The progeny of the oldest beetles were 

either adults or pupae, whereas the progeny from 

the one-month-old beetles were only in the larval 

stage, which indicates that they may not have been 

ready to oviposit when released. Th is experiment 

suggests that releasing mature, ovipositing S. tsugae 

beetles in the spring would be more successful 

than releasing beetles that became adults that 

same spring. With both S. tsugae and the Scymnus 

beetles, better reproduction occurred with adults 

that had been overwintered in the laboratory. 

While sleeve cages are useful in defi ning release 

parameters and impact of lady beetles, the cages 

seem to inhibit their reproduction. In all the trials, 

the cages with the most progeny were the cages that 

Table 8. Production of progeny in sleeve cages 

by Sasajiscymnus tsugae varying 

in age. 

 Adult Age 

(Months) 

Produced 

Progeny 

Progeny 

(Avg. No.) 

Progeny

(max.)

 0.5 0% – –

 1 3% 16 16

 1+1* 0% – –

 >8 30% 32 64 

*Held one month at 18 °C and then refrigerated one month 

had the most HWA at the end of the trial (control 

cages excepted). It seems that all of these lady beetles 

produce more progeny when more food is available. 

Environmental Releases 

Free releases of Ss (Table 9) and to a limited extent 

of Sn (Table 10) have been made, but there is no 

record of recovery of these beetles in the years 

following their release. Following the early spring 

release of adult beetles that had been overwintered, 

both Ss and Sn larvae have been recovered and adults 

found until November, but not in the year following 

release. Thus, there is no evidence that either Ss or 

Sn are established in the eastern United States. 

IMPLICATIONS FOR 

BIOLOGICAL CONTROL 

Th e Scymnus (Neopullus) lady beetles that prey 

on HWA have either delayed reproductive 

maturation in the adult stage or an egg diapause. 

The periods of arrested development in all three 

Neopullus species coincide with the aestival 

diapause of the HWA neosistentes. Th e adults of 

these lady beetles prey on HWA during all periods 

when it is active, and their larvae are present 

in the spring when HWA eggs are present. 

A key question is whether a species with a life 

history where adults exist for several months before 

they lay eggs that quickly hatch, or a species with 

a life history where eggs exist for several months 

before they hatch, is more suitable biological 

control agent for HWA. The adults of Sn and Ss 

spend eight months exposed to predators and 

the elements before they lay eggs that soon hatch 

in spring. On the other hand, Sc begins laying 

eggs soon after eclosion and has an extended 

oviposition period in which to stockpile eggs, 

but these eggs then remain in diapause for many 

months until they hatch in early spring. It is 

not possible to predict which survival strategy 

would result in better HWA control. However, 

in their native range in China, the abundance 

of Sc was more consistent among the sites and 

during the survey period than the other species. 

71 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Table 9. Environmental releases of adult Scymnus sinuanodulus. 

Date No. released Location Rearing Lab. Condition 

April, 2004 150 Rauben Co., GA USFS, Hamden 10-wk-old adults 

October, 2004 320 Rauben Co., GA " 4-month-old adults 

April, 2005 528 Fairfield Co., CT " reproductive adults 

Spring, 2005 1,530 NJ NJ Dept. Agric. new adults

 " 1,210 PA " "

 " 750 NC " "

 " 460 MD " "

 " 460 WV " " 

Spring, 2006 1,500 NJ " "

 " 1,200 PA " "

 " 1,000 NC " "

 " 500 MD " "

 " 1,000 WV " " 

July, 2006 228 Watauga Co., NC Sleeve cages new adults 

Spring, 2007 6,305 NJ NJ Dept. Agric. new and old adults 

April, 2007 496 Avery Co., NC USFS, Hamden reproductive adults 

May, 2007 45 Gt. Smoky Mtn. NP UTK, Knoxville post-reproductive 

Late May, 2008 208 Chattahoochee NF, GA; 1 site UGA, Athens reproductive adults 

Spring, 2008 7,480 Chattahoochee NF, GA; 9 sites UGA, Athens eggs & larvae 

Spring, 2009 8,400 Chattahoochee NF, GA; 15 sites UGA, Athens eggs & larvae 

Late May, 2009 165 Chattahoochee NF, GA; 1 site UGA, Athens reproductive adults 

Spring, 2010 15,741 Chattahoochee NF, GA; 53 sites UGA, Athens eggs & larvae 

Late May, 2010 100 Chattahoochee NF, GA; 1 site UGA, Athens reproductive adults 

Spring, 2011 11,075 Chattahoochee NF, GA; 23 sites UGA, Athens eggs & larvae 

Early Spring, 2011 800 Chattooga River Corridor; 7 sites UGA, Athens reproductive adults 

Th e first predator imported from China for 

biological control of HWA was Sc in 1995. 

Unfortunately, this species has been very 

difficult to rear because of its egg diapause. Th e 

requirements for breaking its egg diapause have 

now been deciphered and colonies can now be 

maintained in the laboratory. This species has 

several characteristics that indicate it would make 

it a good biological control agent: (1) it has a true 

aestival diapause as does HWA; (2) it occurs over 

a broad geographic area and in diverse habitats in 

its native range; (3) its larvae are present at a key 

point in the life cycle of HWA; and (4) its adults 

feed on HWA throughout most of the year. 

Table 10. Environmental releases of reproductive 

adult Scymnus ningshanensis reared 

at the USDA Forest Service lab in 

Hamden, CT. 

Date No. released Location 

April, 2007 300 Hampshire Co., MA 

April, 2007 300 Hampden Co., MA 

April, 2007 300 Hartford Co., CT 

April, 2009 500 Avery Co., NC 

72 
 

Chapter 5: Scymnus (Neopullus) Lady Beetles from China 

The other species of Scymnus (Neopullus) imported 

from China have been environmentally released, but 

appear not to have established. Ss has been released 

in considerable numbers in several areas and does 

not seem to merit further effort at establishment. 

The other species, Sn, has been released only in 

very low numbers and may merit further eff ort. 

A significant problem with this species is that the 

colony was founded on a single collection of a 

low number of specimens, which declined to a 

colony of less than ten individuals before being 

multiplied to sufficient numbers for release. 

In addition to the Scymnus (Neopullus) lady beetles 

imported, there were other predators in China 

that appear when HWA is at its highest densities. 

These predators are mostly opportunists that feed 

on a broad range of prey. Th is non-specifi city 

makes them unsuitable as biological control 

agents, but the significance of opportunistic 

predators in driving down high HWA populations 

where it is indigenous needs to be recognized. 

It is not surprising that native, HWA specifi c 

predators are lacking in the eastern United States 

considering than HWA is a relatively recent 

introduction; what is surprising is the lack of 

native opportunistic predators preying on HWA. 

ACKNOWLEDGMENTS 

Most of the data on HWA and predator biology 

in China were collected by Wenhua Lu, Li Li, 

Jianhua Zhou, Wang Hongbin, Yao Defu, and 

Gouyue Yu, and their diligence and cooperation 

is greatly appreciated. The rearing and study of 

lady beetles in the laboratory would not have been 

possible without the help of Wenhua Lu, Julie 

Slovik, Phetsamon Souphanya, Jenny Ogrodnick, 

Lynn Jones, Gregg Bradford, Jason Biroscak, 

Rebecca Lomanchinsky, Alice Vandel, Vicente 

Sanchez, and Katie Casidy. Th e assistance of 

Nathan Havill, Beth Butin, E. Michael Blumenthal, 

Chris Asaro, and Richard McDonald with 

caged field evaluations is greatly appreciated. 

LITERATURE CITED 

Butin, E.; Montgomery, M.; Havill, N.; Elkinton, 

J. 2002. Pre-release host range assessment for 

classical biological controls: experience with 

predators for the hemlock woolly adelgid. 

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eds. Proceedings: Hemlock Woolly Adelgid in the 

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5-7; East Brunswick, NJ. New Brunswick, NJ: 

New Jersey Agricultural Experiment Station and 

Rutgers University: 205-214. 

Butin, E.E.; Elkinton, J.S.; Havill, N.P.; 

Montgomery, M.E. 2003. Comparison of 

numerical response and predation effects of two 

coccinellid species on hemlock woolly adelgid 

(Homoptera: Adelgidae). Journal of Economic 

Entomology 96: 763-767. 

Butin, E.E.; Havill, N.P.; Elkinton, J.S.; 

Montgomery, M.E. 2004. Feeding preference 

of three lady beetle predators of the hemlock 

woolly adelgid (Homoptera: Adelgidae). Journal 

of Economic Entomology 97: 1635-1641. 

Cheah, C.A.S.-J.; Montgomery, M.E.; Salom, 

S.M.; Parker, B.L.; Costa, S.; Skinner, M. 2004. 

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Fortney, M. 2008. Low temperature in the 

hemlock woolly adelgid system. In: Onken, B.; 

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of America North of Mexico. Journal of the New 

York Entomological Society 93: 1-912. 

Hagen, K.S.; Mills, N.J.; Gordh, G.; Mcmurtry, J.A. 

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and mite pests. In: Bellows, T.S.; Fisher, T.W., 

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Academic Press: 383-503. 

Harris, J.W.E.; Dawson, A.F. 1979. Predator release 

program for balsam woolly aphid, Adelges piceae 

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1960-1969. Journal of the Entomological Society 

of British Columbia 76: 21-26. 

Havill, N.P.; Montgomery, M.E.; Yu, G.; Shiyake, 

S.; Caccone, A. 2006. Mitochondrial DNA 

from hemlock woolly adelgid (Hemiptera: 

Adelgidae) suggests cryptic speciation and 

pinpoints the source of the introduction 

to Eastern North America. Annals of the 

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Havill, N.P.; Montgomery, M.E.2008. Th e role 

of arboreta in studying the evolution of host 

resistance to the hemlock woolly adelgid. 

Arnoldia 65(3): 2-9, inside & back of front 

cover. 

Hoover, K.; Cassidy, K.; Keena, K. 2010. 

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comps. Proceedings of the Fifth Symposium on 

Hemlock Woolly Adelgid in the Eastern United 

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29-30. 

Kawauchi, S. 1997. Life histories of Coccinella 

septempunctata brucki, Propylea japonica and 

Scymnus hoff manni (Col., Coccinellidae) in 

Japan. Entomophaga 42: 41-47 

Keena, M.; Montgomery, M.E. 2010. Eff ects of 

temperature on the eggs and larvae of Scymnus 

camptodromus. In: Onken, B.; Reardon, R., 

comps. Proceedings of the Fifth Symposium on 

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25-28. 

Kovář, I. 2007. Coccinellidae. In: Löbl, I.; Smetana, 

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568-631. 

Li, L.; Lu, W.-H. 2008. Relationships of infestation 

of hemlock woolly adelgids, Adelges tsugae, with 

environmental factors in Lijiang Prefecture, 

Yunnan [in Chinese with English abstract]. 

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Lu, W.; Montgomery, M.E. 2000. Comparative 

biology of three Scymnus lady beetles 

(Coleoptera: Coccinellidae): predators of 

Adelges tsugae (Homoptera: Adelgidae). In: 

McManus, K.A.; Shields, K.S.; Souto, D.R., 

eds. Proceedings of a Symposium on Sustainable 

Management of Hemlock Ecosystems in Eastern 

North America; 1999 June 22-24; Durham, NH. 

Gen. Tech. Rep. NE-267. Newtown Square, PA: 

U.S. Department of Agriculture, Forest Service, 

Northeastern Research Station: 188. 

Lu, W.; Montgomery, M.E. 2001. Oviposition, 

development, and feeding of Scymnus 

(Neopullus) sinuanodulus (Coleoptera: 

Coccinellidae): a predator of Adelges tsugae 

(Homoptera: Adelgidae). Annals of the 

Entomological Society of America 94: 64-70. 

Lu, W.; Souphanya, P.; Montgomery, M.E. 

2002. Description of immature stages of 

Scymnus (Neopullus) sinuanodulus Yu and Yao 

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history. The Coleopterists Bulletin 56: 127-141. 

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Luck, R.F.; Shepard, B.M.; Kenmore, P.E. 

1999. Evaluation of biological control with 

experimental methods. In: Bellows, T.S.; Fisher, 

T.W., eds. Handbook of Biological Control. New 

York: Academic Press: 225-242. 

Lyon, S.M.; Montgomery, M.E. 1995. Scymnus 

(Pullus) suturalis Th unberg (Coleoptera: 

Coccinellidae): New locality records and a 

report on feeding on hemlock woolly adelgid, 

Adelges tsugae Annand (Homoptera: Adelgidae). 

The Coleopterists Bulletin 49: 118. 

Montgomery, M.E.; Lyon, S.M. 1996. Natural 

enemies of adelgids in North America: their 

prospect for biological control of Adelges 

tsugae (Homoptera: Adelgidae). In: Salom, S.; 

Tigner, T.; Reardon, R., eds. Proceedings: Th e 

First Hemlock Woolly Adelgid Review; 1995 

October 12; Charlottesville, VA. FHTET96-10. Morgantown, WV: U.S. Department 

of Agriculture, Forest Service, Forest Health 

Technology Enterprise Team: 89-102. 

Montgomery, M.E.; Lyon, S.M.; Lu, W.; Defu, Y.; 

Wang, H. 1997. Developing biological control 

technology: evaluation of the feeding range 

of predaceous beetles. In: Chinese Academy 

of Forestry, USDA Forest Service, Resource 

Technology Institute, eds. Resource Technology 

1997: Beijing International Symposium 

Proceedings; Beijing: China Forestry Publishing 

House: 28-34. 

Montgomery, M.E.; Yao, D.; Wang, H. 1999. 

Chinese Coccinellidae for biological control 

of the hemlock woolly adelgid: Description 

of native habitat. In: McManus, K.A.; 

Shields, K.S.; Souto, D.R., eds. Proceedings of 

a Symposium on Sustainable Management of 

Hemlock Ecosystems in Eastern North America; 

1999 June 22-24; Durham, NH. Gen. Tech. 

Rep. NE-267. Newtown Square, PA: U.S. 

Department of Agriculture, Forest Service, 

Northeastern Research Station: 97-102. 

Montgomery, M.; Wang, H.; Yao, D.; Lu, W.; 

Havill, N.; Li, G. 2002. Biology of Scymnus 

ningshanensis (Coleoptera: Coccinellidae): 

A predator of Adelges tsugae (Homoptera: 

Adelgidae). In: Onken, B.; Reardon, R.C.; 

Lashcomb, J., eds. Proceedings: Hemlock Woolly 

Adelgid in the Eastern United States Symposium; 

2002 February 5-7; East Brunswick, NJ. 

New Brunswick, NJ: New Jersey Agricultural 

Experiment Station and Rutgers University: 

181-188. 

Montgomery, M.E; McDonald, R.C. 2010. Host 

preferences of Scymnus (Pullus) coniferarum: An 

adelgid. In: Onken, B.; Reardon, R., comps. 

Proceedings of the Fifth Symposium on Hemlock 

Woolly Adelgid in the Eastern United States; 

2010 August 17-19; Asheville, NC. FHTET2010-07. Morgantown, WV: U.S. Department 

of Agriculture, Forest Service, Forest Health 

Technology Enterprise Team: 173-176. 

Wang, C.W. 1961. The Forests of China with a Survey 

of Grassland and Desert Vegetation. Harvard 

University, Cambridge, MA: Maria Moors 

Cabot Foundation, Publication No. 5: 331 p. 

Whitehead, V.B. 1967. Validity of the Higher 

Taxonomic Categories of the Tribe Scymnini 

(Coleoptera: Coccinellidae). Berkeley, CA: 

University of California (Berkeley): Ph.D. thesis. 

Yao, D.; Wang, H. 1999. Biological Control of 

Hemlock Woolly Adelgid. Sino-American 

Cooperative Program, Research Report (1966­

1998). On file at USDA Forest Service, 

Hamden, CT: 12 p. 

Yang, J.; Zheng, R. 1991. A preliminary study of 

bionomics of Scymnus (Neopullus) hoff manni 

Weise. In Huang, B. (ed.). Proceedings of 

the Symposium on Coccinellids in China: Th e 

Centenary of Successful Introduction of Vedalia 

Beetle, Rodolia cardinalis (Mulsant). Shanghai, 

China: Shanghai Scientifi c and Technical 

Publishers: 72-78. (In Chinese with English 

abstract.) 

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Yu, G.; Montgomery, M.E.; Yao, D. 2000. Lady 

beetles (Coleoptera: Coccinellidae) from 

Chinese hemlocks infested with the hemlock 

woolly adelgid, Adelges tsugae Annand 

(Homoptera: Adelgidae). Th e Coleopterists 

Bulletin 54: 154-199. 

Yu, G.; Yao, D.; Liu, H. 1997. Th e Coccinellidae 

collected from Tsuga with Adelges tsuga[e] 

Annand (Homoptera: Adelgidae). Scientia Silvae 

Sinicae 33(5): 432-440. 

Zhou, J.-H.; Xiao, Y.-B.; Xiao, Y.-G.; Lu, W.-H. 

2007. Biology of the hemlock woolly adelgid, 

Adelges tsugae, and its spatial distribution. 

Chinese Bulletin of Entomology 44: 565-569. 

(In Chinese with English abstract.) 

76 
 

 

CHAPTER 6: LARICOBIUS NIGRINUS FENDER 

(COLEOPTERA: DERODONTIDAE) 

D.L. Mausel1, G.A. Davis2, A.S. Lamb3, G.M.G. Zilahi-Balogh4, L.T. Kok2, S.M. Salom2 

1

Department of Forest Management, Menominee Tribal Enterprises, Neopit, WI 


2

University of Massachusetts, Amherst, MA
 

3

Department of Entomology and Plant Pathology, University of Tennessee, Knoxville, TN 


4

Canadian Food Inspection Agency, Kelowna, British Columbia, Canada
 

INTRODUCTION 

The hemlock woolly adelgid (HWA), Adelges tsugae 

Annand, is a native herbivore in western North 

America (Havill et al. 2006). It feeds on western 

hemlock, Tsuga heterophylla (Raf.) Sargent, but it 

is not considered a forest pest (Furniss and Carolin 

1977), in complete contrast to its pest status in 

eastern North America. Observations in Washington 

State suggest that hemlock resistance and predators 

limit HWA densities to innocuous levels (Mausel 

2005, McClure and Cheah 1999). Th e importance 

of predators is underscored by the long-term 

survival of eastern hemlock trees in western 

arboreta (Fig. 1). Clearly, detailed demographic 

studies would be useful to identify the key factors 

that regulate HWA populations in this region. 

To determine what types of biological control agents 

may be needed in the eastern U.S., if any, HWA 

natural enemies were surveyed in Virginia and 

Connecticut, but few were found and they had no 

impact on HWA (Montgomery and Lyon 1996; 

Wallace and Hain 2000). In western North America, 

exploration for potential classical biological control 

agents was carried out in western Washington 

and Oregon to comprehensively document and 

characterize HWA predators (Kohler et al. 2008). Of 

the 55 predator species identifi ed, Laricobius nigrinus 

Figure 1. A healthy eastern hemlock tree in the 

Washington Park Arboretum, Seattle WA 

(No. # 119-49-B, planted in 1949) has 

innocuous HWA densities suggesting that 

factors other than host-resistance, such as 

natural enemies, explain its non-pest status 

Fender and Leucopis spp. (Diptera: Chamaemyiidae) in western North America (photo by 

(see Chapter 8) were the most abundant natural D.L. Mausel in 2002). 

enemies at low and high HWA densities. 

77 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Laricobius nigrinus was prioritized early on for 

evaluation because it was frequently collected from 

HWA-infested trees in Victoria, British Columbia, 

Canada (Humble 1994) and all Laricobius species 

with known biology feed exclusively on adelgids 

(Leschen [in press]). Since 1998, studies on 

L. nigrinus determined major aspects of its life 

history, environmental risk, and potential effi cacy. 

After approval for release in 2000, fi eld releases 

began in 2003 and populations have established 

from the southern Appalachians to New England 

spanning the invaded range of HWA, but remain 

isolated. Beetle population growth has been 

considerable and thousands of adults have been 

mass-collected at some sites for release in nearby 

forests to enhance dispersal. Post-release monitoring 

has detected an L. nigrinus impact on HWA 

populations, but more time is needed to defi nitively 

state in what situations, if any, L. nigrinus saves 

hemlock trees from decline and probable death. 

SYSTEMATICS AND TAXONOMY 

Within the Polyphaga supergroup of beetles, the 

Derodontidae family has archaic morphological 

features and is one of the earliest lineages (Hunt 

et al. 2007). The Derodontidae is a small family 

known as the tooth-necked fungus beetles due 

to the dentate or flattened lateral margins of the 

pronotum (der = neck; odon = tooth). Th ere 

are four genera in the family, with three being 

fungus feeders (Derodontus Leconte, Peltastica 

Mannerheim, Nothoderodontus Crowson) and 

one, Laricobius Rosenhauer, that is predaceous on 

adelgids (Lawrence 1989, Leschen 2000). To date, 

21 Laricobius spp. have been described worldwide 

in the temperate latitudes (Leschen [in press]). 

Laricobius nigrinus, L. rubidus Leconte, and L. 

laticollis Fall are native to North America and L. 

erichsonii Rosenhauer was introduced from Europe 

(Downie and Arnett 1996, Hatch 1962). Laricobius 

species feed exclusively on the Adelgidae, and all 

adelgid species feed on Pinaceae hosts (Havill 

and Foottit 2007). The etymology of the genus 

Laricobius is rooted in Laric-, referring to Larix 

spp. or Larch, which was historically used as a 

generalized term for trees in the Pinaceae family. 

Laricobius nigrinus was first collected and described 

from Bear Springs, Oregon (Fender 1945). Adults 

are small (2-3 mm), shining black, covered with 

fine ashy hairs, have striate elytra (10 rows, oval), 

11-segmented antennae (scape, pedicel, and nine 

annuli), and 5-5-5 tarsal segmentation (Fig. 2). 

The life stages have been described by ZilahiBalogh et al. (2006). Other beetles that look 

similar to L. nigrinus and commonly collected from 

hemlock include L. rubidus, silken fungus beetles 

(Coleoptera: Cryptophagidae), and minute brown 

scavenger beetles (Coleoptera: Lathridiidae). 

RANGE AND COLLECTION AREAS 

Laricobius nigrinus is native to the western 

United States and Canada with records from 

British Columbia, Alberta, Idaho, Washington, 

Oregon, and California (Bright 1991, Fender 

1945; Hatch 1962, Lawrence 1989). Specimens of 

Figure 2. 	Laricobius nigrinus is a predator native to 

western North America, where it feeds and 

reproduces primarily on HWA (illustration 

by Terry Lawrence). 

78 
Chapter 6: Laricobius nigrinus Fender (Coleoptera: Derodontidae) 

L. nigrinus in the U.S. National Museum of Natural 

History, Washington D.C. have been collected 

from Wyoming and the southeastern Yukon (M. 

Montgomery, pers. comm.). The distribution of L. 

nigrinus likely overlaps the range of western hemlock 

and other western conifers that host adelgids. 

Initial collections of L. nigrinus from a western 

hemlock seed orchard in Victoria, British Columbia 

in 1997 were identified by Donald E. Bright 

(Agri-Food Canada, Systematic Entomology 

Section). From 1997 to 2004, 2,710 beetles were 

collected from this site for quarantine evaluation, 

mass rearing and releases in the eastern United 

States. From 2005 to 2011, 54,878 beetles were 

collected from western hemlock in Washington 

State (principally Seattle, WA) for mass rearing and 

releases (D. McDonald, pers. comm.). In addition, 

from 2007-2010, 4,601 beetles were collected in 

northern Idaho and northwest Montana. Equal 

numbers of beetles were collected from western 

hemlock and western white pine (Callahan 

et al. 2008), and the majority of beetles were 

collected in Idaho (97%). Th e identifi cation of L. 

nigrinus from these locations was morphologically 

confirmed by G. Zilahi-Balogh (Canadian Food 

Inspection Agency) and N. Vanderberg (U.S. 

Department of Agriculture, Agriculture Research 

Service), and by mtDNA by N. Havill (U.S. 

Department of Agriculture, Forest Service). 

BIOLOGY AND ECOLOGY 

L. nigrinus has one generation per year (Fig. 3), 

which is synchronized with the phenology of 

HWA in its native and introduced range (Mausel 

et al. 2008, Zilahi-Balogh et al. 2003a). Both the 

beetles and HWA are active in the fall, winter, 

and spring, and dormant in the summer. Lamb 

et al. (2007) investigated the factors infl uencing 

summer dormancy and determined that temperature 

was the most important cue for its termination, 

while photoperiod was a modifying factor. Adult 

beetles emerge from the soil in the fall, disperse to 

hemlock branches, and feed on sistens (summerFigure 3. 	Generalized life-cycle of Laricobius nigrinus. Development rate varies based on temperature as 

influenced by latitude, physiography, and elevation. Arrows indicate major dispersal events and arboreal 

adults search tree canopies for HWA from fall-early spring to feed and lay eggs. Adults are dormant in 

earthen cells during the summer months. 

79 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

early spring HWA generation) nymphs, reproduce 

in early spring, and then die (Zilahi-Balogh et al. 

2003b, c). The adults pierce HWA nymphs with 

pointed mandibles and consume their hemolymph 

and organs. In the lab, females oviposit up to 396 

eggs (mean = 101), typically one per HWA sistens 

ovisac, but more than one is common if prey 

density is high. The larvae have four instars and 

feed on hundreds of progrediens (Spring HWA 

generation) eggs to complete larval development, 

which may require more than one ovisac. Th e most 

apparent sign of L. nigrinus feeding is the disturbed 

ovisacs due to larval feeding (Fig. 4). In spring, 

mature larvae drop to the soil, enter a pre-pupal 

and pupal stage in an earthen cell. Development 

from egg to adult takes 65 days at 15 °C. 

 Laricobius nigrinus is cold hardy and active during 

fall-early spring as long as temperatures are above 

freezing (Zilahi-Balogh et al. 2003a). Immature 

stages have minimum developmental thresholds 

at or below 5.0 °C (Zilahi-Balogh et al. 2003b). 

Adults from Victoria, BC are physiologically able 

to avoid dying down to -16 to -19 °C (Humble 

and Mavin 2005). Eggs can survive -27.5 to 

-26.9 °C. Larval survival diminished from -22.1, 

-17.0, -15.0, and -13.0 °C for each consecutive 

instar. Beetles from northern Idaho appear even 

more cold tolerant (Mausel et al. [in press]-b). 

Studies on L. nigrinus host searching behavior is in 

its infancy. The basic sequence of adult behaviors 

and its visual ability have been described in the 

Figure 4. HWA ovisacs that were fed on by Laricobius nigrinus larvae from a release site in western Massachusetts 

(left), compared with intact HWA ovisacs from a non-release site (right). 

80 
Chapter 6: Laricobius nigrinus Fender (Coleoptera: Derodontidae) 

laboratory (Flowers et al. 2007, Mausel et al. [in 

press]-a). Antennal morphology and preliminary 

behavioral studies suggest that L. nigrinus use host 

volatiles in long-range host searching behavior 

and/or mate location (Broeckling and Salom 2003). 

Few natural enemies are known to aff ect L. 

nigrinus in the eastern United States. Generalist 

predators (i.e., spiders and ground beetles) likely 

prey on beetles in the canopy and in the soil, 

respectively. Other previously released HWA 

predators probably do not drastically compete or 

prey on L. nigrinus due to phenological diff erences 

(Flowers et al. 2006). In Europe, L. erichsonii was 

reported as a host of Earobia sp. and Phrudus spp. 

(Hymenoptera: Ichneumonidae) parasitoids (Franz 

1958). A pathogen that causes fatal disease in larvae 

was observed by Franz (1958). No parasitoids 

have been observed in North America to date, 

but a microsporidian parasite caused mortality 

in a mass-rearing laboratory (see Chapter 12). 

NATIVE RANGE STUDIES 

Field studies were conducted in western North 

America to determine if L. nigrinus has biological 

characteristics that could be damaging to HWA 

populations. The phenology of L. nigrinus was 

monitored for 2 yr and it was highly synchronized 

with HWA (Zilahi-Balogh et al. 2003a). Th is 

suggests that L. nigrinus is host-specific and showed 

that the beetle has a unique niche, as it fed on both 

HWA generations during fall-spring when most 

other predators are inactive. Adult and egg/larvae 

density appears to increase with increasing HWA 

density (Kohler et al. 2008, Mausel 2007) and 

this relationship is an important trait observed 

in successful biological control agents (Huff aker 

1974). The actual impact of L. nigrinus on HWA 

populations in western North America has not been 

experimentally tested (i.e., predator exclusion study). 

However, an exclusion study may not accurately 

predict L. nigrinus impact where it is introduced, as 

that will depend on the abundance that it reaches. 

HOST RANGE EVALUATION 

 During field collections in the native range of L. 

nigrinus, adults are primarily collected from HWAinfested western hemlock in the Puget Trough 

Region (Seattle, WA and Victoria, BC) and western 

hemlock and western white pine in northern Idaho. 

During informal surveys of other hosts in the 

Okanogan region of British Columbia (G.M.G. 

Zilahi-Balogh, unpub. data), western Washington 

(D. McDonald, unpub. data), and northern Idaho 

(D.L. Mausel, unpub. data), adults have been 

collected from Pineus similis (Gillette)-infested 

western white pine, Adelges lariciatus (Patch)-infested 

western larch, and Adelges cooleyi (Gillette)-infested 

Douglas-fir and Engelmann spruce. Larvae have 

been collected from P. similis-infested western white 

pine and Adelges lariciatus-infested western larch. 

Th e L. nigrinus adults used in quarantine host 

range tests were field collected from HWAinfested western hemlock from Victoria, BC 

and imported to Virginia in 2000 for evaluation 

(Zilahi-Balogh et al. 2002). The test prey, six 

hemipteran species in three families (Adelgidae, 

Aphididae, Diaspididae), were used in choice and 

no choice host acceptance and suitability tests 

(Table 1). Host acceptance tests determine whether 

a candidate biological control agent will feed 

and/or oviposit on a host. Host suitability tests 

determine whether the biological control agent is 

able to complete development to the adult stage 

and produce viable offspring on a particular host. 

In both the no-choice and paired-choice oviposition 

tests, L. nigrinus females laid signifi cantly more 

eggs in HWA ovisacs over other test prey. In the 

paired-choice tests, no eggs were laid on host 

plant twigs housing the non-adelgid test prey, 

Chionaspis pinifoliae (Diaspididae), Cinara pilicornis 

(Aphididae) and Myzus persicae (Aphididae). 

Oviposition was more than five times greater on 

HWA than on the adelgid test prey, Adelges piceae, 

Adelges abietis or Pineus strobi in the paired-choice 

tests. Th ese differences indicate an ovipositional 

81 
 

 

 

 

 

 

 

 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Table 1. Summary of Laricobius nigrinus host-range testing results with adelgid, aphid, and 

scale prey (Hemiptera)

 Acceptancea Suitability 

Test prey Oviposition Adult feeding Larval development Final host statusb 

Adelgidae 

Adelges tsugae + + + Yes 

Adelges piceae + + – No 

Adelges abietis + + – No 

Pineus strobi + + – No 

Aphididae 

Cinara pilicornis + x – No 

Myzus persicae – x x No 

Diaspididae 

Chionaspis pinifoliae + x – No 

a + = positive response on test prey; – = negative response on test prey; x = test not conducted. 

b Whether the species could serve as a host to L. nigrinus. 

preference for HWA over these other adelgids. 

In no-choice adult feeding tests, eggs of all the 

test adelgids were fed on by adult L. nigrinus. 

Significantly more HWA eggs were consumed than 

A. piceae and P. strobi eggs, but not A. abietis eggs. 

Though not statistically signifi cant, L. nigrinus 

adults consumed on average of two times more 

eggs of HWA (48) than A. abietis eggs (25). 

Laricobius nigrinus only completed development to 

the adult stage on a diet of HWA (Table 1). Adelges 

piceae and P. strobi supported larval development 

to the fourth instar, providing evidence of larval 

feeding, but did not support further development. 

Laricobius nigrinus larvae that fed on A. abietis, C. 

pilicornis or C. pinifoliae did not survive beyond 

the first instar. Host suitability tests with HWA 

and P. strobi were repeated using L. nigrinus 

collected from western hemlock and western 

white pine in northern Idaho (D.L. Mausel, 

unpub. data). As in Zilahi-Balogh et al. (2002), 

beetles fed and developed to the fourth instar on 

either host, but only completed development on 

HWA. Survival from the egg to 4th instar was 

significantly greater on HWA than on P. strobi. 

Previous reports describing members of the genus 

Laricobius as adelgid specialists (Leschen [in press]) 

support the quarantine test results. Based on these 

data, L. nigrinus is host specific on the family 

Adelgidae and prefers HWA over other adelgids 

tested in this study. These results contributed to 

approvals for release of L. nigrinus from quarantine 

by APHIS, and for field releases by the U.S. 

Forest Service, U.S. Fish and Wildlife Service, 

National Park Service, and several eastern States. 

FIELD EVALUATION 

Field studies at high elevation sites in southwestern 

Virginia showed that L. nigrinus survived the winter, 

oviposited, fed and reduced HWA densities on 

eastern hemlock within mesh cages (Lamb et al. 

2005, Lamb et al. 2006). Furthermore, interactions 

among L. nigrinus, Sasajiscymnus tsugae (see Chapter 

4), and Harmonia axyridis did not decrease survival, 

feeding, or reproduction indicating that these 

species are complementary (Flowers et al. 2006). 

An open release of L. nigrinus adults on planted 

hemlock trees artificially infested with HWA 

established successfully, increased in abundance 

over three years, had a significant impact on HWA 

populations, and revealed a density-dependent 

relationship between the predator and prey 

(Mausel et al. 2008). 

82 
 

 

 

Chapter 6: Laricobius nigrinus Fender (Coleoptera: Derodontidae) 

RELEASES MADE 

Th e fi rst L. nigrinus release utilized a strategy that 

liberated an estimated 10,344 eggs in early spring 

2003 (Lamb et al. 2006). In fall 2003, an open 

release of 258 adults at a “field insectary” was 

conducted to produce field-acclimated beetles to 

supplement mass rearing operations (Mausel et al. 

2008). From 2003-2005, a large-scale exploratory 

release of 9,225 adults at 22 sites was conducted 

to investigate the effect of climate, release size, 

and release season on establishment (Mausel et al. 

2010). With increased mass-rearing productivity 

(see Chapter 11) and collections of adults in Seattle, 

WA, 102,069 adults were released at hundreds of 

sites in 14 States from 2003-2010 (see Chapter 

18). From 2007 to 2010, 2,686 adults from Idaho 

and Montana were released in the northeast and 

mid-Atlantic States. Over 258,747 eggs have been 

released in the extreme southern range of eastern 

hemlock to further evaluate this release approach 

(M. Dalusky and L. Burgess, pers. comm.). 

EVALUATION OF PROJECT OUTCOMES 

Establishment and Spread 

Th e fi rst L. nigrinus egg release resulted in recovery 

of adults two years post-release (Lamb et al. 2006). 

Th e field insectary release successfully established as 

well and after three years, hundreds of adults were 

relocated to HWA-infested forests in Pennsylvania 

and Maryland (Mausel et al. 2008). Th e large-scale 

exploratory release study established 13 L. nigrinus 

populations from Tennessee to Pennsylvania and 

release size and minimum winter temperature were 

positively related with establishment (Mausel et 

al. 2010). A site in western North Carolina and 

western Maryland were sampled intensely from 

the 3rd to 7th year post-release and thousands of 

adult beetles have been collected and redistributed 

to nearby HWA-infested sites to enhance dispersal 

(D. McDonald and B. Thompson, pers. comm.). 

Program releases are considerably adding to the 

number of established sites and results can be 

accessed via an online database (see Chapter 18). 

According to the database at this time, L. nigrinus 

has been recovered in at least 11 states, from 

northern GA to coastal ME. A cold-tolerant 

biotype of L. nigrinus from Idaho and Montana 

was evaluated for release in cold-regions where 

beetles sourced from Victoria, BC had a low 

probability of establishment (Mausel et al. [in 

press]-b). Idaho and Montana beetle recoveries 

one or two years post-release have been made at 

4 of 14 sites. One of these sites is in Vermont, 10 

miles south of the most northerly known HWA 

infestation in the region (D.L. Mausel, unpub. 

data). As such, the entire invaded range of HWA 

has established but isolated L. nigrinus populations. 

To determine the rate at which L. nigrinus disperse 

to the upper hemlock crown, vertical movement 

of adults was evaluated during the spring, for 

two years following beetle release, in Virginia and 

Pennsylvania (G. Davis, unpub. data). Beetles were 

capable of dispersing into the upper crown (>15 

m) for oviposition and approximately 88% of the 

offspring recovered from the branch samples were 

located above 7 m. The proportion of secondgeneration larvae collected above 7 m was 98%. If 

monitoring for L. nigrinus is limited to the lower 

hemlock crown, its presence may be overlooked or 

underestimated. Horizontal movement of L. nigrinus 

was monitored for six generations at eight locations, 

one each in Tennessee and North Carolina, two in 

Virginia, and four in Pennsylvania. Each spring 16 

hemlock trees were sampled at various distances 

from the release areas, with limits of 100 m for the 

first two generations and 300 m for third through 

sixth generations. Its rate of spread increased from 

50 m/yr for the first generation to 75 m/yr by the 

fifth. Other observations suggest that L. nigrinus 

can disperse greater distances. For example, D. 

McDonald (pers. comm.) did not limit sampling 

distance and recovered L. nigrinus from at least 1.6 

km from the release area, five years post-release. Th is 

equates to an approximate spread rate of 320 m/yr. 

By the fi fth generation, L. nigrinus was recovered 

more frequently where densities of HWA were 

highest, regardless of distance from the release. 

83 
 

 

 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Non-target and Unanticipated Effects 

Because L. nigrinus has recently established 

permanent populations in the eastern U.S., its 

impact on non-target organisms has not yet been 

comprehensively evaluated in the fi eld. Informal 

monitoring has recovered L. nigrinus from P. strobiinfested eastern white pine (D. McDonald, pers. 

comm.), but it is unclear if they are reproducing 

on this host, as laboratory tests suggest they will 

not (Zilahi-Balogh et al. 2002). An unexpected 

risk from the release of L. nigrinus, is the recent 

field, lab, and DNA evidence that L. nigrinus 

and the native L. rubidus are very closely related, 

hybridize, and have the ability to produce viable 

offspring (Klein et al. [in press]). Th e eff ects of 

this interaction on L. rubidus, native adelgids, 

and the potential for biological control of 

HWA are unknown, but under careful study. 

Suppression of HWA 

In Virginia, a predator exclusion experiment 

detected a signifi cant L. nigrinus impact on HWA 

in the field after a small release two years before 

(Mausel et al. 2008). During beetle dispersal studies, 

HWA density was reduced nearly seven-fold in the 

upper crown stratum, while L. nigrinus abundance 

increased 15-fold (G. Davis, unpub. data). Th e 

direct impact of L. nigrinus (F4-F7 larvae) on HWA 

was also evaluated at two paired release and control 

sites, one each in North Carolina and Pennsylvania. 

Release sites had a greater abundance of Laricobius 

spp. and reduced HWA survival compared with 

the control sites. Laricobius spp. densities at the 

release sites have reached levels observed in its 

native range. However, the ratio of Laricobius spp. 

to HWA remains much lower at the release sites 

than observed in their native range at this time. 

Recovery of Hemlock 

Hemlock canopy health (i.e., crown transparency) 

was assessed from trees that L. nigrinus was released 

upon at 12 paired release and control sites at the 

time of release and either five or seven years postrelease (G. Davis, unpub. data). In addition, before 

and after photographs were taken of release and 

control trees. This assessment of L. nigrinus impact 

on tree health has detected the typical hemlock 

decline and mortality (Figs. 5 and 6). Crown 

transparency was similar between the release and 

control sites at the onset of the study and increased 

at a similar rate five to seven years later. Substantial 

evidence showing that the progression of hemlock 

decline has materially changed at sites where L. 

nigrinus has established has not been observed, to 

date. At beetle release and control stands in NY, 

MA, VT, NH, and ME, permanent plots were 

installed for long-term monitoring of L. nigrinus 

and HWA populations, hemlock health, and forest 

vegetation structure (D.L. Mausel, unpub. data). 

RECOMMENDATIONS FOR FUTURE WORK 

1. Collect L. nigrinus adults in the western 

US and screen for entomopathogens 

and parasitoids before field releases and 

redistribution from established eastern sites. 

2. 	 Continue to evaluate and test existing 

and new release strategies to improve 

program effi ciency and eff ectiveness. 

3. 	 Initiate long-term monitoring of L. nigrinus 

and HWA populations, hemlock health, 

forest structure and vegetation, and nontarget impacts on permanent plots in the 

mid-Atlantic and southern Appalachian 

States where results may differ from plots 

that have been established in the northeast. 

4. Evaluate L. nigrinus impact on HWA and 

non-target organisms via rigorous predator 

exclusion and life table experiments. 

ACKNOWLEDGMENTS 

We are indebted to L. Humble for bringing this 

beetle to our attention; M. Cornwell, H. Benero, 

R. Hughes, M. Roller, M. Beversdorf, E. Fritz, 

B. Eisenback, B. Roessler, R. Flowers, H. Gatton, 

and others for mass rearing L. nigrinus; M. Meade 

for access to collection sites in Seattle, WA; 

D. McDonald, B. Thompson, B. Onken, J. Fidgen, 

S. Cook, T. Eckburg, L. Pederson, J. Elkinton, 

and R. Kimmich for assistance collecting beetles 

in the west; T. McAvoy, W. Mays, Ashley Hillyer, 

84 
Chapter 6: Laricobius nigrinus Fender (Coleoptera: Derodontidae) 

Figure 5. 	 Relatively rapid eastern hemlock decline in the southern Appalachians presents a challenging scenario 

for classical biological control, as seen here in the Pisgah National Forest where 300 L. nigrinus were 

released on 12 Jan and 13 Mar 2005. The beetle established but was not able to prevent these trees 

from dying. The effect of the beetle on surviving trees at the site and future tree cohorts remains to be 

determined (photos by D.L. Mausel and G.A. Davis). 

C. Scannell, D. Hamilton, J. Demeglio, 

L. Mackasmiel, S. Lyons, R. Herrick, C. Urquart, 

C. Huffman, and K. Foley for assistance with 

various laboratory and field experiments; L. Solter, 

A. Lawrence, and J. Becnel for entomopathogen 

screening; N. Havill, E. Day, N. Vanderburg, 

M. Kosztarab, and M. Stoetzel for insect identification; A. Abdel-Salam, T. Coleman, J. Birch, 

M. Lavine for statistical consulting; H. Cowart, 

L. Barnes, M. Dalusky, L. Randolth, J. Compton, 

G. Taylor, S. Skeate, R. McDonald, C. Th omas, 

P. Gilliland, R. Turcotte, T. Elliott, S. Hutchinson, 

K. Kish, M. Fugate, M. Bing, R. Rabaglia, S. Tilley, 

B. Handley, T. Lupp, B. Onken, A. Soir, 

M. Blumenthal, J. Unger, S. Werner, B. Regester, 

B. Laubscher, A. Paradis, K. Gooch, C. Burnham, 

A. Kanoti, C. Teerling, J. Weimer, K. Lombard, 

B. Burns, and J. Esden for locating release sites 

and/or post-release monitoring; and all the 

agencies and staff involved in program releases and 

monitoring. Funding and in-kind support was 

provided by the USDA-Forest Service: Forest Health 

Protection, Forest Health Technology Enterprise 

Team, and Special Technology Development 

Program; USDA-APHIS; Friends of the Blue 

Ridge Parkway; Seattle City Parks and Recreation; 

and the National Biological Control Institute. 

85 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Figure 6. 	 Typical eastern hemlock canopy decline and mortality is slower in the mid-Atlantic and northeast 

States and presents a less challenging scenario for classical biological control, as seen here in the 

Rothrock State Forest where 300 L. nigrinus were released on 4 Dec 2003 and 20 Apr 2004. The beetle 

established, yet tree decline does not appear to have reversed course, to date (photos by D.L. Mausel 

and G.A. Davis). 

86 
Chapter 6: Laricobius nigrinus Fender (Coleoptera: Derodontidae) 

LITERATURE CITED 

Bright, D.E. 1991. Superfamily Dermestoidea, 

family Derodontidae, tooth-necked fungus 

beetles. In: Bousquet, Y., ed. Checklist of Beetles 

of Canada and Alaska. Agriculture Canada 

Publication 1861/E: 195-196. 

Broeckling, C.D.; Salom, S.M. 2003. Antennal 

morphology of two specialist predators 

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Callahan, P.M.; Merickel, F.W.; Cook, S.P.; Fins, 

L. 2008. First report of four beetle species 

(Coleoptera) (Podapion gallicola Riley) 

(Brentidae), Cimberis turbans Hustache 

(Nemonychidae), Chrysobothris californica 

Leconte (Buprestidae) and Laricobius nigrinus 

Fender (Derodontidae) in Idaho. Th e Pan-Pacifi c 

Entomologist 84: 117-120. 

Downie, N.M.; Arnett, R.H. 1996. The Beetles of 

Northeastern North America. Gainesville, FL: 

The Sandhill Crane Press: 1721 p. 

Fender, K.M. 1945. A new Laricobius from Oregon. 

Th e Pan-Pacifi c Entomologist 21: 152. 

Flowers, R.W.; Salom, S.M.; Kok, L.T. 2006. 

Competitive interactions among two specialist 

predators and a generalist predator of hemlock 

woolly adelgid, Adelges tsugae (Hemiptera: 

Adelgidae) in south-western Virginia. Agriculture 

and Forest Entomology 8: 253-262. 

Flowers, R.W.; Salom, S.M.; Kok, L.T.; Mullins, 

D.E. 2007. Behavior and daily activity patterns 

of specialist and generalist predators of the 

hemlock woolly adelgid, Adelges tsugae. Journal 

of Insect Science 7: 44. 

Franz, J.M. 1958. Studies on Laricobius erichsonii 

Rosenh. (Coleoptera: Derodontidae) a predator 

on Chermesids. Part I, distribution, life-history, 

and ecology. Entomophaga 3: 109-164. 

Furniss, R.L.; Carolin, V.M. 1977. Western Forest 

Insects. USDA Forest Service Miscellaneous 

Publication No. 1339. Washington, DC: 654 p. 

Hatch, M.H. 1962. The family Derodontidae. Part 

III. Pselaphidae and Diversicornia I. In: Th e 

Beetles of the Pacifi c Northwest. Ed. Seattle, WA: 

University of Washington Press: 276-278. 

Havill, N.P.; Montgomery, M.E.; Yu, G.Y.; Shiyake, 

S.; Caccone, A. 2006. Mitochondrial DNA 

from hemlock woolly adelgid (Hemiptera: 

Adelgidae) suggests cryptic speciation and 

pinpoints the source of the introduction 

to eastern North America. Annals of the 

Entomological Society of America 99: 195-203. 

Havill, N.P.; Foottit, R.G. 2007. Biology and 

evolution of Adelgidae. Annual Review of 

Entomology 52: 325-349. 

Huffaker, C.B. 1974. Some ecological roots of pest 

control. Entomophaga 19: 371-389. 

Humble, L.M. 1994. Recovery of additional exotic 

predators of balsam woolly adelgid, Adelges 

piceae (Ratzeburg) (Homoptera: Adelgidae), 

in British Columbia. The Canadian Entomologist 

126: 1101-1103. 

Humble, L.M.; Mavin, L. 2005. Preliminary 

assessment of the cold tolerance of Laricobius 

nigrinus, a winter-active predator of the 

hemlock woolly adelgid from western Canada. 

In: Onken, B.; Reardon, R., comps. Th ird 

Symposium on Hemlock Woolly Adelgid in the 

Eastern United States; 2007 February 1-5; 

Asheville, NC. FHTET-2005-1. Morgantown, 

WV: U.S. Department of Agriculture, Forest 

Service, Forest Health Technology Enterprise 

Team: 304. 

Hunt, T.; Bergsten, J.; Levkanicova, Z.; 

Papadopoulou, A.; John, O.S.; Wild, R.; 

Hammond, P.M.; Ahrens, D.; Balke, M.; 

Caterino, M.S.; Gómez-Zurita, J.; Ribera, I.; 

Barraclough, T.G.; Bocakova, M.; Bocak, L.; 

Vogler, A.P. 2007. A comprehensive phylogeny 

of beetles reveals the evolutionary origins of a 

superradiation. Science 318: 1913-1916. 

Klein, J.L.; Caccone, A.; Havill, N.P. [In press]. 

Polymorphic microsatellite loci for Laricobius 

nigrinus and L. rubidus (Coleoptera: 

Derodontidae), predators of the hemlock woolly 

adelgid. Molecular Ecology Resources. 

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Kohler, G.R.; Stiefel, V.L.; Wallin, K.F.; Ross, D.W. 

2008. Predators associated with the hemlock 

woolly adelgid (Hemiptera: Adelgidae) in the 

Pacifi c Northwest. Environmental Entomology 

37: 494-505. 

Lamb, A.B.; Salom, S.M.; Kok, L.T. 2005. Survival 

and reproduction of Laricobius nigrinus Fender 

(Coleoptera: Derodontidae), a predator of 

hemlock woolly adelgid, Adelges tsugae Annand 

(Homoptera: Adelgidae) in fi eld cages. Biological 

Control 32: 200-207. 

Lamb, A.B.; Salom, S.M.; Kok, L.T.; Mausel, D.L. 

2006. Confi ned field release of Laricobius 

nigrinus (Coleoptera: Derodontidae), a predator 

of the hemlock woolly adelgid, Adelges tsugae 

(Hemiptera: Adelgidae), in Virginia. Canadian 

Journal of Forest Research 36: 369-375. 

Lamb, A.B.; Salom, S.M.; Kok, L.T. 2007. Factors 

influencing aestivation in Laricobius nigrinus 

(Coleoptera: Derodontidae), a predator of 

Adelges tsugae (Hemiptera: Adelgidae). Canadian 

Entomologist 139: 576-586. 

Lawrence, J.F. 1989. A Catalog of the Coleoptera of 

America North of Mexico, Family: Derodontidae. 

Agriculture Handbook 529-65. Washington, 

DC: U.S. Department of Agriculture, 

Agricultural Research Service: 5 p. 

Leschen, R.A.B. 2000. Beetles feeding on bugs 

(Coleoptera: Hemiptera): repeated shifts from 

mycophagous ancestors. Invertibrate Taxonomy 

14: 917-929. 

Leschen, R.A.B. [In press]. World review of 

Laricobius (Coleoptera: Derodontidae). Zootaxa. 

Mausel, D.L. 2005. Observations on fecundity and 

natural enemies of hemlock woolly adelgid, 

Adelges tsugae Annand (Hemiptera: Adelgidae) 

in Seattle, Washington. Pan-Pacifi c Entomologist 

81: 97-98. 

Mausel, D.L. 2007. Release and Monitoring of 

Laricobius nigrinus (Coleoptera: Derodontidae) for 

Biological Control of the Hemlock Woolly Adelgid 

in the Eastern US. Blacksburg, VA: Virginia 

Tech: 153 p. Ph.D. dissertation. 

Mausel, D.L.; Salom, S.M.; Kok, L.T.; Fidgen, 

J.G. 2008. Propagation, synchrony, and impact 

of introduced and native Laricobius spp. 

(Coleoptera: Derodontidae) on hemlock woolly 

adelgid in Virginia. Environmental Entomology 

37: 1498-1507. 

Mausel, D.L.; Salom, S.M.; Kok, L.T.; Davis, 

G. 2010. Establishment of the hemlock 

woolly adelgid predator, Laricobius nigrinus 

(Coleoptera: Derodontidae), in the eastern 

United States. Environmental Entomology 39: 

440-448. 

Mausel, D.L.; Salom, S.M.; Kok, L.T. 

[In press]-a. Visual ability and searching 

behavior of the adelgid predatory beetle, 

Laricobius nigrinus. Journal of Insect Science. 

Mausel, D.L.; Van Driesche, R.G.; Elkinton, 

J.S. [In press]-b. Comparative cold tolerance 

and climate matching of coastal and inland 

Laricobius nigrinus (Coleoptera: Derodontidae), 

a biological control agent of hemlock woolly 

adelgid. Biological Control. 

McClure, M.S.; Cheah, C.A.S.-J. 1999. Reshaping 

the ecology of invading populations of hemlock 

woolly adelgid, Adelges tsugae (Homoptera: 

Adelgidae), in eastern North America. Biological 

Invasions 1: 247-254. 

Montgomery, M.E.; Lyon, S.M. 1996. Natural 

enemies of adelgids in North America: their 

prospect for biological control of Adelges tsugae 

(Homoptera: Adelgidae). In: Salom, S.M.; 

Tigner, T.C.; Reardon, R.C., comps. Proceedings 

of the First Hemlock Woolly Adelgid Review; 

1995 October 12; Charlottesville, VA. 

FHTET-96-10. Morgantown, WV: U.S. 

Department of Agriculture, Forest Service, 

Forest Health Technology Enterprise Team: 

89-102. 

Wallace, M.S.; Hain, F.P. 2000. Field surveys and 

evaluation of native and established predators 

of the hemlock woolly adelgid (Homoptera: 

Adelgidae) in the southeastern United States. 

Environmental Entomology 29: 638-644. 

88 
Chapter 6: Laricobius nigrinus Fender (Coleoptera: Derodontidae) 

Zilahi-Balogh, G.M.G.; Kok, L.T.; Salom, S.M. 

2002. Host specificity of Laricobius nigrinus 

Fender (Coleoptera: Derodontidae), a potential 

biological control agent of the hemlock woolly 

adelgid, Adelges tsugae Annand (Homoptera: 

Adelgidae). Biological Control 24: 192-198. 

Zilahi-Balogh, G.M.G.; Humble, L.M.; Lamb, 

A.B.; Salom, S.M.; Kok, L.T. 2003a. Seasonal 

abundance and synchrony between Laricobius 

nigrinus (Coleoptera: Derodontidae) and its 

prey, the hemlock woolly adelgid (Hemiptera: 

Adelgidae). The Canadian Entomologist 135: 

103-115. 

Zilahi-Balogh, G.M.G.; Salom, S.M.; Kok, L.T. 

2003b. Temperature-dependent development 

of the specialist predator Laricobius nigrinus 

(Coleoptera: Derodontidae). Environmental 

Entomology 32: 1322-1328. 

Zilahi-Balogh, G.M.G.; Salom, S.M.; Kok, L.T. 

2003c. Development and reproductive biology 

of Laricobius nigrinus, a potential biological 

control agent of Adelges tsugae. BioControl 48: 

293-306. 

Zilahi-Balogh, G.M.G.; Humble, L.M.; Kok, L.T.; 

Salom, S.M. 2006. Morphology of Laricobius 

nigrinus (Coleoptera: Derodontidae), a predator 

of the hemlock woolly adelgid. Th e Canadian 

Entomologist 138: 595-601. 

89 
 

CHAPTER 7: LARICOBIUS OSAKENSIS, 


A HEMLOCK WOOLLY ADELGID PREDATOR FROM JAPAN
 

Ashley Lamb1, Michael E. Montgomery2, Ligia Cota Vieira3, Shigehiko Shiyake4, and Scott Salom3 

1

University of Tennessee, Department of Entomology and Plant Pathology, Knoxville, TN
 

2

U.S. Forest Service, Northern Research Station, Hamden, CT
 

3

Virginia Tech, Entomology Department, Blacksburg, VA
 

4

Osaka Museum of Natural History, Higashi Sumiyoshi-ku, Osaka, Japan
 

INTRODUCTION 

The approach for the biological control of hemlock 

woolly adelgid (HWA), Adelges tsugae Annand, 

has been to release multiple species of host-specifi c 

predators in order to reduce HWA populations 

below damaging thresholds. Beetles in the genus 

Laricobius prey excusively on adelgids and have 

life histories matched closely to that of their prey 

(Lawrence and Hlavac 1979). Thus, this group has 

high potential for biological control of adelgids. 

The genus has 21 recognized species—1 native to 

Europe, 3 native to North America, and 17 native 

to Asia (Leschen 2011). The European species, 

L. erichsonii Rosenhauer was released widely 

in North America for biological control of the 

balsam woolly adelgid, A. piceae (Ratzenburg), and 

although it initially established, it did not thrive 

and disappeared (Schooley et al. 1984). Th e only 

Laricobius species native to eastern United States, 

the target area for biological control of HWA, is 

L. rubidus Leconte, a predator of pine bark adelgid, 

Pineus strobi (Hartig). A western North American 

species and known predator of HWA, L. nigrinus 

Fender, has recently been introduced in the eastern 

United States (Mausel et al. 2010). Because of the 

presence of two congeners in the eastern United 

States, it is important that distinctions between the 

species and how they would interact is known prior 

to the introduction of another Laricobius species. 

Recently, Laricobius osakensis Shiyake and 

Montgomery was discovered in Japan (Shiyake 

et al. 2008, Montgomery et al. 2011). It is very 

promising for biological control because it comes 

from the same location as the source of the 

population of HWA in the eastern United States 

(Havill et al. 2006). Therefore, this species has 

evolved with the HWA strain aff ecting T. canadensis. 

This chapter will review information on its 

(1) identification, (2) the phenology and 

distribution of the predator in its native 

habitat and (3) experiments conducted in 

a quarantine laboratory to assess its hostspecificity and compatibility with a native 

and introduced congeneric species. 

IDENTIFICATION 

Laricobius osakensis, like other members of the 

family Derodontidae, is a small beetle between 

2-3 mm in length, elongate, dorsally convex and 

ventrally flattened, with a vestiture of fine hair, a 

head with large pores or canals and 11-segmented 

antennae with a 3 segment club, a narrow 

prothorax with esplanate sides, elytra seriate or 

striate, and an abdomen with five visible sternites. 

Like other members of the genus, it has strongly 

lobed tarsi, open procoxal cavities, and elyta with 

a scutellary striole and 10 rows of punctures. 

Distinguishing L. osakensis from the other two 

90 
 

 

Chapter 7: Laricobius osakensis, a Hemlock Woolly Adelgid Predator from Japan 

Laricobius species now in the eastern United States 

can be difficult in the field with live specimens, but 

fairly easy with dead, mounted specimens in the 

laboratory. A detailed description of L. osakensis 

can be found in Montgomery et al. (2011). 

Live adult L. osakensis have two color morphs—one 

has reddish elytrae with darker maculation on the 

lateral edge, similar to L. rubidus (Fig. 1A), whereas 

the other color form has nearly black (not piceous) 

elytrae, similar to L. nigrinus (Fig. 1B). When a 

group of beetles are sorted according to color and 

then dissected to confirm the sex by examination of 

their genitalia, about 90% of the reddish specimens 

were female, while about 80% of the dark-brown 

specimens were male. Following death, the reddish 

form becomes various shades of brown. The eggs are 

light yellow and about 50% larger than an HWA egg 

(Fig. 1C). The larvae are covered by white wax and 

usually feed inside the HWA woolly ovisac (Fig.1D). 

There are several characteristics, visible under a good 

stereomicroscope at high power, that can be used 

to distinguish L. osakensis from the other Laricobius 

species already present in the eastern United States. 

Most Derodontidae have two small ocelli on the 

head, each mesal of the compound eyes. Laricobius 

osakensis, L. taiwanensis Yu and Montgomery, 

and L. kangdingensis (Zilahi-Balogh and Jelenik 

2003) are exceptions to this rule and have no ocelli 

(Montgomery et al. 2011). Laricobius rubidus and 

L. nigrinus have small rudimentary ocelli. Careful 

Figure 1. Laricobius osakensis life stages. (A) Adult female, (B) adult male, (C) two eggs beside dead adult HWA, 

and (D) fourth instar lava feeding on HWA eggs. 

91 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

examination of the frons area of the head is the 

most reliable way to distinguish L. osakensis from L. 

rubidus and L. nigrinus (Fig. 2). The patterns of the 

deep pits and punctures on the head are variable and 

are not a sure way to separate these three species. 

There are small differences in the dimensions and 

shape of the pronotum of the three species, but these 

are difficult to discern (see Montgomery et al. 2011). 

If the genitalia are dissected and the specimen is a 

male, L. osakensis can readily be distinguished from 

the North American species by its narrow, acutely 

pointed parameres and slender median lobe (Fig. 3). 

Based on PCR-based molecular diagnostics, 

L. nigrinus and L. rubidus are closely related sister 

species but L. osakensis is in a separate clad with 

other Asian species (Montgomery et al. 2011). 

Molecular information is not only a way to identify 

adults, but can be also used to identify larvae. 

DISTRIBUTION 

Laricobius osakensis has been collected in 14 

prefectures on the Japanese islands of Honshu, 

Kyushu and Shikoku (Fig. 4) at elevations from 

80-1850 m on both hemlock species native to 

Japan. The southern Japanese hemlock, Tsuga 

sieboldii (Carrière), grows naturally at elevations 

between 200-1500 m on the southern half of 

Honshu and the more southern islands. Th e 

northern Japanese hemlock T. diversifolia (Maxim.) 

Figure 3. 	 Sexual organs of Laricobius males: (A) 

Laricobius osakensis, (B) L. rubidus, and 

(C) L. nigrinus. 

Masters grows at elevations from 700-2100 m, 

mostly in the more northern areas of Honshu 

Island. Both hemlocks are grown as ornamental 

or landscape trees, and HWA and the beetle are 

more common in landscape plantings or seminatural settings compared to forest sites. 

PHENOLOGY 

The seasonal history of L. osakensis and HWA was 

monitored in the Kansai area of Japan to predict 

its ability to thrive in the eastern United States. 

HWA-infested T. sieboldii trees in 3 prefectures 

were sampled weekly for 2 years. At each sample 

period, beat sheet samples were conducted at each 

Figure 2. (A) Laricobius rubidus and (B) L. nigrinus with arrows pointing to ocelli; (C) L. osakensis with arrows 

showing where ocelli are absent. 

92 
Chapter 7: Laricobius osakensis, a Hemlock Woolly Adelgid Predator from Japan 

Figure 4. 	 Map of Japan indicating the prefectures 

where Adelges tsugae has been collected 

(yellow shading) and locations where 

Laricobius osakensis has been collected 

(red dots). 

tree to determine the presence of adult predators. 

In addition, branch samples were removed from the 

trees and examined microscopically for immature 

stages of the predator and to determine the stages 

of HWA. Both species were present and active on 

the hemlock foliage during all months except for 

June through October (Fig. 5). L. osakensis adults 

first appeared on the trees in the Kansai region in 

mid-November, approximately one week after HWA 

had resumed feeding. These adults remained on the 

trees throughout the winter and early spring. In late 

December, adult L. osakensis began laying eggs in the 

woolly masses, in synchrony with HWA oviposition. 

L. osakensis continued to lay eggs throughout the 

winter and early spring. L. osakensis eggs hatched 

from January to April and larvae fed on HWA eggs. 

After developing through four instars, the larvae 

dropped from the hemlock branches in the early 

spring and pupated in the soil. L. osakensis aestivate 

as adults from May through October in the Kansai 

region. Phenology of L. osakensis is synchronized 

with the winter generation of HWA on T. sieboldii 

in Japan. Both species are present and active from 

Figure 5. The number of Laricobius osakensis adults (blue), eggs (red), and larvae (green) found at each sample 

period on Tsuga sieboldii in the Kansai region of Japan. 

93 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

November through May, feeding and ovipositing at 

the same time. In addition, both species undergo a 

dormant period throughout the summer months. 

The genotype of HWA in this area is identical to 

the genotype of HWA in the eastern United States 

(Havill et al. 2006). The seasonal temperatures 

of the study area in Japan are comparable to 

the temperatures of potential release areas in 

North America (Fig. 6). In the Kansai region of 

Japan, L. osakensis adults became active in midNovember, a little later than expected based on 

emergence in British Columbia (Zilahi-Balogh 

et al. 2003) and emergence of L. nigrinus and 

L. rubidus in Virginia (Mausel et al. 2008). Th e 

delay in emergence of L. osakensis in Japan is 

likely because HWA on T. sieboldii in Japan 

breaks dormancy slightly later, probably due to 

the extended summer at low latitudes in Japan. 

Phenology of L. osakensis on T. diversifolia at higher 

altitudes has not been studied in detail. Adults 

have been found on T. diversifolia in late October 

and early May. Trees were extensively sampled in 

early April, but adults were not collected. Since 

temperatures are very low and a large amount 

of snow falls in these high mountain areas, L. 

osakensis may not be active throughout the winter. 

Although more sampling is needed to determine 

the phenology of L. osakensis in colder regions, 

it’s clear that it can survive in areas with average 

winter minimum temperatures below -20° C. 

IMPACT ON HWA IN JAPAN 

Since Tsuga species in Japan do not appear to 

be injured by HWA, the impact of predatory 

species on HWA populations were of interest. 

A predator exclusion study was conducted at 2 

sites in the Kansai area of Japan. At one site, 48 

HWA-infested branches were chosen and adult 

predators were removed by beat-sheeting. Half of 

the branches were caged from January to April. In 

April, 2 branch samples (10 cm in length) were 

removed from each branch and the density of 

-5

0

5

10

15

20

25

Jan Feb Mar Apr May Jun. Jul Aug Sep Oct Nov Dec

Average Monthly Temperarure ( C)

Victoria, BriƟsh Columbia; 42 N,

70m

Takatsuki, Osaka, Japan; 35 N,

235m

° 

Figure 6. 	 Average monthly temperatures (°C) of seasonal phenology study areas of Laricobius nigrinus in British 

Columbia and L. osakensis in Japan compared to a representative, prospective release area for both 

species in the eastern United States. 

94 
 

 

Chapter 7: Laricobius osakensis, a Hemlock Woolly Adelgid Predator from Japan 

HWA was determined. On the caged branches 

(not exposed to predators), approximately half the 

HWA were alive and produced ovisacs, whereas 

only 5% of the adelgids were alive on the open 

branches. Thirty branches were selected at the 

second site. Half of these branches were caged 

after predators were removed by beat-sheeting. 

The cages were left on the branches from May 

until the following June. The branches that were 

caged for the 14-month period were heavily 

infested with adelgids (>3/cm), whereas branches 

left exposed had almost no adelgids (<0.1/cm). 

Additionally, uncaged branches were sampled 

for predators throughout the year. Laricobius 

osakensis was the primary predator during the 

winter months at both sites. During the spring 

and summer, many generalist predators were 

present on the trees (Coccinellidae, Cantharidae, 

Elateridae, Syrphidae, Reduviidae, and others) 

(Shiyake et al. 2008). One Coccinellid species 

of particular interest found in these samples 

was Sasajiscymnus tsugae (Sasaji and McClure), 

a predator that has been released extensively in 

eastern North America since 1997 (Cheah et al. 

2004). S. tsugae was collected fairly consistently 

on several, but not all, of the study trees during 

May and June. This beetle was not consistently 

present throughout the other months of the year. 

The exclosure experiment suggests that the 

predators in Japan contribute to population 

control of HWA in the Kansai region. Th e 

sampling for predators indicates that L. osakensis 

is the primary predator during the winter and 

early spring. In late spring and early summer, 

generalist predators can be abundant, especially 

on hemlock with dense HWA infestations. 

EVALUATION OF HOST RANGE AND 

DEVELOPMENT IN THE LABORATORY 

Development tests were conducted by placing L. 

osakensis eggs on hemlock with HWA or on an 

alternate host. Larvae were observed for survival and 

maximum developmental stage reached. Choice and 

no-choice tests were set up in petri dishes and the 

amount of host consumed and number of eggs laid 

on each host were determined. The alternate hosts 

used were: balsam woolly adelid (BWA), pine bark 

adelgid (PBA), eastern spruce gall adelgid (ESGA), 

Adelges abietis (Linneaus), woolly alder aphid 

(WAA), Paraprociphilus tessellates (Fitch), elongate 

hemlock scale (EHS), Fiorina externa Farris, and 

pine needle scale (PNS), Chionaspis pinifoliae 

(Fitch). In choice tests, single adults were put in 

petri dishes containing a known number of HWA 

and an alternate host. The amount of prey consumed 

and number of eggs laid on each were counted after 

7 days. In the no-choice tests, single adults were put 

in petri dishes with a known number of HWA or 

alternate host and the number of host consumed 

and eggs laid were determined after 5 days. 

Laricobius osakensis consumed more HWA and laid 

more eggs on HWA than on any other host in both 

the choice and no-choice tests. In addition, these 

predators were only able to complete development 

to adults on HWA. They were only able develop up 

to 4th instar larvae on the adelgid alternate hosts 

and no development occurred on the non-adelgid 

alternate hosts. The host range tests indicate that 

this predator is host-specific on HWA and does not 

develop on other species tested (Vieira et al. 2011). 

CURRENT STATUS IN UNITED STATES 

These data suggest this predator poses no risk to 

native fauna within the eastern United States. 

This information was provided in the petition to 

release L. osakensis from quarantine. In May 2010, 

this predator was issued a FONSI (Finding Of 

No Significant Impact) and is no longer required 

to be in quarantine. In October 2010, a large 

field collection of L. osakensis was brought to 

Virginia Tech for mass rearing. The University of 

Tennessee is also rearing a small colony. Further 

research will be conducted at Virginia Tech and 

based on the results, L. osakensis will be tested 

in the field. If field tests are favorable and large 

numbers can be reared, field releases of this beetle 

will begin. Given what we know about its life 

history, L. osakensis has the potential to contribute 

to biological control of HWA in the future. 

95 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

ACKNOWLEDGEMENTS 

The assistance of the Osaka Museum of Natural 

History for laboratory space and logistical support 

is greatly appreciated. Access to the locations where 

field sampling was conducted was provided by 

Kobe Municipal Arboretum (Hyogo Prefecture), 

Myo-on-ji Temple (Osaka Prefecture), Nara Park 

Management Office (Nara Prefecture), Kyoto 

University Kamigamo Experimental Station (Kyoto 

Prefecture). Rearing of L. osakensis at Virginia Tech 

was conducted by J. Beck, N. Snyder, K. Tourje, and 

N. Morris. Funding was provided to Virginia Tech 

by the U.S. Forest Service, Forest Health Protection. 

LITERATURE CITED 

Cheah, C.A.S.-J.; Montgomery, M.E.; Salom, 

S.M.; Parker, B.L.; Costa, S.; Skinner, M. 2004. 

Biological Control of Hemlock Woolly Adelgid. 

Reardon, R.; Onken, B., tech. coords. FHTET2004-04. Morgantown, WV: U.S. Department 

of Agriculture, Forest Service: 22 p. 

Havill, N.P.; Montgomery, M.E.; Yu, G.Y.; Shiyake, 

S.; Caccone, A. 2006. Mitochondrial DNA 

from hemlock woolly adelgid (Hemiptera: 

Adelgidae) suggests cryptic speciation and 

pinpoints the source of the introduction 

to eastern North America. Annals of the 

Entomological Society of America 99(2): 

195-203. 

Lawrence, J.F.; Hlavac, T.F. 1979. Review of the 

Derodontidae (Coleoptera: Polyphaga) with new 

species from North America and Chile. 

The Coleopterists Bulletin 33: 369-413. 

Leschen, R.A.B. 2011. World review of Laricobius 

(Coleoptera: Derodontidae). Zootaxa 2908: 

1-44. 

Mausel, D.L.; Salom, S.M.; Kok, LT.; Davis, G. 

2010. Establishment of the hemlock 

woolly adelgid predator, Laricobius nigrinus 

(Coleoptera: Derodontidae), in the eastern 

United States. Environmental Entomology 39(2): 

440-448. 

Mausel, D.L.; Salom, S.M.; Kok, L.T.; Fidgen, 

J.G. 2008. Propagation, synchrony, and impact 

of introduced and native Laricobius spp. 

(Coleoptera: Derodontidae) on hemlock woolly 

adelgid in Virginia. Environmental Entomology 

37: 1498-1507. 

Montgomery, M.E.; Shiyake, S.; Havill, N.P.; 

Leschen, R.A.B. 2011. A new species of 

Laricobius (Coleoptera: Derodontidae) from 

Japan with phylogeny and a key for native 

and introduced congeners in North America. 

Annuals of the Entomological Society of America 

104: 389-401. 

Schooley, H.O.; Harris, J.W.E.; Pendrel, B. 

1984. Adelges piceae (Ratz.), balsam woolly 

adelgid (Homoptera: Adelgidae). In: Kelleher, 

J.S.; Hulme, M.A., eds. Biological Control 

Programmes against Insects and Weeds in Canada 

1969-1980. Farnham Royal, Slough, England: 

Commonwealth Agricultural Bureax: 229-234. 

Shiyake, S.; Miyatake, Y.; Montgomery, M.E.; 

Lamb, A.B. 2008. Hemlock woolly adelgid 

phenology and predacious beetle community on 

Japanese hemlocks. In: Onken, B.; Reardon, R., 

comps. Fourth Symposium on Hemlock 

Woolly Adelgid in the Eastern United States; 

2008 February 12-14; Hartford, CT. FHTET2008-01. U.S. Department of Agriculture, 

Forest Service, Forest Health Enterprise Team: 

261-266. 

Vieira, L.C.; McAvoy, T.; Chantos, J.; Lamb, A.B.; 

Salom, S.M.; Kok, L.T. 2011. Host Range of 

Laricobius osakensis (Coleoptera: Derodontidae), 

a new biological control agent of hemlock 

woolly adelgid (Hemiptera: Adelgidae). 

Environmental Entomology 40: 324-332. 

Zilahi-Balogh, G.M.G.; Humble, L.M.; Lamb, 

A.B.; Salom, S.M.; Kok, L.T. 2003. Seasonal 

abundance and synchrony between Laricobius 

nigrinus (Coleoptera: Derodontidae) and its 

prey, the hemlock woolly adelgid (Hemiptera: 

Adelgidae). The Canadian Entomologist 135: 

103-115. 

96 
 

 

CHAPTER 8: CHAMAEMYIID PREDATORS OF THE
 

HEMLOCK WOOLLY ADELGID FROM THE PACIFIC NORTHWEST
 

Darrell W. Ross1, Stephen D. Gaimari2, Glenn R. Kohler3, Kimberly F. Wallin1,4,5, and Sarah M. Grubin1 

1

Department of Forest Ecosystems and Society, Oregon State University, Corvallis, OR
 

2

Plant Pest Diagnostics Branch, California Department of Food and Agriculture, Sacramento, CA
 

3

Washington Department of Natural Resources, Olympia, WA
 

4

Rubenstein School of Environment and Natural Resources, The University of Vermont, Burlington, VT
 

5

USDA Forest Service, Northern Research Station, South Burlington, VT
 

The family Chamaemyiidae (Diptera), known 

colloquially as “silver flies” due to the silvery-gray 

adult body color of most species, includes two 

subfamilies, Cremifaniinae and Chamaemyiinae, 

with 28 described genera and subgenera, and 

over 330 described species worldwide, over 1/3 of 

which are in the single subgenus Leucopis (Leucopis) 

Meigen. The family has a cosmopolitan distribution, 

but more than half the described species are 

Palearctic, at least in part. Both subfamilies occur 

in the Nearctic region, with only 10 of the genera 

present, and fewer than 100 described species, 

although three new genera are currently being 

described by the second author, and many new 

species are known. The taxonomy and systematics 

of the family, particularly the genus Leucopis, has 

been challenging due to their small size (1-4 mm) 

and similar appearances, with a lack of clear, external 

morphological characters to distinguish species 

in most genera (Bennett 1961; Brown and Clark 

1956a; Gaimari and Turner 1996a; Greathead 1995; 

McAlpine 1960, 1971, 1977, 1987; McAlpine and 

Tanasijtshuk 1972; Sluss and Foote 1971, 1973). 

Consequently, most species can only be determined 

by examination of the male genitalia, and females 

are usually not possible to identify without close 

association with males. Identification resources are 

most complete for the Palearctic region, where the 

fauna has been extensively studied (Tanasijtshuk 

1986), and Australia, where Tanasijtshuk (1996) 

has made a first pass at describing the fauna. 

Also, several new species of Nearctic Leucopis 

were described by Tanasijtshuk (2003, 2005, 

2006). At the genus level, several regional keys 

are available, including for the Nearctic region 

(McAlpine 1987), the whole New World (Gaimari 

2010), and the Palearctic region (McLean 

1998), although for the latter Cremifaniinae 

was treated as a separate family (Papp 1998). 

Larvae are known to attack members of all 

superfamilies of sternorrhynchous Hemiptera except 

Aleyrodoidea, but most commonly aphids, adelgids, 

scales, and mealybugs. As a group, they display a 

spectrum of feeding strategies from active predation 

within free-living prey colonies to completely 

sessile predation on eggs within a single coccoid 

egg sac. Gaimari (2010) provided a review of the 

known feeding habits and general biology for the 

family, which includes the following generalities 

about the feeding habits for certain genera. Th ese 

include: mealybugs in leaf sheaths of grasses by 

species of Chamaemyia Meigen (Tanasijtshuk 1970, 

Sluss and Foote 1973, Raspi 1983), Parochthiphila 

Czerny (Tanasijtshuk 1963, 1968; Raspi 1983), 

and Pseudodinia Coquillett (Barber 1985); adelgids 

on gymnosperms by most species of Neoleucopis 

Malloch (McAlpine 1971, Sluss and Foote 1973, 

Gaimari et al. 2007), Anchioleucopis Tanasijthuk 

(Tanasijtshuk 1997, 2001), and Cremifania Czerny 

(Delucchi and Pschorn-Walcher 1954, Clark and 

Brown 1962); scales and mealybugs on dicots 

97 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

by species of Melaleucopis Sabrosky (Beingolea 

1957) and Leucopis (Leucopella) Malloch (Malloch 

1927, James 1934, Gaimari and Raspi 2002); 

on eggs within ovisacs of coccoids by species of 

Echinoleucopis Gaimari and Tanasijtshuk (Griot 

1954, Gaimari and Tanasijtshuk 2001) and 

Leucopomyia Malloch (Malloch 1922, Tanasijtshuk 

1959, Babaev and Tanasijtshuk 1971, Kaydan 

et al. 2006). Other genera have less specialized 

feeding habits, including Leucopina Malloch 

which feed on the whole variety of coccoid hosts, 

and Leucopis (Leucopis) which feed on the entire 

range of available prey. Tanasijtshuk (1986) also 

provides considerable biological data, with host 

records for many of the Palearctic species. 

As a result of their specialization as predators 

of Sternorrhyncha, they have been studied as 

natural controls on herbivore populations and as 

biological control agents of pest insects in those 

groups. Biological control efforts against adelgid 

pests are detailed in this paper, but chamaemyiids 

have also been utilized against Russian wheat 

aphid, Diuraphis noxia (Mordvilko) (Hemiptera: 

Aphididae) in the Pacifi c Northwest (Gaimari 

and Turner 1996b, 1996c, 1997; Noma et al. 

2005), and one species, Leucopis verticalis Malloch, 

was recognized as an important predator in the 

galls of grape phylloxera Daktulosphaeira (as 

Phylloxera) vitifoliae (Fitch) (Stevenson 1967). 

Although the biology and ecology of most adelgidfeeding chamaemyiid species are poorly known, 

some generalizations can be drawn from those 

that have been studied (Amman and Spears 1971; 

Brown and Clark 1956b, 1957; Clark and Brown 

1957, 1962; Delucchi and Pschorn-Walcher 1954; 

Eichhorn 1968; Greathead 1995; Grubin 2011; 

Kohler 2007; Kohler et al. 2008a; McAlpine 1971, 

1978; McAlpine and Tanasijtshuk 1972; Mitchell 

1962; Sluss and Foote 1973; Smith 1958; Smith 

and Coppel 1957; Tanasijtshuk 1986, 2001, 

2002; Wilson 1938). Most species appear to have 

1-3 generations per year that are synchronized 

with the seasonal life history of their prey species. 

Larvae feed on eggs, nymphs, and adults of adelgid 

hosts with many species capable of feeding and 

developing on several species of adelgids. Often, 

pupariation and oviposition occurs near larval 

feeding sites, and the puparia are fi rmly attached 

to host trees. Overwintering is in the larval and, 

more commonly, puparial stages, and diapause 

may occur during the summer or winter. 

The potential value of chamaemyiids as biological 

control agents for adelgids has been recognized for a 

long time. Following the seminal work of Trägårdh 

(1931), detailing the life history and eff ectiveness of 

Neoleucopis obscura (Haliday) against Adelges picea 

(Ratzeburg) in Europe, this species became the 

first chamaemyiid introduced to control the exotic 

adelgid in North America in the 1930’s. In another 

case, following the accidental introduction of Pineus 

pini (Macquart) into Australia, Wilson studied the 

natural enemies of P. pini and Pineus strobi Hartig in 

England (Wilson 1938). He identifi ed Neoleucopis 

obscura as the most efficient predator of these species 

and suggested that it “...should be introduced 

[into Australia] at the earliest possible moment.” 

Subsequently, a number of chamaemyiid species 

have been considered for biological control of Pineus 

Shimer and Adelges Vallot species throughout the 

world with varying degrees of success (Greathead 

1995, Mills 1990, Zilahi-Balogh et al. 2002a). 

At least seven species of chamaemyiids were studied 

as biological control agents for balsam woolly 

adelgid, Adelges piceae, in North America, although 

the exact number released is unknown because 

of poor knowledge of the taxonomy of the group 

(McAlpine 1971, 1978). Chamaemyiid species were 

introduced into eastern Canada (Balch et al. 1956; 

Brown and Clark 1956a, 1956b, 1957; McAlpine 

1978; Smith 1958; Smith and Coppel 1957), 

North Carolina (Amman and Spears 1971), Oregon 

and Washington (Mitchell and Wright 1967), 

and British Columbia (Harris and Dawson 1979, 

Humble 1994, McAlpine 1978). With the exception 

of North Carolina, one or two of the following four 

species became established in each geographic area: 

Cremifania nigrocellulata Czerny, Neoleucopis obscura, 

Neoleucopis atratula (Ratzeburg), and Leucopis 

hennigrata McAlpine (Balch et al. 1956; Brown and 

Clark 1956b, 1957; McAlpine 1971, 1978; Mitchell 

98 
 

 

Chapter 8: Chamaemyiid Predators of the Hemlock Woolly Adelgid from the Pacifi c Northwest 

and Wright 1967; Humble 1994; Schooley et al. 

1984). Although none of these species signifi cantly 

reduced A. piceae populations or associated damage 

to host trees, several developed large populations 

and dispersed rapidly. Neoleucopis obscura reportedly 

dispersed an average of 6 miles per generation 

in eastern Canada (Brown and Clark 1957), and 

within 14 years had spread throughout the infested 

areas of the Maritime Provinces (Brown 1947, Balch 

et al. 1956) and into Maine (Thomas 1968), and 

was recovered about one mile from a release point 

in the Pacific Northwest (Mitchell and Wright 

1967). Cremifania nigrocellulata spread 2-3 miles 

from release points in 3-5 years, and was found to 

begin feeding slightly earlier than Neoleucopis obscura 

(Balch et al. 1956). Collectively, the inability of 

these chamaemyiid species to signifi cantly impact 

A. piceae populations was attributed to feeding 

too late in the host developmental period, limited 

searching ability of larvae, slow rate of population 

increase relative to the prey population, higher rate 

of overwintering mortality than the prey, inability to 

persist in light infestations, and/or lack of seasonal 

synchrony of predator and prey populations. 

Although less well documented, some other 

attempts to use chamaemyiids for biological control 

of adelgids have apparently been more successful 

than those with A. piceae. In Hawaii, there was a 

strong correlation between densities of Neoleucopis 

tapiae (Blanchard) (originally reported as Leucopis 

obscura) and an introduced adelgid, Pineus pini over 

a two-year period 3-5 years after introduction of the 

predator (Culliney et al. 1988, Greathead 1995). In 

New Zealand, Neoleucopis tapiae (originally reported 

to be Leucopis obscura) was responsible for the 

control of Pineus boerneri Annand on Pinus radiata 

D. Don (Rawlings 1958, Zondag and Nuttall 1989). 

And, in Chile, Neoleucopis obscura (which may also 

be Neoleucopis tapiae; see Greathead 1995) provided 

effective control of P. boerneri (Francke-Grosmann 

1963; Zúñiga 1985). In all three of these cases, the 

target Pineus species feed primarily on twigs near 

the base of needles, unlike A. piceae which forms 

dense colonies on the main bole of the host trees. It 

may be that chamaemyiids are more important in 

the population dynamics of twig feeding adelgids 

than those that feed on the bole. However, attempts 

in the 1970’s to introduce Neoleucopis tapiae, N. 

nigraluna McAlpine and Leucopis spp. into Kenya 

for control of the twig feeding Pineus boerneri on 

P. radiata and Pinus patula Schiede ex Schltdl. & 

Cham. were unsuccessful (Greathead 1995, Day et 

al. 2003). In this case, Greathead (1995) speculated 

that the numbers of insects released may have been 

too small to ensure establishment. Day et al. (2003) 

also reported that two other species, Neoleucopis 

manii Tanasijtshuk and Leucopis argenticollis 

Zetterstedt, were imported into Kenya but never 

released, although they may have been released in 

Tanzania along with the species released in Kenya. 

The predator community associated with hemlock 

woolly adelgid, Adelges tsugae Annand, infested 

western hemlock, Tsuga heterophylla (Raf.) Sarg., 

in Oregon and Washington was studied by 

Kohler et al. (2008a). In surveys of 116 infested 

trees at 16 sites from January 2005 to November 

2006, chamaemyiids were the second most 

abundant group of predators to the Derodontidae 

(Coleoptera), predominantly Laricobius nigrinus 

Fender. In total, 1,039 adult and immature 

Leucopis spp. were collected compared to 2,723 

derodontids and 531 Hemerobiidae (Neuroptera), 

the third most abundant group. Since we do not 

know the efficiency of beat sampling used to 

collect these specimens, we do not know whether 

this represents the true differences in abundance 

among these predator species. However, the ratio 

of immatures to adults was over three times higher 

for the chamaemyiids (9.2) compared to the 

derodontids (2.6) or hemerobiids (3.1) suggesting 

that beat sampling was less efficient at collecting 

adult chamaemyiids, and that chamaemyiids are 

more abundant relative to the other predators 

than indicated by the counts from beat samples. 

Two species of Leucopis were collected in samples 

from A. tsugae-infested western hemlock, L. 

argenticollis and L. piniperda Malloch (as L. atrifacies 

Aldrich) (Kohler et al. 2008a). Unfortunately, 

the misidentifi cation as L. atrifacies by S.D. 

Gaimari was not recognized until he began to 

study the more extensive materials collected by 

99 
 

 

 

 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Grubin (2011). This was the first record of either 

L. argenticollis or L. piniperda associated with A. 

tsugae. Eighty-seven percent of the chamaemyiids 

that were collected as adults or reared to the adult 

stage were L. argenticollis, which is a Holarctic 

species previously found in colonies of several 

Pineus species in Russia, India, Japan, Canada, 

and the United States (McAlpine and Tanasijtshuk 

1972). Leucopis piniperda is widespread in North 

America, and has been collected from colonies of 

several species of Adelges and Pineus (Tanasijtshuk 

2002). Although never collected associated 

with A. tsugae, Leucopis atrifacies is restricted to 

western North America where it has been found 

associated with several species of Pineus and Adelges 

(Greathead 1995, Tanasijtshuk 2002). Following 

the introduction of A. piceae into North America, 

L. argenticiollis was found associated with A. piceae 

in eastern Canada (McAlpine and Tanasjitshuk 

1972), and both L. atrifacies and L. piniperda 

were found associated with A. piceae in western 

North America (Tanasjitshuk 2002). Th is indicates 

that all three species are capable of searching for, 

locating, and potentially utilizing novel adelgid 

prey species within their native habitats. 

Since Kohler et al. (2008a) were unable to 

distinguish L. argenticollis and L. piniperda larvae, it 

was not possible to record separate observations of 

the biology for each species except in the cases where 

they were reared to the adult stage. There were two 

peaks in larval abundance, one in the spring and 

one in the early summer coinciding with the two 

periods of A. tsugae oviposition, suggesting that 

these Leucopis spp. have two generations that are 

synchronized with the A. tsugae life cycle. Leucopis 

spp. larvae that were brought to the lab fed on 

eggs and nymphs of the progrediens and sistens 

generations of A. tsugae. Puparia were always fi rmly 

attached to the twigs near the larval feeding sites. 

Immature L. argenticollis that were collected in 

November 2005 and held on A. tsugae infested twigs 

in an environmental chamber simulating outdoor 

conditions did not emerge as adults for four months. 

Larvae of both Leucopis spp. that pupariated in 

the spring emerged as adults within 4 weeks. 

Nonparametric analyses of insect community 

structure demonstrated a strong positive correlation 

between both Leucopis spp. larvae and L. argenticollis 

adult abundance and A. tsugae population density 

(Kohler et al. 2008a, 2008b). Among the nine 

predator taxa that were positively correlated to 

A. tsugae population density, the strength of 

the correlation with Leucopis spp. larvae and L. 

argenticollis adults was similar to or greater than 

that of all the others including Laricobius nigrinus 

adults and Laricobius spp. larvae. However, these 

data do not distinguish whether the Leucopis 

spp. are regulating A. tsugae populations or 

simply reaching high densities where their prey 

are abundant. Further studies will be needed to 

elucidate the nature of the ecological relationships 

among Leucopis spp. and A. tsugae populations. 

In 2009 and 2010, several no-choice feeding trials 

were conducted with Leucopis spp. larvae collected 

in Oregon and Washington from A. tsugae-infested 

western hemlock to determine the suitability of 

alternative adelgid prey (Grubin 2011). Survival 

was always higher on A. tsugae than other species 

of Pineus and Adelges, although the diff erences 

were not always statistically signifi cant. Also, 

some Leucopis spp. survived to the adult stage on 

each of the four alternative adelgid prey species. 

Thirty-eight of the Leucopis spp. larvae survived 

to the adult stage and were identified to species. 

Seventy-one percent were L. argenticollis and 

the remaining 29% were Leucopis piniperda. It is 

worth noting that only 15% of the larvae used 

in the feeding trials survived to the adult stage. 

Although species of native and introduced 

chamaemyiids are found in eastern North America, 

none have been shown to signifi cantly impact 

A. tsugae populations (Montgomery and Lyon 

1996, Wallace and Hain 2000). The two species 

of Leucopis found associated with A. tsugae in the 

Pacifi c Northwest, L. argenticollis and L. piniperda, 

also occur in eastern North America (McAlpine 

and Tanasijtshuk 1972, Tanasijtshuk 2002). 

However, populations of these predators in such 

widely separated geographic regions with diff erent 

native hosts (e.g., A. tsugae is native to the West, 

100 
 

 

 

 

Chapter 8: Chamaemyiid Predators of the Hemlock Woolly Adelgid from the Pacifi c Northwest 

but introduced in the East; Havill et al. 2006) are 

likely distinct. Consequently, the introduction of 

individuals from Pacific Northwest populations 

of these Leucopis spp. to the eastern United States 

may be beneficial to the ongoing biological 

control efforts for A. tsugae. Furthermore, the wide 

geographic range of these species suggests that 

individuals from the West would be able to adapt 

quickly to environments in the East (Mills 1990). 

Prior to the introduction of non-native 

chamaemyiids in the Pacific Northwest for 

biological control, Mitchell (1962) found three 

unidentified, native “Leucopis” spp. (one of which 

is Neoleucopis ancilla McAlpine (McAlpine 1971)) 

associated with A. piceae. All three of these species 

were generally found at low densities, but, at one 

site, one species (not Neoleucopis ancilla) was found 

in the “...heaviest predator population ever seen 

associated with the balsam woolly adelgid” (Mitchell 

1962), although this species only seemed to have 

one generation per year. The presence of these native 

species in A. piceae colonies, in some cases at high 

densities, within 30 years of the discovery of this 

introduced species suggests that they have vagile 

host searching behaviors and wide diet breadth 

within the Adelgidae. Other records of the host 

ranges of adelgid-feeding chamaemyiids would 

suggest that these are common attributes among the 

group (McAlpine 1971, McAlpine and Tanasijtshuk 

1972, Tanasijtshuk 2002). More recently, 

Neoleucopis tapiae were collected from both A. piceae 

on grand fi r, Abies grandis (Douglas ex D. Don) 

Lindl., and Pineus coloradensis (Gillette) on western 

white pine, Pinus monticola Douglas ex D. Don, 

in western Oregon in 2007 (Kohler 2007, G.R. 

Kohler, unpublished data). The ability to utilize 

several adelgid species as prey would allow potential 

biological control agents to survive at times and in 

places where their primary host was unavailable. 

There are many known parasitoids of 

Chamaemyiidae, particularly in the Chalcidoidea, 

where 26 species of 13 genera in fi ve families 

(Aphelinidae, Encyrtidae, Eulophidae, Pteromalidae, 

Signiphoridae) have been reported (Noyes 2010), 

including at least nine species of the pteromalid 

genus Pachyneuron Walker. Several parasitioids 

have been reared from the adelgid-feeders, which 

could potentially interfere with their eff ectiveness 

as biological control agents. Wilson (1938) reared 

Dendrocerus carpenteri (Curtis) (Hymenoptera: 

Megaspilidae) (as Lygocerus testaceimanus Kieff er), 

Amblynotus longitarsus Reinhard (Hymenoptera: 

Cynipidae), and Syrphophagus aphidivorus (Mayr) 

(Hymenoptera: Encyrtidae) (as Aphidencyrtus 

aphidivorus) from puparia of Neoleucopis obscura 

collected in England. Pachyneuron altiscutum 

Howard (Hymenoptera: Pteromalidae) was reared 

from puparia of native and introduced chamaemyiid 

species in eastern Canada (Brown and Clark 1956b, 

1957), with parasitism of Neoleucopis obscura by P. 

altiscutum at 21.3% across two generations. Puparia 

of the native species Neoleucopis pinicola (Malloch) 

were parasitized by Pachyneuron virginicum Girault 

in northeastern Ohio (Sluss and Foote 1973), and 

in eastern Canada by P. altiscutum (Brown and 

Clark 1956b) and Melanips iowensis Ashmead 

(Hymenoptera: Figitidae) (Clark and Brown 

1957). In Oregon and Washington, Mitchell 

(1962) reared unidentified species of Pachyneuron 

and Dendrocerus Ratzeburg (as Lygocerus Förster) 

from Neoleucopis ancilla (as Leucopis sp. I) and 

one of the additional unidentifi ed Leucopis species 

collected from A. piceae colonies, and Kohler et 

al. (2008b) reared two Pachyneuron spp. and a 

species of Melanips Giraud from Leucopis spp. 

puparia collected from A. tsugae-infested western 

hemlock. In the latter study, the parasitism rate 

was 21-23% over two years. If Leucopis spp. from 

the Pacific Northwest are ever introduced in the 

eastern United States, care should be taken to avoid 

introducing associated parasitoids by releasing 

only adults or immatures reared in parasitoidfree colonies. Furthermore, any Pacifi c Northwest 

Leucopis spp. populations that become established 

in the eastern United States should be monitored 

to assess parasitism rates by indigenous parasitoids. 

Successful biological control of A. tsugae in the 

eastern United States will likely require a suite of 

predators (Montgomery and Lyon 1996, Cheah et 

al. 2004). In addition to the predators that have 

already been released, including Laricobius nigrinus 

101 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

and two Coccinellidae (Coleoptera), Sasajiscymnus 

tsugae (Sasaji and McClure) and Scymnus 

sinuanodulus Yu and Yao, it is likely that other 

candidate biological control agents will need to be 

identified, tested, and released to achieve ecologically 

and economically significant reductions of A. tsugae 

populations. Since Leucopis spp. found in the Pacifi c 

Northwest feed on both progrediens and sistens 

eggs and nymphs, they will likely compliment 

Laricobius nigrinus imported from the same region 

which feeds only on progrediens eggs and nymphs 

(Kohler et al. 2008a, Zilahi-Balogh et al. 2002b). 

However, species of Scymnus Kugelann may prey on 

Leucopis spp. larvae and reduce their eff ectiveness 

(Mills 1990). Given the widespread occurrence 

and abundance of Leucopis spp. associated with 

A. tsugae in the Pacific Northwest, they should be 

studied in more detail to understand their roles in 

the population dynamics of A. tsugae in the West 

as well as their potential for biological control in 

the East. In particular, it would be valuable to 

develop rearing techniques to facilitate studies of 

the biology and ecology of each of the species that 

have been found in association with A. tsugae. 

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(Chamaemyiidae)]. Fauna of the USSR, new 

series 134, Dipterans. Volume 14, 7. Nauka, St. 

Petersburg: Zoological Institute of the Russian 

Academy of Sciences: 336 p. [in Russian]. 

Tanasijtshuk, V.N. 1996. Silver-fl ies (Diptera, 

Chamaemyiidae) of Australia. International 

Journal of Dipterological Research 7: 1-62. 

Tanasijtshuk, V.N. 1997. A new Holarctic genus of 

Chamaemyiidae (Diptera). International Journal 

of Dipterological Research 8: 113-116. 

Tanasijsthuk, V.N. 2001. On two sister species of 

the genus Anchioleucopis Tanasijtshuk, 1997 

(Diptera, Chamaemyiidae) from the Holarctic 

Region. Entomologicheskoe Obozrenie 80(4): 

901-907. [in Russian; English translation in 

2001, Entomological Review 81(5): 542-547]. 

Tanasijtshuk, V.N. 2002. Studies on Nearctic species 

of Leucopis (Diptera: Chamaemyiidae). I. Th e 

redescription of Nearctic Leucopis published 

before 1965. Zoosystematica Rossica 11: 193-207. 

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of Leucopis (Diptera: Chamaemyiidae). II. New 

species. Zoosystematica Rossica 11: 375-396. 

Tanasijtshuk, V.N. 2005. Studies on Nearctic species 

of Leucopis (Diptera: Chamaemyiidae). III. L. 

annulipes Zetterstedt and a new sibling species 

from the Nearctic. Zoosystematica Rossica 13(2): 

233-238. 

Tanasijtshuk, V.N. 2006. Studies on Nearctic species 

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species. Zoosystematica Rossica 14(2): 281-288. 

Thomas H.A. 1968. Distribution of the balsam 

woolly aphid predator, Leucopis obscura, in 

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Fliegen. Zeitschrift für Angewandte Entomologie 

18: 672-690. 

Wallace, M.S.; Hain, F.P. 2000. Field surveys and 

evaluation of native and established predators 

of the hemlock woolly adelgid (Homoptera: 

Adelgidae) in the southeastern United States. 

Environmental Entomology 29: 638-644. 

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Chermes with particular reference to Pineus 

pini, Koch, and Pineus strobe, Hartig. Bulletin 

of Entomological Research 29: 373-389. 

Zilahi-Balogh, G.M.G.; Kok, L.T.; Salom, S.M. 

2002a. A review of the world-wide biological 

control efforts for the family Adelgidae. In: 

Onken, B.; Reardon, R.C.; Lashcomb, J., eds. 

Proceedings: Hemlock Woolly Adelgid in the 

Eastern United States Symposium; 2002 February 

5-7; East Brunswick, NJ. New Brunswick, NJ: 

Rutgers New Jersey Agricultural Experiment 

Station: 129-140. 

Zilahi-Balogh, G.M.G.; Kok, L.T.; Salom, S.M. 

2002b. Host specifi city of Laricobius nigrinus 

Fender (Coleoptera: Derodontidae), a potential 

biological control agent of the hemlock woolly 

adelgid, Adelges tsugae Annand (Homoptera: 

Adelgidae). Biological Control 24: 192-198. 

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(Maskell), pine twig chermes or pine woolly 

aphid (Homoptera: Adelgidae). In: Cameraon, 

P.J.; Hill, R.L.; Bain, J.; Th omas, W.P., eds. 

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Pests and Weeds in New Zealand 1874 to 1987. 

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Oxon, United Kingdom: CAB International 

Institute of Biological Control: 295-297. 

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in Chile. Historical review and evaluation of the 

projects undertaken (1903-1983). Agricultura 

Tecnica 45: 175-183. 

106 
 

 

 

CHAPTER 9: INSECT-KILLING FUNGI FOR HWA MANAGEMENT: 


CURRENT STATUS
 

Scott D. Costa
 

Department of Plant and Soil Science, University of Vermont, Burlington, VT
 

ECOLOGICAL CONSIDERATIONS 

Fungi play an active and diverse role in forest and 

other ecosystems. Some are decomposers, breaking 

down and recycling formerly living material so 

that the nutrients are again available for plant 

growth. Some can cause serious diseases of living 

plants, while others are even cultivated by ants 

in subterranean gardens. A number of fungal 

species impact insect populations, sometimes 

causing major disease outbreaks (epizootics) 

that drastically reduce insect numbers. Insectkilling fungi, called entomopathogens, are 

found naturally in most terrestrial habitats. 

The ultimate goal of a biological control program 

based on the use of insect-killing fungi is to have the 

fungus circulating within a target insect population 

and naturally achieving sufficient levels of pest 

suppression. Fungi and other pathogens, such as 

viruses and bacteria, are registered through EPA 

as biopesticides, but when living propagules are 

present, they may better reflect biological control 

approaches instead of traditional insecticides. 

Like introduction or augmentation strategies 

using predators or parasitoids, ideally, after several 

releases the biological agent should integrate and 

work to bring about a check to insect outbreaks 

that damage or kill their hosts. Additionally, like 

predators and parasitoids, insect-killing fungi 

often work best as members of a complex of agents 

effective upon different stages and densities of an 

insect lifecycle. What makes insect-killing fungi 

unique is that most are readily mass produced, and, 

when developed and registered as biopesticides, 

can be released in large amounts to obtain some 

degree of immediate pest suppression. Th is strategy 

in turn can benefit adoption of other biological 

measures that have longer lag times to build up 

to effective levels. This is particularly critical in 

the case of eastern and Carolina hemlocks that 

are rapidly overcome by explosive outbreaks of 

HWA populations soon after initial invasion. 

The successful implementation of insect-killing 

fungi for HWA suppression requires close 

attention to biological properties and limitations 

of fungal agents, their interaction with the 

insect target and potential non-target hosts, and 

important influence of abiotic factors, particularly 

moisture, temperature and sunlight. Eff ectiveness 

of a management program could be seriously 

impaired by simple factors, such as targeting of 

an insect life stage less vulnerable to infection, the 

susceptibility of fungi applied midday to intense 

UV light (Fargues et al. 1996), or the lack of 

sufficient moisture for spore germination after 

application. Practical considerations include the 

need for sufficient amounts of fungus to come 

into direct contact with the target insect to initiate 

infection. In the case of HWA, this means having 

spray droplets penetrate through the canopy to 

deposit at the base of the needle underside where 

HWA are located when non-motile, or timing the 

application to coincide with emergence of HWA 

crawlers so that fungal/insect contact is achieved 

as the motile crawlers seek a suitable place to 

settle and continue development. As with most 

biological products, e.g., Bacillus thuringiensis 

and the nucleopolyhedrosis virus (Gypchek) for 

gypsy moth suppression, adherence to concrete 

application guidelines will be critical to fruitful 

implementation of fungi for HWA management. 

107 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

FUNGAL DISEASE DYNAMICS 

When viable fungal spores (conidia) come in contact 

with a susceptible insect host they adhere to its 

skin/cuticle and are stimulated to germinate (Fig. 

1). The germinating spore sends out a growing 

tube that penetrates into the insect. It does this 

through a tremendous amount of force focused 

on the growing point, often using enzymes that 

digest a hole though the surface. Once the fungus 

enters the body cavity it rapidly proliferates as it 

overcomes insect immune responses. Within several 

days, depending on temperature, the insect dies, 

after which the fungus emerges from the cadaver 

to release large amounts of new spores that can go 

on to infect other insects. If enough susceptible 

insects come into contact with the spores under 

favorable environmental conditions, the prevalence 

of disease can spread rapidly, initiating an epizootic 

and subsequent collapse of the insect population. 

Insect-killing fungi are less abundant in the 

environment between epizootic events, causing 

Figure 1. 	 Generalized infection cycle of an insectkilling fungus. Spores (smaller circles) adhere 

to the surface of insect cuticle (larger circle) 

and then germinate. Germ tubes penetrate 

the insect and fill the body cavity, causing 

insect death. The fungus grows out of the 

dead insect and releases new spores to 

continue the infection cycle in new insects. 

relatively low levels (enzootic) of insect infections. 

Reid (2003) found several insect-killing fungi, 

including Beauveria bassiana, Paecilomyces farinosus, 

and Verticillium lecanii, associated with fi eld 

collected hemlock woolly adelgid populations. 

However, in examining over 6,100 cadavers, these 

3 species of fungi were recovered from barely 3% 

of the specimens. An adequate level of fungal 

inoculum, i.e., the number of spores available for 

causing infection, is a requisite factor leading to 

the development of a fungal epizootic in insect 

populations. As the invasion front of HWA enters 

new hemlock stands, there appears to be insufficient 

time for the level of fungi to increase and suppress 

HWA before serious tree damage occurs. However, 

in some northern regions where lower winter 

temperatures periodically impact HWA population 

growth (Costa et al. 2005), fungi appear to be 

causing significant reductions in survival of HWA 

sistens during summer aestivation (Gouli et al. 

2011; Costa, personal observations). Interestingly, 

after several years of conducting fungal trials at 

Mount Tom State Reservation, Holyoke, MA, high 

mortality in untreated controls made further use 

of this site unreliable for experimental applications 

of fungi (Costa, personal observation). Although 

insect-killing fungi are ubiquitous in most terrestrial 

environments, augmenting their prevalence in 

hemlock forests through inundative applications is 

a first step toward facilitating timely development 

of disease outbreaks. This can only be legally 

accomplished through the use of fungi registered as 

biopesticides by the U.S. Environmental Protection 

Agency that are labeled for the intended use. 

In addition to a sufficient level and adequate 

distribution of fungi in the environment, there must 

be adequate moisture and temperature to initiate 

spore germination and fungal growth, with an insect 

that is susceptible to infection. A relative humidity 

between 85 and 100% is generally required for 

most species of fungi to germinate. In hemlock 

forests, these levels, if not apparent during daytime, 

can be achieved as temperature declines overnight 

(Fig. 2), which may also lead to extended periods 

of needle wetness that favor fungal germination. 

Optimal temperatures for insect-killing fungi 

108 
Chapter 9: Insect-Killing Fungi for HWA Management: Current Status 

Figure 2. 	 Temperature (dotted line) and relative humidity (RH – solid line) recorded in a hemlock forest at 

Tennessee Wildlife and Recreation Area, Titus Creek, during a critical period post fungal treatment 

(2010 Pilot Study). Note extended periods of high humidity and repeated increases in humidity as 

temperatures cycle lower. 

generally range from 15-30 °C depending upon the 

species and isolate of fungus examined. Most fungi 

are intolerant of higher temperatures, which is a 

factor contributing to their general safety for warm 

blooded animals and humans (Vestergaard et al. 

2003). Although several fungal isolates have 

been found to be active at lower temperatures 

(De Croos and Bidochka 1999; Fargues et al. 1997), 

temperature considerations and experience suggest 

that fungal applications against HWA should be 

directed against progredien adults and their sisten 

offspring as temperatures warm in mid to late 

spring, but well before temperatures decline in fall. 

FUNGAL BIOPESTICIDE DEVELOPMENT 

While all insect-killing fungi by defi nition are 

considered pathogenic to at least some species of 

insects, there is variability in the susceptibility 

of different insect species to a particular fungus. 

Certain insect-killing fungi, such as Beauveria 

bassiana, have a broad spectrum of activity against 

insects, in that they kill many different kinds of 

insects from a variety of insect orders. Others have 

relatively narrow or even very restricted host ranges, 

only infecting a small number of closely related 

insect species, as is the case with Entomophaga 

109 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

miamaiga, a pathogen that periodically devastates 

gypsy moth populations. Even within one species 

of fungus, a single isolate may be more virulent 

in that it kills insects at lower concentrations 

than other related isolates. In the laboratory, 

the virulence of fungi as biopesticides can be 

characterized similarly to chemical pesticides 

by determining LC50 and LD50 values against a 

target pest population. While initially valuable in 

earlier stages of research, the artifi cial conditions 

generally used in laboratory assays favor the 

fungus and give only a limited indication of how 

well it will perform under fi eld conditions. 

Considerable effort has been made not only to 

identify fungi for suppression of HWA, but also 

to develop operational formulations and evaluate 

application technology and timing for delivery into 

hemlock forests. In earlier stages, domestic and 

foreign explorations were carried out to identify 

potential candidates to target HWA (Gouli et al. 

1997; Ried 2003). Later small scale field tests on 

single branches or small trees (<3 meters) found 

fungi in the genus now classifi ed as Lecanicillium 

(formerly Verticillium lecanii) to hold the greatest 

promise (Reardon et al. 2004); strains of Beauveria 

bassiana and Metarhizium anisopliae, some 

commercially available in the United States, were 

ineffective. Field testing for non-target eff ects 

on adults of the predatory beetle Sasajiscymnus 

tsugae also found the Lecanicillium sp. (ARSEF 

6010) to be a promising candidate, whereas M. 

anisopliae caused significant predator mortality 

(Reardon et al. 2004). Regretfully, ARSEF 6010 

was an experimental isolate and did not possess 

any background research for environmental safety 

that would allow a US biopesticide registration 

without considerable expense and delay. 

An alternative was found in Mycotal, a commercial 

formulation of L. muscarium registered in several 

European and Asian countries by Koppert 

Biological Systems (Netherlands). Th e biopesticide 

registration package for Mycotal has been recently 

updated for the European Union. Th is extensive 

package of environmental assessment, a previous 

history of release for testing in the United States, 

and documented isolation of the same species in 

hemlock forests in the Eastern United States made 

Mycotal a favorable candidate for importation 

and release. After careful review, permits were 

secured through USDA Plant Protection and 

Quarantine for importation and experimental 

release into hemlock forests. The availability of a 

full registration package also dramatically reduces 

the costs associated with obtaining a biopesticide 

registration within the United States, which is a 

small fraction of the cost for synthetic chemical 

pesticides. Hemlock being a riparian species with 

close proximity to forest streams makes the health 

and environmental safety of Mycotal particularly 

attractive for suppression of hemlock woolly adelgid. 

Historically, fungal biopesticides are generally 

applied using high volumes of water with the 

intention of enhancing spray coverage and providing 

adequate moisture to allow spore germination. 

The use of high volumes of carrier is operationally 

unacceptable for aerial application in forest 

systems. Adoption of rotary atomizers (Micronair 

AU6539, Micron Sprayers Ltd, UK) to generate 

fine mists allowed dramatic reductions in spray 

volumes (10 liters (2.6 gallons)/acre) (Costa et 

al. 2011) and facilitated aerial (helicopter) spray 

penetration into the hemlock canopy, even onto the 

underside of hemlock needles (Reardon and Costa, 

unpublished data). At the same time, frequent 

occurrence of high levels of relative humidity 

within the hemlock forests supplanted the need 

to apply additional moisture to support fungal 

germination (for example, Figure 2). Th ese factors 

contribute to the likelihood that Mycotal could be 

suitable for operational use in HWA suppression. 

Insect-killing fungi as a tool for pest management 

has faced a major hurdle in delivering an efficacious 

amount of fungi over large acreage at a reasonable 

cost. As part of the program to deploy fungi for 

HWA management, an operational formulation 

was developed based on the incorporation of 

sweet whey, an inexpensive byproduct from 

cheese production. The whey additive in this 

fungal microfactory formulation (patent pending: 

MycoMaxTM fungal enhancer) serves as a nutrient 

110 
Chapter 9: Insect-Killing Fungi for HWA Management: Current Status 

to encourage the growth and reproduction of L. 

muscarium in spray deposits. The result is a dramatic 

increase (10- to100-fold) in the number of spores 

and a corresponding reduction in the amount 

of fungal biopesticide that needs to be applied 

(Grassano 2008). In a sense, the use of MycoMaxTM 

fungal enhancer transfers a significant portion of 

fungal mass production out of the factory into the 

forest, with the potential for cost savings through 

reduction in biopesticide application rates. 

PILOT STUDY EFFICACY TRAILS— 

DESIGN OVERVIEW 

In 2009 and 2010, pilot studies using aerial 

applications of Mycotal into hemlock forests were 

conducted at the Tennessee Wildlife and Recreation 

Area, Titus Creek. An additional ground based 

hydraulic application (where individual 

trees were treated) was conducted in 2010 at 

Townsend State Park, VT. A complete description 

of these field trials will be published elsewhere. 

Without issuance of an EPA Experimental Use 

Permit, Federal regulation restricts acreage treated 

with non-registered pesticides to no more than 

10 acres/year. In both years, replicated plots 

(1-1.25 acres) of mixed hemlock/hardwood forest 

were treated aerially (10 liters/acre) by Helicopter 

Applicators, Inc. using a Bell Jet-Ranger helicopter 

with a mounted AU6539 Micronair nozzle spray 

system. Targeting of treatments was accomplished 

with an on-board navigation system and groundtruthed plot locations. Two applications were made 

each year to enhance spray coverage; sampling 

of deposits within the hemlock forest indicated 

penetration of sprays through the canopy. 

In 2009, treatments were made in the evening 

and the following morning and consisted of a no 

spray control, Mycotal alone ( L. muscarium at 

1 x 108

 spores/ml), and Mycotal formulated with 

MycoMaxTM fungal enhancer (5% w/v). Th e oil 

adjuvant Addit (0.25% v/v : Koppert Biological 

Systems) and the sticker Hyperactive (0.05% v/v: 

Helena Chemical) were also added to both fungal 

treatments. In 2010, the rate was quadrupled: 

the treatment using Mycotal alone was dropped, 

and applications were only made in the evening. 

In Vermont, only a single hydraulic application 

in evening was made (38 liters/tree at 1 x 107 

spores/ml). Treatments are made later in the day 

to reduce effects of UV radiation on survival of 

fungal spores. Assessment of treatment eff ects 

were made for preselected individual trees (10/ 

plot) by evaluating HWA population levels in 

the lower and mid canopy (5 samples/elevation) 

both before and after treatment. The timing of 

the sampling provides evaluation of the overall 

change in the population of HWA sistens from 

one year to the next so that suppression by fungal 

treatments was assessed relative to populations 

in controls. A similar criterion was used for the 

ground based trial at Townsend State Park, VT. 

PILOT STUDY EFFICACY TRAILS— 

EFFICACY SUMMARY 

Aerial application of Mycotal reduced hemlock 

woolly adelgid population growth. The 2009 pilot 

study found that after nearly a year, plots treated 

with the MycoMax enhanced fungal formulation 

had less than half the growth in population than 

untreated plots (Fig. 3); those treated with the 

standard Mycotal formulation were intermediate 

(Costa, 2011). A subsequent application at a higher 

rate to a portion of these plots the following year, 

along with appropriate controls, did not provide 

any additional benefit (Fig. 4). However, the data 

did suggest that while HWA populations remained 

higher in the controls over the 2 year period, in the 

treated plots population growth had been arrested. 

A separate pilot study in 2010 found more than 

75% reduction in HWA population growth (Fig. 5). 

The actual population in control plots increased 

from an average of 8 HWA/branchlette to nearly 12, 

whereas with almost 9 HWA/branchlette in treated 

plots the increase was only 10%. In both years, these 

differences were statistically signifi cant (P≤0.05) 

when trees were evaluated as experimental units, 

which is justified given the similarity in variance 

between trees and plots. This approach was taken to 

compensate for variability associated with densitydependant effects on HWA population levels. 

111 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Figure 3. 	 Response of HWA populations to aerial 

application of insect-killing fungi (Mycotal, 

L. muscarium), with and without a fungal 

enhancer (MycoMax), into hemlock forest 

plots for the 2009 Pilot Study. Bars represent 

the differences between pre- and posttreatment HWA counts (sisten generation); 

capped lines indicate the standard error and 

different letters denote signifi cant (P≤0.05) 

differences between treatment means. HWA 

populations in the no spray and fungus 

alone plots grew at a faster rate than when 

enhanced fungus was applied. 

Figure 5. 	 Response of HWA populations to aerial 

application of insect-killing fungi (Mycotal, 

L. muscarium), formulated with a fungal 

enhancer (MycoMax), into hemlock forest 

plots for the 2010 Pilot Study. Bars represent 

the differences between pre- and posttreatment HWA counts (sisten generation); 

capped lines indicate the standard error and 

different letters denote a signifi cant (P≤0.05) 

difference between treatment means. HWA 

population growth was suppressed by 

treatment with enhanced fungi. 

Figure 4. 	 Response of HWA populations in a subset 

of the 2009 Pilot Study plots following a 

second year of aerial application of insectkilling fungi (F: Mycotal, L. muscarium), 

with and without a fungal enhancer (M: 

MycoMax), into hemlock forest plots. 

The “initial effect” represents changes in 

HWA population from pre-treatment levels 

after the first year and the “overall effect” 

is the change between the second year 

HWA counts and those taken before the 

year 1 treatment. Only the “F+M 2 years” 

treatment had a second aerial application, 

which was at 4 times the initial application 

rate; the data for those plots is broken out 

in the “initial effect” section for comparison. 

The treatment effect was signifi cant (P≤0.05) 

for both initial and overall response data, but 

a significant interaction with HWA density 

precludes presenting mean separations. 

Application for a single year at a lower rate 

was sufficient to suppress HWA populations. 

112 
 

 

Chapter 9: Insect-Killing Fungi for HWA Management: Current Status 

Ground-based treatment in VT using the enhanced 

fungal formulation arrested (P≤0.05) HWA 

population growth, while those on control trees 

continued to expand (Fig. 6). Th e actual number 

of HWA in control samples more than tripled 

from 15 to 50 per tree sample, whereas on fungal 

treated trees their number declined slightly after 

treatment; before treatment populations in the 

control trees and those to be treated did not diff er 

significantly (P>0.05). Gouli et al. (2008) also did 

ground applications with whey and/or oil enhanced 

formulations of B. bassiana and L. muscarium, but 

employed an ULV sprayer. The total level of fungi 

delivered was similar (1x1011 spores/tree) to the 

VT study, although the trees were approximately 

1/10 the size, and HWA mortality ranged from 

85-90 percent. The ability to suppress HWA using 

fungi applied from the ground helps corroborate 

the findings from aerial applications and holds 

promise for application to landscape plantings 

and those not suitable for aerial treatment. 

INSECT-KILLING FUNGI— 

MANAGEMENT POTENTIAL 

Over the course of development, multiple strategies 

have been attempted to increase the opportunity 

for successful deployment of insect-killing fungi 

for HWA management. Applications that target 

the sistentes generation of crawlers seem to hold 

the most promise. Ambient temperatures are more 

suitable for fungal germination and growth when 

this generation emerges in late May through June. 

Treatments applied in late summer have proved 

inconsistent because declining temperatures are 

less suitable, particularly for benefi ting from 

the microfactory approach with whey-based 

formulations. Although targeting the crawlers 

arising from overwintering sisten populations has 

not been attempted, temperature records suggest 

conditions are less favorable for fungal development. 

Another advantage in targeting sisten crawlers is 

their movement across the foliage as they seek to 

settle undoubtedly increases opportunity for insect 

fungal contact. The growth of L. muscarium from 

microfactory spray droplets should also facilitate 

HWA contact with a level of fungal inoculum that 

might not otherwise be sufficient to cause mortality. 

Figure 6. 	 Response of HWA populations to ground 

application of insect-killing fungi (Mycotal, 

L. muscarium), formulated with a fungal 

enhancer (MycoMax), onto individual 

hemlock trees in Vermont (2010). Bars 

represent the differences between preand post-treatment HWA counts (sisten 

generation); capped lines indicate the 

standard error and different letters denote 

a signifi cant (P≤0.05) difference between 

treatment means. HWA population growth 

was suppressed by treatment with enhanced 

fungi. 

Th e influence of the observed effects of L. 

muscarium (Mycotal) on longer-term HWA 

population dynamics is uncertain. Costa (2011) 

reported the results of an elementary modeling 

exercise incorporating the fungal impact obtained 

in the 2009 pilot study (~50% suppression in 

expansion) into a Hemlock Woolly Adelgid 

Population Simulator (Trotter 2011). Appreciable 

inhibition to HWA populations over a 4-year 

cycle was projected, although biannual spikes in 

crawler population still occurred. Interestingly, 

with an increase in fungal impact to the level 

observed in 2010, crawler spikes begin to subside 

and overall populations decreased over time! 

Modeling exercises cannot be substituted for 

real world evaluations, but they can provide 

insights into potential interaction of mortality 

factors on longer-term population dynamics. 

Insufficient time has passed since the 2009 and 2010 

pilot studies to determine if the observed impact 

on HWA populations will result in concurrent 

113 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

improvement in tree health and survival; data 

collected during 2011 should begin to address this 

question. While dramatic knockdown is hoped for 

through making “inundative” applications of fungal 

spores, the benefits of broadly inoculating HWA 

populations to enhance prospects for future disease 

outbreaks should not be ignored. It may be that 

without other biological agents integrated into the 

system, the effect of fungus alone will be insufficient. 

One hypothesis is that enduring HWA populations 

will provide food for predatory beetles to maintain 

stability of their populations. Ecologically 

relevant non-target effects from L. muscarium, 

particularly to introduced predators, have not been 

identified and additional testing is underway. 

There is a pressing need for implementation of 

effective tactics for protecting hemlock forests 

from the rapid decline experienced, particularly 

in southern range, as HWA becomes established. 

Currently, an experimental use permit is being 

sought from the EPA to allow expanded testing 

of Mycotal on larger acreage—approval seems 

probable. Biopesticide registration of Mycotal 

for forestry and other uses is likely only a minor 

hurdle due to its overseas regulatory history and 

a parent company committed to its development 

for HWA management. Recently, the economics 

of various management approaches was evaluated 

in a structured decision making exercise 

(Blomquist et al. 2010). A fungal biopesticide 

approach compared favorably with release of 

predators and silvicultural practices. However, 

the economic viability and efficacy of Mycotal 

seems dependant on adopting microfactory 

formulation technology to reduce application 

rates and to enhance insect/fungal contact. 

The inaccessibility and rugged terrain often 

accompanying growth of hemlock makes aerial 

application a requisite tactic. Recent changes 

in EPA-NPDES regulations that specifi cally 

target forestry application of biopesticides 

highlight the need for forward planning if use 

of insect-killing fungi is to be broadly adopted, 

particularly because of riparian habitats hemlock 

occupy. A realistic appraisal suggests that with 

a concerted effort toward expanded testing, 

obtaining federal and state biopesticide label 

registrations, and early incorporation into 

strategic management plans, insect-killing fungi 

will become available for HWA management. 

ACKNOWLEDGMENTS 

The development work has been accomplished with 

the steadfast intellectual and practical support of at 

least the following individuals: Brad Onken, Karen 

Felton, Dick Reardon, Rusty Rhea, Dennis Souto, 

Michael Bohne, Bob Tichenor, Brant Miller, Jerome 

Grant, Joe Elkins, Wes Pilley, Justin Walden, Ashley 

Lamb, Bud Mayfield, Brad Register, and the Costa 

family. The author is also indebted to Helicopter 

Applicators Inc. for their exceptional service and 

Koppert Biological Systems, Inc. for supply of 

Mycotal. The Pilot Studies were funded under 

USFS Cooperative Agreement Number: 07-CA11420004-117, Whey-Based, Fungal Micro-Factory 

Technology for Enhancement Mycopathogen 

Deployment in Forest Pest Management. 

LITERATURE CITED 

Blomquist, S.M.; Johnson, T.D.; Smith, D.R.; 

Call, G.P.; Miller, B.N.; Th urman, W.M.; 

McFadden, J.E.; Parkin, M.J.; Boomer, G.S. 

2010. Structured Decision-Making and Rapid 

Prototyping to Plan a Management Response to 

an Invasive Species. Journal of Fish and Wildlife 

Management, June 2010, Vol. 1, No. 1: 19-32. 

Costa, S.D. 2011. Hemlock woolly adelgid 

suppression with aerial application of Mycotal 

in a microfactory formulation, Can it work? In: 

Onken, B.; Reardon, R., comps. Proceedings of 

the Fifth Symposium on Hemlock Woolly Adelgid 

in the Eastern United States; 2010 August 17-19; 

Asheville, NC. FHTET-2010-07. Morgantown, 

WV: U.S. Department of Agriculture, Forest 

Service, Forest Health Technology Enterprise 

Team: 36-42. 

114 
Chapter 9: Insect-Killing Fungi for HWA Management: Current Status 

Costa, S.; Felton, K.; Onken, B.; Reardon, R.; 

Rhea, R. 2011. Aerial application of the insectkilling fungus Lecanicillium muscarium in a 

microfactory formulation for hemlock woolly 

adelgid suppression. In: McManus, K.A.; 

Gottschalk, K.W., eds. Proceedings, 21st U.S. 

Department of Agriculture Interagency Research 

Forum on Invasive Species 2010; 2010 January 

12-15; Annapolis, MD. Gen. Tech. Rep. NRSP-75. Newtown Square, PA: U.S. Department 

of Agriculture, Forest Service, Northern 

Research Station: 80-81. 

Costa, S.D.; Skinner, M.; Parker, B.L. 2005. 

Hemlock woolly adelgid cold-hardiness: 

towards defing the limits of range expansion. 

In: Gottschalk, Kurt W., ed. 2005. Proceedings, 

XV U.S. Department of Agriculture Interagency 

Research Forum on Gypsy Moth and Other 

Invasive Species 2004; 2004 January 13-16; 

Annapolis, MD. Gen. Tech. Rep. NE-332. 

Newtown Square, PA: U.S. Department of 

Agriculture, Forest Service, Northeastern 

Research Station: 12-22. 

DeCroos, J.N.A.; Bidochka, M.J. 1999. Eff ects 

of low temperature on growth parameters in 

the entomopathogenic fungus Metarhizium 

anisopliae. Canadian Journal of Microbiology 

45: 1055-1061. 

Fargues, J.; Goettel, M.S.; Smits, N.; Ouedraogo, 

A.; Vidal, C.; Lavey, L.A.; Lomer, C.J.; 

Rougeir, M. 1996. Variability in susceptibility 

to simulated sunlight of conidia aomong 

isolates of entomopathogenic Hyphomycetes. 

Mycopathologia 135: 171-181. 

Fargues, J.; Goetel, M.S.; Smits, N.; Ouedraogo, 

A.; Rougier, M. 1997. Effect of temperature on 

vegetative growth of Beauveria bassiana isolates 

from diff erent origins. Mycologia 89: 383-392. 

Gouli, V.V.; Gouli, S.Y.; Parker, B.L.; Skinner, M. 

2011. Fungi as epizootic factor in populations 

of sucking alien insects in north-eastern region 

of the USA. In: Proceedings of the International 

Conference - Integrated Pest Management: 

Strategy and Tactics. Minsk, Belarus: National 

Academy of Science of Belarus and International 

Organization for Biological Control. 

Gouli, S.; Gouli, V.; Teillon, H.B.; Kassa, A.; 

Skinner, M.; Frank, C.; Parker, B. 2008. 

Controlling hemlock woolly adelgid with new 

forms of Mycoinsecticides. In: Onken, B.; 

Reardon, R., eds. Fourth Symposium on Hemlock 

Woolly Adelgid in the Eastern United States: 

Proceedings of the Meeting; 2008 February 12-14; 

Hartford, CT. FHTET-2008-01. Morgantown, 

WV: U.S. Department of Agriculture, Forest 

Service, Forest Health Technology Enterprise 

Team: 179-187. 

Gouli, S.; Parker, B.L.; Skinner, M. 1997. Isolation 

of fungi associated with hemlock woolly adelgid 

(Homoptera: Adelgidae). Journal of Invertebrate 

Pathology 70: 76-77. 

Grassano, S. 2008. Whey-based Fungal Microfactories 

for In Situ Production of Entomopathogenic Fungi. 

Burlington, VT: University of Vermont. 

67 pages. M.S. thesis. 

Reardon, R.; Onken, B.; Cheah, C.; Montgomery, 

M.E.; Salom, S.; Parker, B.L.; Costa, S.; 

Skinner, M. 2004. Biological Control of Hemlock 

Woolly Adelgid. FHTET-2004-04. Burlington, 

VT: U.S. Department of Agriculture, Forest 

Service, Forest Health Technology Enterprise 

Team: 22 p. 

Reid, W.R. 2003. Isolation and characterization 

of entomopathogenic fungi associated with the 

hemlock woolly adelgid, Adelges tsugae Annand. 

Burlington, VT: University of Vermont. 

99 pages. Ph.D. dissertation. 

Trotter, R.T. 2011 Hemlock Woolly Adelgid 

Population Simulator V2.0. http://www.nrs. 

fs.fed.us/disturbance/invasive_species/hwa/ 

biology_ecology/ 

Vestergard, S.; Cherry, A.; Keller, S.; Goettel, 

M. 2003. Safety of Hyphomycete fungi as 

microbial agents. In: Hokkanen, H.M.T.; 

Hajek, A.E., eds. Environmental Impacts of 

Microbial Insecticides: Needs and Methods for Risk 

Assessment. The Netherlands: Kluwer Academic 

Publishers: 277 p. 

115 
CHAPTER 10: OTHER SPECIES CONSIDERED 

Michael E. Montgomery1, Thomas J. McAvoy2, and Scott M. Salom2 

1

U.S. Forest Service, Northern Research Station, Hamden, CT 

2

Virginia Tech, Department of Entomology, Blacksburg, VA 

INTRODUCTION 

This chapter reports on predators which have only 

preliminary information or were found to have 

a host range that was too broad to be considered 

as safe biological control agents. Th e predators 

reviewed here include predaceous anthocorid 

bugs from China and western North America, a 

Laricobius beetle from China, and a lady beetle 

native to western North America. For each of the 

species, information on their taxonomy, biology, 

and host range is provided and their potential as a 

biological control agent for Adelges tsugae Annand, 

the hemlock woolly adelgid (HWA), is evaluated. 

Key questions for the reader to ponder are how 

host specific should HWA predators be and can 

successful biological control be achieved using 

only predators that are specific only to HWA. 

SPECIES CONSIDERED 

Tetraphleps galchanoides Ghauri 

(Hemiptera: Anthocoridae) 

The Anthrocoridae is a family of 500-600 

worldwide species that are mostly predaceous, a 

few of which have been deliberately introduced as 

biological control agents (Lattin 1999). Within the 

Anthrocoridae, the genus Tetraphleps has about 14 

described species worldwide which feed primarily 

on aphids and adelgids (Lattin 2000). Th ere are 

five species of Tetraphleps native to North America 

and four of these have been observed to prey on the 

balsam woolly adelgid (Kelton 1978). Tetraphleps 

raoi Ghauri and T. abdulghani Ghauri, both native 

to Pakistan and India, were introduced to North 

America for biological control of the balsam woolly 

adelgid, but apparently did not establish. Bu and 

Zheng (2001) attributed the failure to establish 

to a lack of climatic suitability. While T. raoi was 

not successfully introduced into North America, it 

was established in Kenya for control of Pineus pini 

(Macquart), where it is believed to be responsible for 

a decline in the density of the adelgid (Lattin 2000). 

Tetraphleps galchanoides (Fig. 1) is known from India 

and China (Bu and Zheng 2001). Th e first report in 

China was a specimen collected from Tsuga chinensis 

in Baoxing County, Sichuan Province (Yao and 

Wang 1998). They reported that an adult could eat 

2.7 nymphs and 5.6 eggs of HWA per day. In the 

spring of 2007, a very dense population of HWA 

was observed in Yunnan Province that was preyed on 

heavily by both adults and nymphs of T. galchanoides 

(McAvoy et al. 2008). Laboratory experiments 

showed that T. galchanoides preying on HWA was a 

Type II Holling functional response—predation rate 

increased as prey densities increased (Li et al. 2008). 

Figure 1. Adult Tetraphleps galchanoides. 

116 
 

Chapter 10: Other Species Considered 

From 2002-2008, basic biology and host feeding 

studies were conducted in quarantine to determine 

the potential of T. galchanoides as a biological 

control agent of HWA (McAvoy et al. 2007, 

McAvoy et al. 2008). In 2002, eggs and nymphs 

were recovered from hemlock foliage collected in 

the fall in Sichuan, China that were shipped to the 

quarantine facility at Virgina Tech. When newly 

hatched nymphs were fed only the balsam woolly 

adelgid, Adelges piceae Ratz., none develped to 

the next nymphal stage; but when fed exclusively 

pine bark adelgid, Pineus strobi (Hartig), or woolly 

alder aphid, Paraprociphilus tessellatus (Fitch), they 

successfully developed to the adult stage. Research 

(unpublished) in China indicated that T. galchinoides 

preyed on the bean aphid placed in Petri dishes. 

Tetraphleps galchanoides females normally oviposit 

their eggs into the lower side of the hemlock needle 

tissue. When offered a choice of foliage of white 

pine (Pinus strobus), red spruce ( Picea rubens), and 

Fraser fi r ( Abies fraseri), T. galchanoides oviposited in 

white pine and fir but not spruce needles. However, 

in a no-choice test with spruce only, T. galchanoides 

did lay eggs in spruce needles. In a choice test with 

all four conifer species, T. galchanoides oviposited 

only in hemlock needles, indicating that hemlock 

is the preferred host, but will lay eggs in the 

other three non-target species, spruce being the 

least preferred species. Eggs were also oviposited 

in leaves of smooth alder ( Alnus serrulata) in a 

no-choice test. Unlike the eggs inserted into the 

conifer needle tissue, eggs in the alder test were 

laid on the surface of the leaf and not inserted 

into the leaf tissue. Several of these eggs hatched. 

Tetraphleps galchanoides fed and oviposited on 

non-target adelgids and aphids. Additional feeding 

tests found that T. galchanoides adults will feed 

on Laricobius osakensis Montgomery and Shiyake 

(Coleoptera: Derodontidae) larvae. Due to the 

suitability of several non-target homopteran 

species as hosts and its feeding on the larvae of 

a potential biological control agent, this species 

is not considered at this time to be a suitable 

candidate for release as a biological control agent. 

Anthocoris nemoralis Fabricius 

and A. antevolens White 

Th e genus Anthocoris occurs primarily in the 

Northern Hemisphere and consists of about 70 

species. The Palearctic has 47 species, and 12 

species occur in Canada and the United States 

(Lattin 2000). Only one species that is native to 

North America, Anthocoris antevolens White, is 

known to use adelgids as prey and this is considered 

incidental (Lattin and Stanton 1992). In North 

America, the hemipterans Daecocororis pinicola, D. 

piceicola, and D. nubilus in the family Miridae prey 

on adelgids on pine and spruce (Wheeler 2001). 

Several species of Anthocoridae were collected 

by R. McDonald from October to February, 

2008-2011, in Seattle, WA while he was collecting 

L. nigrinus for release in the eastern United 

States. The most abundant species was Anthocoris 

nemoralis (Fab.) with 55 adults collected or 89% 

of the total Anthocoris collected. Th is species 

was intentionally introduced from Europe as a 

biological control agent in orchards against aphids, 

psyllids, thrips, and moth eggs and larvae (Kelton 

1978). It became established in the 1950s and is 

found in British Colombia, Ontario, Washington, 

Oregon, and California (Lewis et al. 2005). Th e 

second most common species was A. antevolens 

White, with seven adults found (11%). Th is species 

is native to North America and is found across 

Canada and western United States. This is the fi rst 

report of fi nding Anthocoris species on Tsuga. 

Adults shipped to the Virginia Tech quarantine 

facility were reared on Tsuga canadensis foliage 

infested with HWA at 15 °C. The mean length of 

time that the adults lived was 36 days. Th e number 

of females collected was low with only 1 female for 

every 6 males, and no oviposition was observed on 

hemlock. No nymphs were found during beat-sheet 

sampling of hemlock. It appears that the adults of 

these two species of Anthocoris may only use HWA 

as an occasional food source, but not for oviposition. 

117 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Laricobius kandingensis Zilahi-Balogh 

and Jelínek 

Th e genus Laricobius (Coleoptera: Derodontidae) 

has 21 described species, all of which prey only on 

adelgids (Leschen 2011). Two of these species were 

discovered as a result of an expedition to China 

that specifically targeted this genus. In April 2002, 

an expedition to Sichuan Province, China found a 

new species in Baoxing County and another new 

species in Kangding County. These were described, 

and named according to where they were found: 

L. baoxingensis Zilahi-Balogh and Jelínek, and L. 

kangdingensis Zilahi-Balogh and Jelínek (ZilahiBalogh et al. 2007). Both species were found on 

adelgid infested hemlock (T. chinensis (Franchet) 

Pritzel) in April 2002. Only female adults (n=5) of 

L. baoxingensis were collected and these died without 

reproducing after being shipped to the quarantine 

laboratory at Virginia Tech in Blacksburg, Virginia. 

Only larvae (n=23) of L. kangdingensis were 

collected and these completed development on 

T. canadensis infested with HWA in the quarantine. 

Quarantine studies of L. kangdingensis at Virginia 

Tech (Gatton et al. 2009) found it to be univoltine, 

have 4 larval instars, a low temperature development 

threshold for eggs (0 °C), larvae (1.6 °C), and 

pre-pupae (5.8 °C), yet completed development only 

at temperatures between 12-15 °C. Host-range 

studies showed it preferred HWA over all other 

species tested. 

This species has not been pursued as a biological 

control agent due to problems with rearing the 

beetles in quarantine. The largest challenge was the 

result of the small size of the founding colony and 

ultimately the colony could not be maintained. 

Efforts to start new colonies were not successful 

because only small numbers could be collected 

in China, and collected beetles did not survive 

shipment to the quarantine in the United States. 

In the meantime, another species, Laricobius 

osakensis Montgomery and Shiyake, was discovered 

in Japan and it was much more abundant and 

widespread in its native habitat; thus, work with 

the Chinese Laricobius was discontinued. 

Scymnus (Pullus) coniferarum (Crotch) 

Scymnus is the largest genus of lady beetles 

(Coccinellidae) with over 600 species worldwide. 

This large genus is divided into six subfamilies 

of which the subgenus Pullus is the largest with 

more than 300 described species. Although a 

large subgenus, Scymnus (Pullus) has only three 

species that are known to be specialists on adelgids 

(Whitehead 1967). Two of these species, S. (P.) 

impexus (Mulsant) and S. (P.) suturalis (Th unberg), 

are native to Eurasia and have been introduced 

to North America, and one species, discussed 

here, is native to the western North America. 

Scymnus (Pullus) coniferarum Crotch (Coleoptera: 

Coccinellidae) was described in 1874 from 

specimens collected from pine in California. It is 

a small lady beetle, about 2 mm in length, that is 

clothed in fine, short pubescence with a black head 

and pronotum and reddish-yellow brown elytra 

that is piceous along the suture and at the base 

(Fig. 2a). Its larvae are covered in a white woolly 

wax (Fig. 2b). Whitehead (1967) and Gordon 

(1976) provide full descriptions with fi gures. 

The known native geographical range of S. (P.) 

coniferarum is western North America (Gordon 

1985). Specimens have been collected from 

various species of pines in British Columbia, 

Arizona, California, Colorado, Idaho, Nevada, 

New Mexico, Oregon, South Dakota, Utah, 

and Wyoming (Gordon 1976). Recently several 

hundred specimens have been collected from 

western hemlock in Washington State (Montgomery 

et al. 2009, McDonald 2010). It has recently 

been recovered from Monterey pine in Chile and 

Peru (Gonzalez 2006), probably an accidental 

introduction since pine and adelgids are not native 

to South America. There is a report of it in the 

eastern United States (Malkin 1945), but this may 

be a misidentification. Gordon (1976) recorded 

it in Pennsylvania, but later clarified that these 

were S. (P.) suturalis (Gordon 1985). Considering 

that the species has spread intercontinentally, 

has been found as far east as the Black Hills of 

South Dakota, and occurs in a variety of habitats 

118 
Chapter 10: Other Species Considered 

a b 

Figure 2. Adult (a) and larva (b) of Scymnus (Pullus) coniferarum. 

in western North America, it is possible that it is 

already established in eastern North America. 

The seasonal occurrence of S. (P.) coniferarum and 

another predator, L. nigrinus, as well as their prey, 

HWA, in the Seattle, WA area is shown in Table 1. 

Other information indicates that the production of 

progrediens eggs continues during July in western 

Washington (Kohler et al. 2008). Th e adults of 

both predators are present and presumably feed 

on HWA during the late fall and winter months. 

Larvae of L. nigrinus appear in March and 

complete development by mid-May. Th e larvae 

of S. coniferarum appear near the end of May and 

continue to feed on the progrediens and their sistens 

eggs into July. In severe winter weather, adults seek 

shelter in bark crevices on the bole of the tree. 

The only hosts of S. (P.) coniferarum reported 

in the literature are adelgids that feed on pine 

(Pineus spp.) (Whitehead 1967, Gordon 1976). 

Recent observations in the Seattle area have 

found it abundant on hemlocks infested with 

HWA, and to a lesser extent on western white 

pine, but it was not found on several species of 

hard pines and spruce trees (Montgomery et al. 

2009, Montgomery and McDonald 2010). 

Table 1. Seasonal occurrence of HWA progediens stages (blue shading) and HWA sistens stages 

(yellow shading) and their predators Laricobius nigrinus (Ln) and Scymnus coniferarum 

(Sc) in the Seattle, WA area (light pink = relatively few, dark pink = peak abundance). 

Observations not made July-Sept. 

Month Oct Nov Dec Jan Feb Mar Apr May Jun 

HWA adults
 

HWA eggs
 

HWA nymphs
 

Ln adults
 

Ln larvae
 

Sc adults
 

Sc larvae
 

119 
 

 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Laboratory studies indicate that S. (P.) coniferarum 

feeds specifically on adelgids and does not attack 

other Homoptera to a significant extent. In the 

laboratory, the beetle completed development from 

egg to adult on HWA and the pine bark adelgid, 

but mortality was greater on the latter host. In nochoice tests S. (P.) coniferarum adults did not feed on 

the native woolly alder aphid, Prociphilus tessellatus 

(Fitch) or the lime aphid, Eucallipterus tiliae (L.). 

In the laboratory, S. (P.) coniferarum has been 

reared for several successive generations on HWA 

growing on eastern hemlock, and studies on its 

potential to feed on non-target species are ongoing. 

IMPLICATIONS FOR 

BIOLOGICAL CONTROL 

Tetraphleps galchanoides is an example of an 

opportunistic predator. These bugs migrate to 

high prey densities where they can capture prey 

efficiently. Because of their feeding effi ciency at 

high prey densities and relatively large size, these 

opportunistic predators are very effective in reducing 

outbreak populations in the adelgids’ native 

environment. After the food supply is depleted, 

they disperse in search of another abundant source 

of suitable prey. In Japan, several opportunistic 

predators in the beetle families Elateridae, 

Cantharidae, Melyridae, and Coccinellidae also 

attack the progrediens/sexupara generations 

during May (Shiyake et al. 2008). Thus, we see in 

Asia that late spring predation by opportunistic 

generalists may play a critical role in reducing high 

outbreak densities of HWA to below damaging 

thresholds. Why similar predation on HWA by 

opportunistic generalist predators does not occur 

in the eastern United States is unclear. Th ere are 

several species of native predators in the families 

Anthocoridae, Miridae, Elateridae, Cantharidae, and 

Melyridae that potentially could prey on HWA. 

Scymnus (P.) coniferarum merits further study to 

better assess its potential as a biological control 

of HWA. Its larvae appear after L. nigrinus larvae 

have stopped feeding and have migrated to the 

soil. The late spring feeding of S. (P.) coniferarum 

indicates its impact on HWA population dynamics 

should complement that of L. nigrinus. Late spring 

predation of HWA may be critical in obtaining 

suppression of HWA below damaging levels 

(see Chapter 2). A question that remains to be 

addressed is the extent that S. (P.) coniferarum 

would feed on other adelgids, including native 

species present in the eastern United States. 

Except for Pineus floccus Patch, most of the adelgid 

species present in the eastern United States also 

occcur in the western United States. Its feeding 

on alternative prey, particularly species such as 

Pineus coloradensis Fitch that are active during the 

summer, may allow it to sustain higher populations 

and thus be a more effective predator of HWA. 

LITERATURE CITED 

Bu, W.J.; Zheng, L.Y. 2001. Fauna Sinica Insecta 

Vol. 24. Hemiptera Lasiochilidae, Lyctocoridae, 

Anthocoridae. Beijing, China: Science Press: 

267 p. 

Gatton, H.; Zilahi-Balogh, G.M.; Salom, S.M.; 

Kok, L.T. 2009. Biology and temperaturedependent development of Laricobius 

kangdingensis, a newly discovered predator 

of Adelges tsugae. BioControl 54: 219-228. 

Gonzalez, G. 2006. Los Coccinellidae de Chile. 

Accessed online September 20, 2011 at 

http://www.coccinellidae.cl/paginasWebChile/ 

PaginasOriginal/scymnus_coniferarum.php 

Gordon, R.D. 1976. The Scymnini (Coleoptera: 

Coccinellidae) of the United States and Canada: 

Key to genera and revision of Scymnus, Nephus 

and Diomus. Bulletin of the Buff alo Society of 

Natural Sciences 28: 1-362. 

Gordon, R.D. 1985. The Coccinellidae (Coleoptera) 

of America north of Mexico. Journal of the New 

York Entomological Society 93: 1-912. 

Kelton, L.A. 1978. The Insects and Arachnids of 

Canada. Part 4. The Anthocoridae of Canada 

and Alaska (Heteroptera: Anthocoridae). 

Ottowa, Canada: Agriculture Canada Research 

Publication 1639: 1-101. 

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Chapter 10: Other Species Considered 

Kohler, G.R.; Stiefel, V.L.; Wallin, K.F.; Ross, D.W. 

2008. Predators associated with the hemlock 

woolly adelgid (Hemiptera: Adelgidae) in the 

Pacifi c Northwest. Environmental Entomology 

37: 494-504. 

Lattin, J.D. 1999. Bionomics of the Anthocoridae. 

Annual Review of Entomology 44: 207-231. 

Lattin, J.D. 2000. Minute pirate bugs 

(Anthocoridae). In: Schaefer, C.W.; Panizii, 

A.R., eds. Heteroptera of Economic Importance. 

New York, NY: CRC Press: 607-637. 

Lattin, J.D.; Stanton, N.L. 1992. A review of 

the species of Anthocoridae (Hemiptera: 

Heteroptera) found on Pinus contorta. Journal of 

the New York Entomological Society 100: 

424-479. 

Leschen, R.A.B. 2011. World review of Laricobius 

(Coleoptera: Derodontidae). Zootaxa 2908: 

1-44. 

Lewis, T.M.; Horton, D.R.; Broers, D.A. 2005. 

New state and United States records for 

Anthocoridae (Hemiptera: Heteroptera). 

Th e Pan-Pacifi c Entomologist 81: 59-67. 

Li, L.; Montgomery, M.E.; Yu, G. 2008. Functional 

responses of Tetraphleps galchanoides preying 

on different stages of hemlock woolly adelgid. 

Chinese Bulletin of Entomology 46: 72-76. 

Malkin, B. 1945. A supplement to the New York 

State list of Coleoptera, No. 6, additions 

and corrections. Journal of the New York 

Entomological Society 53: 91-116. 

McAvoy, T.J.; Zilahi-Balogh, G.M.G.; Salom, S.M.; 

Kok, L.T.; Guoliang, Z. 2007. Development 

and feeding of Tetraphleps galchanoides, a 

predator of hemlock woolly adelgid. BioControl 

52: 339-350. 

McAvoy, T.J., Li Li, Guoyue Yu, Montgomery, 

M.E.; Salom, S.M.; Kok, L.T. 2008. Phenology, 

biology, and collections of Tetraphleps 

galchanoides (Hemiptera: Anthocoridae), a 

predator of hemlock woolly adelgid in China. 

In: Onken, B.; Reardon, R., eds. Fourth 

Symposium on Hemlock Woolly Adelgid in the 

Eastern United States: Proceedings of the Meeting; 

2008 Feb. 12-14; Hartford, CT. FHTET2008-01. Morgantown, WV: U.S. Department 

of Agriculture, Forest Service, Forest Health 

Technology Enterprise Team: 253-258. 

McDonald, D. 2010. Key cues and factors for 

improving HWA predator recovery eff orts. In: 

Onken, B.; Reardon, R., comps. Proceedings of 

the Fifth Symposium on Hemlock Woolly Adelgid 

in the Eastern United States; 2010 August 17-19; 

Asheville, NC. FHTET-2010-07. Morgantown, 

WV: U.S. Department of Agriculture, Forest 

Service, Forest Health Technology Enterprise 

Team: 7-16. 

Montgomery, M.E.; McDonald, R.; Schwartzberg, 

L. 2009. Scymnus (Pullus) coniferarum 

(Coleoptera: Coccinellidae): an adelgid 

predator native to the western United States. 

In: McManus, K.A.; Gottschalk, K.W., eds. 

Proceedings 20th U.S. Department of Agriculture 

Interagency Research Forum on Invasive Species 

2009; 2009 January 13-16; Annapolis, MD. 

Tech. Rep. NRS-P-51. Newtown Square, PA: 

U.S. Department of Agriculture, Forest Service, 

Northern Research Station: 89. 

Montgomery, M.E.; McDonald, R. 2010. Host 

preferences of Scymnus (Pullus) coniferarum: an 

adelgid predator. In: Onken, B.; Reardon, R., 

comps. Proceedings of the Fifth Symposium on 

Hemlock Woolly Adelgid in the Eastern United 

States; 2010 August 17-19; Asheville, NC. 

FHTET-2010-07. Morgantown, WV: U.S. 

Department of Agriculture, Forest Service, 

Forest Health Technology Enterprise Team: 

173-176. 

121 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Shiyake, S.; Miyatake, Y.; Montgomery, M.E.; 

Lamb, A. 2008. Hemlock woolly adelgid 

phenology and predacious beetle community 

on Japanese hemlocks. In: Onken, B.; Reardon, 

R., eds. Fourth Symposium on Hemlock Woolly 

Adelgid in the Eastern United States: Proceedings 

of the Meeting; 2008 Feb. 12-14; Hartford, CT. 

FHTET-2008-01. Morgantown, WV: U.S. 

Department of Agriculture, Forest Service, 

Forest Health Technology Enterprise Team: 

261-266. 

Wheeler, Jr., A.G. 2001. Biology of the Plant Bugs 

(Homoptera: Miridae). Ithaca, NY: Cornell 

University Press: 528 p. 

Whitehead, V.B. 1967. The Validity of the Higher 

Taxonomic Categories of the Tribe Scymnini 

(Coleoptera: Coccinellidae). Berkeley, CA: 

University of California, Department of 

Entomology: 215 p. Ph.D. dissertation. 

Yao, Defu; Wang, Hongbin. 1998. Sino-American 

Cooperative Program: Biological Control of 

Hemlock Woolly Adelgid—Research Report 

1996-1998. Beijing, China: Chinese Academy 

of Forestry. 

Zilahi-Balogh, G.M.G.; Jelinik, J.; McAvoy, T.J.; 

Salom, S.M.; Kok, L.T. 2007. Two new species 

of Laricobius (Coleoptera: Derodontidae) 

from China, and a key to Laricobius in 

the southeastern Palaearctic. Proceedings 

Entomological Society of Washington 109: 

377-384. 

122 
SECTION III LABORATORY REARING
 

FOR FIELD RELEASE
 
 

 

 

 

 

 

 

 

 

CHAPTER 11: REARING LABS AND DISTRIBUTION OF PREDATORS
 

FOR RELEASE
 

Carrie S. Jubb
 

Department of Entomology, Virginia Tech, Blacksburg, VA
 

REARING LABS 

There are currently eight primary labs rearing fi ve 

different predators of hemlock woolly adelgid. 

The labs include Clemson University, New Jersey 

Department of Agriculture (Philip Alampi Benefi cial 

Insect Laboratory), North Carolina Department 

of Agriculture and Consumer Services, North 

Georgia College & State University, University 

of Georgia, University of Tennessee (Lindsay 

Young Beneficial Insect Laboratory), Virginia 

Tech, and Young Harris College. Species currently 

being reared include Laricobius nigrinus Fender, 

Laricobius osakensis Montgomery and Shiyake, 

Scymnus coniferarum Crotch, Scymnus sinuanodulus 

Yu et Yao and Sasajiscymnus tsugae Sasaji and 

McClure. Table 1 details the species currently being 

reared by each lab as of fall 2011. This has been a 

dynamic list and will likely continue to change. 

Table 1. Rearing labs 

DISTRIBUTION OF PREDATORS 

FOR RELEASE 

Predators of hemlock woolly adelgid have been 

released across 17 states in the Eastern United 

States. Operational releases began as early as 1998 

and have continued steadily since then. Releases 

take place primarily on public lands, including 

state and national forests, parks and Hemlock 

Conservation Areas. Figure 1 depicts release 

locations of three primary predators ( S. tsugae, S. 

sinuanodulus, and L. nigrinus) which have been 

entered into the newly created HWA Predator 

Release and Monitoring Database. Entry of this data 

is currently ongoing and many releases have yet to 

be represented. Additional species will also be added. 

Rearing Lab L.n.* L.o. S.c. S.s. S.t. 

Clemson University x x 

New Jersey Department of Agriculture x 

North Carolina Dept. of Agriculture and Consumer Services x 

Northern Georgia College & State University x 

University of Georgia x x 

University of Tennessee x x x 

Virginia Tech x x x 

Young Harris College x 

*L.n. = Laricobius nigrinus Fender; L.o. = Laricobius osakensis Montgomery and Shiyake; S.c. = Scymnus coniferarum Crotch; 

S.s. = Scymnus sinuanodulus Yu et Yao; S.t. = Sasajiscymnus tsugae Sasaji and McClure 

125 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Figure 1. Map of the Eastern United States showing S. tsugae, S. sinuanodulus, and L. nigrinus releases which have 

been entered into the HWA Predator Release and Monitoring Database. 

Laricobius nigrinus Release Locations 

Clemson: Georgia, North Carolina, 

and South Carolina 

New Jersey Department of Agriculture: 

Pennsylvania and New Jersey 

University of Georgia: U.S. Forest Service Hemlock 

Conservation Areas throughout Georgia 

University of Tennessee: Tennessee – Great Smoky 

Mountains National Park, Cherokee National 

Forest; Tennessee State Parks and Natural Areas, 

and Tennessee Wildlife Management Areas 

Virginia Tech: Connecticut, Georgia, Maine, 

Maryland, Massachusetts, New Hampshire, 

New York, North Carolina, Pennsylvania, Rhode 

Island, Vermont, Virginia, West Virginia 

Sasajiscymnus tsugae Release Locations 

Clemson University: 

South Carolina - Sumter National Forest, 

Jocassee Gorges, Mountain Bridge 

Wilderness Area, Oconee State Park; 

North Carolina – Nantahala National Forest 

Georgia – Chattahoochee National Forest 

New Jersey Department of Agriculture: 

Connecticut, Maryland, Massachusetts, 

Maine, New Hampshire, New Jersey, 

North Carolina, New York, Pennsylvania, 

Rhode Island, South Carolina, Tennessee, 

Vermont, Virginia, West Virginia 

North Carolina Department of Agriculture and 

Consumer Services: North Carolina, Maine 

North Georgia College & State University: 

Georgia – U.S. Forest Service Hemlock 

Conservation Areas throughout North 

Georgia on National Forest and State lands 

University of Georgia: U.S. Forest Service Hemlock 

Conservation Areas throughout Georgia 

University of Tennessee: 

Tennessee – Great Smoky Mountains National 

Park, Cherokee National Forest; Tennessee 

State Parks and Natural Areas, and 

Tennessee Wildlife Management Areas 

Georgia – U.S. Forest Service study 

Young Harris College: 

Georgia – Chattahoochie National Forest 

and State properties in North Georgia 

126 
 

 

 

Chapter 11: Rearing Labs and Distribution of Predators for Release 

Scymnus sinuanodulus Release Locations 

Clemson University: Chattooga River – Bull 

Sluice Site (Georgia/South Carolina border) 

New Jersey Department of Agriculture: Maryland, 

New York, North Carolina, Pennsylvania, 

Tennessee, Vermont, West Virginia 

Northern Georgia College & State University: 

Georgia – Hemlock Conservation 


Areas throughout North Georgia on 


National Forest and State lands
 

University of Georgia: U.S. Forest Service Hemlock 

Conservation Areas throughout Georgia 

University of Tennessee: Tennessee – Great 

Smoky Mountains National Park 

Scymnus coniferarum Release Locations 

No releases yet. 

A summary of release totals is shown in Table 2. 

REARING PROCEDURES 

Laricobius nigrinus 

Dr. Ashley Lamb developed Laricobius nigrinus 

rearing procedures at Virginia Tech from 2000­

2005 while working toward her Ph.D. and later 

as a postdoctoral associate. They were adapted 

Table 2. Release totals 

for mass rearing, and have since been adjusted to 

maximize production. Each lab has manipulated 

their rearing protocols to fit their individual facility 

limitations such as space availability and equipment. 

In general, mass rearing begins in late January or 

early February when both HWA and L. nigrinus 

begin to lay eggs. New adults are collected each 

year in their native habitat, the Pacifi c Northwest 

area of the United States. These adults are brought 

back to rearing labs and serve as founders for 

that year’s colony. Collecting fresh beetles each 

year helps add to the genetic diversity of released 

populations. Additionally, wild-caught beetles 

are often more fecund which results in higher 

laboratory production numbers. Once adult L. 

nigrinus beetles are brought back to the lab, they are 

held in clear, plastic feeding containers on hemlock 

bouquets (Fig. 2). These bouquets usually consist of 

roughly 20-25 branches of highly infested hemlock 

roughly 20-25 cm in length. These are nested in 

Parafilm wrapped floral foam for hydration. 

Adults are randomly placed in containers, since sex 

cannot be determined through visual observation. 

They are held at densities of roughly 20 beetles 

per container to ensure minimal competition 

between individuals for feeding and egg laying 

sites. Adults will remain on these bouquets for a 

Rearing Lab L. nigrinus S. coniferarum S. sinuanodulus S. tsugae 

Clemson University 14,605 (adults) + 

44,678 (eggs) 52 (adults) 940,721 (adults) + 82,256 (eggs) 

New Jersey 

Department of 

Agriculture 

75 (adults) 23,348 (adults) 

536,290 (adults) + 

22,744 (eggs) + 360 

(larvae) 

North Carolina Dept. 

of Agriculture and 

Consumer Services 

484,800 (adults) 

Northern Georgia 

College & State 

University 

2,080 (adults) 204,206 (adults) 

University of Georgia 141,580 (eggs) 32,089 (eggs) Minimal egg releases 

made in 2006 

University of 

Tennessee 17,656 (adults) 98 (adults) 736,733 (adults) + 89,084 (eggs) 

Virginia Tech 72,489 (adults) 

Young Harris College 120,034 (adults) + 

100,092 (eggs) 

127 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

period of one week at specifi c temperatures and 

daylength depending on the time of year (Fig. 3). 

Adults are removed after one week and the 

foliage is placed into floral foam blocks with 

fresh twigs. These blocks are then nested into 

galvanized steel funnels or other larval rearing 

systems where eggs are allowed to hatch 

and subsequent larvae develop (Fig. 4). 

If labs are releasing eggs, they are placed in the 

field at this time. Typically, temperatures during 

this phase of development are between 12-15 °C 

with a 12:12 L:D daylength. It takes approximately 

4-6 weeks for larvae to hatch and develop through 

4 instars. Since their natural tendency is to drop 

to the soil when development is complete, larval 

rearing systems are designed to have a catch area 

such a drawer or jar under the foliage, where the 

larvae can congregate. These are checked daily until 

roughly July when production typically stops. 

Collected larvae are immediately placed into 

summer aestivation containers. Th ese containers 

are relatively small (approximately 15 cm × 

15 cm) plastic, and ventilated on at least two 

sides for adequate air circulation (Fig. 5). 

Approximately 6-7.5 cm of soil is placed into 

these containers with moisture levels at or close 

to 35% since the pre-pupae and pupae prefer 

a wet soil. A 2:1 ratio of peat moss and sand is 

used to create the most optimal soil mix. Mature 

larvae are simply placed on the soil surface 

and they eventually burrow down to create a 

pupation cell. Once in the soil, containers are 

held at roughly 12-15 °C for approximately 

6 weeks while pupae develop into adults. 

After pupae eclose, containers are then moved into 

19 °C (16:8 L:D) temperatures for the summer 

months. These higher temperatures again mimic 

conditions in their native habitat and encourage 

the adults to stay in the ground. Soil moisture 

levels are kept very low at this point. Containers 

are watered very lightly on a weekly basis. 

Figure 2: L. nigrinus feeding containers. 

Figure 3. L. nigrinus adult feeding temperatures and 

daylength during oviposition. 

0

2

4

6

8

10

12

January

(10:14)

February

(11:13)

March

(12:12)

April

(12:12)

Day Temperature

Night Temperature

Month (L:D) 

Temperature (°Celsius) 

128 
Chapter 11: Rearing Labs and Distribution of Predators for Release 

Figure 4: 	 L. nigrinus larval rearing system utilizing steel 

funnels. 

When HWA has broken its summer diapause in 

the field in early October, L. nigrinus aestivation 

containers are then moved to lower temperatures to 

encourage emergence (12-15°C; 12:12 L:D). Adults 

will typically begin emerging in large numbers about 

2 weeks after the temperature is dropped and will 

continue to emerge from these containers until mid 

to late December. If labs are doing adult releases, 

they are typically placed in the field during this time. 

Laricobius osakensis 

Protocols for the mass rearing of L. osakensis are 

currently being developed at Virginia Tech. 

Dr. Ashley Lamb has completed initial groundwork, 

which is being followed by Ph.D. student 

Ligia Cota Viera and the Virginia Tech mass 

rearing staff. Many procedures will be identical 

to that of L. nigrinus; however, oviposition 

temperatures will likely be slightly lower. 

Sasajiscymnus tsugae 

Carole Cheah and the staff at the Connecticut 

Agricultural Experiment Station (CAES) developed 

initial small-scale rearing procedures for S. tsugae. 

These methods were then built upon and adjusted 

for mass rearing by Dan Palmer and Jennifer DeSio 

at the New Jersey Department of Agriculture 

(PABIL) and now stand as the foundation for 

procedures utilized at other rearing labs today. Each 

lab has tweaked protocols to suit their facilities’ 

Figure 5. 	 L. nigrinus soil aestivation containers. 

capabilities and limitations. Ovipositioning adults 

are typically held on hemlock bouquets placed 

in soaked floral foam in 1 gallon glass jars with 

modified, ventilated lids. Eight to ten twigs, 

15-20 cm in length, comprise one bouquet (Fig. 6). 

Gauze is placed between foliage to help estimate 

egg totals, as roughly 50% of the total number of 

eggs will be laid there. Beetles are typically held at 

densities of 15 per container with a 10:5 female: 

male ratio. Honey is added as a supplemental 

diet. Glass jars are held on their sides on racks 

at 24-25 °C, 55-65% RH, 16:8 L:D (Fig. 7). 

Adults are removed after one week and the 

foliage and gauze are placed into a rearing cage. 

Adults are placed on fresh bouquets and the 

process is repeated. If egg releases are being made 

and if outdoor temperatures are consistently 

above 15 °C, they are done at this time. 

129 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Figure 6. S. tsugae ovioposition jars. 

Figure 7. S. tsugae oviposition jars on racks. 

Foliage and gauze containing eggs are placed in 

rearing tents (BugDorm – 152 cm ×152 cm 

× 152 cm) until an estimated 1500 eggs 

(750 from gauze) is reached (Fig. 8). 

Twigs are placed into foam blocks with the 

gauze sandwiched between. Fresh, infested 

hemlock branches are added 2 times per 

week and after 20 days a honey strip is 

placed in each cage 1 time per week. 

Any adults found during this time are collected 

and placed into storage cages. Rearing cages are 

broken down after roughly 30-50 days. All foliage is 

examined and any beetles found are collected. Up to 

400 adults are placed into storage cages 

at 18-20 °C, 55-65% RH, 16:8 L:D 

at least 3 weeks prior to release to ensure they 

are sexually mature. Adult releases begin when 

outdoor temperatures stay above freezing. 

Figure 8. BugDorm rearing tents. 

130 
 

 

Chapter 11: Rearing Labs and Distribution of Predators for Release 

Adults are stored in summer with protocols similar 

to those used for the rearing cages. Roughly 

400 adults are held in each cage at temperatures 

of 14-20 °C, 55-65% RH, 12:12 L:D, and 

foliage blocks are replaced every 45-60 days. 

Scymnus sinuanodulus 

Since it is so similar to S. tsugae, protocol 

development for the mass rearing of S. sinuanodulus 

was again based on the foundations built by 

Carole Cheah at CAES and Dan Palmer and 

Jennifer DeSio at PABIL. Also, as with S. tsugae, 

individual labs have tweaked various procedures 

to meet their particular needs. More specifi cally, 

a multi-lab collaborative effort to fi ne-tune 

protocols for operational egg releases was headed 

up by the staff at the University of Georgia rearing 

facility. Rearing begins in February after HWA 

is confirmed to be developing eggs in the fi eld. 

Adults are held at densities of 9 (6:3 female:male) 

on 5 twigs of infested hemlock. Temperatures run 

from roughly 12-14 °C, RH 65%, 12:12 L:D. 

Gauze is also used as egg indicator in oviposition 

jars but S. sinuanodulus will only lay roughly 1⁄4 

of its eggs there. Twigs are removed and refreshed 

weekly through June. Egg releases are done with 

the resulting twigs. If rearing to the adult stage, 

resulting twigs will be placed in BugDorms for larval 

development. The use of ample quantities of fresh, 

heavily infested hemlock is critical since larvae can 

become cannibalistic. Fresh foliage is added weekly. 

Larval rearing temperatures are set to 20-22 °C, 65% 

RH and 16:8 L:D. Adults will begin to appear in 

7-8 weeks. They are collected and stored at 20-22 °C 

for 4 weeks at 16:8 L:D, 65% RH. Th ese conditions 

allow maturation and mating to occur. Th ey are 

then moved to 12-14 °C at higher densities (50-60 

adults) until October when beetles are released. 

Scymnus coniferarum 

S. coniferarum is currently being studied in 

quarantine at Virginia Tech to confi rm suitability 

as a biological control agent of HWA. Preliminary 

rearing techniques are being developed; however, 

rearing will likely be similar to other Scymnus 

species. Adults will typically lay more eggs at 26 

°C but will die off sooner. Adults are therefore 

held at 18-20 °C as a compromise. Th ey’re held 

at densities of roughly 12 with a 2:1 female:male 

sex ratio, although it is difficult to distinguish 

between sexes. Gauze is also used in oviposition 

bouquets to estimate the number of eggs laid. 

Larvae are reared at 18 °C and adults emerge 

in roughly 50 days at this temperature. 

GENERAL ISSUES 

AND PRODUCTION LIMITATIONS 

Mortality before Release 

(S. sinuanodulus and S. tsugae) 

Adults are typically held for several weeks before 

release to allow for mating and feeding; however, 

mortality is often high during this time. 

Early Emergence ( L. nigrinus) 

A significant challenge when rearing L. nigrinus 

is that of early emergence of adults before HWA 

has broken diapause in the fi eld. L. nigrinus adults 

will not feed on aestivating HWA nymphs so a 

great deal of mortality can occur if beetles are 

emerging from the soil early in the lab. A small 

level of early emergence typically occurs at all 

labs each year; however, some years have been 

particularly high. This is thought to happen when 

L. nigrinus larvae do not receive ample nutrients 

during their maturation process. Other unknown 

factors likely play into this as well. Mortality levels 

can be devastating. Holding early-emerged adults 

at low temperatures and off ering supplemental 

artificial diets helps to stave off high mortality 

levels before HWA breaks. Holding temperatures 

of at least 4 °C greatly increased survival. 

Staffi ng 

High turnover rates have been shown to negatively 

affect success rates at the various labs. The rearing of 

each species is very labor intensive and challenging. 

L. nigrinus, for example, only has one generation 

per year. It takes several seasons to fully understand 

what is necessary to be successful. With high 

turnover, you lose that gained knowledge and 

must start over with training. Some labs also have 

trouble finding funding to hire ample staff (Fig. 9). 

131 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Figure 9. Rearing lab staff. 

Equipment/Rearing Space 

Because biocontrol agents of HWA are winter 

active, this presents a unique challenge for rearing 

in terms of temperatures. Some predators require 

temperatures as low as 4 °C (39 °F) for extended 

periods of time. Temperatures of roughly 12 °C 

(54 °F) must be available in large spaces for rearing 

of L. nigrinus larvae. Most rearing facilities have 

faced the challenge of retrofitting old buildings and 

equipment to create these large spaces. Even with 

the best equipment, there are failures, which can 

cause temperature spikes or drops. Th ese fl uctuations 

can be devastating to colony health. Several labs 

have seen high mortality due to this reason. 

Food Quality 

As the HWA infestation front moves westward 

and trees continue their rapid decline, it has 

become increasingly diffi cult to fi nd heavy 

adelgid infestations on relatively healthy trees 

with which to feed colonies. Signifi cant travel 

time is involved for some labs to find the best 

infestations, often requiring overnight trips. Other 

labs must have food shipped in, which presents 

problems in terms of quality of prey health. 

Since such a high volume of food is necessary 

for rearing, especially in the spring, this becomes 

the primary challenge for most labs (Fig. 10). 

Pathogens 

As touched upon in other chapters of this 

publication, pathogens such as microsporidia are 

Figure 10. Infested hemlock used for predator rearing. 

a significant issue for rearing predators in the lab. 

Colonies are routinely tested for the presence of 

microsporidia since this organism is found occurring 

naturally in wild populations. Since most L. nigrinus 

rearing begins with yearly introductions of insects 

collected in the wild, it is important to do these 

tests. Infections in the lab can be devastating to 

fecundity levels and very diffi cult to eradicate. 

ACKNOWLEGEMENTS 

Paul Arnold, LayLa Burgess, Carole Cheah, 

Hugh Conway, Mark Dalusky, Jen DeSio, 

Cera Jones, Kathy Kidd, Ashley Lamb, Pat 

Parkman, Andy Roberts, Ligia Cota Viera. 

LITERATURE CITED 

Cheah, C.A.S.-J.; Montgomery, M.E.; Salom, 

S.M.; Parker, B.L.; Costa, S.; Skinner, M. 2004. 

Biological Control of Hemlock Woolly Adelgid. 

Reardon, R.; Onken, B., tech. coords. FHTET2004-04. Morgantown, WV: U.S. Department 

of Agriculture, Forest Service: 22 p. 

Lamb, A.B. 2005. Evaluating the Suitability 

of Laricobius nigrinus Fender [Coleoptera: 

Derodontidae] as a Biological Control Agent for 

Hemlock Woolly Adelgid, Adelges tsugae Annand 

[Hemiptera: Adelgidae]. Blacksburg, VA: Virginia 

Tech: 166 p. Ph.D. dissertation. 

132 
CHAPTER 12: MICROSPORIDIAN DISEASE IN PREDATORY BEETLES
 

Leellen Solter1, Wei-Fone Huang1, Brad Onken2 

1

Illinois Natural History Survey, Prairie Research Institute, University of Illinois, Champaign, IL 

2

U.S. Forest Service, Northeastern Area State and Private Forestry, Morgantown, WV 

INSECT PATHOGENS IN THE FIELD 

AND LABORATORY 

Like all other living organisms, insects continually 

interact with a variety of natural enemies that 

include predators, parasites and pathogens. Th ese 

interactions stabilize insect populations over 

the long term (Anderson and May 1981), but 

immediate effects on individuals or populations 

can be severe when the prevalence of natural 

enemies in the host population increases. Pathogens 

typically persist in insect populations at low, 

enzootic levels, but may increase exponentially, 

often when host population density increases, 

causing local or large-scale and, typically, cyclic 

host population declines (Fuxa and Tanada 1987). 

Insects reared in laboratory colonies are usually 

protected from most of their naturally occurring 

predators and parasites, but pathogens are more 

insidious because often they are not detectable or 

easily identifiable without the use of specialized 

microscopic and molecular tools. Pathogens that 

initially occur in colonies at low prevalence or at low 

infection intensity in the hosts may not immediately 

produce noticeable effects, and a low level of host 

mortality is generally accepted as “normal” in a 

colony. Nevertheless, a single insect harboring a 

highly infectious pathogen can, when introduced 

into the colony or is used to found a colony, initiate 

an epizootic. The vast majority of pathogens are 

orally transmitted via feces, oral exudates and/or 

decomposing tissues, and some are additionally 

transmitted from infected females to their off spring. 

The laboratory colony provides a highly favorable 

environment for transmission of pathogens 

because of the high densities of insects being 

reared in confined spaces, and build-up of disease 

can quickly compromise expensive, long-term 

efforts to establish and maintain insect colonies. 

IMPORTANCE OF PATHOGEN-FREE 

COLONIES FOR BIOLOGICAL CONTROL 

PROGRAMS 

Under the authority of the Plant Protection Act 

of 2000, natural enemies of pest insects that are 

collected for use outside their native environments 

in biological control programs are regulated by 

USDA APHIS Plant Protection and Quarantine. 

A PPQ 526 permit is required and natural enemies 

are held in quarantine for a period of time after 

arrival in the U.S. to assess their specificity to the 

host and determine whether they carry diseases. Any 

unusual mortality is investigated but infections may 

be latent in the host at the time of collection, or 

produce chronic effects that are diffi cult to observe, 

allowing diseased individuals to escape detection and 

resulting in much higher levels of prevalence and 

infection intensity than typically occurs in the fi eld. 

Because biological control programs usually require 

mass rearing to increase numbers of biological 

control agents for release, it is critical that costly 

and time-consuming efforts are not destroyed by the 

build-up of pathogens in laboratory colonies. Early 

detection and mitigation can avoid debilitation 

or complete loss of the colony (reviewed by Etzel 

and Legner 1999), as well as introduction of exotic 

diseases into new environments where other natural 

enemies of the targeted pest may be compromised. 

133 
 

 

 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

It is nearly impossible to ensure that insect colonies 

are completely pathogen-free, particularly because 

chronic submicroscopic viruses are extremely 

difficult to detect and may not cause acute 

mortality, but many pathogens are detectable 

and, if prevalence levels are initially low, it is 

sometimes possible to “cure” the colony. 

MICROSPORIDIAN INFECTIONS 

Some of the most commonly detected pathogens 

in laboratory insect colonies are the Microsporidia. 

Single-cell organisms related to Fungi, microsporidia 

are obligate pathogens—they can only reproduce 

within the cells of their hosts (Tanada and 

Kaya 1991) (Fig. 1). Of the > 1,200 species 

that have been described from vertebrate and 

invertebrate animals, over half are pathogens of 

insects (Becnel and Andreadis 1999). Th ey are 

typically chronic pathogens causing slow larval 

development, increased larval mortality, decreased 

adult lifespan and reduced fecundity (Brooks 

1988, Becnel and Andreadis 1999). Th ese eff ects, 

while they may not be immediately devastating, 

eventually reduce colony growth and prevent 

production of sufficient numbers of natural 

enemies to support a biological control program. 

Microsporidia typically infect the gut tissues or are 

systemic, and mature infective spores are passed 

in the feces and/or when dead infected insects 

decompose, contaminating the food source. 

Many species are also transmitted to the off spring 

of infected females in or on the surface of the 

egg, and mortality is typically high for larvae 

that are infected as embryos or neonates (Solter 

2006). Infections are rarely detectable via visual 

inspection of the host and the chronic nature of 

the disease may allow the pathogen to build up 

to very high levels before noticeable mortality 

occurs. Several generations may appear to be 

healthy and then, quite suddenly, the adults fail 

to reproduce and the colony declines or is lost. 

Figure 1. Developing microsporidian stages in midgut 

epithelial tissues of Sasajiscymnus tsugae. 

Microsporidian disease is detectable using light 

microscopy at 250x-500x. The oval or “jellybean” 

shaped infective spores are typically 3-5 microns 

long and are brightly refractive, particularly 

when viewed with a phase contrast microscope 

(Fig. 2). Detection usually requires dissection 

of the host and examination of tissues, but if 

prevalence is high in the colony, the spores 

can sometimes be detected in the feces. 

Figure 2. Mature spores of a microsporidium isolated 

from Laricobius nigrinus. 

134 
Chapter 12: Microsporidian Disease in Predatory Beetles 

MICROSPORIDIA IN SASAJISCYMNUS 

TSUGAE AND LARICOBIUS NIGRINUS 

In 2002, an unprecedented high level of mortality 

occurred in a laboratory colony of the HWA 

predator Sasajiscymnus tsugae and was attributed to 

a microsporidian infection. The infection prevalence 

increased over one year from 12% to approximately 

50% of the colony beetles. Subsequent examinations 

of other laboratory-reared S. tsugae colonies 

revealed that microsporidia were common and 

were associated with high levels of mortality. As 

investigations intensified, microsporidia were 

isolated from several additional laboratory colonies 

of S. tsugae and were epizootic in one of the 

colonies. Screening of the F1

 generation of fi eld 

collected S. tsugae adult beetles from Japan resulted 

in one detectable infection. A microsporidian 

species was also isolated from the HWA predator, 

Laricobius nigrinus, collected from western hemlock 

near Seattle, Washington in 2005. Microsporidia 

were not observed again in the field or in laboratory 

colonies of L. nigrinus until late 2010 when 

isolates were found in another site near Seattle, 

Washington and in a laboratory colony of beetles 

originating from Idaho. Prevalence of infection in 

the L. nigrinus laboratory colony exceeded 50%. 

Because the 2005 observations suggested a fi eld 

prevalence of 20%, microsporidia may also be 

important in field populations of L. nigrinus. 

There are several reasons to be concerned about 

microsporidian infection in the HWA predators. 

These typically chronic pathogens have signifi cant 

deleterious effects on their hosts. In the laboratory, 

infected beetles easily transmit the pathogen due 

to high density rearing, and infection levels and 

prevalence increase quickly. Infected adults may 

appear to be “normal” and carry on their usual 

activities, but oviposition may be reduced and 

infected larvae fail to mature. Infections in L. 

nigrinus were systemic (Fig. 2) and male testes 

were filled with spores. The colony failed to 

reproduce and mortality was high. In the fi eld, 

infection may result in high winter mortality, 

compromising the release project. A preliminary 

semi-field experiment in one laboratory resulted 

in 90% winter mortality of infected S. tsugae. 

Laboratory host specificity testing suggested that 

one S. tsugae microsporidium can infect several 

species of predatory beetles that are either being 

reared for release or are under study for use in the 

HWA biological control program. Whether this 

physiological susceptibility is important in the fi eld 

is unknown; studies are in progress to determine 

whether the microsporidia infect and are persistent 

in reproducing populations of released beetles. 

SCREENING OF PATHOGENS IN 

COLONIES OF HWA PREDATORY BEETLES 

Molecular studies are underway to determine 

the identity of microsporidia infecting HWA 

predatory beetles. Current data suggest that several 

microsporidian species infect beetles in mass rearing 

programs and those under study for use in the HWA 

biological control program. These include at least 

two distinct Nosema-type species and a Tubilinosema 

sp. in S. tsugae laboratory colonies; a diff erent 

microsporidian species yet to be characterized in an 

F1

 generation S. tsugae specimen reared from adults 

collected in Japan; a Nosema-type species in a fi eldcollected Scymnus coniferarum from Washington; 

a Nosema similar to one of the S. tsugae isolates 

in S. sinuanodulus reared in two laboratories; and 

at least one species in field and laboratory-reared 

L. nigrinus. Three of these microsporidian species 

produced laboratory epizootics in three diff erent 

colonies and compromised the rearing programs. 

True entomopathogenic fungi have not been 

observed frequently in the HWA program, although 

some S. tsugae individuals in one colony were 

apparently infected with a Beauveria bassianalike fungus (Fig. 3). No entomopathogenic fungi 

have been reported to cause serious epizootics in 

laboratory colonies. Occasionally, saprophytic 

fungi have apparently overgrown some of the 

cultures, resulting in higher than usual mortality. 

Parasites such as mites and nematodes have not 

been reported, nor have less common pathogens 

such as protozoans (e.g. trypanosomes, amoebae 

135 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Figure 3. Entomopathogenic fungus infecting 

Sasajiscymnus tsugae. 

and gregarines). Rearing lab managers should be 

watchful for unexplained mortality; viral diseases 

of insects, for example, are very diffi cult to 

recognize because most are not detectable under 

light microscopy and some, like the microsporidia, 

may cause chronic, relatively low intensity 

infections, or occur at low prevalence levels until 

laboratory rearing conditions stress the hosts 

and allow for unusual levels of transmission. 

MITIGATION OF MICROSPORIDIAN 

DISEASE IN LABORATORY COLONIES 

To reduce the possibility of contaminating 

laboratory colonies with infectious organisms 

and, thus, compromising mass rearing and fi eld 

releases, a series of protocols for handling and 

evaluating colonies should be developed and 

carefully followed. Broadly considered, individual 

insects should not be crowded into cages during 

field collection. Insects collected from diff erent 

populations should be held in separate collecting 

cages and equipment such as aspirators, vials and 

cages should be cleaned with a microbiocide (5% 

bleach is adequate) and rinsed well before collecting 

from different populations. If possible, diff erent 

field populations should be reared in isolated groups 

until screened for pathogens. Field collected insects 

also should be isolated from the main laboratory 

colonies until they are screened for pathogens and 

parasites, and for at least one generation before 

being added to the main colony. Screening of fi eld 

collected insects to be introduced to the colony 

should include evaluation of all beetles that die 

during transport or in the laboratory, adults after 

mating and oviposition, and all larvae reared from 

field collected adults that die during development. 

Random samples of the offspring of fi eld collected 

insects, even if apparently healthy, should be 

screened before being added to the main colony. 

In addition, the main colony should be regularly 

screened by evaluating dead and post-ovipositional 

adults, and larvae that die during development. 

Cages should be cleaned and sterilized with diluted 

bleach after each rearing. Hygienic practices and 

reducing the overall stress on the beetles during 

the collection, handling and rearing of beetles 

in the laboratory will greatly reduce the risk of 

contamination and spread of many pathogens. 

While individual insects infected with microsporidia 

are seldom curable, colonies that are infected at 

relatively low prevalence levels sometimes can be 

saved. There are few treatments for most insect 

diseases; microsporidia may or may not respond 

to Fumidil (fumagillin, an anti-microsporidia 

treatment sometimes used in honeybee colonies) 

and effects on the host species would need to 

be evaluated. This chemical typically suppresses 

development of the microsporidia (the response 

of microsporidia to the drug is species-specifi c) 

but it rarely, if ever, cures the colony. Removal 

of the treatment usually results in a resurgence 

of the pathogen that can occur very quickly, a 

situation that could have negative implications 

for success of field releases. Heat treatment of 

eggs is sometimes successful for some species 

of microsporidia, but may not completely 

eliminate the pathogen and often causes 

unacceptable levels of mortality in the hosts. 

136 
Chapter 12: Microsporidian Disease in Predatory Beetles 

It is possible to mitigate microsporidian infection 

in insect colonies by initiating new colonies from 

uninfected females, a technique employed in the 

1800’s by Louis Pasteur to control “pebrine disease” 

(Nosema bombycis) in silkworm colonies (Undeen 

and Vavra 1997). It requires a very labor-intensive 

process but is frequently successful when initial 

microsporidian prevalence is relatively low. It is best 

utilized when first establishing a colony or when 

adding field collected insects into an established 

colony. This is a useful protocol even if pathogens 

are not initially observed in the fi eld collected 

insects. For this procedure, males and females are 

mated in isolated single pairs (or sometimes one 

female and two males to ensure mating) and, after 

oviposition is completed or at death, the adults are 

evaluated for infection by destructive sampling. If 

any adult is infected, all eggs produced by the pair 

(or mating group) are destroyed. If the adults are 

uninfected, the offspring are reared in isolation 

from those of other pairs and other colony insects, 

and are evaluated by random sampling during 

development. In addition, all larvae that die are 

checked for disease. If any one F1

 insect from 

the rearing group is infected, the entire F1

 stock 

produced by the breeding pair or group is destroyed. 

(This procedure also can be used for single gravid 

females collected from the field.) If no infections are 

observed, the F1

 generation can be combined with 

other putatively uninfected F1

 off spring. Ideally, 

an F2 generation is produced from these insects 

and is evaluated for infection prior to addition to 

the main colony, but this may not be possible due 

to time constraints, particularly with univoltine 

insects. The “Pasteur Technique” can also be used 

when microsporidia are found in low prevalence 

in a colony by pairing newly eclosed adults. Use 

of this technique should be accompanied by 

scrupulous cleaning and sterilization of the rearing 

facility and of all rearing cages and materials. 

Modern biotechnology can support use of the 

Pasteur Technique, especially for early and rapid 

diagnosis and detection of microsporidian infection 

in large numbers of adults or dead larvae, and 

also for detection of infection in colonies after 

single pair matings have produced off spring. Th is 

group of predatory beetles, however, appears to 

harbor a complex of microsporidia similar in 

diversity to those found in Lepidoptera, so to 

use PCR detection to its full extent, pathogens 

must be identified and stored properly, DNA 

extraction methods need to be refi ned, and 

primer construction is needed when “universal” 

primers for microsporidia do not amplify DNA. 

SUMMARY 

Pathogens are perhaps the most serious of the issues 

associated with mass rearing of insects in biological 

control programs. Because of the chronic nature of 

some of the more insidious disease organisms, they 

are often not noticed until the colony is in danger 

of being completely decimated. Field collected 

insects that will be used to found colonies or will 

be added to existing laboratory colonies should 

always be isolated and evaluated for pathogens so 

that extant rearing programs are not compromised. 

Frequent assessments of colony health that 

include presence and prevalence of pathogens 

should be routine activities for establishment 

and maintenance of mass-reared insects. 

LITERATURE CITED 

Anderson, R.M.; May, R.M. 1981. Th e population 

dynamics of micro parasites and their 

invertebrate hosts. Philosophical Transactions of 

the Royal Society of London. Biological Sciences 

291: 451-524. 

Becnel, J.J.; Andreadis, T.G. 1999. Microsporidia in 

insects. In: Wittner, M.; Weiss, L.M., eds. Th e 

Microsporidia and Microsporidiosis. Washington, 

DC: ASM Press: 447-501. 

Brooks, W.M. 1988. Entomogenous protozoa. 

In: Ignoffo, C.M., ed. Handbook of Natural 

Pesticides. Boca Raton, FL: CRC Press: 1-149. 

137 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Etzel, L.K.; Legner, E.F. 1999. Culture and 

colonization. In: Bellows, T.S.; Fisher, T.W.; 

Caltagirone, L.E.; Dahlsten, D.L.; Gordh, G.; 

Huffaker, C.B., eds. Handbook of Biological 

Control. San Diego, CA: Academic Press: 

125-197. 

Fuxa, J.R.; Tanada, Y. Epizootiology of Insect Diseases. 

New York, NY: John Wiley & Sons: 555 p. 

Solter, L.F. 2006. Transmission as a predictor of 

ecological host specificity with a focus on 

vertical transmission of microsporidia. 

Journal of Invertebrate Pathology 92: 132-140. 

Solter, L.F.; Becnel, J.J. 2007. Entomopathogenic 

microsporidia. In: Lacey, L.; Kaya, H.K., eds. 

Field Manual of Techniques in Invertebrate 

Pathology; 2nd Edition. Dordrecht, Th e 

Netherlands: Springer: 199-221. 

Tanada, Y.; Kaya, H.K. 1991. Insect Pathology. New 

York, NY: Academic Press, Inc.: 666 p. 

Undeen, A.H.; Vavra, J. 1997. Research methods for 

entomopathogenic Protozoa. In: Lacey, L., ed. 

Manual of Techniques in Insect Pathology. New 

York, NY: Academic Press (Elsevier): 117-151. 

138 
CHAPTER 13: DEFINING PC/QC STANDARDS
 

FOR MASS-REARING HWA PREDATORS
 

Allen C. Cohen1, Carole Cheah2, Kathleen Kidd3, and Th om Hodgson4 

1

Department of Entomology, North Carolina State University 


2

The Connecticut Agricultural Experiment Station, Windsor, CT
 

3

NC Department of Agriculture and Consumer Services
 

4

Department of Industrial and Systems Engineering, North Carolina State University
 

ABSTRACT 

We have defined guidelines to help mass-rearing 

laboratories establish programs of process 

control (PC) and quality control (QC) in the 

production systems where Sasajiscymnus tsugae 

and Laricobius nigrinus are reared. We have 

included process and quality characteristics 

that span a range of technological diffi culty and 

organizational levels. The tests for measuring 

these characteristics include biomass, morphology, 

biochemistry, and behavior. We emphasize the 

importance of using statistically-based QC and 

PC systems according to the protocols established 

by the industrial engineering community. 

INTRODUCTION 

Systems of process control (PC), quality assessment 

(QA) and quality control (QC) have been in use 

in industry for nearly 100 years, and they have 

been more recently applied to insect rearing 

over the past several decades. Using PC/QA/QC 

systems properly greatly adds to the value of the 

products in question and to the economy of their 

production and to consumer/user satisfaction. 

Chambers’ (1977) excellent review of quality control 

explains the biological aspects of quality and how 

they can be applied in mass rearing systems. 

MASS-REARING FRAMEWORK 

Importantly, Chambers points out that numbers 

of insects produced in true mass-rearing systems 

over time must exceed 10,000 → 1,000,000 times 

the average fecundity for an individual female 

over the cycle of one generation. Along with the 

numerical constraints (quantities) that Chambers 

sets forth, he also treats the quality assessments 

that result in successful programs. Th ese quality 

considerations have served as the basis of many 

mass-rearing systems and include biological 

features such as weight, mobility measurements, 

search capacity, fecundity, fertility, development 

rate, longevity, and Chambers also reviews other 

biological aspects such as pheromone production 

and response, sound production, and other features 

that reflect the health and vigor of insect to be used 

in large scale programs such as biological control or 

genetic pest management. We note that the rearing 

systems for HWA predators do not produce the 

numbers to qualify as mass-rearing by the standards 

of Chambers; however, the tenets of process 

control and quality control can still be applied 

profitably to these smaller-scale rearing programs. 

Therefore, in this chapter, we focus on development 

of a practical system of process control and quality 

control. We distinguish these concepts as follows: 

1) process is the series of events, procedures, and 

materials involved in the production of the product 

(in this case, the HWA predators); 2) the quality 

of the end-product ( S. tsugae and L. nigrinus) 

139 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

means the relative ability (or capability) of the 

product to do the job for which it is intended. 

In this chapter and in the PC/QC program that 

we are developing, we try to present a systematic 

analysis of the processes involved in mass-rearing 

these HWA predators to allow early detection of 

flaws in the process. We further try to defi ne the 

standards that characterize the end product as 

capable of controlling HWA in hemlock forests. 

We also try to make the PC/QC system one that 

harmoniously fits the current eff orts, minimizes 

the efforts of production teams, and adds to the 

value of the product without adding substantially 

to the cost. In accord with other PC/QC systems 

in industry, all these goals can be achieved by 

application of appropriate techniques that are 

based on careful study of the existing system of 

production. Several authors have treated quality 

control including reviews by Boller (1979), Boller 

and Chambers (1977), Chambers (1977), Calkins 

et al. (1996), and a comprehensive work that 

updates modern QC standards, Dyck et al (2005). 

Reviews of the genetics of mass-production of 

insects are provided by Bartlett (1984, 1985) and 

Mackauer (1976). Finally, an excellent introductory 

statement about the need for quality control and 

process control is provided by Bigler (1989). He 

expressed the treatment of quality control and 

process control in earlier applications to insects: 

Boller and Chambers (1977) divided the overall 

quality of fruit flies reared for sterile insect release 

programmes into major quality components, traits 

and measurable parameters. The question remains 

whether laboratory assessed traits or attributes have 

a predictive value for the performance of an insect in 

the field. Mackauer and Van Den Bosch (1973) and 

Messenger et al. (1976) concluded that it is hardly 

possible to identify attributes which will “precisely” 

characterize an effective biocontrol agent for a 

particular situation. Th e first problem is the clear 

defi nition of what attributes are to be measured. 

PRODUCTION PROCESS 

FOR SASAJISCYMNUS TSUGAE 

An analysis of the rearing process must be 

performed, starting with a listing of all the elements 

of the rearing system. In the case of S. tsugae, the 

components of the process are as follows: 

1) collection of insects to start colonization; 

2) holding P generation adults in containers; 

3) feeding them adelgid prey presented as 

infestations on hemlock; 4 ) supplementing natural 

diet with honey, Wheast, or other supplements; 

5) adding water as a spray or in some other manner; 

6) allowing oviposition and either collecting eggs or 

allowing them to remain in adult cages; 

7) harvesting F1

 generation to start new cage; and 

8) continuing process by repeating steps 1)-8) to 

produce subsequent generations (F2, F3...Fn). At 

some point in the process P → Fn, a harvesting 

step is added where some stage (usually adults) are 

removed from the colony and prepared for release. 

As Chambers (1977) points out, the harvesting/ 

preparation/release step is very important and 

can be the point of great losses in quality and 

failure of the system. However, the scope of this 

chapter is confined to the production steps. 

PRODUCTION OF LARICOBIUS NIGRINUS 

The steps in production of L. nigrinus are similar 

to those involved in S. tsugae production, except 

that the former species includes a step that 

involves a complex life stage where L. nigrinus 

larvae enter the soil to pupate, aestivate for several 

months, then emerge as adults in the fall. 

In Figure 1, we see the five major factors that can 

contribute to the loss of quality in a production 

program for an HWA predator: 1) microbial 

factors, 2) containers, 3) soil factors, 4) diet 

quality, and 5) environment. For example, if the 

production process allows microbial contaminants 

or pathogens to enter the target insects, these 

microbial components can either kill or sicken the 

incipient products (Cohen 2003). One microbe 

that has gained considerable attention in HWA 

predators is a species of protozoan known as a 

microsporidia. Although viruses, bacteria, and fungi 

have received lesser attention in HWA predators 

than microsporidia, they can be equally destructive. 

140 
Chapter 13: Defining PC/QC Standards for Mass-Rearing HWA Predators 

Figure 1. 	 A fishbone diagram that covers fi ve major 

facets of rearing of Laricobius nigrinus. 

Although containers seem to be a simple 

component, their design can lead to many 

problems in rearing HWA predators (and other 

insects as well). Frequently, in rearing situations, 

improvements in cages can influence gas exchange, 

heat exchange, moisture retention, and many other 

factors that can make the difference between a highly 

successful, economically-sound program and a fail. 

For soil-dwelling insects, including L. nigrinus, soil 

features, including texture, moisture content, pH, 

microbial profile, etc. can be of huge signifi cance 

in survival and health of the insects (Johnson 

et al. 2007), and this includes the predators 

that are products of our rearing systems. 

In Figure 2, we extended the hypothesis that overwatering could be a substantial cause of mortality 

by causing the larvae to drown. Conversely, 

desiccation could also be a source of mortality, 

but in our measurements of rearing soils provided 

by the PABIL laboratory, all soil samples tested 

had a water activity of close to 1.00 (equivalent to 

100% relative humidity). The sphagnum moss in 

the artificial soil is known to hold moisture to help 

keep soil air spaces humid. We also had input from 

rearing labs that they felt that larval nutrition had 

a strong impact on survival of larvae in the soil. 

Figure 2. 	 A hypothetical Pareto plot that “dissects” or 

demonstrates the various potential causes of 

mortality (failure) in the soil phase of rearing 

L. nigrinus. The possible causes of failure 

or mortality were derived from discussions 

with workers in L. nigrinus rearing labs and 

from the literature such as Johnson et al. 

(2007). 

In the soil choice pupation experiment, soil sizes 

represented by coarse and fine hemlock soil, 

approximated coarse sand and larger particles 

vs. fine sand, silt, and clay-sized particles, sifted 

out from soil collected under a planting of urban 

hemlock trees. Results indicated that when L. 

nigrinus had a choice of different soil textures to 

pupate in, all pupae were located in coarse hemlock 

soil, and none in either the standard soil mixture 

used in laboratory rearing of this species or fi ne soil 

(Fig. 3). In all cases of successful pupation, new 

adults emerged from pupal chambers in the fall. 

141 
 

 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

a 

b 

c 

Figure 3. 	 Plexiglass soil sandwiches used to test choices of Laricobius nigrinus mature larval responses to soil 

moisture, soil texture, organic content, pH, or other soil parameters. The dimensions of the sandwiches 

are about 9 cm x 10 cm and about 1 cm in width. (a) The soils used in these chambers are, from left to 

right, standard sand and milled sphagnum mix used in rearing L. nigrinus, fine and coarse hemlock soils 

sifted to particle sizes of ≤ 0.60 mm and > 0.60 mm, respectively; (b) depth at which a pupal chamber 

of L. nigrinus (circled in red) was detected in coarse hemlock soil; (c) close-up of a newly formed pupa 

of L. nigrinus in pupal chamber in coarse hemlock soil (photos by C. Cheah). 

As we see in Figure 4, the collection and 

presentation of data allows us to treat quality in 

an objective and quantifiable way. Th e control 

chart in Figure 4 shows the weights measured in 

female S. tsugae produced over a 45 week period, 

where collections of 100 beetles were weighed, 

and average weights calculated. It is evident from 

this chart that during two periods over the whole 

rearing interval were notable for weights dipping 

below the lower control limit (LCL). Th ese declines 

in weight during Week 24 and Week 38 must be 

considered indications that the process was out 

of control and that the products were of inferior 

and unacceptable quality during these periods. 

It is important to note that the biomass or weights 

of S. tsugae have not definitively been shown 

to be indicators of quality, i.e. to be related to 

the desirable characteristics of voracity, large 

search capacity, longevity, and high fecundity, 

but we are assuming that the correlations 

exist. Therefore, the underlying hypothesis 

of this portion of our study is that a certain, 

minimal mean body weight is correlated with 

the biological characteristics stated above. 

Figure 4. 	 Range charts based on data from 

Sasajiscymnus tsugae program at the North 

Carolina Department of Agriculture’s 

Biological Control Laboratory. 

142 
Chapter 13: Defining PC/QC Standards for Mass-Rearing HWA Predators 

This leads to the question of “upstream” issues 

having potential effects on production. Clearly, 

in light of the high cost of the HWA predators, 

it would be advantageous to catch the problems 

that cause defects before they are manifested in 

the final product. This is especially important in 

predators such as S. tsugae and L. nigrinus, which 

have very long life cycles with a large input of 

materials (hemlocks infested with HWA, cages, 

soil) and labor. The entire rearing process from 

adults in one generation to adults in the next 

generation takes months (nearly a year, in the 

case of L. nigrinus). Figure 5 and the ensuing 

discussion explain the distinctions between 

product control and production control. 

Strictly speaking, when we discuss product 

quality, we are focusing on the characteristics of 

the end-product and how they meet the needs for 

which the product was intended. In our case, this 

product must have the characteristics that lead to 

the control of HWA to reduce pest populations 

below a biological threshold, which translates into 

preventing HWA from killing hemlock trees. In the 

discussions by Leppla and Fischer (1989) and Penn 

et al. (1998), there is a separation of “process” and 

“production” control where the process is analyzed 

by measurement of the materials, including the 

biological materials such as the immature stages 

of the insects being produced, are examined and 

evaluated. We suggest that the diagram could be 

simplified for convenience, with Production Control 

and Process Control fused into one category. In our 

model, overall quality control would be divided 

into Process Control (where all elements of the 

production are potential elements of scrutiny) 

and Quality Control (where the final product is 

measured in a context of standards developed to 

assure product capability to perform as expected). 

RECOMMENDED PROCESS CONTROL 

AND QUALITY CONTROL SYSTEM 

FOR HWA PREDATORS 

This leads to the specifics of the process control 

measures in rearing Laricobius and Sasajiscymnus. 

Figure 5. 	 The components of a quality control system 

for mass-reared natural enemies, adopted 

from Leppla and Fischer (1989) and from 

discussions by Penn et al. (1998). 

The areas that we have identified for Process 

Control are 1) environment, 2) diet materials, 

3) microbial factors, 4) soil factors, 5) container 

factors, and 6) genetics. We note that genetics 

would be important to assess, but there has 

been little background established for assessing 

genetic factors. Therefore, we summarize our 

recommendations for the further development 

of measurement of the fi ve components 

mentioned here and illustrated in Figure 1. 

1) 	 Environmental Factors can be evaluated 

by the use of a temperature, humidity, and 

light measurement data logger (or similar 

device). We have found that Hobo-type data 

loggers are small enough to fit into rearing 

containers, and they can be programmed 

to report temperature, humidity, and light 

intensity over a one-week period. Th e 

accumulated data can be downloaded to a 

computer so that deviations from standard 

conditions can be detected by examination 

of the graphs that the data loggers and their 

software support. This technique is discussed 

in detail by Cohen (2003). It is important to 

mention that for under $500, a laboratory 

can be equipped with four data loggers and 

the software required to read the loggers. We 

143 
 

 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

further suggest that the nearly continuous 

output of the data loggers gives insectary 

workers a much more comprehensive sense of 

deviations in the rearing system. Other kinds 

of sensing systems can be used to detect other 

parameters such as CO2

 or O2 concentrations. 

2) 	 Diet Factors: There is common agreement 

that predators’ diets are among the most 

important determinants of quality. Yet, there 

is no clear definition of what is meant by 

high or low quality diets (Cohen, personal 

observation). Basing the following discussion 

on interviews with rearing personnel, 

judgments of prey quality are based on a) 

the condition of the tree, b) the number of 

woolly masses present, c) the size of the woolly 

masses, and d) the numbers of eggs present 

in egg masses. We have tried to elaborate on 

these factors to include biochemical/biomass 

factors including: a) weight of excised woolly 

masses, b) protein content, c) lipid content, 

d) carbohydrate content, and e) antioxidant 

content of woolly masses. We developed or 

modified tests for measuring 1 to 10 HWA 

for each test. We found that the presence of 

HWA “wool” complicated the analysis of the 

nutritional factors, especially lipid analysis. 

Protein: To determine protein content, 

we refined a dye-binding test according to 

modifications of the method of Heller and 

Sherbon (1976), (see Udy website). Th is 

method involves the homogenization of 

HWA tissue in a solution of Acid Orange 

12 Dye, then centrifuging and measuring 

with a spectrophotometer at 480 nm. When 

the disappearance of color is compared 

with a standard curve established with 

authentic proteins of known concentrations, 

the protein content of individual HWA 

woolly masses can be determined. 

Lipids: The total lipid concentration of 

egg masses is determined by the vanillin 

method, which is a colorimetric procedure 

performed similarly to the protein test, 

using a spectrophotometer. As with the 

protein determination, authentic standards 

are used to establish a standard curve. 

The method is explained by van Handel 

(1988) as is the following analysis of 

carbohydrate concentrations. In the vanillin 

method, the materials to be tested, such as 

insects, are homogenized in concentrated 

sulfuric acid and phosphoric acid, then 

reacted with the vanillin reagent. 

Carbohydrates: Both soluble (free) sugars 

and glycogen can be determined by using 

the anthrone test with samples of HWA 

and comparison with standard curves as 

described by van Handel (1988). Like 

the vanillin/lipid test, the anthrone test is 

performed with sulfuric acid, which breaks 

down (hydrolyzes) the organic components, 

including all kinds of carbohydrates, which 

then react with the anthrone molecules to 

form a colored product whose optical density 

can be read colorimetrically and compared 

with known carbohydrate standards. 

Free-Radical Scavengers: We have 

determined that the most simple and 

comprehensive test of free radical scavengers 

(anti-oxidants) in diet materials and in 

the insect products is the colorimetric 

DPPH method described by Cohen (2003) 

and Cohen and Crittenden (2004). 

3) 	 Microbial Factors: Although there is a potential 

that any of several taxa of pathogens may 

adversely affect the HWA predators in our 

production systems, Dr. Lee Solter (personal 

communication) has stated that the most 

common and serious microbial threats to 

HWA predator-quality are the protozoan 

pathogens known as microsporidia (Phylum: 

Microspora, by the classification of Undeen 

and Vavra 1997). In light of Solter’s fi ndings, 

we strongly recommend that HWA production 

facilities include a search for pathogens. 

A trained technician can perform tests of 

predators by making wet-mounts or Giemsa 

or Gram stains of dry mounts (Undeen and 

Vavra 1997). With these stains and a phase144 
Chapter 13: Defining PC/QC Standards for Mass-Rearing HWA Predators 

contrast microscope, microsporidia-infected 

individuals can be effi ciently recognized. 

We must add, however, that the number of 

specimens that must be examined expressed 

as a percentage of the population that is 

to be released has yet to be determined. 

4) 	 Soil Factors: Predators belonging to the genus 

Laricobius spend more than half of their lifecycle (4-5 months) in the soil as pre-pupal 

larvae, pupae, and newly-eclosed adults, 

which must dig their way out of the soil to 

seek populations of HWA prey and mates. 

We found that L. nigrinus larvae burrow 

about 4-5 cm into the soil (Figure 3b). Once 

larvae reach the appropriate depth, they 

pupate and remain in their pupal cocoons 

(Figure 3c) until they are ready to emerge as 

adults. In Figure 1, we list several soil-related 

factors that we hypothesize as related to 

biological fitness and survival of Laricobius. 

5) 	 Container Factors: The environmental factors 

can be major forces in determining the quality 

or loss of quality in any insect, but researchers 

who study HWA predators have discovered 

that these insects are especially attuned to 

temperatures and light/dark cycles that signal 

seasons and potential prey availability. Given 

the importance of environmental conditions, 

we recommend a rigorous attention to light, 

temperature and humidity conditions in 

insectaries and especially within cages. Th e 

technology of the cages, including sites of 

foliage placement, degree of crowding of foliage 

and beetles, and mechanisms for harvesting 

can greatly affect numbers of predators 

being produced and also the quality of these 

predators. When we consider the architecture 

of a hemlock tree in nature, it becomes clear 

that the arrangement of shorn branches in cages 

can become a maze, rather than a natural series 

of corridors for beetles to discover their prey. 

As far as predator density is concerned, the 

derodontids and the coccinellids in the HWA 

predator programs are not strongly cannibalistic 

as are some predators, but they clearly can 

become competitors when resources such as 

high quality prey are scarce or inadequate. 

6) 	 Genetic Factors: Too often, declines in 

production and/or quality of insects is attributed 

to genetic truncation or deterioration, but 

too seldom has the genetic deterioration 

hypothesis been confirmed as being causative 

(Bartlett 1985). In fact, Cohen (2003) has 

summarized a number of failings in diet or 

environment, or simply personnel errors that 

were documented to cause declines in quality 

or production numbers. Hopefully, with 

molecular methods having become readily 

available, insect rearing programs will be able 

to develop a clearer understanding of causes 

and effects in genetic truncation or genetic 

shifts that are clearly inherent in mass-rearing. 

THE FINAL PRODUCT: 

QUALITY CONTROL OF PREDATORS 

THAT ARE TO BE RELEASED 

The tests of quality that we recommend for both 

species are 1) weights of adult beetles, 2) sex ratios, 

3) internal morphology/development, 4) protein 

content, 5) carbohydrate content, and 6) voracity. 

1) 	Weights of individual adults can be determined 

with a balance sensitive enough to read to 

0.01 mg (10 μg), or if a less sensitive balance 

must be used (such as analytical balances 

that read to 0.10 mg (100 μg), collections of 

either 10 individuals or 100 can be weighed 

in pools. Weights can be evaluated for their 

fit to process control charts (Figure 4). 

2) 	Sex Ratio: For S. tsugae, sex ratios can be 

determined externally, using live beetles. 

For L. nigrinus, sexing would have to be 

done by examining internal morphology. 

Normal sex ratios of both species are 

approximately 1:1 (males: females). 

3) 	Internal morphology/development: Th ese tests 

must be performed with dissected insects, and 

we have determined that a sampling of 6-10 

insects is adequate to reflect the condition 

of the population as a whole (Fig. 6). 

145 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Fat body 

Ovarioles 

Gut 

Figure 6. 	 The internal structures of a female 

Sasajiscymnus tsugae, showing the poorly 

developed ovaries and fat body. It is 

evident from this image that the insect had 

recently fed, but the internal organs are not 

developed to a point where the insect could 

soon reproduce. On a scale of 0-10, with 10 

being fully developed and ready to lay eggs, 

this insect would be rated as a 2-3 (photo by 

A. Cohen). 

4) 	Protein content: We recommend use of the 

dye-binding test known as the Acid Orange 

Test, which is outlined above under “Process 

Control.” In Figure 7, we present a standard 

curve for authentic proteins measured as a 

comparison with the proteins from either 

diet materials (HWA) or predators. 

5) Carbohydrate content: We suggest the anthrone 

test, which is same type of analysis used for 

HWA (above in section on Process Control). 

6) 	Voracity: A voracity or feeding vigor test of a 

sub-sample of beetles that are to be released is 

important. Each beetle should be given a twig 

with 30 HWA adults with eggs confined in a 

9 cm diameter Petri dish at optimal 

temperatures and light cycles for each species. 

After 72h, the number of prey consumed 

should be measured by visual observation of 

disturbed woolly masses and/or consumed 

adelgid stages. The numbers consumed 

should be compared with a control chart 

to determine whether or not the voracity 

is comparable to the established mean. 

Figure 7. 	 A graph of absorbance at 482 nm vs. 

protein concentration (1-4 mg). 

LITERATURE CITED: 

Bartlett, A.C. 1984. Genetic changes during 

insect domestication. In: King, E.G.; Leppla, 

N.C., eds. Advances and Challenges in Insect 

Rearing. New Orleans, LA: U.S. Department of 

Agriculture, Agricultural Research Service: 2-8. 

Bartlett, A.C. 1985. Guidelines for genetic diversity 

in laboratory colony establishment and 

maintenance. In: Singh, P.; Moore, R.F., eds. 

Handbook of Insect Rearing. Vol. I. Amsterdam: 

Elsevier: 7-17. 

Bigler, F. 1989. Quality assessment and control 

in entomophagous insects used for biological 

control. Journal of Applied Entomology 108: 

390-400. 

Boller, E.F. 1979. Behavioral aspects of quality in 

insectary production. In: Hoy, A.; McKelvey, 

J.J., eds. Genetics in Relation to Insect 

Management. The Rockefeller Foundation: 

153-170. 

Boller, E.F.; Chambers, D.L. 1977. Quality 

aspects of mass-reared insects. In: Ridgway, 

R.L.; Vinson, S.B., eds. Biological Control by 

Augmentation of Natural Enemies. New York, 

NY: Plenum Press: 219-235. 

146 
Chapter 13: Defining PC/QC Standards for Mass-Rearing HWA Predators 

Calkins, C.O.; Ashley, T.R.; Chambers, D.L. 1996. 

Implementation of technical and managerial 

systems for quality control in Mediterranean 

fruit fl y (Ceratitis capitata) sterile release 

programs. 4th International Symposium on 

Fruit Flies of Economic Importance; 1994 June 

05-10; Sand Key, FL. Fruit Fly Pests - A World 

Assessment of Their Biology and Management: 

399-404. 

Chambers, D.L. 1977. Quality control in mass 

rearing. Annual Review of Entomology 22: 

289-308. 

Cohen, A.C. 2003. Insect Diets: Science and 

Technology. Boca Raton, FL: CRC Press: 324 p. 

Cohen, A.C.; Crittenden, P. 2004. Deliberately 

added and “cryptic” antioxidants in three 

artificial diets for insects. Journal of Economic 

Entomology 97: 265-272. 

Dyck, V.A.; Robinson, A.S.; Hendrichs, J. 2005. 

Sterile Insect Technique: Principles and Practice 

in Area-wide Integrated Pest Management. 

Dordrecht, The Netherlands: Springer: 787 p. 

Heller, S.N.; Sherbon, J.W. 1976. Interlaboratory 

study of acid orange 12 dye binding for 

measuring protein in meat. Journal of the 

Association of Offi cial Analytical Chemists 59: 

62-66. 

Johnson, S.N.; Zhang, X.; Crawford, J.W.; Gregory, 

P.J.; Young, I.M. 2007. Egg hatching and 

survival time of soil-dwelling insect larvae: 

A partial differential equation model and 

experimental validation. Ecological Modelling 

202: 493-502. 

Leppla, N.C.; Fisher, W.R. 1989. Total quality 

control in insect mass production for insect 

management. Journal of Applied Entomology 

108(5): 452-461. 

Mackauer, M. 1976. Genetic problems in the 

production of biological control agents. 

Annual Review of Entomology 21: 369-385. 

Penn, S.L.; Ridgway, R.L.; Scriven, G.T.; Inscoe, 

M.N. 1998. Quality assurance by the 

commercial producer of arthropod natural 

enemies. In: Ridgway, R.L.; Hoff man, M.P.; 

Inscoe, M.N.; Glenister, C.S., eds. Mass-Reared 

Natural Enemies: Application, Regulation and 

Needs. Proceedings Thomas Say Publications in 

Entomology: 202-230. 

Udy website at: http://www.udyone.com/udydocs/ 

udysys2.shtml 

Undeen, A.H.; Vavra, J. 1997. Research methods 

of entomopathogenic Protozoa. In: Lacey, L.A., 

ed. Manual of Techniques in Insect Pathology. San 

Diego, CA: Academic Press: 117-151. 

vanHandel, E. 1988. Assay of lipids, glycogen and 

sugars in individual mosquitoes: correlations 

with wing length in fi eld-collected Aedes vexans. 

Journal of the American Mosquito Control 

Association 4: 549-550. 

147 
CHAPTER 14: DEVELOPMENT OF ARTIFICIAL DIETS
 

FOR PREDATORS OF HEMLOCK WOOLLY ADELGIDS
 

Allen C. Cohen1 and Carole Cheah2 

1

Department of Entomology, North Carolina State University 

2

The Connecticut Agricultural Experimental Station, Windsor, CT 

ABSTRACT 

We report the results of the process that we 

undertook to develop a rearing system based on 

artificial diets or factitious prey for two species of 

specialist predators of hemlock woolly adelgid, 

Adelges tsugae (HWA). The predators in these 

studies were beetles (Coleoptera), Sasajiscymnus 

tsugae (Coccinellidae) and Laricobius nigrinus 

(Derodontidae). Besides testing more than 50 

diff erent artificial diets, we also attempted to use 

factitious prey, including insect eggs, insect larvae, 

and annelid worms. We also experimented with 

a variety of diet presentation systems that were 

designed to fulfill the feeding requirements of the 

two beetle species and to meet the needs to preserve 

the diets to prevent desiccation and deterioration. 

Although we had little success with most of the 

artificial diets and none of the factitious prey, we 

succeeded in developing several forms of a hen’s 

egg-based diet and a diet presentation system 

that involved both gels made from alginate and 

slurry diets that were made from adhering liquid 

materials to a proprietary solid/capture medium. 

The most successful diets and diet-presentation 

systems allowed adults of both species of predators 

to stay alive and active for several months and 

to return to egg production after being returned 

to natural hosts (HWA) for a few days. Larvae 

fed readily on these chicken egg-based diets, 

but they failed to develop on any of the diets. 

INTRODUCTION 

Biological control of invasive adelgids remains one 

of the most promising means of control for these 

threatening pests. Predators have been the most 

emphasized biological control agents (Grenier et al 

1994). Currently, rearing programs for two widelyused predators ( Sasajiscymnus tsugae Sasaji and 

McClure, Coleoptera: Coccinellidae, and Laricobius 

nigrinus Fender, Coleoptera: Derodontidae) use 

hemlock woolly adelgids, Adelges tsugae Annand 

(Hemiptera: Adelgidae), collected from eastern 

hemlock, Tsuga canadensis. The collection of 

branches infested with adelgids is a costly process 

in terms of labor, travel, and destruction of large 

portions of the trees. Clearly, this method of 

rearing predators imposes severe limitations to 

the scale of production. A further complicating 

factor is that the complexity of the HWA life cycle 

(Figure 1) imposes further limitations on how 

many predatory beetles can be produced and even 

more constraints on the quality of the predators. 

Therefore, an artificial means of supplying high 

quality nutrition to HWA predators would be a 

tremendous advantage to HWA control programs. 

When we specify “artificial nutrition”, we imply 

either 1) insect prey that are not natural hosts to 

the predators, known also as factitious hosts, or 2) 

artificial diet that is composed partially or entirely 

of non-insect derived materials. Several species of 

predators (ladybeetles, pirate bugs and lacewings) 

148 
HWA

Chapter 14: Development of Artificial Diets for Predators of Hemlock Woolly Adelgids 

Laricobius nigrinus 

FALL

SUMMER

WINTER

SPRING

SISTENS

Nymphs

Adults + Eggs

Hatch

PROGREDIENS

Nymphs

Adults +

Eggs

Hatch

Sistens N1

aestivate

Winged

sexuparae

Sasajiscymnus tsugae 

Overwintering adults 

Larvae 

enter soil 

for pupation 

until fall 

HWA

Eggs 

and 

larvae 

New 

Adults 

Eggs and 

larvae 

Adults 

New adults 

emerge 

from soil 

Eggs and larvae 

Figure 1. 	 Life cycles of the hemlock woolly adelgid ( ------ ) and its introduced predators, Sasajiscymnus tsugae 

and Laricobius nigrinus in eastern North America. Solid lines indicate relative active periods of adult and 

larval predator feeding on the adelgid ( S. tsugae; L. nigrinus); Cheah 2011. 

and parasitoids (such as Trichogramma) have 

been reared successfully on factitious hosts such 

as the eggs of brine shrimp (Arijs and de Clercq 

2001, Castane et al. 2006) and the eggs of various 

grain moths such as the Mediterranean grain 

moth Ephestia kuehniella (Pyralidae) (Bonte et al. 

2010) and the Angoumois grain moth, Sitotroga 

cerealella (Gelechiidae) (Abdel-Salam et al. 2001). 

The advantages of using factitious hosts are that 

they are more conveniently available and generally 

less costly than natural hosts (Cohen 2003), but 

the disadvantages are that they are not always the 

most nutritious prey, and they are more expensive 

than artificial diets. Besides nutritional value and 

palatability, a further consideration in various foods 

used in mass-rearing is the packaging of predators’ 

foods. The cuticle of natural and factitious hosts is a 

rather incredible packaging material, being made of 

chitin. While the cuticle is strong and water-proof, 

it can be as thin as 5-10 μM. This means that even 

predators with very small, short mouthparts can 

penetrate the cuticle and gain access to the foods. 

Furthermore, it is important to understand that 

many predators (including those in the current 

study) feed by extra-oral digestion where they 

inject digestive enzymes into the host to pre-digest 

149 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

it (Cohen 1990). The highly nutritious slurry 

that exists as a digestive product is then ingested, 

leaving behind the nearly intact cuticle empty of its 

previous contents. Also relevant to this discussion 

of packaging is the fact that the adelgids as prey are 

very small “packages” of nutrients (about 1-10 μg) 

which the specialist predators are fully adapted to 

feed on and thrive. Likewise, factitious prey, such as 

Ephestia eggs, are small (ca 20 μg), and they resist 

microbial contamination and chemical deterioration, 

in part by being so small and separate from one 

another, that contaminating micro-organisms do 

not spread through an egg mass. All this being said, 

it is clear that a mass rearing system for S. tsugae 

and L. nigrinus would be well-served if it could 

utilize factitious prey instead of natural hosts. In 

Figure 2 (a-d), S. tsugae are pictured feeding both 

on the natural prey (HWA) and artificial diet. In 

this case, the diet presentation is either in the form 

of natural eggs of HWA or the chicken egg diet 

offered as an alginate gel whose surface was made 

into a film by allowing the diet to interact with a 

calcium compound used as a cross-linking agent. 

The description of natural and factitious prey leads 

to a comparison of the issues with artifi cial diets 

and the requirements of a diet presentation system 

for S. tsugae and L. nigrinus. As explained by Cohen 

2003, for an artificial diet and diet-presentation 

system (called “diet system” for this chapter) to 

be considered completely successful it must: 

1. 	 Stimulate robust feeding 

2. Support survival 

3. 	 Support growth and development 

4. Support reproduction 

5. 	 Allow production of continuous 

generations indefi nitely 

6. 	 Support high quality insects that are fully useful 

for their intended purpose (biological control, 

genetic pest management, food for other 

species, conservation, education, research, etc.) 

Relatively few diet systems have been developed to 

meet all these specification, with a rough estimate 

of about 20 basic diets that have been shown to 

support about 300 species (Cohen 2003, Singh 

1977). The accepted diets developed for S. tsugae 

and L. nigrinus, have supported survival in the 

absence of adelgids for several months ( S. tsugae) and 

one month (L. nigrinus), with negligible mortality. 

Adults of S. tsugae fed exclusively on diet were able 

to commence normal reproduction on return to 

adelgid-infested foliage, although eggs were not laid 

while on diet only. Use of the most successful diets 

as supplements have improved laboratory survival 

of adult predators when quality and or quantity of 

adelgid-infested foliage was degraded or depleted. 

Even fewer diets have been developed successfully 

for predaceous lady beetles (Kariluoto et al. 1976, 

Racioppi et al. 1981, Hodek 1996), the most 

notable success being that of Attalah and Newsom 

(1966), who reared eight successive generations 

of Coleomegilla maculata De Geer on a diet void 

of insect materials. Several authors reported using 

factitious diets such as formulations with powder 

honey bee brood (reported in Singh 1977). 

The most successful factitious host diet for a 

coccinellid in our experience was the use of pink 

bollworm eggs to rear multiple generations of 

a b c d

Figure 2. Sasajiscymnus tsugae (a) adult feeding on HWA egg sac, (b) larva feeding on artificial diet, and (c & d) 

adults feeding on artificial diet (photos by C. Cheah). 

150 
 

Chapter 14: Development of Artificial Diets for Predators of Hemlock Woolly Adelgids 

Serangium parcesetosum in the USDA, APHIS Pink 

Bollworm Facility in Phoenix, AZ in the 1990s in 

a program developed by Dr. Robert T. Staten. 

SPECIALISTS VS. GENERALISTS 

In most programs and research efforts to control 

forest pests, specialist predators or parasitoids 

have been used in classical biological control 

systems (Pschorn-Walcher 1977). Because hemlock 

woolly adelgid is an exotic pest, only host specifi c 

natural enemies have been selected, such as the 

HWA specialists, Laricobius and Sasajiscymnus. In 

accord with the complex life cycle of HWA in the 

Northeast (Fig. 1), both species of predators have 

complex life cycles (Cheah and McClure 1998, 

2000; Zilahi-Balogh et al. 2003), which track the 

prey’s periods of dormancy, or reductions or surges 

in nutritional availability (Fig. 1). Th is “tracking” 

and “meshing” of the predators’ life cycle with 

that of the prey is an indication of the degree of 

specialization of the predator. A further indication 

of the high degree of specialization is the fact that 

L. nigrinus has been shown to be restricted either 

nutritionally or in terms of feeding stimulation 

to require HWA to complete its life cycle. Th is 

fastidious feeding response, which excludes 

acceptance of substitute (factitious) prey, raises 

the question about whether rearing L. nigrinus 

can be possible on foods other than HWA. 

The life cycle of the hemlock woolly adelgid is 

complex, with progrediens and sistens generations, 

along with phases where the insects actively feed, 

develop, and reproduce, and other periods when it 

is inactive, e.g., during summer aestivation. Along 

with the morphological and behavioral diff erences 

in HWA during these different phases, there are also 

biochemical differences. Our preliminary work on 

lipid, carbohydrate, and protein content indicates 

that during certain periods, HWA nutritional value 

greatly drops, especially during the late phases of 

their torpor and early stages of feeding activity 

onset. We have found the nutritional composition 

of HWA in their inactive phases drops to less 

than half of what is present in actively feeding 

individuals. This helps explain the periods of 

dormancy in predators that are specially adapted to 

feed on HWA, where their nutritional needs must 

mesh with their hosts. This concept of predator/ 

prey ecological meshing is depicted in Figure 1. 

We have divided our experiments and observations 

into three major categories to cover all aspects 

of our research on development of artifi cial diets 

and rearing systems for predators of HWA: 

I. Feeding on Natural Hosts (Prey) 

II. Feeding on Factitious Hosts 

III. Feeding on Artifi cial Diet 

a. Various artifi cial diets 

b. Diet presentation techniques 

c. Factors in diet stability 

Natural Hosts 

Th e different life stages of HWA off er diff erent 

nutrient rewards to predators, especially with respect 

to 1) overall biomass, 2) protein content, 3) lipid 

content, 4) carbohydrate content, and 5) vitamins 

and minerals. We have begun analysis of these 

factors in several of the life-stages, but because these 

studies are preliminary, we can provide only partial 

results. Using the analytical techniques outlined 

in the chapter (Chapter 13 in this volume), we 

have found that eggs that have been oviposited, 

as well as eggs that are inside females, have a high 

lipid content, with approximately 50% of the dry 

weight of an egg being lipid. The protein content 

ranges from about 30-40% of dry weight and the 

carbohydrate content is less than 10%, leaving 

about 4-5% ash (minerals) and a small biomass 

(less than 3%) composed of other components 

such as nucleic acids and components derived 

from host plants. Th ese findings are in accord with 

Cohen and Patana (1985) regarding the nutritional 

composition of eggs. However, adelgid eggs and 

neonate larvae have a higher lipid content than 

comparable lepidopteran eggs and neonates. 

It is evident from Figure 3 that a great deal of 

lipid material, especially oil, is stored in the eggs 

and remains present in the neonate crawlers. 

The oil is present as a storage material providing 

151 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Stylets inside 

Oil droplets leaked out 

Figure 3. 	 A pre-hatched 1st instar adelgid nymph that 

had a gentle pressure applied to express the 

droplets of oil that are present inside the 

insect (photo by A. Cohen). 

energy and bio-materials for crawlers as they 

settle on host plants and begin their feeding 

forays. The lesson that we learn from analysis of 

natural prey is that HWA predators, especially 

those that feed on adelgid adults with eggs, 

derive a large lipid component from their diet. 

Factitious Prey 

The use of factitious prey or hosts has a long 

history in rearing entomophagous insects. For 

example, the Mediterranean grain moth Ephestia 

kuehniella (Pyralidae) and the Angoumois grain 

moth Sitotroga cerealella (Gelechiidae) have been 

used extensively in commercial predator and 

parasitoid production of green lacewings and 

egg-parasites such as Trichogramma spp. (Cohen 

and Debolt 1983; Cohen and Smith 1998). 

In Figure 4a, an egg that had been fed on by S. 

tsugae is seen with its characteristic depletion of 

materials removed by the predator. The egg is also 

dark as a result of the polyphenol oxidase action that 

takes place during the extra-oral digestion process 

(Cohen 1995). Because S. tsugae fed minimally on 

Ephestia eggs, we also tried to “disguise” the Ephestia 

eggs by placing fresh adelgid wool and exudates 

around the eggs, to determine whether or not the 

HWA materials would enhance predation. All 

uses of factitious prey met with little success and 

a very limited amount of feeding on such prey. 

Feeding on Artifi cial Diet: 

a. Various artifi cial diets 

Starting with the Cohen and Smith 1998 diet, 

we tested more than 100 different diets. Th e 

formulations that we have deemed most successful 

are proprietary combinations of cooked chicken 

egg mixture with functional diet components 

that are suspended in a freeze-dried (proprietary) 

carrier material. The formulation is again freezedried and stored until used with appropriate 

re-hydrating agents, which are discussed below 

(under “diet presentation techniques” and 

“diet preservation or stability techniques”). 

Exudate 

Ephestia eggs 

HWA wool 

Eaten egg 

a b 

Figure 4. Eggs of Ephestia kuehniella that were exposed to Sasajiscymnus and Laricobius, indicating (a) minimal 

feeding and (b) eggs wrapped in adelgid wool (photos by A. Cohen). 

152 
Chapter 14: Development of Artificial Diets for Predators of Hemlock Woolly Adelgids 

b. Diet presentation techniques 

It has long been realized that diet texture and 

presentation techniques can be as equally important 

as the nutritional composition of the diet itself 

(Singh 1977, Cohen 2003). Diet researchers have 

often resorted to the “missing nutrient hypothesis” 

(Cohen 2003) to explain the reasons for a diet’s 

failure. This hypothesis was advanced in early studies 

of insect nutrition by such eminent researchers 

as G. Fraenkel and S. Beck, who had discovered 

cryptic nutrients that were essential factors for 

insects as a whole or target species. For example, 

the discovery that sterols were essential to insects 

went far to explain why earlier formulations that 

lacked sterols failed to support insect growth. 

Similarly, the discoveries of carnitine and choline 

as essential to some insect species further supported 

the “missing nutrient hypothesis”. However, 

extensive analysis and empirical trials during many 

diet-development studies did not reveal hidden 

nutrients that could result in successful artifi cial 

diets for a large number of insect species. Such 

failures may very well be explained by alternative 

hypotheses such as texture failure or packaging (or 

presentation) failure. For example, if an insect uses 

extra-oral digestion, presentation of a complete diet 

that is in liquid form can be unsuitable (Cohen 

1985, Cohen 1995, Cohen and Smith 1998). 

Other issues in diet presentation include whether 

1) the diet is covered with a material that can 

be penetrated by the target insect’s mouthparts, 

2) the diet covering is capable of preventing 

excessive water loss or other degradation factors 

such as contamination with microbes. In 

response, diet researchers have used fi lms such 

as ParafilmTM or Whatman Laboratory Film, the 

dipping of diet in molten wax, or encapsulation 

techniques such as those illustrated in Figure 5. 

Some feeding responses of S. tsugae are shown in 

Figures 6 through 8. Laricobius nigrinus adults 

also appear to accept similar diet formulations 

originally developed for S. tsugae (Fig. 9). 

c. Factors in diet stability 

Once a palatable and nutritious diet has 

been developed and the presentation system 

(encapsulation, film, uncovered diet, etc.) has 

been established, the next stage concerns the 

preservation or stability of the diet. In addition 

to using coverings that protect the diet from 

contamination, techniques that employ heat, 

water activity, extremely low or high pH, and/or 

chemical prophylaxis are conventionally employed. 

a 

b 

Figure 5. A technician (a) making wax-coated capsules (b) with liquid/slurry centers (according to the method of 

Cohen 1983; photos by A. Cohen). 

153 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Timed Observations (over 2 h)

%

0 5 10 15 20 25

S. tsugae Feeding on Egg Diet + Honey

0

20

40

60

80

100

Rep. 1 (n=20)

Rep. 2 (n=20)

Figure 6. Mature adult S. tsugae feeding response 

to egg diet and honey presented on fi lter 

paper. 

Time over 3 Days

0 2 4 6 8 10 12 14 16 18 20

Total no. of adults feeding

0

2

4

6

8

10

12

14

16

18

20

Feeding on pollen diet

Feeding on egg diet

Feeding on HWA

Figure 7. 	 The changes in adult S. tsugae feeding 

preferences over time when presented 

simultaneously with artificial diets and 

HWA. 

a b c 

Figure 8. 	 Sasajiscymnus tsugae adult and larval feeding on an egg-based diet which has been mixed with honey 

or an alginate/calcium base. The diet was presented on (a) filter paper with honey, or as an alginate 

formulation on hemlock twigs for (b) adults & (c) late instar larva (photos by C. Cheah). 

a b c 

Figure 9. Laricobius nigrinus adults feeding on (a) egg diet mixed with honey, (b & c) egg diet-alginate 

formulations (photos by C. Cheah). 

154 
Chapter 14: Development of Artificial Diets for Predators of Hemlock Woolly Adelgids 

The use of heat is nearly universal in diet synthesis 

where diets are heated to the point where all 

microbes are killed (by autoclaving or other 

temperature/pressure treatments) or where vegetative 

stages of microbial contaminants have been killed 

(Pasteurization). We have found that either method 

produces diets that can be made acceptable to the 

predators in terms of maintaining palatability. We 

have demonstrated this fact with the egg-based 

formulation that we call FDFE3 diet. However, 

because the predators can inoculate the diet with 

microbes while they are feeding, heat treatment 

alone cannot continuously protect the diet from 

microbial contamination. Therefore, we resorted to 

lowered water activity and chemical preservatives. 

The concept of water activity (a w) is a useful way of 

expressing the tendency of a diet to suff er microbial 

deterioration and other degradation processes such 

as oxidation. Water activity is expressed as a unitless 

ratio of the actual vapor pressure divided by the 

potential vapor pressure at a given temperature. Th is 

can be conveniently thought of as relative humidity 

divided by 100. Common examples of foods 

with naturally low water activity are honey and 

molasses. Although both of these are liquids at room 

temperature, and although both contain ample 

nutrients, neither supports microbial growth. Water 

activity below 0.60 does not support microbial 

growth of any of the common environmental 

microbes that habitually contaminate our diets. Th e 

range of water activity is between 0 and 1.0, with 

the lower value being associated with concentrated 

sulfuric acid and the higher number being associated 

with distilled water. Most common diets for insects, 

including most of those that we have tested, have 

a water activity of 0.98-0.99 (Cohen 2003). Th is 

means that the diets are well above the minimal 

threshold for support of microbial activity, and 

unless we used special measures to lower water 

activity, we must expect the diets to be subject to 

unimpeded microbial growth. In contrast with 

common insect diets, honey has a water activity of 

about 0.50-0.55; therefore, it is very uncommon 

for microbial growth to take place in honey. 

Therefore, we have incorporated several measures 

to lower the water activity of our diets, including 

the use of freeze drying diets for preservation, then 

hydrating the diets with honey or similarly low 

water activity hydrating sources (such as glycerol 

or sorbitol solutions). We have found honey to be 

an especially suitable medium for presentation of 

diets to adult S. tsugae. In keeping with our eff orts 

to lower water activity of test diets, we have used 

freeze drying (lyophilization or cryodesiccation) 

to prepare diets for storage, transport, then rehydration with appropriate mixtures to help 

retain low water activity. We have adopted freeze 

drying extensively because it not only preserves 

diet components that are highly perishable (such 

as eggs), but it also lends itself to diet presentation 

techniques that maintain low water activity. To 

clarify this concept, we offer this example. If we 

lower the water activity of our egg diet from 0.98 

to 0.10 by freeze drying, then if we re-hydrate the 

diet with honey (water activity of 0.50), the overall diet/honey mixture has a water activity of 0.50 

or less. This mixture will not support microbial 

growth, and it has an extremely long shelf life (well 

over 6 months without refrigeration). We must add 

that the concept of lowering water activity with 

materials that have high concentrations of dissolved 

small molecules (such as sugars, sugar alcohols, 

or salts) is known as the “humectant” strategy. 

However, the two drawbacks of the use of 

humectants is that 1) the material making up the 

humectant/water mixture may be unpalatable to the 

target insects (for example, glycerol is not palatable 

to the predators thus far tested), 2) the mixture can 

be very sticky and cause insects to become trapped 

(we have found this with S. tsugae larvae), and 3) the 

humectant/diet mixture can become hydrated (for 

example, by watering the cage contents to provide 

free water to the predators or to raise the humidity), 

and this hydration can raise water activity to above 

the minimal threshold to prevent microbial growth. 

This leads to the 3rd strategy: the use of low 

pH. We have used various pH lowering agents 

155 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

such as acetic acid, propionic acid, phosphoric 

acid, and citric acid (all fairly well-established in 

insect diets) to lower the diets’ pH below 5.0. 

We have found that acetic acid and citric acid are 

fairly well-tolerated by the predators, and they 

are fairly, but not totally, effi cient at lowering 

microbial growth. For example, we have found 

little problem with bacterial contamination and 

deterioration of our diets, but fungal contamination 

remains a problem in our formulations. 

Therefore, we have had to resort to the 4th 

strategy of using chemical agents to prevent 

fungal growth. We have experimented with 

several anti-fungal chemicals (benzoic acid, 

nystatin, methyl paraben, sodium propionate, 

and potassium sorbate, to mention a few), and 

we have discovered that the most well-tolerated 

agents are propionate and sorbate in their salt 

forms, which makes them soluble in our diets. 

We also have found that the combination of the 

acids and the antimicrobial chemicals have added 

to the efficiency of mold-prevention and the 

duration of the time frame that diets can be kept 

in cages. However, we must add that because of 

the highly nutritious character of the diets, they 

still attract mold growth after several days or 

several weeks’ exposure to the predators. Th erefore, 

mold prevention remains one of the barriers to 

completely successful diets for HWA predators. 

LITERATURE CITED 

Abdel-Salam, A.H.; Abdel-Baky, N.F. 2001. Life 

table and biological studies of Harmonia axyridis 

Pallas (Col.: Coccinellidae) reared on the grain 

moth eggs of Sitotroga cerealella Olivier (Lep.: 

Gelechiidae). Journal of Applied Entomology 

125: 445-462. 

Arijs, Y.; De Clercq, P. 2001. Rearing Orius 

laevigatus on cysts of the brine shrimp Artemia 

franciscana. Biological Control 21: 79-83. 

Atallah, Y.; Newsom, L.D. 1966. Ecological and 

nutritional studies on Coleomegilla maculata 

DeGeer (Coleoptera: Coccinellidae). I. 

The development of an artificial diet and 

a laboratory rearing technique. Journal of 

Economic Entomology 59: 1173-1179. 

Bonte, M.; Samih, M.A.; De Clercq, P. 2010. 

Development and reproduction of Adalia 

bipunctata on factitious and artifi cial foods. 

BioControl 55: 485-491. 

Castane, C.; Quero, R.; Riudavets, J. 2006. Th e 

brine shrimp Artemia sp. as alternative prey 

for rearing the predatory bug Macrolophus 

caliginosus. Biological Control 38: 405-412. 

Cheah, C.A.S.-J.; McClure, M.S. 1998. Life history 

and development of Pseudoscymnus tsugae 

(Coleoptera: Coccinellidae), a new predator 

of the hemlock woolly adelgid (Homoptera: 

Adelgidae). Environmental Entomology 27: 

1531-1536. 

Cheah, C.A.S.-J.; McClure, M.S. 2000. Seasonal 

synchrony of life cycles between the exotic 

predator, Pseudoscymnus tsugae (Coleoptera: 

Coccinellidae) and its prey, the hemlock woolly 

adelgid, Adelges tsugae (Homoptera: Adelgidae). 

Agricultural and Forest Entomology 2: 241-251. 

Cohen, A.C. 1983. Improved method of 

encapsulating artificial diet for rearing predators 

of harmful insects. Journal of Economic 

Entomology 76: 957-959. 

Cohen, A.C. 1990. Fatty acid distribution as related 

to adult age, sex and diet in the phytophagous 

heteropteran, Lygus hesperus. Journal of 

Entomological Science 25: 75-84. 

Cohen, A.C. 1995. Extra-oral digestion in 

predaceous terrestrial arthropoda. Annual 

Review of Entomology 40: 85-103. 

Cohen, A.C.; Debolt, J.W. 1983. Rearing Geocoris 

punctipes on insect eggs. Southwest Entomology 

8: 61-64. 

156 
 

Chapter 14: Development of Artificial Diets for Predators of Hemlock Woolly Adelgids 

Cohen, A.C.; Patana, R. 1985. Chemical 

composition of tobacco budworm eggs during 

development. Comparative Biochemistry and 

Physiology 81: 165-169. 

Cohen, A.C.; Smith, L. 1998. A novel concept in 

artificial diets for Chrysoperla rufi labris: the 

efficacy of solid diet. Biological Control 13: 

49-54. 

Cohen, A.C. 2003. Insect Diets Science and 

Technology. Boca Raton, LA: CRC Press: 324 p. 

Grenier, S.; Greany, P.; Cohen, A.C. 1994. Potential 

for mass release of insect parasitoids and 

predators through development of artifi cial 

culture techniques. In: Rosen, D.; Bennett, 

F.D.; Capinera, J., eds. Pest Management in 

the Subtropics: Biological Control: A Florida 

Perspective. Andover, Hants, U.K.: Intercept 

Press: 181-205. 

Kariluoto, K.; Junnikkala, E.; Markkula, M. 1976. 

Attempts at rearing Adalia bipunctata L. (Col. 

Coccinellidae) on diff erent artifi cial diets. 

Annales Entomologici Fennici 42: 91-97. 

Hodek, I. 1996. Food Relationships. Chapter 6. 

In: Hodek, I.; Honek, A. 1996. Ecology of the 

Coccinellidae. Series Entomologica. Volume 54. 

Kluwer Academic Publishers: 143-238. 

Pschorn-Walcher, H. 1977. Biological control of 

forest insects. Annual Review of Entomology 

22: 1-22. 

Racioppi, J.V.; Burton, R.L.; Eikenbary, R. 1981. 

Th e effects of various oligidic diets on the 

growth of Hippodamia convergens. Entomologia 

Experimentalis et Applicata 30: 68-72. 

Singh, P. 1977. Artifi cial Diets for Insects, Mites 

and Spiders. New York, NY: IFI/Plenum Data 

Company: 594 p. 

Zilahi-Balogh, G.M.G.; Salom, S.M.; Kok, L.T. 

2003. Development and reproductive biology of 

Laricobius nigrinus, a potential biological control 

agent of Adelges tsugae. BioControl 48: 293-306. 

157 
SECTION IV ESTABLISHMENT
 

AND MONITORING
 
CHAPTER 15: WHOLE-TREE ENCLOSURES: A TOOL TO ASSESS
 

INTRODUCED PREDATORS OF HEMLOCK WOOLLY ADELGID, 


ADELGES TSUGAE 

Jerome F. Grant1, Rusty Rhea2, Gregory J. Wiggins1, Abdul Hakeem1, and Paris L. Lambdin1 

1

Department of Entomology and Plant Pathology, The University of Tennessee, Knoxville, TN 

2

U.S. Forest Service, Forest Health Protection, Southern Region, Asheville, NC 

WHY CAGE A TREE? 

Many experimental (laboratory and fi eld) methods 

exist to evaluate the Chapter 15: Whole-Tree Enclosures: A Tool To Assess Introduced Predators of Hemlock Woolly Ad

against insect pests (Grant and Shepard 1985). 

Several of these methods (such as Petri dishes, small 

arenas, growth chambers, greenhouse studies, and 

sleeve cages) have been used to assess predators 

of hemlock woolly adelgid (HWA), Adelges 

tsugae Annand (Hemiptera: Adelgidae), since its 

introduction into North America (Cheah and 

McClure 2000; Cheah et al. 2004; Flowers et al. 

2005; Grant et al. 2005; Lamb et al. 2005, 2006; 

McClure 2001; Zilahi-Balogh et al. 2002, 2005). 

These types of evaluative approaches provide 

important and essential information on the 

predatory capabilities of introduced predators, 

as well as better define their biology, life history, 

predator/prey interactions, etc. However, the small 

size, extremely confined spatial resources, and 

reduced scale of these arenas relative to naturallyoccurring hemlock systems limit their use in fully 

characterizing the impact of introduced natural 

enemies on field-established populations of 

HWA on eastern hemlock, Tsuga canadensis (L.) 

Carrière. New tools to complement the existing 

methods would further enhance and strengthen 

our abilities to assess predatory capabilities of 

introduced predators of HWA. Can existing 

methods be improved or modified to further 

evaluate natural enemies of HWA? Can a wholetree assessment approach be developed to better 

define predatory performance in the fi eld? To 

address these questions, a pilot study was conducted 

to evaluate the feasibility of using whole-tree 

canopy enclosures (i.e., cages) to assess: 1) survival, 

colonization, interactions, and establishment 

of introduced predators on HWA on eastern 

hemlock, and 2) impact of introduced predators on 

population densities of HWA and on tree health. 

This pilot study focused on the use of large cages 

to qualitatively and quantitatively assess the 

successful field application (colonization and 

impact) of three introduced biological control agents 

(Laricobius nigrinus Fender, Sasajiscymnus tsugae 

Sasaji and McClure, and Scymnus sinuanodulus 

Yu and Yao) on populations of HWA in the 

southeastern United States (Tennessee). Th is 

chapter provides a general overview of this project 

and describes how these cages could enhance 

our biological control activities against HWA. 

HOW DID WE DO IT? 

This study was conducted at Blackberry Farm in 

Walland, Tennessee, near the boundary of the 

Great Smoky Mountains National Park. HWA 

was first documented in the Park in 2002 

(Lambdin et al. 2006). Cages were designed in 

2007 through cooperation of personnel from the 

University of Tennessee, U.S. Forest Service, and 

Camel Manufacturing (Pioneer, Tennessee). 

The resulting nylon-screened cages (ca. 8 m tall; 

161 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Fig. 1) were constructed by Camel Manufacturing 

and erected in the field from October through 

December 2007. Fifteen HWA-infested eastern 

hemlock trees (ca. 7-8 m tall) were selected, 

and 12 cages were erected over HWA-infested 

hemlock trees (one cage/tree). Th e following 

treatments (three replications per treatment) 

were applied beginning in December 2007: 

a) releases of L. nigrinus (190 adults/caged tree; 

January 2008), 

b) releases of S. tsugae (300 adults/caged tree; 

March 2008), 

c) releases of S. sinuanodulus (90 adults/tree; 

March 2008), and 

d) control (no releases of predatory beetles). 

In addition, three non-caged, HWA-infested 

trees also were included as control trees. Adults 

of L. nigrinus and S. tsugae were obtained from 

Lindsay Young Beneficial Insects Laboratory, 

University of Tennessee, and adults of 

S. sinuanodulus were obtained from the 

University of Georgia and the U.S. Forest Service 

Northern Research Station, Connecticut. 

Cages remained over the trees for more than 11⁄2 

years and were removed in July 2009. Trees were 

sampled for beetles and HWA densities were assessed 

every three to four months after cages were removed. 

WHAT DID WE LEARN? 

Survival and Colonization 

Following their releases, all three species of 

introduced beetles survived, colonized, and 

reproduced inside the cages, as larvae of each 

species were recovered in 2008 (Grant et al. 

2010b). Adult L. nigrinus were found inside the 

cages in March and November 2008 (about one 

year after initial placement in cages), adult S. 

sinuanodulus were found in April, June, and July 

Figure 1. Nylon-screened cages erected over eastern hemlock, Blackberry Farm, Walland, Tennessee, 2007. 

162 
 

 

Chapter 15: Whole-tree Enclosures: A Tool to Assess Introduced Predators 

of Hemlock Woolly Adelgid, Adelges tsugae 

2008, and adult S. tsugae were found in April, 

May, June, July, and November 2008 (about 8 

months after initial placement in cages). No S. 

sinuanodulus was found during the November 2008 

sampling period. In 2009, prior to cage removal, 

adult and larval L. nigrinus and S. tsugae were 

recovered from trees inside the cages, but no S. 

sinuanodulus was recovered (Grant et al. 2010b). 

Impact on HWA Densities 

In July 2009, HWA densities had declined about 

62% on trees caged with S. tsugae and about 85% on 

trees caged with L. nigrinus, suggesting that both of 

these predatory species reduced HWA populations 

during this 11⁄2-year period (Grant et al. 2010b). 

Establishment and Interactions 

In 2010, on previously caged trees, adult and larval 

S. tsugae (F3 to F5 generations) were recovered 

from February to November (Wiggins et al. 2010a). 

In 2010, adult and larval L. nigrinus (F2 to F3 

generations) also were recovered. Following removal 

of the cages, L. nigrinus and S. tsugae dispersed 

throughout the site (Hakeem et al. 2010b, Hakeem 

et al. 2011). The lack of recovery of S. sinuanodulus 

may be due to the low numbers of adults released 

inside each cage and/or the low vigor of many of 

the released adults. These results also suggested 

that predation of HWA by L. nigrinus and S. tsugae 

benefited tree health (Wiggins et al. 2010b). 

WHAT CONDITIONS ARE APPROPRIATE? 

Several conditions are necessary and certain needs 

must be considered in using whole-tree canopy 

enclosures in forest settings (Fig. 2). Th ese include: 

the availability of a suitable location for 2+ years 

(the location must be easily available for use, 

generally secure, few activities in area, etc.), trees 

of appropriate height (7-9 m tall) for canopy 

enclosures, trees that are healthy and consistently 

shaped, trees infested with appropriate densities 

of HWA (a relatively new infestation is best), trees 

with new growth, trees that are easily accessible 

using a bucket truck or lift, trees that are “solitary” 

(none intermingled/side-by-side), a ground surface 

that is relatively flat or slightly slanted, and wholetree canopy enclosures. Once these conditions and 

needs are satisfied and a design has been developed, 

canopy enclosures can be constructed and deployed 

into the field for use in biological assessments. 

WHAT ARE THE BENEFITS AND 

CHALLENGES OF USING TREE CAGES? 

Once deployed, researchers must consider the 

potential benefits and challenges associated with 

the use of these enclosures. Benefi ts include: 

• 	 a more realistic and complementary 

field assessment of introduced biological 

control agents (than other methods) 

• 	 long-term monitoring of the impact of natural 

enemies on an invasive pest and on tree health, 

as well as predator performance and survival 

• 	 a tool to assess single species or combination 

of species of natural enemies 

• 	 a better understanding of compatibility 

and interactions of introduced and/ 

or native predatory species 

• 	 a better understanding of actual predatory 

expectations in the fi eld 

Figure 2. 	 Whole-tree canopy cage in understory of 

forest/urban interface, Walland, Tennessee, 

2007. 

163 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Th e benefits, however, are accompanied by 

several challenges to the use of these wholetree enclosures. These challenges include: 

• 	 enclosures are still a “controlled” or artifi cial 

environment (how similar is “inside” 

to “outside”? In our cages, however, 

temperature and humidity levels varied 

little between open and caged trees.) 

• 	 nontarget organisms cannot be 

removed thoroughly from the 

trees before they are “caged” 

• 	 environmental stresses (e.g., high winds, 

snow, rain, hail, animals, humans, etc.) that 

can damage cages are diffi cult to control 

• 	 installation of tree cages may require 

specialized equipment and/or be 

labor intensive/time consuming 

• 	 high costs associated with initial 

construction and deployment of cages 

WHAT DOES IT ALL MEAN? 

The use of whole-tree canopy enclosures provides a 

viable technique to assess single or multiple species 

of indigenous or introduced predators against HWA 

on eastern hemlock. Populations of L. nigrinus 

and S. tsugae were successfully established using 

whole-tree canopy enclosures and their impact 

on HWA was documented. Several implications 

can be drawn from the results of this study. First, 

these enclosures allow the use of whole trees 

in natural, field situations and provide a better 

understanding of actual predatory expectations in 

the field. Similar to sleeve cages, tree cages limit 

the dispersal of predators to other trees, while 

limiting influx/colonization of pests from outside 

sources. Unlike sleeve cages, tree cages allow the 

evaluation of predators on a whole-tree scale and 

allow predators and prey to move freely throughout 

the entire tree canopy. Second, whole-tree canopy 

enclosures provide a tool to acclimate introduced 

natural enemies to field situations and conditions, 

as well as a mechanism to establish natural enemies 

in the field. Recoveries of established populations of 

S. tsugae using standard release protocols generally 

occurred five to seven years after release in the Great 

Smoky Mountains National Park and, in some areas, 

these predators have not yet been recovered (Grant 

2008, Grant et al. 2010a, Hakeem et al. 2010a); 

however, introduced predators were recovered from 

hemlock trees after a single release in only two to 

three years using whole-tree canopy enclosures. 

Finally, in addition to providing an assessment of 

predatory performance and survival, this technique 

facilitates the monitoring of long-term impacts 

of these predators on HWA and tree health. By 

confining predators to specific trees for an extended 

amount of time, tree cages allow the comparison 

of direct and indirect impacts of predators on 

the short- and long-term tree health between 

caged trees and those in the open environment. 

The use of whole-tree canopy enclosures should 

enhance our knowledge of the establishment and 

effectiveness of introduced predators of HWA 

to improve their use in management eff orts. 

WHAT DOES THE FUTURE HOLD? 

Whole-tree canopy enclosures also could be used to 

assess other types of tree/organismal interactions, 

and cages can be modified for other predators or 

for other tree pests or tree species. For example, 

a new, more durable tree cage for use in further 

research of predators of HWA has been designed 

and constructed (Fig. 3), and these cages will 

be used in future studies to assess complexes 

of predatory species and to rear large numbers 

of natural enemies for re-release. In summary, 

whole-tree canopy enclosures are a new and 

innovative approach to assessing natural enemies 

for release against HWA, they complement other 

evaluative methods, and can be a vital tool in the 

management of this invasive pest in forest systems. 

164 
Chapter 15: Whole-tree Enclosures: A Tool to Assess Introduced Predators 

of Hemlock Woolly Adelgid, Adelges tsugae 

Figure 3. 	 Newly modifi ed field cage erected over 

eastern hemlock, Great Smoky Mountains 

National Park, 2011. 

ACKNOWLEDGEMENTS 

Appreciation is expressed to Sam Beall and the 

personnel of Blackberry Farm for permitting the use 

of trees and access to their property, and to Mark 

Lester and Camel Manufacturing for construction 

of the tree cages. Many thanks to Pat Parkman, 

Lindsay Young Beneficial Insect Rearing Laboratory 

(University of Tennessee), Mark Dalusky, University 

of Georgia, and Mike Montgomery, USDA Forest 

Service (Northern Research Station) for providing 

the predatory beetles used in this study. Funding for 

this study was provided in part by the U.S. Forest 

Service, Northeastern Area/Northern Research 

Station and U.S. Forest Service, Southern Region. 

LITERATURE CITED 

Cheah, C.; McClure, M. 2000. Seasonal synchrony 

of life cycles between the exotic predator 

Pseudoscymnus tsugae (Coleoptera: Coccinellidae) 

and its prey, the hemlock woolly adelgid Adelges 

tsugae (Homoptera: Adelgidae). Agricultural and 

Forest Entomology 2: 241-251. 

Cheah, C.; Montgomery, M.E.; Salom, S.M.; 

Parker, B.L.; Costa, S.; Skinner, M. 2004. 

Biological Control of Hemlock Woolly Adelgid. 

Reardon, R.; Onken, B., tech. coords. 

FHTET-2004-04. Morgantown, WV: U.S. 

Department of Agriculture, Forest Service: 22 p. 

Flowers, R.; Salom, S.; Kok, L. 2005. Competitive 

interactions among two specialist predators and 

a generalist predator of hemlock woolly adelgid, 

Adelges tsugae (Homoptera: Adelgidae), in the 

laboratory. Environmental Entomology 34: 

664-675. 

Grant, J.F. 2008. Establishing Sasajiscymnus tsugae 

in the South. In: Onken, B.; Reardon, R., eds. 

Fourth Symposium on Hemlock Woolly Adelgid 

in the Eastern United States: Proceedings of the 

Meeting; 2008 February 12-14; Hartford, CT. 

FHTET-2008-01. Morgantown, WV: U.S. 

Department of Agriculture, Forest Service, 

Forest Health Technology Enterprise Team: 

39-44. 

Grant, J.F.; Shepard, M. 1985. Techniques for 

evaluating predators for control of insect pests. 

Journal of Agricultural Entomology 2: 99-116. 

Grant, J.F.; Palmer, D.; Rhea, J.; Taylor, G.; 

Lambdin, P.L.; Deal, I. 2005. Assessment of egg 

releases for establishment of Sasajiscymnus tsugae 

on eastern hemlock. In: Onken, B.; Reardon, 

R., comps. Proceedings of the Th ird Symposium 

on Hemlock Woolly Adelgid in the Eastern United 

States; 2005 February 1-3; Asheville, NC. 

FHTET-2005-01. Morgantown, WV: U.S. 

Department of Agriculture, Forest Service, 

Forest Health Technology Enterprise Team: 

297-303. 

165 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Grant, J.F.; Hakeem, A.; Rhea, J.; Wiggins, G.; 

Lambdin, P.L.; Taylor, G. 2010a. Recovery 

and establishment of introduced predators 

of hemlock woolly adelgid in the Southern 

Appalachians. In: Onken, B.; Reardon, R., 

comps. Proceedings of the Fifth Symposium on 

Hemlock Woolly Adelgid in the Eastern United 

States; 2010 August 17-19; Asheville, NC. 

FHTET-2010-07. Morgantown, WV: U.S. 

Department of Agriculture, Forest Service, 

Forest Health Technology Enterprise Team: 

21-23. 

Grant, J.F.; Hakeem, A.; Wiggins, G.; Rhea, J.; 

Lambdin, P.L. 2010b. Assessment of introduced 

predator species against hemlock woolly adelgid 

on eastern hemlock using whole-tree canopy 

enclosures. In: Onken, B.; Reardon, R., comps. 

Proceedings of the Fifth Symposium on Hemlock 

Woolly Adelgid in the Eastern United States; 

2010 August 17-19; Asheville, NC. FHTET2010-07. Morgantown, WV: U.S. Department 

of Agriculture, Forest Service, Forest Health 

Technology Enterprise Team: 

53-55. 

Hakeem, A.; Grant, J.F.; Lambdin, P.L.; Buckley, 

D.; Hale, F.A.; Rhea, J.R.; Wiggins, G.J.; Taylor, 

G. 2010a. Recovery of Sasajiscymnus tsugae, 

released against hemlock woolly adelgid, Adelges 

tsugae, in the southern Appalachians. Biocontrol 

Science and Technology 20: 1069-1074. 

Hakeem, A.; Wiggins, G.J.; Rhea, R.; Lambdin, 

P.L.; Grant, J.F. 2010b. Co-existence of two 

introduced predatory beetle species of hemlock 

woolly adelgid on eastern hemlock. In: Onken, 

B.; Reardon, R., comps. Proceedings of the 

Fifth Symposium on Hemlock Woolly Adelgid in 

the Eastern United States; 2010 August 17-19; 

Asheville, NC. FHTET-2010-07. Morgantown, 

WV: U.S. Department of Agriculture, Forest 

Service, Forest Health Technology Enterprise 

Team: : 157-158. 

Hakeem, A.J.; Grant, F.; Wiggins, G.J.; Lambdin, 

P.L.; Rhea, J.R. 2011. Establishment and 

coexistence of two predators, Laricobius nigrinus 

and Sasajiscymnus tsugae, introduced against 

hemlock woolly adelgid on eastern hemlock. 

Biocontrol Science and Technology 21: 687-691. 

Lamb, A.; Salom, S.; Kok, L.; and Mausel, D. 2006. 

Confi ned field release of Laricobius nigrinus 

(Coleoptera: Derodontidae), a predator of 

the hemlock woolly adelgid, Adelges tsugae 

(Hemiptera: Adelgidae), in Virginia. Canadian 

Journal of Forest Research 36: 369-375. 

Lamb, A.; Salom, S.; Kok, L. 2005. Survival and 

reproduction of Laricobius nigrinus Fender 

(Coleoptera: Derodontidae), a predator of 

hemlock woolly adelgid, Adelges tsugae Annand 

(Homoptera: Adelgidae) in fi eld cages. Biological 

Control 32: 200-207. 

Lambdin, P.; Grant, J.; Paulsen, D.; Saxton, A. 

2006. Invasion of the hemlock woolly adelgid 

in the Great Smoky Mountains National Park. 

Journal of the Tennessee Academy of Science 81: 

49-54. 

McClure, M. 2001. Biological Control of Hemlock 

Woolly Adelgid in the Eastern United States. 

FHTET-2000-08. Morgantown, WV: U.S. 

Department of Agriculture, Forest Service: 10 p. 

Wiggins, G.J.; Hakeem, A.; Follum, R.; Rhea, R.; 

Lambdin, P.L.; Grant, J.F. 2010a. Seasonality 

of established populations of Sasajiscymnus 

tsugae on eastern hemlock in the southern 

Appalachians. In: Onken, B.; Reardon, R., 

comps. Proceedings of the Fifth Symposium on 

Hemlock Woolly Adelgid in the Eastern United 

States; 2010 August 17-19; Asheville, NC. 

FHTET-2010-07. Morgantown, WV: U.S. 

Department of Agriculture, Forest Service, 

Forest Health Technology Enterprise Team: 

203-205. 

Wiggins, G.J.; Hakeem, A.; Follum, R.; Paulsen, 

D.; Rhea, R.; Lambdin, P.L.; Grant, J.F. 

2010b. Assessment of tree health characteristics 

following releases of predatory beetles on eastern 

hemlock. In: Onken, B.; Reardon, R., comps. 

Proceedings of the Fifth Symposium on Hemlock 

Woolly Adelgid in the Eastern United States; 

2010 August 17-19; Asheville, NC. FHTET2010-07. Morgantown, WV: U.S. Department 

of Agriculture, Forest Service, Forest Health 

Technology Enterprise Team: 206-209. 

166 
Chapter 15: Whole-tree Enclosures: A Tool to Assess Introduced Predators 

of Hemlock Woolly Adelgid, Adelges tsugae 

Zilahi-Balogh, G.; Kok, L.; Salom, S. 2002. 

Host specifi city of Laricobius nigrinus Fender 

(Coleoptera: Derodontidae), a potential 

biological control agent of the hemlock woolly 

adelgid, Adelges tsugae Annand (Homoptera: 

Adelgidae). Biological Control 24: 192-198. 

Zilahi-Balogh, G.; Broeckling, C.; Kok, L.; Salom, 

S. 2005. Comparison between a native and 

exotic adelgid as hosts for Laricobius rubidus 

(Coleoptera: Derodontidae). Biocontrol Science 

and Technology 15: 165-171. 

167 
 

 

 

CHAPTER 16: A CASE STUDY OF A RELEASE OF THE PREDATOR
 

LARICOBIUS NIGRINUS FENDER AGAINST
 

HEMLOCK WOOLLY ADELGID, ADELGES TSUGAE, ANNAND, 


AT THE URBAN COMMUNITY FOREST INTERFACE: HEMLOCK HILL, 


LEES-MCRAE COLLEGE, BANNER ELK, NORTH CAROLINA
 

Richard McDonald1, David Mausel2, Scott Salom3, and Loke T. Kok3 

1

Symbiont Biological Pest Management, Sugar Grove, NC
 

2

Dividion of Entomology, University of Massachusetts, Amherst, MA
 

3

Department of Entomology, Virginia Tech, Blacksburg, VA
 

WHY RELEASE AT THE URBAN 

COMMUNITY FOREST INTERFACE? 

For the purpose of effi cient collection and 

redistribution of large numbers of predators of the 

hemlock woolly adelgid, Adelges tsugae, Annand 

(HWA), the urban community forest interface 

has many advantages: easy access by auto and 

by foot; urban hemlock trees have skirts with 

lower branches, making them more accessible for 

collecting/monitoring; hemlocks in parks, golf 

courses, schools, and cemeteries are sometimes 

fertilized and irrigated; urban hemlocks can occur 

singly in open areas exposed to winter sun, which 

favors predator presence; there is convenient 

nearby overnight shipping in urban areas, and 

easy access to collecting/shipping resources 

(boxes, blue ice, pint cartons, excelsior, etc). 

BACKGROUND OF HIGH COUNTRY 

REGION OF NC 

The High Country of northwestern North Carolina 

is one of the most biologically diverse areas in 

the world. This area includes Alleghany, Ashe, 

Avery, Mitchell, Watauga, Wilkes and Yancey 

Counties in northwestern North Carolina. Th e 

headwaters of five major river systems begin in 

this mountainous seven-county area, including 

the New (both North and South Forks of the 

New), Watauga, Yadkin, Catawba, and French 

Broad Rivers. Grandfather Mountain was the fi rst 

privately owned property that has been designated 

as a biosphere by the U.N.; it is now owned by 

the state of North Carolina. There are extensive 

native stands of Carolina and eastern hemlock, as 

well as Eastern White Pine ( Pinus strobus), Fraser 

Fir ( Abies fraseri), and Red Spruce ( Picea rubens) 

in this region. Abundant landscape plantings of 

Blue Spruce ( Picea pungens) and other landscape 

conifers like Mugo Pine ( Pinus mugo) are also 

present. The diversity of conifers gives this area 

abundant alternate adelgid hosts for predators as 

well. The HWA was first found in Avery County 

in 2002. The entire northwestern corner of North 

Carolina’s “High Country”—Avery, Ashe, Alleghany, 

Mitchell, Watauga, Wilkes, and Yancy counties—is 

now considered generally infested with HWA. 

The HWA is causing region-wide loss of hemlock 

populations, resulting in an environmental disaster. 

BACKGROUND OF RELEASE SITE 

Hemlock Hill is a 25-acre old-growth hemlock 

forest remnant area that is part of Lees-McRae 

College and is next to the town of Banner Elk, NC 

(elev. 1100 meters) (Fig. 1). It had moderate to 

heavy HWA infestations in 2003 (Mausel 2007). 

168 
Chapter 16: A Case Study of a Release of the predator Laricobius nigrinus Fender against hemlock woolly adelgid,
 

Adelges tsugae, Annand, at the Urban Community Forest Interface: 


Hemlock Hill, Lees-McRae College, Banner Elk, North Carolina.
 

Figure 1. 	 Location of Laricobius nigrinus release trees (in green - 10 trees, 30 beetles per tree) at Hemlock Hill 

in 2003. The yellow boxes are F1 recoveries during the 2004 season. Note that releases one through 

five were in a shaded area during winter, while releases six through ten were in areas that received 

abundant winter sun. 

169 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

RELEASE SITE - HEMLOCK HILL 

300 L. nigrinus adults released 31 December 

2003, plus 300 adults augmented near 

release tree #9 in late March 2006 

In late December 2003, 300 adult L. nigrinus reared 

in the laboratory at Virginia Tech University (British 

Columbia strain) were released at Hemlock Hill, at 

Lees-McRae College, Banner Elk, NC, into a stand 

of old growth hemlock infested with HWA. Th e 

release, first- and second-year monitoring of the 

site was done in conjunction with David Mausel’s 

research for his Ph.D., with guidance from Brad 

Onken and Rusty Rhea, U.S. Forest Service, and 

Professors Scott Salom and Loke Kok (Mausel 

2007, McDonald et al. 2005). One hundred fi fty 

beetles were released along the Elk River, a shady, 

cool, flood-prone area (5 trees with 30 beetles each), 

and the remaining 150 beetles were released near 

or at the top of the ridge on 5 trees, 30 per tree in 

an area which received winter sunshine (Fig. 1). 

This gave us two release areas to compare: a shaded, 

cool, wet river area, and a sunny, drier, ridge area. 

Initial Recovery and Dispersal Patterns 

of L. nigrinus from Hemlock Hill 

We sampled the release trees and nearby trees twice 

monthly from October until April, using 1 meter 

squared beat sheets. We sampled approximately 

30 trees (10 release trees and 20 nearby trees) 

each sample date. We were initially fortunate in 

that we divided the release of 300 beetles into 

two halves. The following fall/winter after the 

release (fall 2004), we found a total of 3 adults (F1 

generation) and 10 F2 larvae (April 2004 sample). 

The next year (Fall 2005), 12 L. nigrinus adults 

were found, along with 314 F3 larvae from branch 

samples by Mausel during April 2005 (Table 1) 

(Mausel 2007). During years one and two, we 

only found beetles on release trees; by the third 

year, beetles began dispersing to nearby trees. 

Determining Predation Rates of L. nigrinus 

on HWA ovisacs at Hemlock Hill 

HWA-infested branches were collected at Hemlock 

Hill in February and April 2006 and 31 March 

2007. Branch samples were taken along the river 

and on the ridge above. We focused our samples 

on hemlocks in an area with prior high larval 

and adult beetle recoveries (trees 9 & 10) from 

branch sampling work done by Mausel (2007). 

HWA-infested hemlock branches were bagged 

into gallon Ziploc bags and brought back into 

the laboratory, where we dissected ovisacs in 

order to determine presence of a L. nigrinus egg/ 

larva. From samples taken between release trees 

9 and 10, we found 10% of HWA ovisacs had 

a L. nigrinus egg or larva during April of 2006. 

We found an increase to 31% predation rate of 

ovisacs in the same area a year later, during late 

March 2007 (Figs. 2, 3, and 4). After this date, 

we began to sample using beat sheet method 

only to determine the presence of adult beetles at 

various areas distant to the original release site. 

Table 1. Recovery of L. nigrinus adults by season and place at Hemlock Hill 2004-2008. “River” is a 

shaded area during winter; “ridge” is a sunny area at the top of the Hemlock Hill (see Fig. 1). 

Larval numbers are from Mausel (2007). 

Season or place 

F1 Adults 

(‘04/’05) 

F2 Larvae 

(April ‘05) 

F2 Adults 

(‘05/’06) 

F3 Larvae 

(April ‘06) 

F3 Adults 

(‘06/’07) 

F4 Adults 

(‘07/’08) 

Fall 

Winter 

3 

0 10 

12 

0 314 

93 

109 

80 

23 

River 

Ridge (fall) 

2 

1 

1 

11 

4 

89 

41 

39 

170 
Chapter 16: A Case Study of a Release of the predator Laricobius nigrinus Fender against hemlock woolly adelgid,
 

Adelges tsugae, Annand, at the Urban Community Forest Interface: 


Hemlock Hill, Lees-McRae College, Banner Elk, North Carolina.
 

Bet. 9 & 10 Trees 1-5 Bet. 9 & 10 Tate Field Bet. 9 & 10

24-Feb-06 20-Apr-06 20-Apr-06 20-Apr-06 31-Mar-07

# Ovisacs 108 55 231 33 235

# Ovisacs/Ln 2 0 25 0 105

0

50

100

150

200

250

Number of Ovisacs

Date & Location

Predation Rate of HWA Ovisacs at Hemlock Hill

# Ovisacs

# Ovisacs/Ln

Figure 2. 	 Predation rate of HWA ovisacs at the Hemlock Hill L. nigrinus release site during 2006 and 2007. 

During 2006, we found 10% of ovisacs with a Laricobius egg or larva present; during 2007 we found 

31% of ovisacs with a Laricobius egg or larva. (Bet. 9 & 10 = sample taken between release trees 9 

and 10.) 

171 
 

 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Figure 3. 	 Laricobius nigrinus egg (yellow) above a 

cluster of HWA eggs. The distinct size, 

shape, and yellow color of the L. nigrinus 

egg versus HWA eggs are evident (photo 

by David Mausel). 

Establishment and spread of 

Laricobius nigrinus at Hemlock Hill, 

Banner Elk, North Carolina 

As we began to recover beetles over the fi rst three 

years post-release (2004-2007), we noticed a pattern 

of distribution by the beetles during the late fall 

and winter months. We initially recovered more 

beetles in sunny areas with HWA (trees 8, 9 and 

10), compared to cooler, shaded areas with HWA. 

For example, we recovered 101 beetles over the fi rst 

three years in the upper ridge area, which received 

winter sun, versus only 7 beetles in the release trees 

1 through 5 along the river, which did not receive 

winter sun (Table 1). From beat sheet samples taken 

during 2004 to 2008, we began to find that more 

beetles were dispersing in a southerly direction 

(Fig. 5, blue boxes), following the presence of winter 

sun on HWA-infested hemlocks. This pattern of 

beetles dispersing into areas having winter sun 

continued for the next 2 years (Fig. 6); with beetles 

now (2011) present more than 3 kilometers in every 

direction from the original release site. During 

fall 2006-April 2007, we recovered over 200 F3 

adults, giving us establishment of L. nigrinus at the 

Hemlock Hill release site and over 100 F4 adults 

(Fall 2007) from an area of over 1⁄2 square mile. 

Figure 4. 	 Laricobius nigrinus larva inside a dissected 

HWA ovisac; a single larva consumes 200­

250 HWA eggs to complete development 

(photo by David Mausel). 

Collection numbers of L. nigrinus adults that were 

taken from the Hemlock Hill area and redistributed 

to other sites are as follows: 2007 – 46; 2008 

– 189; 2009 – 581; 2010 - 1,838 (Figure 7). Th ese 

beetle recovery data were entered into the Forest 

Service’s HWA database, maintained by Virginia 

Tech, at http://hwa.ento.vt.edu/hwa/hwa.cgi. 

SUMMARY 

We can see three distinct phases (early, middle, 

and mature phases) of colonization, establishment 

and dispersal of L. nigrinus at the Hemlock Hill 

release site. During the “early phase” of colonization 

(Years 1 (2004) and 2 (2005)) by L. nigrinus, beat 

samples showed that beetles stayed on the release 

hemlocks trees with HWA. During the “middle 

phase” of establishment and dispersal (Years 3 to 5, 

2006-2008) we had establishment of populations 

of L. nigrinus (recovery 3 years in a row), signifi cant 

predation rates on sampled ovisacs near release trees 

9 & 10 (31% ovisacs with Ln), and limited dispersal 

to the south, into sunlit areas. During the “mature 

phase”—5 to 7 years (2009-onward)—we were 

able to collect and move hundreds to thousands of 

6th and 7th generation beetles to new areas, both 

172 
 

 

Chapter 16: A Case Study of a Release of the predator Laricobius nigrinus Fender against hemlock woolly adelgid,
 

Adelges tsugae, Annand, at the Urban Community Forest Interface: 


Hemlock Hill, Lees-McRae College, Banner Elk, North Carolina.
 

N. Elk River 

Oct. ‘07 

Nov. ‘06 

Mar./Dec. ‘07 

Jan. ‘08 

L. nigrinus 

Figure 5. 	 Pattern of dispersal of L. nigrinus adults from 2004 through January 2008 (light blue boxes). Adults 

appear to be more prevalent on south facing slopes during the coldest times of their season (Dec.-Feb.). 

Dispersal recoveries shown in 4 blue boxes: Mid-November of 2006: recovery of a single L. nigrinus 

adult on the main ridge of Hemlock Hill, 300+ meters from the nearest release tree. March/December 

2007: recovery of a single L. nigrinus adult more than 3/8ths mile from the closest release site during 

March; December 2007: 2 adults in same area. January 2008: 1 L. nigrinus adult below Tate Field 

scoreboard. This was 200 yards beyond prior known dispersal at that time, in a southerly direction. 

173 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Figure 6. 	 Current distribution of L. nigrinus adults in the Hemlock Hill area as of December 2010; compare to 

Figure 2. Adults are now present in a 3-kilometer radius from the original release. Beetles appear to have 

moved mainly to the south and west. We have collected a total of 2,757 L. nigrinus adults since 2007 in 

the Banner Elk area. Beetles have dispersed westward and are now found in the Pisgah National Forest. 

Figure 7. 	 Sum of beetles caught by beat sheet 

method at Hemlock Hill, Banner Elk, NC 

each year since 2004 (x axis years post 

release). Collection numbers of L. nigrinus 

adults that were redistributed to other areas: 

2007 – 46; 2008 – 189; 2009 – 581; 

2010 - 1,838. 

174 
 

 

Chapter 16: A Case Study of a Release of the predator Laricobius nigrinus Fender against hemlock woolly adelgid,
 

Adelges tsugae, Annand, at the Urban Community Forest Interface: 


Hemlock Hill, Lees-McRae College, Banner Elk, North Carolina.
 

locally and regionally as need drives the system. 

For example, we collected and redistributed 1,838 

beetles during fall of 2010 & spring of 2011 (Fig. 

7). We are also seeing sustained hemlock tree 

regrowth evident in the release areas. Beetles are now 

common on hemlocks with HWA throughout the 

Banner Elk, NC area. L. nigrinus is extinguishing 

populations of HWA locally and fragmentation of 

HWA populations and outbreaks is beginning— 

similar to patterns we see at the field insectary at 

Virginia Tech and in the field in Seattle. From our 

field studies and collections, we have found that 

L. nigrinus beetles do best in the following areas: 

1) Release beetles in an area with 

good levels of HWA; 

2) Release site must have hemlocks that 

receive abundant winter sun; 

3) Hemlocks need to have needle duff under 

tree for L. nigrinus beetles to pupate; 

4) Site must be undisturbed (no logging, 

mowing, pesticide spraying, etc.); and 

5) 	More conifer species in the area is 

desirable; the increased biodiversity and 

alternate adelgid species as food. 

Two thirds of the hemlock forest in the eastern 

USA is privately held. Thus for the future, we need 

to interface with the public by publishing a guide 

sheet for L. nigrinus focusing on landowner interest 

in the program. The urban community forest 

interface has the potential to produce high numbers 

of HWA predatory beetles that can be released into 

forest ecosystems to assist in bringing the hemlock 

woolly adelgid into balance much more rapidly 

than if we had to rely on natural dispersal alone. 

RECOMMENDATIONS 

BASED ON THIS STUDY 

To accelerate the establishment of HWA predators 

through the use of field insectaries, we suggest using 

abandoned hemlock nurseries or other favored 

areas in the urban/forest community interface 

with hemlocks: parks, cemeteries, schools, golf 

courses, playgrounds, colleges, universities, and 

similar areas that have hemlocks trees or hedges. 

These areas must also have resource managers that 

are willing to protect beetle release areas from 

spraying, cutting, disturbance of needle duff , etc. 

ACKNOWLEDGEMENTS 

Thanks to Brad Onken for helping us with this 

work; special thanks to Gina Davis for suggesting 

Hemlock Hill as a release site, her assistance and 

her many valuable suggestions; Stan Steury and 

Cliff Vinson of the USDA Blue Ridge Resource 

Conservation and Development Council 

(BRRC&D); through the BRRC&D we were able 

to receive funding for this project with an Urban 

and Community Forestry Grant from the North 

Carolina Division of Forest Resources, Department 

of Environment and Natural Resources, working 

with Don Rogers and Nancy Stairs, who were 

particularly helpful in the early stages of this 

program; in cooperation with Rusty Rhea of the 

U.S. Forest Service, Southern Region, whose 

suggestions and knowledge are much appreciated. 

LITERATURE CITED 

Mausel, D.L. 2007. Release and Monitoring of 

Laricobius nigrinus (Coleoptera: Derodontidae) for 

Biological Control of the Hemlock Woolly Adelgid 

in the Eastern US. Blacksburg, VA: Virginia 

Polytechnic Institute and State University: 

153 pages. Ph.D. dissertation. 

McDonald, R.; Mausel, D.; Salom, S.; Kok, L.T.; 

Steury, S.; Luker, G.; Spears, G.; Skeate S.; 

Graham, J. 2005. F1 adults of the HWA 

predator, Laricobius nigrinus Fender, recovered 

in October and November 2004 at Hemlock 

Hill, an old-growth hemlock site at Lees-McRae 

College, Banner Elk, North Carolina, from a 

release made in December 2003. In: Onken, 

B.; Reardon, R., comps. Proceedings of the 

Third Symposium on Hemlock Woolly Adelgid in 

the Eastern United States; 2005 February 1-3; 

Asheville, NC. FHTET-2005-01. Morgantown, 

WV: U.S. Department of Agriculture, Forest 

Service, Forest Health Technology Enterprise 

Team. 

175 
CHAPTER 17: THE HWA PREDATOR RELEASE AND RECOVERY
 

DATABASE
 

Andy Roberts1, Ashley Lamb2, Brad Onken3, and Scott Salom1 

1

Department of Entomology, Virginia Tech, Blacksburg, VA 24060 

2

Department of Entomology, University of Tennessee, Knoxville, TN 

3

U.S. Forest Service, Northeastern Area Forest Health Protection, 180 Canfield St., Morgantown, WV, 26505 

INTRODUCTION 

Hemlock woolly adelgid (HWA), Adelges tsugae 

Annand (Hemiptera: Adelgidae) is an introduced 

insect that attacks and kills eastern hemlock, Tsuga 

canadensis (L.) Carriere, and Carolina hemlock, T. 

caroliniana Engelmann, trees in the eastern United 

States (Cheah et al. 2004). Importation of its 

natural enemies is the most promising option for 

controlling HWA on a landscape level (Wallace and 

Hain 2000). Since the late 1990s, state and federal 

agencies have released lab reared or wild predatory 

beetles of HWA in an effort to control this pest. 

Until recently, data pertaining to the release, 

monitoring, and recovery of the predators Laricobius 

nigrinus Fender (Coleoptera: Derodontidae), 

Scymnus sinuanodulus Yu et Yao (Coleoptera: 

Coccinellidae), and Sasajiscymnus tsugae Sasaji and 

McClure (Coleoptera: Coccinellidae) have been 

maintained on paper data forms or in small local 

databases and as a result have been inaccessible 

to HWA scientists and managers at regional and 

national levels. The management, accessibility, and 

usability of these data are made more cumbersome 

because of differences in how various agencies 

performed releases and monitoring and in how 

data were collected and stored, with multiple data 

forms and protocols among agencies being the 

norm. In 2007 we initiated the development and 

implementation of the HWA Predator Release 

and Recovery Database (PDB) which is housed 

in the Department of Entomology at Virginia 

Tech in Blacksburg, VA. The goal of the project 

is to include all historic release and monitoring 

information as well as provide a mechanism 

for field personnel to enter and update current 

and future records. Inherent in this program 

is an attempt to standardize field protocols as 

well as data forms. The PDB will impose an 

organizational structure on the data and serve 

as a central repository for information collected 

on release and recovery eff orts. Implementing 

the PDB will facilitate improved access to and 

use of the data; provide project-wide reporting, 

mapping, and analysis; ensure that data from all 

cooperators are maintained, archived and available; 

and improve decision-making for future actions. 

DATABASE STRUCTURE 

The database is developed in Oracle® 10g database 

management system installed on a Windows 2003 

Server platform in the Department of Entomology 

at Virginia Tech. Each release site record contains 

approximately 85 data fields distributed among 

four predator related database tables plus an 

additional table containing information on 

database users (Fig. 1). These data are obtained 

from the forms used when beetles are released 

(Appendix 17-1) or monitored (Appendix 17-2). 

In addition to information on HWA predator 

release sites the database contains data on the 

individual and organization performing the 

activity, site and stand conditions, weather, origin 

of beetles released, release tree information, 

post-release survey, and predators recovered. 

176 
Chapter 17: The HWA Predator Release and Recovery Database 

Figure 1. 	 Database table structure and relationships in 

the HWA PDB. 

The database is organized around release sites. 

Each site is geographically referenced and 

contains from one to several release trees. Data 

for post-release monitoring, sometimes referred 

to as recoveries, are in almost all cases associated 

with release sites. A subsequent beetle release 

at an initial site is called an augmentation, 

and the database includes a mechanism to 

attach an augmentation to an existing site. 

Each record in the PDB must have a unique 

identifi er. This unique ID is assigned by the database 

at time of data entry and consists of the species 

(LN, SS, or ST), state abbreviation, year (2 digits), 

and a sequential number. Thus, LNTN06007 is 

the ID for the seventh recorded L. nigrinus release 

in Tennessee in 2006. Post-release survey records 

are associated with their initial release record; their 

unique ID, assigned at data entry, consists of the 

state abbreviation, year, and a sequential number. 

Currently the standard release form specifi es 

that geographic coordinates be recorded in the 

Universal Transverse Mercator (UTM) coordinate 

system. Recently, however, we altered the database 

to include data entry in any of the three standard 

formats for latitude and longitude (decimaldegrees, degrees minutes seconds, degrees decimalminutes). We found this necessary because of the 

large amount of historical information collected in 

lat/long as well as the fact that this is the preferred 

format for many agencies conducting release and 

survey work. Conversion from lat/long to UTM 

can be convoluted and tedious on a user’s desktop 

computer but is straightforward in the database. 

The PDB is relatively unconstrained with regard 

to restrictions placed on data fields. Because there 

have been no standard protocols and forms to which 

different agencies adhered, there is variability in 

many of the data values as well as the absence of 

data in many fi elds. This is especially true when 

integrating older data with the most recently 

collected information. To address this looseness in 

the data, it was necessary to construct the database 

with a minimum of data rules and restrictions. For 

example, of the roughly 85 possible data fi elds, 

only nine are required at data entry. Th is results 

in a somewhat messy database, but the decision 

was to capture as much data as possible, including 

historical records, rather than expend resources 

on cleaning up data which easily could be over 

a decade old. It is hoped that consistency and 

clarity will emerge as the database matures and 

begins to consist primarily of current records. 

DATABASE FUNCTIONALITY 

The PDB is accessed through a web portal 

(http://hwa.ento.vt.edu)(Fig. 2). 

Two types of user accounts are available. Th e 

“VT User” account is reserved for persons who 

require both read and write privileges for the 

entire database. This includes database managers 

and technicians (usually at Virginia Tech) who are 

responsible for entering or correcting data from 

multiple agencies. A “Remote User” account is 

assigned to non-VT personnel whose responsibility 

is to enter and maintain data for a specifi c agency. 

Currently, the site is publicly available and anyone 

can log in as “guest” without a password. Guest 

logins may view all site content but have no editing 

capability. Presently there are approximately 48 

registered users, about 30 of whom are actively 

involved with data entry and management. 

177 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Figure 2. The login screen to the PDB web portal. 

The functionality of the database and associated 

web site is fairly standard and consists of data 

entry and editing for both initial release data, 

post-release monitoring and predator recovery, 

augmentative release, database query and reporting, 

map query and display, and data download. In 

addition, there is access to documents addressing 

field protocols, data entry protocols, and user 

account administration. The main page for the 

remote user interface is shown in Figure 3. 

The VT User main page is similar to the Remote 

User page except it includes access to data entry 

screens for earlier versions of the release data forms 

and access to database user statistics. It also allows 

someone with VT user privileges to review and 

approve data submitted by a remote user. Records 

submitted by a remote user are marked as pending 

until they are reviewed and approved by a VT user. 

These records enter the database, are marked for 

review, and are available for most database functions 

and products, including reports and summaries. 

Data entry is straightforward (Fig. 4). One area 

where we have imposed quality control at the 

data entry stage is in geographic coordinates. Th e 

database accepts coordinates in either Universal 

Transverse Mercator (UTM) or in latitude/ 

longitude, which may be in any of the most 

common formats: decimal-degrees, degrees decimalminutes, or degrees minutes seconds (Fig. 4). 

Constraints have been placed on these coordinates 

so that values out of range are not allowed. Th e 

database stores only UTM coordinates (and the 

UTM zone), but all web display and output 

includes both latitude/longitude and UTM. Th e 

178 
Chapter 17: The HWA Predator Release and Recovery Database 

Figure 3. The primary page on the Remote User database site. 

179 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Figure 4. The predator release data entry screen. Entry fields for trees 2 through 5 and 

6 through 10 are not shown. 

180 
 

Chapter 17: The HWA Predator Release and Recovery Database 

conversion between UTM and lat/long is performed 

in database memory, and to accommodate 

different protocols, latitude and longitude are 

displayed in the three formats mentioned above. 

Database actions which may occur on a site record 

subsequent to the initial beetle release include 

editing release information, adding post-release 

monitoring data, or adding augmentative release 

data. To perform these tasks, the user must fi rst 

access the initial release data through a database 

query and then select the appropriate action, 

either Update, AddRecovery, or AddAugmentative 

(Fig. 5). When post-release data are entered 

in this way, the database populates the online 

data forms with pertinent information from the 

release records and thus expedites data entry. In 

addition, this reduces possible omissions or data 

entry errors by the user. An example is shown 

in Figure 6 where relevant release information 

for the Gauley River site has been pre-loaded 

into the post-release monitoring data form. 

DATABASE CONTENT 

Reliable estimates place current HWA 

predator releases at above 2.5 million for 

S. tsugae, 30,000 S. sinuanodulus, and over 

150,000 L. nigrinus (B. Onken, pers. comm.), 

and only a fraction of the estimated fi eld 

releases have been recorded in the PDB. 

Adding previously existing data can be more timeconsuming than adding newly collected data; it can 

appear daunting to consider adding hundreds of 

historical records especially if data formats and/or 

collection methodologies do not fit nicely with 

the data entry structure of the PDB. It is for this 

very reason that we relaxed data entry constraints 

Figure 5. The results returned by a query of the release data. This template is used to access data entry for 

subsequent actions at the site. Only the top portion of the form is shown. 

181 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

as well as incorporated the ability to enter either 

UTM or lat/long coordinates. It is possible to 

enter a large amount of data in one batch process 

if those data currently exist in a digital format, 

such as an Excel spreadsheet. In this case, the data 

may require some manipulations to accommodate 

the new database structure, but these eff orts are 

minor compared to entering records manually. 

For example, with only a modicum of eff ort we 

recently batch loaded 236 historical records from 

Great Smokey Mountains National Park. Eff orts 

are ongoing to remedy the discrepancy between the 

contents of the database and existing information. 

Figure 6. The post-release monitoring and predator recovery data entry form. Only the top portion of the form 

is shown. 

182 
 

Chapter 17: The HWA Predator Release and Recovery Database 

Accessing data through the web portal allows more 

refined searches than a simple export from the 

database. One can perform queries based on the 

values of specific variables, such as Location Name, 

State, Species, and Year. The default Summary 

Report generates a table by species of the number 

of unique locations and beetles released to date 

and a yearly time series of post-release monitoring 

results (Fig. 7). Numbers reported for the total 

number of release locations in the Summary 

Report differ from those in Table 1 because the 

Summary Report does not include augmentative 

releases in this statistic. Of the 164,381 L. 

nigrinus released at 345 locations, there were 51 

post-release surveys in the following year, 24 of 

which were positive for beetle recoveries (Fig.7). 

To date, there were a total of 109 post-release 

surveys at these sites and beetles were recovered 

at 54 of those. Because some locations have been 

surveyed multiple times over the years the grand 

total does not equal the sum of each yearly total. 

The PDB has the ability to “drill down” through the 

data to reveal more detail. For example, clicking on 

the 54 sites with positive recoveries will bring up a 

list of those sites with associated data as well as the 

number of beetles recovered at each site (Fig. 8). 

Further, selecting the 1067 individuals recovered 

for the Rocky Gap release site from the previous 

query (Fig. 8) will return a list of the results for 

each individual survey (Fig. 9). In the returned 

data table, selecting the value in the Date fi eld will 

return the original survey record for that activity. 

MAP QUERY 

The PDB incorporates a simple map display 

developed in a Google Maps® application which 

serves as a front end to the database. As with the 

reporting section of the web portal, the mapping 

function supports parameterized database queries 

using several data fields (Fig. 10). Symbology 

is simplified to differentiate among species and 

between release and post-release monitoring 

sites. Data in the map are grouped by unique 

geographic coordinates, and these unique locations 

Figure 7. The default Summary Report from the database. 

183 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Table 1. Numbers of predator release sites and beetles released in the database by species 

and state. 

Figure 8. Example of records returned through the site query (top of table only). 

Figure 9. Example of monitoring records returned from the database for a specific site (top of table only). 

184 
 

Chapter 17: The HWA Predator Release and Recovery Database 

are numbered and listed in the panel to the left 

of the map. Release sites as well as post-release 

monitoring sites are grouped under each unique 

location. For example, site 3 in Figure 10 references 

four S. tsugae release records from TN that may 

or may not occur on the same date. Obviously, 

only data for which geographic coordinates 

are available are displayed on the maps. 

As with all Google® maps, the base layer can 

be either a street layer, a terrain layer (Fig.11) 

or remotely sensed imagery. Each location has 

a hyperlinked reference to all activities at that 

site. For example, in Figure 11 data for an initial 

release of 1000 L. nigrinus (ID=LNWV08005) 

and a subsequent survey (ID=WV090002) are 

referenced by the point in the center of the map. 

Clicking the ID of the one of the referenced events 

will open up the database record for that event. 

DATA EXPORT 

All data in the PDB can be exported in text fi le 

format. Information on releases, release trees, 

recoveries, recovery trees, and predators are 

downloaded in separate files which will easily load 

into spreadsheets such as Excel® and GIS software 

such as ArcGIS®. One planned enhancement is 

to implement .kmz file formats so PDB output 

can be exported directly to Google Earth® . 

DATABASE ISSUES 

There are a number of important issues to address 

in the continued development of the PDB: 

Completeness. The necessity to incorporate past 

survey data which exists on paper and/or local 

data files has been discussed above. This is one 

of the most pressing issues facing the PDB. 

Cooperator buy-in. Equally important is to 

obtain buy-in from groups that are releasing and 

monitoring beetles. Cooperators who are not 

invested in the project will find it more diffi cult to 

take the additional step of updating the database as 

they collect field data. This is especially pertinent 

to the task of submitting historical data. 

Data Correction. The unconstrained nature of 

the PDB facilitates data entry from disparate 

sources, but it allows null or erroneous data to pass 

unchallenged into the database. Detecting and 

correcting these data after the fact is challenging 

from both a technical, practical, and human 

nature aspect. In its current state, the PDB has 

a large amount of missing or erroneous data 

including geographic coordinates. Since inception, 

we have added or corrected coordinates for over 

200 release sites, but there still remain dozens 

of records with no geographic information. 

Location name standards. Standardizing the 

naming procedures of release sites such that 

location names provide more consistent location 

information is one methodological change that 

would dramatically improve database value. 

Not surprisingly, releases at identical or nearby 

locations performed by different persons or 

at different times appear in the database with 

different Location Names. These often are due 

to spelling or abbreviation inconsistencies, 

but the result is that it reduces the accuracy 

and effectiveness of database queries. 

Map display. There are more advanced options for 

the mapping component of the PDB which would 

provide more flexibility and power. Development in 

this area is contingent on user demand and response. 

In conclusion, the HWA predator release and 

recovery database is an ongoing project to 

add meaning to historical, current, and future 

biocontrol efforts directed toward this pest. 

Hopefully, increased knowledge regarding release 

activity numbers and locations will supplement 

associated research and control activities as 

well as increase access to data and awareness of 

the extent of these activities in the country. 

185 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Figure 10. Results of map query displaying all georeferenced records in the database. 

Figure 11. Map display with linked data records and terrain background. 

186 
Chapter 17: The HWA Predator Release and Recovery Database 

ACKNOWLEDGMENTS 

In addition to those listed above as contributors, 

Jiang Wu and Ian Firkin, both at Virginia Tech, 

have devoted much time to the database and 

mapping applications. Karen Felton with the 

U.S. Forest Service in Morgantown greatly 

assisted in developing the database structure 

and schema. Funding for this project is supplied 

by the U.S. Forest Service HWA Initiative. 

LITERATURE CITED 

Cheah, C.; Montgomery, M.E.; Salom, S.M.; 

Parker, B.L.; Costa, S.; Skinner, M. 2004. 

Biological Control of Hemlock Woolly Adelgid. 

Reardon, R.; Onken, B., tech. coords. 

FHTET-2004-04. Morgantown, WV: 

U.S. Department of Agriculture, Forest 

Service: 22 p. 

Wallace, M.S.; Hain, F.P. 2000. Field surveys and 

evaluation of native and established predators 

of the hemlock woolly adelgid (Homoptera: 

Adelgidae) in the southeastern United States. 

Environmental Entomology 29: 638-644. 

187 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Appendix 17-1. Data collection forms and instructions for predator release. 

188 
Chapter 17: The HWA Predator Release and Recovery Database 

Appendix 17-1 (continued). Data collection forms and instructions for predator release. 

189 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Appendix 17-2. Data collection forms and instructions for post-release monitoring. 

190 
Chapter 17: The HWA Predator Release and Recovery Database 

Appendix 17-2 (continued). Data collection forms and instructions for post-release monitoring. 

191 
SECTION V ADDITIONAL TOPICS
 
CHAPTER 18: FIELD INSECTARY: CONCEPT FOR
 

FUTURE PREDATOR PRODUCTION
 

Scott Salom1, Loke T. Kok1, Tom McAvoy1 and Richard McDonald2 

1

Department of Entomology, Virginia Tech, Blacksburg, VA 24060-0319 

2

Symbiont Biological Pest Management, Sugar Grove, NC 28679 

INTRODUCTION 

Predators for use in biological control can be 

obtained one of three ways: 1. collect them 

from their native habitat and ship them to 

their destination (lab or field); 2. rear them in 

a laboratory or other type of controlled setting 

(Lamb et al. 2005); and 3. use a natural or 

planted setting, in an area close to the targeted 

release locations, to encourage prey production to 

build up the predator populations for harvesting 

and redistribution on a consistent basis. Th is 

is called a field insectary. For hemlock woolly 

adelgid biological control, importing the predator 

Laricobius nigrinus and rearing them in labs have 

been the most common approaches used. But it 

is understood that both approaches are expensive. 

The concept of employing field insectaries are 

especially helpful after a biological control 

program has matured and the knowledge of how 

to rear successful agents has been obtained. 

Reports of use of field insectaries/nurseries for 

rearing of predators, parasitoids, and weed biological 

control agents have been well documented, but few 

have been in the forest ecosystem (Kok and Salom 

2002). The biological control lab at Virginia Tech 

created a hemlock plantation with the long-term 

objective of developing a sustainable fi eld insectary. 

What follows is a description of the effort to date. 

ESTABLISHMENT OF A FIELD INSECTARY 

An eastern hemlock plantation was established at 

Virginia Tech’s Kentland Farm, McCoy, Virginia 

in October 2001 (Kok and Salom 2002, Mausel 

et al. 2008). A field that was left fallow with 

naturally occurring wild grass on a northeastfacing slope (10 to 15%) was selected for the 

location of a 0.4 ha field insectary. Adjacent to 

the field insectary was a young (10-12 yr old) 

white pine, Pinus strobus L., plantation that was 

naturally infested with pine bark adelgid. For the 

hemlock plantation, twelve 12 × 20 m blocks were 

marked in a 4 × 3 block rectangle and spaced 5 m 

apart. Trees were spaced 2.4 m within and 3.7 m 

between rows that were oriented northeast to 

southwest. Six rows with five tree locations were 

marked in each block (30 trees per block) (Fig. 1). 

Figure 1. 	 Eastern hemlock field insectary soon after its 

establishment in Kentland Farms, 

McCoy, VA. 

195 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Three hundred 1.2 to 2.4 m tall hemlock trees 

with 0.6 m diameter root balls wrapped in burlap 

were purchased from a local nursery. Ten of the 

12 blocks were planted with these large hemlocks. 

The other two blocks were planted at the same 

spacing but with 60 potted 0.6 m tall hemlocks 

purchased from a nursery in Pennsylvania, for a 

total of 360 trees. Augers (60 and 15 cm diameter) 

were used to make holes for the root balls of 

the large and small hemlocks, respectively. Bark 

mulch was applied 10 to 15 cm deep and 4 to 

8 liters of water were poured to the drip line of 

each tree immediately after planting. Th ey were 

watered again at 3 and 6 wk after planting. 

The trees were inoculated with HWA three times in 

March 2002 and again in April 2003 to supplement 

existing populations. Two hemlock clippings (~30 

cm long) infested with progrediens eggs from local 

forests were implanted in the mid-crown of each 

living tree every 2 wk until the onset of egg hatch 

(i.e. on the 1st, 15th, and 30th day of each month). 

We estimated the density of HWA on 193 trees 

greater than 1.0 m tall to determine which trees 

had enough HWA for a predator release. On 12 

November 2003, the number of new shoots infested 

with one or more HWA sistens and total number of 

new shoots were counted on 30 cm segments at the 

middle third of a branch’s length. One branch was 

measured at each cardinal point and the percentage 

of infested shoots from the four branches was 

averaged for each tree. On 18 November 2003, 

predators were released on each tree with the 

numbers released based on the estimated density of 

HWA as follows: six adult L. nigrinus were released 

on each of four heavily-infested trees (≥ 76% of 

new shoots with at least one HWA), four were 

released on 42 moderately-infested trees (25 ≤ x ≤ 

75%) and one was released on 66 lightly-infested 

trees (≤ 25%). This open release amounted to 258 

L. nigrinus adults (unknown sex ratio) on 112 

hemlocks that ranged in height from 1.8 to 3.0 m. 

Population estimates of L. nigrinus and L. rubidus 

were conducted in winter of 2005 through 2009 

on the hemlocks using canvas (71 cm2

) beat sheets 

(Bio-quip, Rancho Dominguez, CA). We sampled 

the most heavily HWA-infested branches of a 

tree first (typically at the top of each tree’s crown) 

followed by lower branches. We spent ~1 min. per 

tree hitting branches with a 1.0 m long bamboo 

stick in the afternoon (from 13:00-17:00) and 

counting dislodged L. nigrinus and L. rubidus 

adults on the sheet. The species were diff erentiated 

on the basis of color and then carefully returned 

to the trees. In 2007, both species were collected 

from the beat sheets with an aspirator, transferred 

to plastic containers with HWA-infested foliage 

and transported to the insectary for processing. 

They were sorted and L. nigrinus adults were 

packaged in ventilated plastic containers for 

release in HWA-infested natural forests. 

In winter of 2004 through 2007, HWA 

infestation level was categorized on each tree as 

heavy, moderate, light, or absent (after ~1 min. 

of searching). These estimates were based on 

the quantitative estimates taken in 2003, when 

the number of L. nigrinus to release per tree 

was initially determined (described previously). 

Data were summarized as the total number of 

trees in each infestation category and presented 

graphically. In 2007, tree decline was classifi ed 

subjectively into categories as healthy (<10% 

HWA damage symptoms), light (10-25%), 

moderate (26-50%), severe (51-100%), or dead 

(no green foliage). Damage symptoms included 

no new shoot growth, twig dieback, foliage 

discoloration, and needle loss (Young et al. 1995). 

MONITORING THE FIELD INSECTARY 

After collecting over 200 F3 L. nigrinus in 2007, 

the number of L. nigrinus and L. rubidus dropped 

in 2008 and 2009. L. rubidus is present because the 

hemlock plantation supporting HWA is located 

next to a 20-yr old white pine plantation that 

supports pine bark adelgid, the primary host of L. 

rubidus. The decrease in predators collected (Fig. 

2) coincides with a reduction in HWA found on 

hemlock trees (Fig. 3). Such a high percentage of 

trees exhibiting none to light infestation provides 

some circumstantial evidence that the predators are 

playing a role in suppressing HWA populations. 

196 
 

 

300

350

400

450

Chapter 18: Field Insectary: Concept for Future Predator Production 

140

120

L. nigrinus

L. rubidus

None

Light

Moderate

Heavy

100

Tota l num ber of beetles

250

200

150

Number of trees

8 0

6 0

4 0

100

2 0

5 0

0 0 Jan 2004 Jan 2005 Jan 2006 Jan 2007 May 2008 Mar2009

2005 2006 2007 2008 2009 Sam pling season Year sam p led

Figure 3. HWA infestation levels at Kentland Research Figure 2. Population trends of Laricobius nigrinus and Farm, in Montgomery County, VA, 

L. rubidus at the field insectary at Kentland 2004-2009.
 

Research Farm, in Montgomery County, VA, 

2005-2009. 

REINFESTING AND EXPANDING 

THE INSECTARY 

In spring 2010, hemlocks were reinfested with 

HWA sistens and progrediens when each stage 

was ready to hatch from eggs. In October, 2010, 

hemlocks were sampled for L. nigrinus. Eighty-two 

L. nigrinus were collected from 17 trees identifi ed 

as modestly infested. Thirteen additional beetles 

were collected from 10 trees considered uninfested. 

In fall 2010, the insectary was doubled in size 

to 24 blocks with 1.5 m tall trees. Additionally, 

1 m tall trees were added to the old blocks 

where trees had died early on due to drought. 

NATURAL INSECTARIES 

In northwestern North Carolina, where L. nigrinus 

was first released in 2003 and has been established 

since 2006, healthy hemlocks remain. Beetles (F7) 

from our 2003 release have dispersed more than 

1.5 miles in every direction from the original site 

in Banner Elk, NC. Collections of L. nigrinus 

adult beetles have been made each year since 2007, 

when 46 F4 beetles were collected and released 

in Sugar Grove, NC. Collections of beetles have 

increased each year; 2008 – 189 beetles (collected 

by Salom, Story, and McDonald; all beetles 

returned to Virginia Tech); 2009 – 581 beetles 

(100 to Mausel, 381 released behind Mast Store, 

42 beetles to Mountain Aire near Celo and 58 

beetles to Stewart Skeate); 2010 – 1,094 beetles 

during fall of 2010 (468 to Brad Onken, 200 

beetles to Mausel, 320 beetles to Calloway Ridge 

road, Foscoe, NC and 106 to Blowing Rock, NC). 

We are now able to collect L. nigrinus beetles in 

significant numbers from our 3 oldest release sites. 

Collection data show that we are able to recover 

more beetles in the urban community forest 

interface because of ease of access, edge factors, 

consistently high HWA pest populations, and 

tree characteristics in open grown sites. For more 

detailed information on natural insectaries, please 

see the Case Study, Chapter 16 in this publication. 

LITERATURE CITED 

Kok, L. T., and S. M. Salom. 2002. Possible use of 

field insectaries to rear hemlock woolly adelgid 

predators. In: Onken, B.; Reardon, R.C.; 

Lashcomb, J., eds. Proceedings: Hemlock Woolly 

Adelgid in the Eastern United States Symposium; 

2002 February 5-7; East Brunswick, NJ. New 

Brunswick, NJ: Rutgers, New Jersey Agricultural 

Experiment Station: 225-232. 

197 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Lamb, A.; Salom, S.M.; Lok, L.T. 2005. Guidelines 

for rearing Laricobius nigrinus Fender. In: 

Onken, B.; Reardon, R., comps. Proceedings of 

the Third Symposium on Hemlock Woolly Adelgid 

in the Eastern United States; 2005 February 1-3; 

Asheville, NC. FHTET-2005-01. Morgantown, 

WV: U.S. Department of Agriculture, Forest 

Service, Forest Health Technology Enterprise 

Team: 309-318. 

Mausel, D.L.; Salom, S.M.; Kok, L.T.; Fidgen, J.G. 

2008. Propagation, synchrony, and impact 

of introduced and native Laricobius spp. 

Environmental Entomology 37: 1497-1507. 

Young, R.F.; Shields, K.S.; Berlyn, G.P. 1995. 

Hemlock woolly adelgid (Homoptera: 

Adelgidae): stylet bundle insertion and feeding 

sites. Annals of the Entomological Society of 

America 88: 827-835. 

198 
CHAPTER 19: INTEGRATING CHEMICAL AND BIOLOGICAL CONTROL
 

Scott Salom1, Albert Mayfi eld2, and Tom McAvoy1 

1

Department of Entomology, Virginia Tech, Blacksburg, VA 24060-0319 

2

Southern Research Station, U.S. Forest Service, Asheville, NC. 28804-3454 

INTRODUCTION 

Research and management efforts to establish an 

effective biological control program against HWA 

has received significant support by the U.S. Forest 

Service over the past 17 years. Other federal and 

state agencies, universities, and private entities 

have also contributed to this overall research and 

management effort. Although a number of HWAspecific predator species from Asia and western 

North America have been studied in quarantine, 

mass reared, and released, the work discussed here 

will focus on Laricobius nigrinus (Coleoptera: 

Derodontidae). This predator, from western North 

American hemlock forests, has become established 

throughout the mid-Atlantic region (Mausel et 

al. 2010). Also current studies by G. Davis (Ph.D 

student at Virginia Tech) show that the beetle does 

not disperse very far the year they are released and 

only about 300 m 5 years after release. Long-term 

impact studies of the predator are ongoing, but it is 

apparent that at many of the release locations where 

L. nigrinus has established, older mature trees have 

succumbed to HWA. The younger, more vigorous 

understory trees do not decline as quickly, and 

appear to sustain growing populations of L. nigrinus. 

Imidacloprid has been the standard insecticide 

for application against HWA in urban and other 

settings where individual trees are highly valued. 

Merit 75WP and, more recently, Advance Tree and 

Shrub (Bayer) for homeowners have been used 

effectively in soil applications. Stem injections of 

various imidacloprid formulations have also found 

a niche for treating high value hemlocks. Recent 

formulation advances by Bayer have included 

CoreTect®, slow-release tablets placed under the 

organic layer around the root collar of trees. Th is 

recently registered product allows for a much 

easier application of imidacloprid and makes 

treatment of trees in remote areas more feasible. 

Laricobius nigrinus susceptibility to imidacloprid was 

recently studied by Eisenback (2008). While acute 

toxicity was demonstrated in the laboratory from 

topical application and from feeding on poisoned 

prey (Eisenback et al. 2009), results were much 

less conclusive in the field. At sub-lethal dosage 

applications, predator mortality and fi tness impacts 

from feeding on HWA settled on previously treated 

trees were minimal. Furthermore, HWA is extremely 

sensitive to imidacloprid (Cowles et al. 2006) and 

the presence of HWA on previously treated trees 

should indicate that imidacloprid concentrations in 

those branches are low or absent. A greater source of 

negative effects of imidacloprid on HWA predators 

was therefore predicted to be a result of reduced prey 

quality and density (Eisenback, 2008). Although 

imidacloprid exposure through feeding on adelgids 

on treated trees is possible, most HWA available to 

predators should be located on untreated trees or 

trees with little risk of exposing predators to toxicity. 

Therefore, the new chemical technologies, the 

limited dispersal ability of the predator, and the 

predicted limited impact that systemically applied 

insecticides may have on predators in the fi eld all 

lead toward the idea of developing a strategy that 

uses both chemical and biological tactics in the 

same stands. One integration scenario is to maintain 

the health of a select number of large hemlocks 

with insecticide applications, and at the same time 

199 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

release and allow the biological control agents to 

become established on understory trees, increase, 

and serve as long-term suppressers of HWA. We 

hypothesize that this integrated approach could 

save more hemlock trees over time in a given area 

than the use of either control treatment (biological 

or chemical) in isolation. If shown to be an 

improvement over current strategies, it can become 

the standard approach to area-wide IPM for HWA. 

TESTING THE HYPOTHESIS 

A study has been initiated at Kentucky Ridge 

State Forest, near Middlesboro, KY. Th ree blocks, 

each containing one replicate of four treatments, 

have been established (Fig. 1). Th e treatments 

are: 1. treat a cohort of co-dominant to dominant 

trees with imidacloprid; 2. release L. nigrinus 

on a cohort of understory hemlock trees; 3. 

combine insecticide and beetle release treatments 

as in 1 and 2; and 4. do nothing (control). 

In plots assigned chemical-only and chemical plus 

predator treatments, 6 dominant or co-dominant 

eastern hemlock trees were chosen for chemical 

treatment. Chemical treatments consist of soil 

injection of imidacloprid (Merit 2F) applied with 

a Kiortz soil injector at a recommended rate of 

0.2 ounces of product per inch dbh (0.5 g [AI] 

/cm dbh). Insecticide treatments were applied 

Figure 1. Chemical and biological control plots in Kentucky Ridge State Forest, KY. 

200 
Chapter 19: Integrating Chemical and Biological Control 

in May 2010. In plots assigned beetle-only and 

beetle plus chemical treatments, 6 intermediate/ 

suppressed trees were chosen as predator ( L. 

nigrinus) release trees. A total of 125 lab reared 

L. nigrinus adults were released per tree (750 per 

plot) in October 2010. An additional 6 untreated 

trees within each plot were randomly selected for 

data collection to contrast with treated trees. 

Tree health and HWA population measures 

were made for all chosen trees before treatment 

and will be assessed annually for 3 more years. 

Tree health measures include percent live crown 

ratio, foliage transparency, new growth, and tip 

dieback. HWA populations will be measured by 

randomly selecting 10 branches per tree. Th e 

terminal 30 cm will be examined and the number 

of HWA counted until ten HWA are found 

and the next branch will be examined. Th e total 

number of HWA found on the ten branches will 

be summed and this number will be recorded 

as an index of HWA density for that tree. 

Sampling for adult predators every fall using 

beat-sheet methods and larvae every spring by 

clipping infested branches and rearing will be 

carried out to assess predator establishment. 

Predator exclusion sleeve-cage evaluations may 

also be carried out to determine if the impact 

from predation differs between predator only 

and predator plus chemical treatments. 

Additional sites over a wide geographic range will 

be added to this study in an attempt to evaluate 

the proposed strategy along the active front of 

HWA movement. We will choose locations that 

normally would be chosen for HWA predator 

releases (i.e., stands with building HWA populations 

and none to minimal decline in tree health). In 

these situations, HWA populations are often 

sporadically present throughout the stands. 

While this design will provide more answers over a 

longer time period, it is likely to yield measurable 

results within 3 years of application. We will be able 

to compare overall stand health for each treatment 

tested. We will also be able to compare predator 

establishment under the two predator treatment 

regimes to determine if presence of chemicallytreated trees impacts predator success. Perhaps when 

this is done, we will be able to recommend a new 

coordinated integrated control treatment that can 

save a higher percentage of trees at any one location. 

LITERATURE CITED 

Cowles, R.S.; Montgomery, M.E.; Cheah, C.A. 

2006. Activity and residues of imidacloprid 

applied to soil and tree trunks to control 

hemlock woolly adelgid (Hemiptera: Adelgidae) 

in forests. Journal of Economic Entomology 99: 

1258-1267. 

Eisenback, B.M. 2008. Prey-mediated Eff ects of 

Imidacloprid on Laricobius nigrinus (Coleoptera: 

Derodontidae) and Sasajiscymnus tsugae 

(Coleoptera: Coccinellidae), Two Predators of 

Hemlock Woolly Adelgid. Blacksburg, VA: 

Virginia Tech: 175 p. Ph.D. dissertation. 

Eisenback, B.M.; Salom, S.M.; Kok, L.T.; Lagalante, 

A.F. 2010. Lethal and sublethal eff ects of 

imidacloprid on hemlock woolly adelgid 

(Adelges tsugae, Hemiptera: Adelgidae) and two 

introduced predator species in the laboratory. 

Journal of Economic Entomology 103: 

1222-1234. 

Mausel, D.L.; Salom, S.M.; Kok, L.T.; Davis, 

G. 2010. Establishment of the hemlock 

woolly adelgid predator, Laricobius nigrinus 

(Coleoptera: Derodontidae), in the eastern 

United States. Environmental Entomology 39: 

440-448. 

201 
 

CHAPTER 20: INTEGRATING THE EARLY STEPS OF HOST SELECTION
 

BEHAVIOR INTO BIOLOGICAL CONTROL OF HWA 

Kimberly F. Wallin1,2 

1

The University of Vermont, The Rubenstein School of Environment and Natural Resources, 


Aiken Center, Burlington, VT
 

2

U.S. Forest Service, Northern Research Station, South Burlington, VT
 

INTRODUCTION 

When developing new programs, biological control 

practitioners have sometimes overlooked the role of 

behavior in the successful introduction of potential 

biological control agents. This is surprising since 

both host herbivore and natural enemy behavior can 

influence the overall efficacy of the biological control 

effort. To increase the effi ciency and effi cacy of using 

biological control agents to manage populations of 

non-native invasive organisms, biological control 

practitioners need to better understand the direct 

and indirect effects of the control agent. One way 

of predicting the potential direct and indirect 

effects of the control agent is to understand host 

specificity and behavioral processes involved in the 

multiple steps of host selection that end in host use. 

Host selection behavior is a series of direct 

and indirect behavioral responses to hosts. Th e 

steps in host selection behavior include habitat 

location, host location within the habitat, host 

acceptance, and host use (Kennedy 1965). 

This sequence of behavioral steps in host 

selection raises a number of questions that have 

consequences for host specificity testing in 

biological control programs. Perhaps the most 

important consequences are those that originate 

from lack of understanding of the early steps in 

the host selection sequence. For example, if habitat 

location and host location are absent in testing 

host specificity then the lack of that information 

may result in false positives and therefore, lead 

to an overestimation of host use in the fi eld. 

The same steps that dictate the acceptance of plants 

as hosts for herbivores also apply to natural enemies 

of the herbivores, such as predators (Vinson 1998). 

Natural enemies also use visual and chemical cues 

associated with the food plant to locate their hosts or 

prey. Known chemical attractants that are common 

in plants and attractive to insects include green leaf 

volatiles and terpenoids (Bukovinszky et al. 2005). 

Once at the plant, natural enemies rely on chemical, 

tactile, gustatory, and visual cues to locate their prey. 

In the case of specialized natural enemies, chemical 

cues used to locate suitable plants may diff er 

between exotic and native plant species. Predators 

may not initially respond to novel odors associated 

with the exotic plant, and these behaviors should 

be evaluated and incorporated into control eff orts. 

Testing to assess host use of potential biological 

control agents (arthropods) needs to incorporate as 

many steps of the host selection behavior sequence 

as possible. Options include using large assay arenas, 

outdoor arenas, or open fi eld-testing. Unfortunately, 

false positives and false negatives can occur in cage 

experiments due to the lack of behavior fi lters that 

occur early in the sequence. For example, Turanli 

and Schaffner (2004) reported that the sessiid moth 

Tinthia myrmosaeformis (Herrich-Schaff er) showed 

little host specificity in contact bioassays with test 

plants, great specificity in multiple-choice cage tests, 

and the highest level of specificity in open fi eld 

tests. Habitat location and pre-aligning cues often 

rely heavily on olfaction (Bernays and Chapman 

1994) and still air or cage assays do not allow 

202 
 

Chapter 20: Integrating the Early Steps of Host Selection Behavior

 into Biological Control of HWA 

testing insect responses to cues experienced during 

these early steps in the host selection sequence. 

Therefore, laboratory tests need to include wind 

tunnels, olfactometers, and/or airflow though cages. 

Olfactometers are well-known tools used to 

identify host finding cues in parasitoids (Mills and 

Wajnberg 2008), phytophagous insects (Bernays 

2001), and mosquitoes (Gillies 1990) but are less 

used in the development of biological control with 

predatory natural enemies. Nonetheless, there 

is no evidence that suggests that the behavioral 

responses of predators to host cues would be any 

different to that of parasitoids or phytophagous 

insects (Nakashima et al. 2002, Vanas et al. 

2006). Therefore, including olfactometer assays to 

complement methods currently used in biological 

control program could lead to greater success due to 

increased establishment in the novel environment. 

In addition to using multiple behavioral biological 

assays, it is important to incorporate external 

and internal factors that interact to infl uence the 

behavioral responses to host selection and use 

(Vinson 1984, Vet and Dicke 1992, Turlings et al. 

1993, Powell et al. 1998). External stimuli are often 

prey or plant derived (Misell et al. 1984, Rhoades 

1985, Price 1991), and may relate directly to host 

nutritional suitability or defensive capacity of the 

preys host plant (Hugentobler and Renwick 1995; 

Chambers et al. 1997; Frankfater and Scriber 1999, 

2003). Insects use visual (Prokopy and Owens 

1983), auditory (Drosopoulos and Claridge 2006), 

olfactory (Wood 1982), gustatory (Glendinning et 

al. 2009), and tactile stimuli (Rehman and Powell 

2010) to identify and locate potential hosts. Once 

received by the insect, they are fi ltered, integrated, 

and interpreted as an attractant or deterrent at each 

step in the host selection sequence. Individual insect 

condition such as lipids (Mayhew 1997, Wallin 

and Raffa 2000), age (Stienberg et al. 1992), prior 

experience (Cunningham et al. 2001, deBoer and 

Dicke 2006), and hosts associated with pre-adult 

development (Tamo et al. 2006) influence an insect’s 

response(s) to the external stimuli encountered 

during each step of the host selection sequence. 

Several natural enemies discriminate between 

volatiles emitted by infested or uninfested trees 

(Vet et al. 1990, Harmel et al. 2007). Th e source of 

these chemical stimuli can be from the herbivore, 

the plant, or from the interaction of the two. In the 

early steps of host selection sequence stimuli from 

the host plant of the prey may be more reliable 

for the insect predator. Understanding the role of 

habitat or hosts infested with the prey in the host 

selection process is an important but often difficult 

step in evaluating the overall efficacy of a biological 

control agent. Stimuli generated by the herbivore 

prey are the most reliable source of information to 

a predator because they can inform the predator of 

the presence, identity, availability, and suitability 

of the prey (Whitman 1988), but they may not 

be apparent or available. Herbivore-derived 

information has two inherent constraints that limit 

its detectability and, therefore, its use as stimuli for 

prey location. Generally, herbivore prey are a small 

component of a complex environment and any 

information they provide will be in small amounts. 

Additionally, prey tend to be inconspicuous to 

avoid predation. Stimuli from the host plant of the 

herbivore prey are usually more readily available 

because of the plants comparatively large biomass, 

but are less reliable predictors of herbivore prey 

presence and suitability (Lima and Dill 1990, 

Dicke 1999, Cortesero et al. 2000). Understanding 

interactions among host plants, herbivore prey, 

and predator behavior may uncover important 

aspects of the biology of the predator that would 

otherwise be unnoticed, such as the infl uence of 

pre-release handling (e.g., age, mating status, level 

of satiation), the response of predators to plant 

odors, and herbivore host-induced plant odors. 

Natural enemies are often held without exposure 

to their prey before release with unknown 

consequences to their host selection behavior. 

Physiological changes within an individual 

insect have been shown to affect orientation to 

hosts and responses to external cues (Kennedy 

1977, Miller and Strickler 1984, Wallin and 

Raffa 2000). Physiological changes predict that 

factors linked to time limitation (e.g. age, time 

203 
 

 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

since last meal) should increase the selection of 

suboptimal resources. However, in Papilio glaucus 

(Lepidoptera: Papilionida) the apparent direction 

of age effects are towards a greater specifi city 

(Scriber 1993). A reversal from time limitation 

to egg limitation appears counterintuitive as 

age increases, unless there is an increased cost 

of egg production associated with aging. 

Th e influence of food deprivation on host 

acceptance behavior suggests that satiated 

predators introduced into choice tests are not 

as likely to attack and feed on less preferred but 

otherwise acceptable test species than insects 

that are previously deprived of food (Withers et 

al. 2000, Barton and Whithers 2002). Predators 

that are satiated might reject test species in nochoice tests if the period without food or feeding 

on a non-target prey was short. Consequently, 

such tests would fail to reveal the fundamental 

host range (sensu Nechols and Kikuchi 1985, 

van Klinken 2000) therefore producing a false 

negative result. It is lesser known how recent food 

deprivation and lipid content of individual insects 

might influence the early steps of host selection 

sequence. Hence determining an optimal period 

of food deprivation prior to initiating testing host 

location and specificity is likely a good practice. 

Understanding consequences of food deprivation 

on host acceptance behavior may guide the 

release of potential biological control agents. 

Change in preference by ovipositing or feeding 

in adults due to previous experience with a host 

has been documented in various insects (e.g., 

Vinson et al 1977, Prokopy et al. 1982, Jaenike 

1983, Rausher 1983, Stanton 1984, Szentesi and 

Jermy 1990, Bjorksten and Hoffmann 1998; see 

also Agrawal et al. 2002). The phenomena of host 

selection being altered by previous experience with 

a host has been well reviewed for hymenopteran 

parasitoids by Turlings et al. (1995) and Vet et 

al. (1995). The general prediction is that there is 

a positive relationship between time of the last 

feeding and responsiveness to a lower ranked 

host. However, this is not always supported in 

laboratory tests. For example, Scriber (1993) 

exposed P. glaucus adults from the same population 

for two days to four differently preferred hosts. 

Oviposition specificity was infl uenced only 

on the first day after being placed in the arena 

with the four hosts present and not after. 

There are fewer studies that have investigated the 

effects of experience on host selection behavior 

of predators. However, this is a very important 

step in developing a biological control program 

because one challenging effect to avoid is any 

enhanced responsiveness to the rearing host or 

environment. For example, increased attraction 

following experience with a host plant volatiles was 

demonstrated in the introduced predator Anthocoris 

nemoralis (F.) (Heteroptera: Anthocoridae) (Drukker 

et al. 2000). Specifically, wild-caught predators 

preferred odors from the host plant when off ered 

either clean air, whereas laboratory reared fi rst 

generation predators did not. However, after the 

laboratory reared predators were exposed to the 

host plant or host plant volitiles they showed a 

strong preference for the volatiles. Th e predatory 

mite Phytoseiulus persimilis Athias-Heriot is more 

attracted to the host that the prey was reared on 

when previously exposed to volatiles emanating 

from the prey’s host (Dicke et al. 1990, de Boer 

et al. 2005). These results are similar to those seen 

in some parasitoids and suggest that experienceinduced changes in host selection behavior could 

be expected to occur with predators. Any bias 

for the previously consumed species and/or its 

substrate can be avoided or reduced by parallel 

experimental approaches used for parasitoids. 

A particular difficulty in identifying larval 

conditioning is separating early adult experience 

from larval experience. Th is difficulty led to a period 

in which the presence of Hopkins’ Host Selection 

Principle was largely discounted (e.g. Courtney 

and Kibota 1990, van Emden et al. 1996, Barron 

2001). However, several studies in which larvae 

have been exposed to specific host plant chemicals 

have indicated that induction of adult ovipositional 

preference may occur during the larval period (e.g., 

Del Campo et al. 2001, Ikkei at al. 2010; see also 

Jaenike 1983, Tully 1994). The capacity to alter 

behavior with experience is now well documented 

in parasitoid insects, with associative learning in 

204 
 

 

Chapter 20: Integrating the Early Steps of Host Selection Behavior

 into Biological Control of HWA 

adults and learning through contact or exposure to 

host-derived chemicals during development (Vet 

et al. 1990). Parasitoids use of a variety of host and 

host-plant cues in host selection behavior sequences 

was thought to be fixed and innate. However, it is 

now clear that behavior is often plastic and varies 

between individuals with different exposure histories 

as well as their genetic composition. Identifying 

how past exposure to host herbivore and hostplant derived chemicals as a source of intraspecifi c 

behavioral variation is of interest for the utilization 

of parasitoids as biological control agents (Vet and 

Groenewold 1990). The host selection behavior 

of the generalist predatory paper wasp, Polistes 

dominula (Hymenoptera: Vespidae), is infl uenced 

by larval experience (Rayor and Munson 2002). 

They tested the role that larval experience with 

unpalatable prey played in subsequent foraging 

choices by adult predators. The results of the 

study demonstrate that previous experience with 

deterrent chemicals during larval development 

altered patterns of prey acceptability to the adult 

insects. It is not clear if these trends will also be 

seen with additional predatory insects but they 

are observed in herbivores and may also apply to 

natural enemies of the herbivores, such as predators. 

Understanding these cues, interactions between 

host plant and herbivore prey, and behaviors 

associated with predation are especially important 

with the increased attention on biological control 

of invasive organisms. We have conducted and are 

in the process of conducting host selection behavior 

studies of several potential predators of hemlock 

woolly adelgid, Adelges tsugae Annad (Homoptera: 

Adelgidae) (HWA) and provided information to 

increase overall efficacy in the efforts to control 

HWA in hemlock forests in eastern United States. 

OVERVIEW OF METHODOLOGY 

Behavioral bioassays were conducted to test the 

responses of adult beetles to HWA and several tree 

species (Table 1). Bioassays are ongoing. Methods 

are fully described in Wallin et al. (in press). Briefl y, 

beetles were brought to the laboratory and held at 

16 °C; 12:12 on HWA infested hemlock branches 

for 24-48 hours. All beetles were randomly chosen 

from one laboratory population, tested once, 

and moved to a separate laboratory population. 

The four-arm olfactometer (Analytical Research 

Systems, #OLFM-4-C-2440PE. Gainesville, FL 

USA) was used in all bioassays. Th e olfactometer 

consisted of three parts: the base with the air 

output, the intermediate part, which delimited 

the walking chamber with a 9-cm circular 

central opening to introduce insects, with the 

four air inputs, and the transparent lid. Air was 

drawn in at a flow rate of -0.1Mpa. Air fl ow was 

controlled at 1.2Mpa in all four arms into the 

glass chambers containing the test material and 

carrying volatiles into the olfactometer. Arms with 

chambers receiving air but without test material 

were regarded as “blank” or control chambers. 

All ambulatory responses were tested at 20 °C with 

artificial overhead lighting. For each experimental 

run an individual beetle was introduced at the 

center of the olfactometer. Each test lasted 15 

min. Two criteria quantified behavior: (1) time 

spent in each field and (2) if the insects chose 

an odor field at the beginning of the test and 

remained in that fi eld (final position). When air 

passed through the chambers into the olfactometer 

stage each of the airfields were considered to be a 

separate field and an additional field, central fi eld 

(CF), at the center of the arena. We considered 

that an insect entered a given field when its 

entire thorax crossed the field boundary. A 

test was not retained when an insect remained 

motionless in the CF for more than 5 minutes. 

SUMMARY OF FINDINGS 

Laricobius nigrinus, S. coniferarum, and S. tsugae 

responded to odors from HWA’s host trees, but 

not to odors associated with HWA alone. HWA 

being inconspicuous to all three predators suggests 

that HWA is extremely difficult to detect. Th is 

suggests these predators rely on volatiles produced 

by HWA’s host trees to locate potential prey. 

This is true for many parasitoid species (Lima 

and Dill 1990, Vet et al. 1990, Tumlinson et al. 

1993, Dicke 1999, Cortesero et al. 2000). 

205 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Table 1: Series of laboratory experiments quantifying the interactions of internal physiology and 

external stimuli on an early step in host selection sequence. The behaviors of Laricobius 

nigrinus (Ln), Scymnus coniferarum ( Sc), Sasajiscymnus tsugae (St), Laricobius rubidus 

(Lr), and Laricobius hybrids (Ln x Lr) were tested using four-armed olfactory behavioral 

bioassays. 

Internal physiology 

External stimuli Starvation/lipids Sex Host during pre-adult development 

HWA Ln1

, Sc2, Lr4, LnxLr4, 

LrxLn4 

Ln1, Sc2, Lr4, LnxLr4, 

LrxLn4 Ln1

, St3,5 

Eastern Hemlock Ln1

, Sc2, Lr4, LnxLr4, 

LrxLn4 

Ln1, Sc2, Lr4, LnxLr4, 

LrxLn4 Ln1

, St3,5 

Western Hemlock Ln1, Lr4, LnxLr4, LrxLn4 Ln1, Lr4, LnxLr4, LrxLn4 Ln1, St3,5 

Eastern White Pine Ln1

, Sc2, Lr4, LnxLr4, 

LrxLn4 

Ln1, Sc2, Lr4, LnxLr4, 

LrxLn4 Ln1

, St3,5 

Western White Pine Ln1, Lr4, LnxLr4, LrxLn4 Lr4, LnxLr4, LrxLn4 Ln1, Lr4, LnxLr4, LrxLn4 

Douglas-Fir Ln1 Ln1 

Spruce Ln1 Ln1 

Ponderosa Pine Ln1 Ln1 

Eastern Hemlock with HWA Ln1

, Sc2, St3, Lr4, 

LnxLr4, LrxLn4 

Ln1, Sc2, St3, Lr4, 

LnxLr4, LrxLn4 Ln1

, St3,5 

Western Hemlock with HWA Ln1

, Sc2, St3, Lr4, 

LnxLr4, LrxLn4 

Ln1, Sc2, St3, Lr4, 

LnxLr4, LrxLn4 Ln1

, St3,5 

1

Collected by Glenn Kohler from 16 sites in WA and OR (see Kohler et al. 2008, Wallin et al. in press).
 2

Collected by Dr. Richard McDonald from 1 site in WA (see Wallin et al. in prep).
 3

Reared and supplied by Virginia Poly Technical Institute and State University Biological Rearing facility.
 4

Collected by Dr. Richard McDonald from release sites in NC (assays to begin in 2011).
 5

Collected by Dr. Ashley Lamb from sites in Japan.
 

Host odors are less reliable than odors generated 

by the prey. This challenge may be overcome in 

systems where the predator, prey, and host have 

co-evolved with plants (Vet et al. 1990, Harmel 

et al. 2007). Volatiles induced by feeding of 

the prey may provide specific information to 

the predator and greatly increase its reliability 

(Harmel et al. 2007). However, the presence of 

feeding HWA did not increase the attractiveness 

of hemlock branches to L. nigrinus (Wallin et al. 

in press), S. coniferarum (Wallin et al. in prep) or 

S. tsugae (Wallin et al. in prep) suggesting that 

these predators are responding to hemlock volatiles 

produced by the cut branch, but not specifi c 

odors produced in response to adelgid feeding. 

The condition of individual beetles infl uenced the 

response to host plant odors in the olfactometer. 

All three beetle species were more likely to respond 

to host plant odors if they were starved. Th e eff ects 

of starvation on response to host plant odors 

manifested within 24 hr without access to HWA 

for L. nigrinus and 4 hr without access to HWA for 

both S. coniferarum and S. tsugae (Wallin et al. in 

prep). However, the response to host plant odors 

decreased as the length of starvation increased for 

L. nigrinus, S. coniferarum, and S. tsugae. Total lipid 

content declined with the duration of starvation; 

however, there was not a linear relationship 

between lipid content and the mean time response 

time of L. nigrinus, S. coniferarum, and S. tsugae 

to host plant odors (Wallin et al. in prep). 

206 
 

Chapter 20: Integrating the Early Steps of Host Selection Behavior

 into Biological Control of HWA 

Another way predators can overcome the low 

reliability of host odors is through conditioning 

during pre-adult development to respond to volatiles 

from the host plant they developed on (Tamo et 

al. 2006) or conditions in which they were reared. 

We found that L. nigrinus collected from western 

hemlock walked toward the chamber with western 

hemlock, whereas those reared in the laboratory on 

eastern hemlock showed no preference. However, S. 

tsugae collected from hemlock in Japan responded 

to both eastern and western hemlock, whereas 

those reared in the laboratory on eastern hemlock 

did not respond in the assay arena (Wallin et 

al. in prep). It is not clear why adults raised on 

eastern hemlock did not walk toward their natal 

hemlock species. Multi-generational studies on 

the effect of rearing on predator-prey location 

behavior are needed as are studies on associative 

learning. However, it should be noted that our 

results apply to an enclosed environment where 

chemosensory cues are highly concentrated and 

easily discernible. Whether or not the diff erence in 

volatiles would be detectable by insects in a fi eld 

situation is unknown and requires further study. 

Overall, our study suggests that these three potential 

predators of HWA that have been tested to date 

use chemosensory cues from HWA host trees and 

that these responses are plastic and depend on both 

season, external, and internal cues. Th ese factors 

should be taken into consideration when planning 

releases of predators as biological control agents. 

ACKNOWLEDGEMENTS 

Support from Oregon State University, University of 

Vermont, The University of Sydney is acknowledged. 

Funding for this research was from U.S. Forest 

Service-FHTET and U.S. Forest Service, Northern 

Research Station. I thank Glenn Kohler, Dan Ott, 

Dr. Darrell Ross, Dr. Tanya Latty, Dr. Stephen 

Cook, Dr. Richard Mc Donald, Dr. Scott Salom, 

Schroder Seed Orchard, Hoyt Arboretum, and 

others for their assistance in coordinating, collecting, 

and shipping material. I am grateful that Brad 

Onken and Richard Reardon invited me to write 

about the importance of host selection behaviors in 

biological control and predator-prey interactions. 

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grandis) II. Associative learning. Physiological 

Entomology 2: 157-164. 

Vinson, S.B.; Elzen, G.W.; Williams, H.J. 1987. 

Th e influence of volatile plant allelochemics 

on the third trophic level (parasitoids) and 

their herbivorous hosts. Insects-Plants. In: 

Labeyrie,V.; Fabres, G.; Lachaise, D. eds. 

Proceedings of the 6th International Symposium on 

Insect-Plant Relationships (Pau 1986): 109-114. 

Vinson, S.B. 1998. The general host selection 

behavior of parasitic Hymenoptera and a 

comparison of initial strategies utilized by 

larvaphagous and oophagous species. Biological 

Control 11: 79-96. 

Wallin, K.F.; Latty, T.M.; Ross, D.W. 201x. 

Orientation behavior of the predator Laricobius 

nigrinus (Coleoptera: Derodontidae) to 

hemlock woolly adelgid and host tree odors in 

a multi-chambered olfactometer. Environmental 

Entomology. Accepted. 

Wallin, K.F.; Raffa K.F. 2000. Influences of host 

chemicals and internal physiology on the 

multiple steps of postlanding host acceptance 

behavior of Ips pini (Coleoptera: Scolytidae). 

Environmental Entomology 29: 442-453. 

Whitman, D.W. 1988. Allelochemical interactions 

among plants, herbivores, and their predators. 

In: Barbosa, P.; Letourneau, D., eds. Novel 

Aspects of lnsect-Plant Interactions. New York, 

NY: John Wiley & Sons: 11-64. 

210 
Chapter 20: Integrating the Early Steps of Host Selection Behavior

 into Biological Control of HWA 

Withers, T.M.; Barton Browne, L.; Stanley, J. 

2000. How time-dependent processes can 

effect the outcome of assays used in host 

specificity testing. In: Van Driesche, R.G.; 

Heard, T.A.; McClay, A.S.; Reardon, R., eds. 

Proceedings: Host Specifi city Testing of Exotic 

Arthropod Biological Agents: Th e Biological 

Basis for Improvement in Safety. X International 

Symposium on Biological Control of Weeds; 

1999 July 4-14; Bozeman, Montana, 

USA. USDA Forest Service Bulletin 

FHTET-99-1. Morgantown, WV: U.S. 

Department of Agriculture, Forest Service, 

Forest Health Technology Enterprise Team: 

27-41. 

Wood, D.L. 1982. The role of pheromones, 

kairomones, and allomones in the host selection 

behavior of bark beetles. Annual Review of 

Entomology 27: 411-446. 

211 
 

 

 

 

 

 

CHAPTER 21: THE INTRODUCTION OF LARICOBIUS NIGRINUS AS A
 

BIOLOGICAL CONTROL AGENT FOR THE HEMLOCK WOOLLY ADELGID: 


IS THERE A THREAT TO THE NATIVE CONGENER, L. RUBIDUS? 


Nathan Havill1, Gina Davis2, Melissa Fischer2, Scott Salom2, Dave Mausel3, and Brad Onken4 

1

U.S. Forest Service, Northern Research Station, Hamden, CT
 

2

Department of Entomology, Virginia Tech, Blacksburg, VA
 

3

Division of Entomology, University of Massachusetts, Amherst, MA
 

4

U.S. Forest Service, Northeastern Area, State and Private Forestry, Morgantown, WV
 

BACKGROUND 

Laricobius Rosenhauer is one of four genera in the 

family Derondontidae (Coleoptera) that occupies 

the temperate regions of the Northern Hemisphere 

(Lawrence 1989). Members of this genus are only 

known to prey on adelgids (Hemiptera: Adelgidae) 

(Lawrence and Hlavac 1979, Lawrence 1989). 

There are three species native to North America: L. 

nigrinus and L. laticollis are native to western North 

America, and L. rubidus is native to eastern North 

America. Laricobius nigrinus is being used in the 

eastern United States as a biological control of the 

hemlock woolly adelgid. Previously, L. erichsonii 

was introduced to both coasts of North America 

from Europe for control of the balsam woolly 

adelgid (summarized in Montgomery et al. 2011). 

This species was reported to have established, but 

its most recent recorded recovery was in 1978 

(Schooley et al. 1984). A molecular study of the 

relationships among the four Laricobius species 

reported in North America, plus L. kangdingensis 

and L. osakensis from Asia, showed that, surprisingly, 

the two species from western North America 

were not the most closely related (Montgomery 

et al. 2011). Instead, it was found that L. nigrinus 

is very closely related to the eastern species, L. 

rubidus. In fact, the genetic distance between 

these species (using a portion of the mitochondrial 

COI gene) was only slightly higher than within 

each species (Davis et al. 2011). Th is suggests 

that they diverged very recently and may not be 

reproductively isolated. Recent work (described 

in more detail below) has shown that L. nigrinus 

and L. rubidus are in fact interbreeding at sites in 

the eastern U.S. where L. nigrinus was released. 

It is not yet known if this will enhance or hinder 

hemlock woolly adelgid (HWA) biological control. 

ACKNOWLEDGING RISK 

It is important to weigh the benefits and risks 

when making decisions for natural resource 

management (Loomans and van Lenteren 2005). 

Benefits of hemlock woolly adelgid are nil, while 

the risks and costs are great. Loss of hemlock 

timber and pulpwood (Burns and Honkala 

1990, Ward et al. 2004) and residential property 

values (Holmes et al. 2006) can be tabulated, 

while calculating costs associated with intangible 

environmental and aesthetic benefits are much 

more difficult (Anders 1977, McConnachie et al. 

2003). Left uncontrolled, HWA has the potential 

to cause hemlock mortality within all 25 forest 

cover types of which it is a component (Burns 

and Honkala 1990, Orwig et al. 2002). Loss of 

hemlock alters eco-hydrological systems (Ford 

and Vose 2007) and accelerates growth of invasive 

plants (Eschtruth and Battles 2008). It can also 

negatively impact temperature-sensitive streams 

(Snyder et al. 2002, Ross et al. 2003) and habitat 

212 
 

Chapter 21: The Introduction of Laricobius nigrinus as a Biological Control Agent for the Hemlock 

Woolly Adelgid: Is There a Threat to the Native Congener, L. rubidus? 

for numerous wildlife species (Yamasaki et al. 1999, 

Onken and Souto 2000, Lishawa et al. 2007). 

Some of the risk associated with a biological control 

agent can be evaluated by laboratory experiments 

prior to its introduction, yet it is recognized that 

environmental variability and other sources of 

uncertainty are cause for continued post-release 

assessment (Louda et al. 2003, Hopper et al. 2006). 

An unexpected risk that was recently discovered in 

association with the release of L. nigrinus is its ability 

to hybridize with a native species, L. rubidus. In this 

report, we summarize what is currently known about 

interbreeding between L. nigrinus and L. rubidus, 

and we discuss research directions to evaluate the 

implications for biological control of HWA. 

Laricobius nigrinus Fender 

Laricobius nigrinus Fender is a small (2-3 mm), 

black beetle native to western North America 

(Fender 1945, Zilahi-Balogh et al. 2006) where it 

has been found to be a widespread and abundant 

natural enemy of HWA, at both low and high 

densities of the pest (Kohler et al. 2008). 

Both adults and larvae feed on A. tsugae eggs, 

nymphs, and adults. Eggs of L. nigrinus are laid 

in late winter and early spring. Larvae develop 

through four instars, feeding on HWA progrediens 

eggs, and drop to the forest floor to pupate. Adults 

diapause during summer in the soil and emerge 

in fall to feed on HWA sistens nymphs in the 

fall and winter (Zilahi-Balogh et al. 2003). Th e 

life cycles of L. nigrinus and HWA are highly 

synchronized (Zilahi-Balogh et al. 2003). 

Laricobius nigrinus was imported into the eastern 

United States from Victoria, British Columbia for 

further evaluation and was determined to be hostspecific in the laboratory (Zilahi-Balogh et al. 2002). 

Federal and State approval for environmental release 

of L. nigrinus was granted in 2000. Laboratory 

mass-rearing methods were developed for L. 

nigrinus and adults are currently being reared in a 

number of laboratories (Lamb et al. 2005). Free 

releases of L. nigrinus began in 2003. As of 2009, 

L. nigrinus adults were released in 15 eastern states, 

spanning USDA plant hardiness zones 5a to 7a 

(Roberts et al. 2010). It was found to establish 

in 13/22 (59%) of initial release sites (Mausel et 

al. 2010). The probability of establishment was 

greater at sites with higher minimum annual 

temperatures and where more beetles were released. 

Additional L. nigrinus from Idaho has been released 

in several New England states in an attempt to 

establish a more cold-hardy strain in the north. 

Laricobius rubidus 

Laricobius rubidus is the only species of Laricobius 

native to eastern North America (Clark and Brown 

1960; Lawrence 1989). Its known distribution 

extends from the District of Columbia, north to 

New Brunswick, west to Minnesota, and south to 

North Carolina (Brown 1944, Raske and Hodson 

1964, Lawrence 1989, Wallace and Hain 2000). Its 

primary host is the pine bark adelgid (PBA), Pineus 

strobi Hartig (Clark and Brown 1960). Laricobius 

rubidus has also been found to occasionally feed 

on the balsam woolly adelgid, Adelges piceae 

Ratz. (Lawrence and Hlavac 1979) and has been 

collected from eastern hemlock infested with HWA 

throughout its introduced range (Montgomery 

and Lyon 1996, Wallace and Hain 2000, Mausel 

et al. 2008). Laboratory studies have shown that 

it can reproduce and complete development on 

HWA, but has an ovipositional preference for pine 

bark adelgid (PBA) (Zilahi-Balogh et al. 2005). 

The life cycle of L. rubidus is well synchronized with 

that of PBA (Clark & Brown 1960). Adults are 

active between late March and early June with peak 

activity between mid-April to mid-May (Clark and 

Brown 1960, Zilahi-Balogh et al. 2005). Four instars 

are present late April through early June (Clark 

and Brown 1960), migrating to the soil to pupate 

by late June (Zilahi-Balogh et al. 2005). Emerging 

adults undergo an aestival diapause, becoming 

active in October through early November (ZilahiBalogh et al. 2005). In Virginia, L. rubidus adults 

have been observed migrating from the branches 

to the duff where they are thought to undergo a 

hibernal diapause, but adults associated with HWA 

have also been found to be active in the winter 

(Zilahi-Balogh et al. 2005, Mausel et al. 2008). 

213 
 

 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

POTENTIAL FOR INTERBREEDING 

Adult L. nigrinus can be distinguished 

morphologically from L. rubidus. Laricobius 

nigrinus (Fig. 1) has unicolorous (black) elytra, 

the distance across the posterior of the pronotum 

is greater than across the anterior, and the apices 

of the lateral parameres of the male genitalia 

are narrowly acute. In contrast, L. rubidus 

(Fig. 2) has bicolor (red and black) elytra, the 

distances across the posterior and anterior of 

the pronotum are subequal, and the apices of 

the lateral parameres are truncate (Montgomery 

et al. 2011, Leschen 2011). The immature life 

stages are morphologically indistinguishable. 

Several observations prompted questions about the 

potential of L. nigrinus and L. rubidus to interbreed: 

1) both species are routinely recovered from 

HWA-infested hemlock trees in the eastern U.S. at 

sites where L. nigrinus was released; 2) molecular 

analysis of the genus Laricobius found, surprisingly, 

that L. nigrinus and L. rubidus are very closely 

related suggesting that they are recently diverged 

species that may have the ability to produce viable 

offspring (Klein et al. 2010, Montgomery et al. 

2011); 3) members of the two species were observed 

copulating with each other on HWA infested 

hemlock at the Virginia Tech field insectary, which 

neighbors a white pine stand infested with pine bark 

adelgid (Mausel et al. 2008); and 4) morphological 

and molecular species identification were found 

to be in conflict for two beetles collected from a 

L. nigrinus release site in Maryland, suggesting 

that these individuals could be of hybrid origin. 

This prompted the development of microsatellite 

markers that could be used to distinguish L. nigrinus 

and L. rubidus from their hybrids (Klein et al. 

2010). This method exposed a trend of an increasing 

proportion of hybrids recovered at L. nigrinus 

release sites in Pennsylvania, North Carolina, and 

Tennessee between 2007 and 2009 (Havill et 

al. 2010). It was also used to identify Laricobius 

adults collected from HWA-infested hemlock the 

Virginia Tech field insectary where PBA-infested 

white pine grows in close proximity (N. Havill, 

unpublished data). Data from six microsatellite 

loci analyzed with the software NEWHYBRIDS 

(Anderson & Thompson 2002) were used to classify 

beetles. In 2008 we collected 27 L. nigrinus, 15 

L. rubidus, and 13 hybrids. In 2010 we collected 

87 L. nigrinus, 4 L. rubidus, and 8 hybrids, and in 

2011 we collected 87 L. nigrinus, 7 L. rubidus, and 

9 hybrids. Further confirmation that these species 

can interbreed was shown in a 2009 laboratory 

study where three interspecific pairs produced 

viable offspring (T. Dellinger, unpublished data). 

Figure 1. Laricobius nigrinus (photo by Gina Davis). Figure 2. Laricobius rubidus (photo by Gina Davis). 

214 
 

Chapter 21: The Introduction of Laricobius nigrinus as a Biological Control Agent for the Hemlock 

Woolly Adelgid: Is There a Threat to the Native Congener, L. rubidus? 

Field collected beetles that were identifi ed as 

having mixed parentage had morphological 

characters that resembled either parent species 

or were intermediate—i.e., they had black 

or bicolored elytra, and the parameres of the 

male genitalia were either accute, truncate, or 

intermediate (Fig. 3). It is therefore not possible 

to use morphology to distinguish beetles of 

mixed parentage from the parent species. 

The ecological niches occupied by Laricobius species 

and their offspring may affect the geographic 

distribution and extent of interbreeding. Laboratory 

host range studies show that L. nigrinus prefers 

HWA on hemlock (Zilahi-Balogh et al. 2002), 

and L. rubidus prefers PBA on white pine (ZilahiBalogh et al. 2005). It may therefore be more likely 

for the two species to encounter each other in areas 

where hemlock and white pine co-occur than in 

areas with only one host species is present. Th e 

extent to which Laricobius adults migrate between 

stands would also affect the rate of interbreeding. 

Laricobius nigrinus was found to be common 

within 300 m of the original release trees by the 

fourth generation (G. Davis, unpublished data). 

Other observations suggest that L. nigrinus can 

disperse greater distances. For example, McDonald 

(2010) recovered L. nigrinus from at least 1.6 

km from the release area, five years post-release. 

Preliminary data suggest that the geographic overlap 

of hemlock and white pine may indeed aff ect the 

rate and incidence of interbreeding between the 

species. Recovery of L. rubidus on hemlock was 

lower where eastern white pine was sparse or absent 

from stands in which L. nigrinus was released (G. 

Davis, unpublished data). In addition, we collected 

Laricobius from white pine at the Virginia Tech 

field insectary in 2011, and all 47 were classifi ed 

as pure L. rubidus (Havill, unpublished data). 

Additional samples from hemlock and white pine 

in L. nigrinus release sites, as well as laboratory 

choice tests with hybrid beetles will help to further 

predict the importance of ecological factors in 

determining the outcome of interbreeding. 

POSSIBLE HYBRIDIZATION SCENARIOS 

Introductions of nonnative species can have 

large impacts on the genetics of native species 

through hybridization and introgression (i.e. 

gene flow) (Mooney and Cleland 2001, Mallet 

2007). Hybridization between L. rubidus and L. 

nigrinus could have several outcomes, including: 

• Hybrid Incompatibility 

▪ Sterility of hybrids 

▪ Outbreeding depression 

▪ Reinforcement of premating isolation 

• Hybrid vigor 

▪ Speciation 

▪ Genetic assimilation 

Figure 3. 	 Examples of elytra (left) and slide-mounted male genitalia (right) of L. nigrinus, L. rubidus and their 

hybrids. Arrows point to the lateral parameres that are acute in L. nigrinus and truncate in L. rubidus. 

Hybrids can resemble either parent species or can be intermediate. 

215 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

Hybrid Incompatibility 

Reproductive isolation between populations 

can result in the accumulation of genetic 

incompatibilities over time. This could make 

reproductive isolation permanent, even if the cause 

of isolation were removed (Palmer and Feldman 

2009). Hybridization produces recombinant 

genotypes that have not previously been subjected 

to selection. These genotypes will typically be less 

well adapted than those of their parents, resulting in 

selection against hybrids (Burke and Arnold 2001). 

Sterility 

Selection against hybrids is often exhibited as 

sterility or inviability (Haldane 1922, Mallet 

2007). The production of sterile or inviable 

offspring would result in a decrease in fi tness of 

the parental species due to an overall decrease 

in reproductive output. Although Laricobius 

hybrid sterility is a possibility, there is evidence 

of F2 hybrid individuals and backcrosses in the 

field (N. Havill, unpublished data), suggesting 

that at least some of the F1 hybrids are fertile. 

Outbreeding depression 

Outbreeding depression is a reduction in hybrid 

fitness, possibly due to the hybrid off spring being 

less well adapted to environmental conditions than 

the parental species (Klug and Cummings 2003). 

Outbreeding could result in lower reproductive 

potential (Arnold 1997). The reduction in 

reproductive output may occur as a result of a 

decrease in the number of off spring produced 

or as a result of lower levels of fertility or vigor 

among the hybrid progeny (Arnold 1997). We 

do not know whether Laricobius hybrids are 

less fit than their parents. Ongoing laboratory 

and field studies are explicitly testing this. 

Reinforcement of pre-mating isolation 

Hybridization can lead to an increase in 

reproductive isolation between parent species when 

mating barriers evolve due to selection against 

unfit hybrids (Mallet 2007). If reinforcement is 

occurring as L. nigrinus is released into sites where 

L. rubidus is present, over time we will see a decrease 

in hybridization and eventually a termination of 

hybridization as reinforcement becomes more 

powerful. This would also allow the parent 

species to remain genetically intact. Assessment of 

whether this is occurring will require long-term 

monitoring of the frequency of interbreeding. 

Hybrid Vigor/Heterosis 

Hybrids are often assumed to be less fi t than 

their parents, but this is not always the case 

(Arnold 1997). A review by Arnold and 

Hodges (1995) found that hybrids were not 

uniformly less fit than parental genotypes. 

Speciation 

Unique adaptations might arise from combining 

divergent genomes (Arnold 1997, Mooney and 

Cleland 2001, Mallet 2007). The increased genetic 

variability that results from crossing divergent 

genotypes can result in offspring that are better 

adapted to changed and changing environments, 

allowing them to occupy a habitat that was not 

available to the parents (Arnold 1997, Mooney 

and Cleland 2001). If hybrids tend to assemble 

in habitats different than the parents by means of 

seasonality, drift in small populations, or change 

in host preference, then gene flow between hybrids 

and parents will be reduced, and hybrid speciation 

(the origin of a new species) can occur (Mooney 

and Cleland 2001, Mallet 2007). Adelgids on 

hemlocks in the eastern U.S. is a new niche 

that was created when HWA was introduced 

from Japan. Laricobius hybrids could be better 

adapted to this niche than their parents if they 

receive a preference for hemlock woolly adelgid 

from their L. nigrinus parents, and hardiness in 

eastern climates from L. rubidus. Th is possibility 

is being evaluated in laboratory and fi eld studies. 

Genetic assimilation 

Open niches are not the only possible habitats 

for hybrids to invade (Arnold and Hodges 1995). 

If hybrids have an equivalent or higher fi tness 

than the parents in their own habitat, the hybrids 

may replace the “pure” parental species due to 

competition (Arnold 1997, Mallet 2007). For 

216 
 

 

 

 

Chapter 21: The Introduction of Laricobius nigrinus as a Biological Control Agent for the Hemlock 

Woolly Adelgid: Is There a Threat to the Native Congener, L. rubidus? 

example, if hybrids were to show greater feeding 

efficiencies than those of the parental species, this 

could result in a greater reproductive capacity 

of hybrids and the displacement of the parental 

species locally (Grant and Grant 1996). 

HYBRIDIZATION IN OTHER CLASSICAL 

BIOLOGICAL CONTROL PROGRAMS 

There are very few examples in the literature of 

introduced biocontrol agents interbreeding with 

native species. We are aware of just three systems in 

which this was investigated in the laboratory, one of 

which also tracked hybridization in the fi eld. Naka 

et al. (2005, 2006) found that a Chrysoperla carnea 

(Chrysopidae) introduced from Germany was able 

to produce fertile F1, F2, and backcrossed off spring 

with native Japanese C. nipponensis in the laboratory, 

but concluded that they were unlikely to hybridize 

extensively in the field because hybrid fertility was 

low, and the parent species have diff erent courtship 

songs. Davies et al. (2009) used DNA sequence data 

to confirm that introduced Diadegma semiclausum 

(Ichneumonidae) can hybridize with native Japanese 

D. fenestrale in the lab, and encouraged fi eld studies 

to follow up. Finally, Moriya et al. (1992) showed 

that an introduced parasitoid of chestnut gall 

wasps, Torymus sinensis (Torymidae), from China 

can hybridize with a native Japanese species, T. 

benefi cus. The native species has an early-spring 

and a late-spring strain. Using fi eld-collected wasps 

from a single chestnut orchard, Yara et al. (2010) 

found that the early-spring strain was displaced 

by the introduced species without evidence of 

hybridization, while the late-spring strain showed 

increasing frequency of hybrids over time. Th e 

effects on pest control were not evaluated. 

CONCLUSIONS 

Hybridization between L. nigrinus and L. rubidus 

has been confirmed in several L. nigrinus release 

sites where eastern white pine and eastern hemlock 

co-occur. We know hybrids are feeding on HWA 

and are capable of reproducing but there is no 

indication as yet whether hybridization will 

negatively or positively affect the HWA biocontrol 

program. Laboratory tests are underway to assess 

the feeding preferences and fitness of hybrids 

relative to the parent species. Both species readily 

feed and reproduce on hemlock woolly adelgid 

although laboratory studies indicate L. nigrinus 

is not able to successfully reproduce on pine bark 

adelgid. Laboratory studies have also shown each 

predator species to have a preference for one or 

the other adelgids when presented with a choice. 

Based on preliminary results of genotyping more 

than 1700 specimens collected from across the 

landscape where L. nigrinus has been released, 

the rate of hybridization has thus far been shown 

to be approximately 7 percent. These results and 

the known differences in host preference suggest 

that species separation is likely to be maintained 

with infrequent gene flow between the two 

species. We will continue to monitor this unusual 

hybridization event as it plays out over time. 

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Leschen, R.A. 2011. World review of Laricobius 

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Louda, S.M.; Pemberton, R.W.; Johnson, M.T.; 

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Chapter 21: The Introduction of Laricobius nigrinus as a Biological Control Agent for the Hemlock 

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Entomologist 138: 595-601. 

221 
CHAPTER 22: AN OVERVIEW AND OUTLOOK
 

FOR BIOLOGICAL CONTROL OF HEMLOCK WOOLLY ADELGID
 

Brad P. Onken1 and Richard C. Reardon2 

1

U.S. Forest Service, Northeastern Area, Morgantown, WV
 

2

U.S. Forest Service, Forest Health Technology Enterprise Team, Morgantown, WV 


Surveys in the eastern United States indicated 

that the native natural enemy complex associated 

with hemlock woolly adelgid (HWA) consisted 

primarily of generalist predators which did not 

effectively control HWA populations. Since no 

known parasitoids are associated with Adelgidae, 

the classical biological control program for HWA 

has focused on prey-specifi c predator species 

and entomopathogens. Although the earliest 

investigations into HWA biological control dates 

back to the early 1990s, the program has been 

going strong for the past decade and has involved 

28 federal and state agencies, 24 universities, 

seven institutions in China and Japan, and 

numerous private industries. Th e integration 

and focus of biological control into an integrated 

pest management program remains the option 

of primary interest for suppression of HWA. 

OVERVIEW 

In the early 1990s, the lady beetle Pseudoscymnus 

n. sp (now Sasajiscymnus tsugae) and the oribatid 

mite Diapterobates humeralis were discovered in 

association with HWA in Japan by Dr. Mark 

McClure (retired, CT Agricultural Experiment 

Station). The mite fed on the woolly material 

surrounding the eggs, thereby dislodging the eggs 

from the trees to the ground. However, the mite 

is an opportunistic predator that occurs in many 

areas where the adelgid is absent. Even though it 

was already widely distributed in coniferous forests 

throughout the northern hemisphere, it was released 

in 1993 at two sites in CT but proved diffi cult to 

mass culture and ineffective for HWA. Sasajiscymnus 

tsugae was first reared and released in Connecticut in 

1995 and, beginning in 1999 through 2011, 

over 2.5 million adults have been reared by fi ve 

laboratories and released in 15 states in the East. 

S. tsugae has overwintered, established, reproduced, 

and spread at a limited number of release sites, but 

generally beetle recovery in most areas has been 

disappointing. Although the reason for sporadic 

establishment is unknown, several explanations 

have been put forward including: 1) poor sampling 

methodology involving the use of beat sheets to 

sample for adults in the lower crown while adult 

beetles tend to be phototrophic; 2) the tendency 

for adult beetles to widely disperse following 

release; and 3) the original rearing colony was 

small and a genetic bottleneck may have occurred 

over the 30+ generations of beetles that have 

been lab reared since 1995. S. tsugae was recently 

rediscovered in association with HWA in Japan, 

and mass rearing colonies have been established 

at North Carolina Department of Agriculture 

and University of Tennessee (Lindsay Young 

Beneficial Insect Laboratory) using only this new 

genetic stock. Studies are underway to determine 

if there are any apparent behavioral diff erences 

between the new and original genetic stock. 

In 1995, Dr. Mike Montgomery (retired, U.S. 

Forest Service, Northern Research Station) began 

exploration in China with efforts focused on the 

diversity of lady beetles present in the forests with 

hemlock. Over 50 species were collected from 

222 
 

 

 

 

Chapter 22: An Overview and Outlook for Biological Control of Hemlock Woolly Adelgid 

adelgid-infested hemlock in China, 21 of which 

were new to science. Th ree species of Scymnus 

(camptodromus, sinuanodulus, and ningshanensis) 

were the most abundant lady beetles feeding on 

HWA. Over 30,000 S. sinuanodulus beetles were 

released in 8 states from 2005-2009, with most 

located in the southeastern United States. As of 

2011, establishment of S. sinuanodulus has not been 

confirmed at any of the release sites and rearing 

colonies of this beetle have been greatly reduced. 

Two small experimental releases of S. ningshanensis 

occurred in 2010 in North Carolina but 

establishment has not yet been confi rmed. Rearing 

this species has proven difficult to maintain and the 

small colony has since been abandoned. Th e biology 

and rearing of S. camptodromus is currently being 

evaluated at Penn State University in an eff ort to 

better understand this species, its obligate diapause, 

and its host range. Based on the distribution of S. 

camptodromus in China, it is likely this beetle will be 

more cold tolerant than the other Scymnus species 

investigated and better suited for the northern range 

and higher elevations in the south. Field releases 

of S. camptodromus are anticipated in 2013. 

In the late 1990s, the Pacific Northwest became the 

focus for predators of HWA with Dr. Lee Humble 

(Canadian Forest Service) observing Laricobius 

nigrinus feeding on HWA in seed orchards in British 

Columbia. The predator was imported by Drs. Lok 

Kok and Scott Salom to the quarantine facility at 

Virginia Tech. Laricobius nigrinus was approved 

for release in 2000 and the first releases were made 

in 2003. Additional surveys and collections of 

predators of HWA in Oregon and Washington have 

resulted in the shipment of thousands of L. nigrinus 

(Seattle biotype) for establishing lab colonies at 

New Jersey Department of Agriculture (Phillip 

Alampi Beneficial Insect Laboratory), Virginia 

Tech, University of Tennessee, Clemson University, 

North Georgia College and State University, and 

field releases in the East. Over 150,000 L. nigrinus 

(Seattle biotype) have been released and are now 

becoming widely established in plant hardiness 

zones 6a and 6b. These establishments span 11 states 

from the southern Appalachians to New England 

and have become the first potential biological 

control agent of HWA to establish in such numbers 

that beetles can be field-collected and redistributed 

elsewhere in the East. An inland biotype of L. 

nigrinus, which is more cold hardy than the Seattle 

biotype, has been collected in northern Idaho and 

northwest Montana since 2007 and released at a 

limited number of sites in New England. Small 

numbers of this biotype have since been recovered, 

indicating its adaptability to the more northerly 

climate and colder plant hardiness zones. 

Two species of Leucopis, L. piniperda and 

L. argenticollis, were recovered from HWA 

infested western hemlocks. One of the species, 

L. argenticollis, was recovered feeding on both 

progredientes and sistentes eggs and nymphs in 

fairly large numbers from several collection sites 

in Washington and Oregon. L. piniperda and L. 

argenticollis also occur in eastern North America but 

are rarely recovered from HWA infested hemlock. 

Additional study and monitoring of these species 

in the West is warranted as Leucopis spp. may be 

adaptable to a range of climates because of their 

wide geographic distribution and host specifi city 

toward HWA. Also, several species are among 

the biocontrol agents that have been responsible 

for measurable control of Adelgidae, specifi cally 

Pineus species in Chile and Hawaii. Th e current 

focus is to continue studies of biology and to 

develop rearing methods for L. argenticollis. 

In 2005, an accelerated three year eff ort was 

initiated to explore for natural enemies of HWA in 

Asia, with a focus on China and Japan, followed by 

evaluation and quarantine studies. It was felt that 

the approach to foreign exploration before this time 

was “restrictive and inefficient.” A full-time person, 

Dr. Wenhua Lu, was hired to be in China 3-6 

months each year for two years. During this period 

she worked with Chinese scientists at intensive 

study sites in Sichuan and Yunnan Provinces 

as well as expanded the search for predators in 

additional provinces. Th is effort provided a more 

detailed assessment of HWA phenology and its 

associated natural enemies and the opportunity 

to study the full range of potential predators 

from the pests’ native range. Unfortunately, this 

223 
 

 

 

 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

effort did not result in the discovery of additional 

promising predator species in China. One species, 

Tetraphleps galchanoides, was investigated but 

soon abandoned as the larvae of T. galchanoides 

are also effective predators of L. nigrinus larvae. 

The continued mass rearing and release of the old 

genetic stock of S. tsugae in the southern range has 

not produced many recoveries and only recently 

is being phased out of the overall program with 

initiation of field releases of the new genetic stock 

of S. tsugae. Also, the mass rearing of L. nigrinus has 

often been difficult due to high mortality during the 

pre-pupae and pupae life stages in the soil. Now that 

populations are becoming established in the East, 

field insectaries supplemented by wild collections 

in the Pacific Northwest will become the primary 

source of beetles which will provide the opportunity 

for laboratories to rear other HWA predators. 

In 2005, the discovery by Drs. Montgomery and 

Shiyake (Museum of Natural History, Osaka, Japan) 

of a new species of Laricobius, now osakensis, in 

Japan created a renewed interest in the classical 

biological control program. In 2008-09, Dr. Ashley 

Lamb (University of Tennessee) conducted extended 

studies at several HWA locations in Japan to assess 

the phenology of HWA and its predators, and the 

impact of natural enemies on HWA in its native 

habitat. It was determined that Laricobius osakensis 

was the most abundant and frequently encountered 

predator of HWA in Japan, sharing many 

similarities to L. nigrinus in western North America. 

In laboratory and field cage studies, L. osakensis has 

shown to be more robust in consumption of HWA 

and in producing a greater number of progeny 

than L. nigrinus. L. osakensis comes from the same 

location as the source population of HWA in the 

eastern U.S.; therefore, this species has evolved 

with the HWA aff ecting T. canadensis. Because 

of the geographically broad range this predator 

is found in Japan (elevations of 80-1850m), it is 

likely it will adapt well across the varied climate 

range in the United States. In 2010, this species 

was approved for release from quarantine. 

The exploration for additional HWA predators 

in western North America continued into the 

early 2010s, but at a much reduced level. Only 

the previously mentioned Leucopis species and the 

coccinellid predator S. coniferarum warrant further 

investigations. S. coniferarum is another adelgid 

specialist often found throughout the year associated 

with HWA but typically in lower numbers than 

L. nigrinus. It readily feeds on both generations 

of HWA but is believed to be a primary predator 

of the spring progredientes generation. Host 

range testing and rearing methodologies for this 

predator are currently under investigation. Field 

releases of S. coniferarum are anticipated in 2012. 

As exploratory efforts wind down, more emphasis 

will go toward the predators currently under 

review by improving the effi ciency of collecting, 

rearing, and releasing the two biotypes of L. 

nigrinus, L. osakensis and S. tsugae. Maximizing 

the production of quality HWA predators requires 

highly skilled laboratory managers, adequate 

facilities and staffing, and an abundance of healthy 

host material (HWA) for food. The mass rearing 

of HWA predators by numerous laboratories with 

the primary goal of maximizing production over 

the shortest time has caused concern about the 

quality of the predators being reared and released. 

Drs. Allen Cohen (Insect Diet and Rearing 

Research LLC, North Carolina) and Carole Cheah 

(Connecticut Agricultural Experiment Station) are 

developing a fitness-based system of quality control 

standards for rearing predators (using L. nigrinus 

and S. tsugae) among the rearing laboratories. 

Efforts to maximize mass rearing of predators 

based on a supply of sufficient quantities of quality 

host material ((HWA) has been problematic 

for all of the rearing laboratories. The impact of 

declining tree health on the nutritional quality 

of HWA as a food source has forced personnel 

at the labs to greatly expand their search to feed 

the laboratory reared beetles. Th e development 

and testing of artificial diet supplements by Drs. 

Cohen and Cheah has helped to mitigate the issue 

but more work in this area is needed. Th e most 

224 
 

 

 

Chapter 22: An Overview and Outlook for Biological Control of Hemlock Woolly Adelgid 

successful diets and diet-presentation systems 

allowed adults of both S. tsugae and L. nigrinus 

to survive for several months, but no oviposition 

occurs in the absence of live host material. When 

provided HWA for a few days following feeding 

experiments on the artificial diet, egg production 

returns. L. nigrinus larvae will feed readily on the 

chicken egg-based diets but fail to develop. 

Maintaining healthy laboratory-reared predators 

is a tremendous challenge that can be easily 

compromised if pathogens are inadvertently 

introduced to the colonies. Th e discovery by 

Dr. Lee Solter (Illinois Natural History Survey, 

Champaign, IL) of microsporidia infections in 

several laboratory and field collected populations 

of predators has resulted in heightened monitoring 

and periodic screening of predators. Microsporidia 

are single-cell organisms related to fungi and are 

obligate pathogens, typically chronic in nature, 

causing slow larval development, increased larval 

mortality, decreased adult lifespan, and reduced 

fecundity. Entire rearing colonies can be easily lost 

if adequate screening for microsporidia infections is 

not addressed. As of 2011, there are 8 laboratories 

mass rearing HWA predators including: Clemson 

University, New Jersey Department of Agriculture, 

North Carolina Department of Agriculture and 

Consumer Services, North Georgia College and 

State University, University of Georgia, University 

of Tennessee, Virginia Tech, and Young Harris 

College in North Georgia. The four predators 

currently being reared for mass release include: 

L. nigrinus (Seattle and Idaho biotypes), L. 

osakensis, S. camptodromus, and S. tsugae. 

Investigations into potential entomopathogens have 

been ongoing since the mid-2000s. Dr. Bruce Parker 

(University of Vermont) conducted one survey in 

China and several surveys in the eastern U.S. for 

entomopathogens of HWA. Even though some 

non-host specific, native fungi have been found 

to infect HWA, naturally occurring epizootics of 

primary pathogens have seldom been observed. 

One particular fungus, Verticillium lecanii (now 

Lecanicillium muscarium), was recovered from 

HWA populations in the East and is currently 

being investigated by Dr. Scott Costa (University 

of Vermont). L. muscarium strain Ve6 is registered 

and commercially available in Europe to control 

whitefly pests in greenhouses. Laboratory, ground, 

and aerial application trials to document its efficacy 

on HWA have thus far been promising but the rapid 

degradation of the fungus once applied remains 

a challenge. Entomopathogenic fungi in general 

tend to be highly sensitive to photo-degradation 

and low humidity conditions. Th e commercially 

available formulation of the fungus Mycotal® 

(Koppert Biological Systems, a Netherlands-based 

company) along with MycoMax, a whey bi-product, 

has been added to the tank mix in an eff ort to 

stimulate fungal growth and spore production 

in the fi eld. This formulation has been aerially 

applied to small replicated plots in Tennessee. 

Preliminary efficacy results are promising, although 

larger scale pilot tests are still needed (planned 

for 2012). Meanwhile, registration of Mycotal® in 

the United States is being pursued by Koppert. 

The classical biological control program has 

encountered several “bumps-in-the-road” along 

with the many successes. A major deterrent from 

the onset was our inability to import predators from 

China into U.S. quarantine facilities in a timely 

manner to prevent large numbers of predators from 

dying in transit or while waiting to be shipped. In 

spite of attempts to resolve some of the specifi cally 

identified issues, this appears to be a major problem 

common to many U.S.-supported biological 

control foreign exploration efforts in China. 

The recent utilization of molecular genetics by 

Dr. Nathan Havill (U.S. Forest Service, Northern 

Research Station) revealed that HWA in the eastern 

U.S. originated from southern Japan, and that 

HWA found in western North America is genetically 

unique from other HWA found worldwide, 

indicating that this biotype is likely native to this 

region. DNA sequencing technology and the use 

of molecular markers such as DNA barcodes and 

microsatellites helped characterize genetic variation 

within species of HWA predators as well as identify 

potential issues involving closely related predator 

species that would have otherwise been diffi cult to 

225 
 

Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

realize. Th e first example of this involved the release 

of L. nigrinus (native to western North America) 

in the East, which was later found by Dr. Gina 

Davis (Virginia Tech) to have hybridized in some 

areas with a closely related species, L. rubidus. L. 

rubidus is a native predator of the pine bark adelgid 

Pineus strobi, an adelgid that attacks eastern white 

pine, Pinus strobus. L. rubidus had been reported in 

association with HWA, but were typically few in 

numbers and only in areas where white pine and 

hemlock were in close proximity. Th e outcome of 

this hybridization event is currently being studied 

in the laboratory and at L. nigrinus release sites. A 

second example of how this technology is being 

used is that it can distinguish the immature stages 

of L. nigrinus from that of L. rubidus, which are 

otherwise indistinguishable morphologically. Prior 

to implementing this technology, larvae needed to 

be reared to adults for identifi cation. This is labor 

intensive, space consuming, and less reliable than 

molecular diagnostics. Finally, the third example 

of incorporating the science of molecular genetics 

into the HWA biological control program involves 

L. osakensis. In this case, a previously undescribed 

species of Laricobius (now L. naganoensis) was 

found contaminating the rearing colony of L. 

osakensis prior to it being released. L. naganoensis 

is separated morphologically as a diff erent species 

primarily by differences in the male genitalia, but 

it is genetically distinct. It was DNA sequencing 

of the parent population of L. osakensis used 

to establish the rearing colony that led to this 

discovery, and this technology will be used to 

help “purify” the rearing colony in the future. 

Assessment of the establishment, spread, and 

effectiveness of the predators released in the East 

is a long-term process with a constant need for 

refinement that begins by locating “ideal” sites 

and optimizing the timing of releases for specifi c 

predators. Unfortunately, the persistent challenge 

in predator field sampling efforts continues to be 

the use of the beat sampling technique, which only 

samples accessible lower canopy foliage of hemlock. 

We suspect the use of this technique might result in 

misinterpretation of the abundance of several species 

of predators, and other means of sampling for 

establishment should continue to be explored. For 

example, shifting from sampling for adult L. nigrinus 

predators to sampling for immature predator life 

stages is generally more labor intensive as it also 

requires laboratory processing, but it has been 

shown to be a more sensitive measure for detection 

of smaller predator populations. Assessment of 

predator impact on HWA and protecting tree 

health at the stand level, however, is even more of a 

challenge. In plant hardiness zones 6a and warmer, 

tree decline often occurs within a few years, and the 

small number of predators released in these areas 

has little chance to populate sufficiently to prevent 

this decline. At best, we will only be able to assess 

tree health recovery in areas where the ratio of 

predator to prey populations begin to balance out. 

There have been numerous field evaluations of 

predator effectiveness as well as competition among 

predators using mesh cages over portions of hemlock 

branches containing various densities of HWA 

and predators. Whole tree canopy enclosures have 

been used by Dr. Jerome Grant et al. (University 

of Tennessee) to enhance understanding of the 

survival, colonization, and establishment of 

predators and to assess the impact of these agents on 

population-densities of HWA and on tree health. 

Prior to 2007, data pertaining to the release, 

monitoring, and recovery of predators were 

maintained on paper data forms or in small local 

databases and, as a result, were inaccessible to 

HWA scientists and managers at regional and 

national levels. In 2007, the HWA Predator Release 

and Recovery Database (PDB) was initiated and 

is to include all historic release and monitoring 

information, as well as to provide a mechanism 

for field personnel to enter and update current 

and future records. The PDB uses standardized 

field protocols and data forms. It will facilitate 

improved access to the data, provide project-wide 

reports and maps, and is a tool for analysis and 

improved decision-making for future actions. 

View-only public access to the PDB is available 

at: http://hwa.ento.vt.edu/hwa/hwa.cgi. 

226 
 

Chapter 22: An Overview and Outlook for Biological Control of Hemlock Woolly Adelgid 

In 2010, Dr. Richard McDonald (Symbiont 

Biological Pest Management) began a 2-year eff ort 

to monitor predators released and assess their ability 

to establish, disperse, and impact HWA across 

the pest’s geographic range at a selected number 

of sites. Only 3+ year old release sites are being 

sampled to confirm establishment and the data will 

be entered into the HWA Predator Database. Th us 

far, at least 5 L. nigrinus release sites in NC, NJ, 

and PA have been identified as having sufficient 

predator densities that allow for collection and 

redistribution of beetles to other HWA infested 

areas. These are also some of the oldest release sites 

that were established between 2004 and 2006. 

The guiding principles for developing a classical 

biological control program have been followed for 

HWA, especially with the release of climatically 

matched species or biotypes, including those 

adapted to local temperature conditions. 

Collections overseas were made not only in forests 

but also in ornamental sites and on susceptible 

exotic hosts, where factors that aff ect herbivore 

population dynamics more closely resemble 

those that characterize introduced habitats (e.g. 

eastern United States). Even though we are still 

collecting data on the ecological requirements 

and performance following introduction, this 

complex of predators has not yet provided a 

detectable regulation of HWA populations and 

corresponding protection of tree health. We 

suspect that, as our sampling techniques improve 

and the predators have additional time to increase 

in density and spread, their role in suppressing 

HWA population will become more apparent. 

OUTLOOK 

High mortality of hemlocks in the eastern U.S. 

has been attributed to a combination of host tree 

susceptibility and lack of effective natural enemies. 

Therefore, the dynamics of adelgid populations 

in the eastern U.S. is believed to be driven mainly 

by weather (cold winter temperatures) and the 

negative density-dependent consequences of host 

deterioration on adelgid survival. Signifi cant eff ort 

has focused on classical biological control in an 

effort to alleviate the lack of natural enemies. Th e 

establishment of these additional predators in 

the northern part of the HWA range, along with 

periodic HWA density reductions due to cold 

temperatures, may provide the additional mortality 

necessary to effectively suppress populations of 

HWA below damaging levels. In the absence of 

these regulating cold winter temperatures in the 

southern part of the HWA range, this is more 

problematic. The intolerance of eastern hemlock 

species to attack by HWA may seriously constrain 

our biocontrol efforts, because exceptionally 

high mortality from natural enemies may be 

needed to maintain HWA at innocuous levels. 

We have realized that natural enemies that 

are effective control agents in native natural 

habitats (where their hosts typically occur at 

low, innocuous densities) may have limited 

ability for pest populations that outbreak in 

introduced habitats. Natural enemies collected 

and evaluated for HWA biocontrol originated 

from trees growing on as well as off their preferred 

growing sites, which often renders them less 

resistant to insect herbivores, presumably due to 

stress from less adequate growing conditions. 

Successful biological control of HWA in the 

eastern U.S. will require a suite of predators that 

attack all of the life stages of HWA. Th e rearing 

of predators in laboratories should continue, 

along with the release of predators into numerous 

geographical areas to establish and promote their 

natural spread. The number of fi eld insectaries 

should be expanded in natural or planted settings 

in an effort to build up predator populations 

for harvesting and redistribution. Th ese “wild” 

individuals are more adapted to climate and 

other local variables and would supplement the 

release of lab-reared individuals. Field insectaries 

resulting from several of our oldest release sites 

are already providing thousands of L. nigrinus 

beetles for release in other infested areas. 

227 
Implementation and Status of Biological Control of the Hemlock Woolly Adelgid 

The classical biological control program needs to 

continue, but, as we understand more about the 

ecological requirements of these predators, we need 

to shift toward manipulation of these predators 

and integration into a management program. Even 

though biocontrol needs to be the focal tactic in an 

integrated program, no single control method will 

enable managers to meet their HWA management 

objectives in all environments. Early detection 

surveys conducted for HWA well in advance of 

symptoms of tree decline is critical if damage is 

to be prevented. Likewise, continued monitoring 

of pest densities throughout the management 

effort is recommended. A combination of control 

methods consistently applied through time will 

be necessary to obtain management objectives for 

HWA, especially as HWA expands its range. 

The integration of chemical and biological control is 

being conducted on a landscape scale and evaluated 

on smaller stands of hemlock. The general strategy 

is to maintain the health of a select number of large 

hemlocks with insecticide applications, and at the 

same time release and allow the predators to become 

established on understory and other non-treated 

trees. The idea is that by protecting the health of and 

reducing HWA population densities on the larger 

higher value hemlock trees, predators will have more 

time to populate suffi ciently to off er long-term 

suppression of HWA. These integrated eff orts are 

urgent for management of HWA in the southern 

United States as hemlocks are dying at a rapid rate 

and an integrated management program is essential. 

228 
C

SPECIES INDEX
 

A

Abies fraseri 117, 168 

Abies grandis 101 

Adelges abietis 81, 82, 95 

Adelges cooleyi 43, 66, 67, 81 

Adelges lariciatus 81 

Adelges laricis 43, 66, 67 

Adelges piceae 9, 11, 32, 40, 43, 54, 61, 73, 74, 81, 82, 

87, 90, 96, 98-105, 117, 213, 218, 220 

Adelges tsugae 3-9, 11, 13-15, 23-25, 38, 39, 43, 45-47, 

51, 53, 54, 58, 67, 74-77, 82, 87-90, 92-94, 99-102, 

105, 106, 115, 116, 120, 132, 148, 156, 157, 161, 

165-168, 176, 201, 205, 213, 219, 221 

Alnus serrulata 43, 117 

Amblynotus longitarsus 101 

Anthocoris antevolens 117 

Anthocoris nemoralis 117, 204 

Aphidencyrtus aphidivorus 101 

Aphis gossypii 66, 67 

B

Bacillus thuringiensis 107 

Beauveria bassiana 108-110, 113, 115, 135 

Chionaspis pinifoliae 81, 82, 95 

Chrysoperla carnea 217 

Chrysoperla nipponensis 217 

Cinara pilicornis 81, 82 

Cinara pinea 66, 67 

Coleomegilla maculata 150, 156 

Cremifania nigrocellulata 98, 99, 102, 103 

D

Daecocororis nubilus 117 

Daecocororis piceicola 117 

Daecocororis pinicola 117 

Daktulosphaeria vitifoliae 98 

Dendrocerus carpenteri 101 

Diadegma fenestrale 217 

Diadegma semiclausum 207, 217 

Diapterobates humeralis 29, 39, 222 

Diuraphis noxia 98, 103, 105 

E

Entomophaga miamaiga 109 

Ephestia kuehniella 149, 152 

Eucallipterus tiliae 66, 67, 120 

F

Feniseca tarquinius 66 

Fiorina externa 95 

Fiorinia externa 23, 48, 65, 67 

H

Harmonia axyridis 66, 82, 156 

L

Lambdina fiscellaria 9 

Laricobius erichsonii 78, 81, 90, 212 

Laricobius kangdingensis 91, 118, 212 

Laricobius laticollis 78, 212 

Laricobius naganoensis 226 

Laricobius nigrinus 2, 7, 8, 10, 13-15, 18, 20, 21-23, 29, 

32, 35, 37-40, 77-92, 94, 96, 99-102, 106, 117, 

119, 120, 125-129, 131, 132, 134, 135, 139-143, 

145, 148-151, 153, 154, 157, 161-164, 166-177, 

181, 183, 185, 195-201, 205-207, 210, 212-219, 

221, 223-227 

Laricobius osakensis 7, 10, 29, 36-38, 90-96, 117, 118, 

125, 129, 212, 224-226 

Laricobius rubidus 8, 35, 37, 38, 40, 78, 84, 87, 90-92, 

94, 167, 196, 197, 206, 212-218, 221, 226 

Laricobius taiwanensis 91 

Larix deciduas 43 

Lecanicillium muscarium 110-115, 225 

Leucopis argenticollis 99, 100, 104, 223 

Leucopis atrifacies 99, 100 

Leucopis hennigrata 98 

Leucopis obscura (see Neoleucopis obscura) 

Leucopis piniperda 99, 100, 223 

Leucopis verticalis 98, 105 

Lygocerus testaceimanus 101 

M

Melanips iowensis 101 

Metarhizium anisopliae 110, 115 

Myzus persicae 81, 82 

229 
N

Neoleucopis ancilla 101 

Neoleucopis atratula 98 

Neoleucopis manii 99 

Neoleucopis nigraluna 99 

Neoleucopis obscura 98, 99, 101, 102, 106 

Neoleucopis pinicola 101, 102 

Neoleucopis tapiae 99, 101 

Nosema bombycis 137 

O

Oenopia signatella 59 

P

Pachyneuron altiscutum 101 

Pachyneuron virginicum 101 

Paecilomyces farinosus 108 

Papilio glaucus 204, 208, 209 

Paraprociphilus tessellatus 43, 66, 95. 117 

Phylloxera vitifolia (see Daktulosphaeria vitifoliae) 

Phytoseiulus persimilis 204, 210 

Picea brachytyla 7 

Picea likiangensis 7, 58 

Picea pungens 168 

Picea rubens 117, 168 

Picea torano 7 

Pineus armandicola 56 

Pineus boerneri 99 

Pineus coloradensis 101, 120 

Pineus floccus 120 

Pineus pini 54, 98, 99, 106, 116 

Pineus similis 81 

Pineus strobi 43, 54, 66, 67, 81, 82, 84, 90, 98, 117, 213, 

218, 220, 226 

Pinus armandii 56, 58 

Pinus mugo 168 

Pinus patula 99 

Pinus radiata 99 

Pinus strobus 43, 68, 117, 168, 195, 226 

Polistes dominula 205 

Prociphilus tessellatus 67, 120 

Pseudoscymnus tsugae (see Sasajiscymnus tsugae) 

Pseudotsuga menziesii 43 

S

Sasajiscymnus tsugae 2, 18, 29, 40, 43-52, 54, 68, 71, 

82, 95, 102, 110, 125-127, 129-131, 134-136, 139, 

140, 142, 143, 145, 146, 148-150, 152-156, 161­

166, 176, 181, 185, 201, 205-207, 222, 224, 225 

Scymnus camptodromus 29, 54-56, 59-66, 68, 71, 72, 74, 

119, 206, 223, 225 

Scymnus coniferarum 54, 75, 118-121, 125, 127, 131, 

135, 205, 206, 224 

Scymnus geminus 58 

Scymnus impexus 54, 61, 63, 118 

Scymnus ningshanensis 29, 54-56, 59-61, 63-68, 70-73, 

75, 223 

Scymnus sinuanodulus 29, 54-56, 58-73, 102, 125-127, 

131, 135, 161, 176, 181, 223 

Scymnus suturalis 54, 66-68, 75, 118 

Scymnus yunshanpingensis 58 

Sitotroga cerealella 149, 152, 156 

Syrphophagus aphidivorus 101 

T

Tetraphleps abdulghani 116 

Tetraphleps galchanoides 59, 60, 116, 117, 120, 121, 224 

Tetraphleps raoi 116 

Tinthia myrmosaeformis 202, 209 

Torymus beneficus 217, 220, 221 

Torymus sinensis 217, 220, 221 

Tsuga canadensis 3-5, 9-11, 13, 14, 68, 90, 117, 118, 148, 

161, 176, 218, 220, 224 

Tsuga caroliniana 3-5, 9, 10, 176 

Tsuga chinensis 4, 5, 7, 11, 54, 116, 118 

Tsuga diversifolia 4, 7, 43, 92, 94 

Tsuga dumosa 54, 56 

Tsuga formosana 4 

Tsuga heterophylla 7, 77, 99 

Tsuga mertensiana 7, 9 

Tsuga sieboldii 4, 7, 10, 43, 47, 92-94 

V

Verticillium lecanii 108, 110, 225 

230