United States Department of Agriculture

M arketing and Regulatory Programs

A nimal and

Plant Health Inspection
Service 

# Field Release of **Aphelinus**
**rhamni** ( Hymen o ptera: Aphelinidae ) for B iological
Control of the Soybean Aphid, **Aphis glycines** ( Hemiptera: Aph ididae ), in
the Continental United
States

E nvironmental A ssessment

, Ju ly 2014

# Field Releas e of **Aphelinus rhamni**  (Hymenoptera: Aphelinidae) for Biological Control of the Soybean Aphid, **Aphis glycines** (Hemiptera: Aphididae), in the Continental United
State s

Environmental Assessment,

June 2014

Agency Contact : Shirley Wager

- Page, Ph.D.

Pest Permitting
Plant Protection and Quarantine Animal and Plant Health Inspection Service U.S. Department of Agriculture 4700 River Road, Unit 133 Riverdale, MD 20737

– 1236

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## Table of Contents

I. Purpose and Need for the Proposed Action

............................ 1

II. Alternatives

................................ ................................ ................ 2

III. Affected Environment

................................ .............................. 5

IV. En vironmental Consequences

................................ ............... 7

V. Other Issues

................................ ................................ ............ 1 2

VI. Agencies, Organizations, and Individuals Consulted

........ 1 3

VII. R eferences

................................ ................................ ............ 1 5

Appendix A

................................ ................................ .................. 2 3

Appendix B

................................ ................................ .................. 2 5

## I. Purpose and Need for the Proposed

Action

The U.S. Department of Agriculture (USDA), Animal and Plant Health Inspection Service (APHIS) , Plant Protection and Quarantine (PPQ) , P est Permitting Branch is proposing to iss ue permit s for release of the insect Aphelinus rhamni ( Hymenoptera : Aphelin idae )

. This organism would be

used by the appli cant for biological control of soybean aphid

, Aphis glycines ( Hemiptera: Aphididae )

, in the c ontinental United States . Before issuing permit s for release of A phelinus rhamni , APHIS

– PPQ needs to analyze the potential impacts of the release of this organism into the

continental United States

.

This en vironmental assessment 1 (EA) was prepared to be c onsistent with

USDA

– APHIS' National Environmental Policy Act of 1969 (NEPA) implementing procedures (Title 7 of the Code of Federal Regulations (CFR), part 372). It examines the potential effects on the quality of the human environment that may be associat ed with the release of the parasit oid wasp
, A. rhamni

, to control soybean aphid in the continental

United States

. A parasitoid is a n insect whose immature stages (larvae and pupae) live as parasites that eventually kill their hosts (typically other

insects )

. This EA considers a “no action” alternative and the potential effects of the proposed action

.

The applicant’s purpose for releasing A. rhamni is to reduce the severity of damage to soybean ( Glycine max ) from infestations of soybean aphid in

the contine ntal United State s

. In 2000

, t he soybean aphid ( native to Asia )

, was found in North America

, and is now a major soybean pest infesting 42

million acres in 2003 which result ed in decreased soybean yields and greatly increased control costs (Ragsdale et al., 2011; Landis et al., 2008; Song and Swinton, 2009) . T he soybean aphid invaded most soybean production regions in North America, including 21 states and three Canadian provinces since originally reported in North A merica in 2000 (Ragsdale et al., 2004)

.

The applicant identified several purposes for releasing this particular insect species for biological control of soybean aph id. F irst, impacts to

non

- target species are expected to be low. Second, this parasitic wasp may exhi bit utility as a biological control agent on the soybean aphid’s alternate buckthorn host, R hamnus species

. Lastly

, the distribution of A.

rhamni in the United States may differ from the expected distribution of A. glycinis , a related biological control ag ent of soybean aphid recently

1 Regulations implementing the National Environmental Policy Act of 1969 (42
United States Code 4321 et seq. ) provide that an enviro nmental assessment “[shall include brief discussions of the need for the proposal, of alternatives as required by section 102(2)(E), of the

e nvironmental impacts of the proposed action and alternatives, and a listing of agencies and persons

consulted” ( 40 CFR § 1508.9 )

. 1approved for release in the United States . Deployment of both insect species has the potential to provide enhanced control of soybean aphid

over a broader area in the United States than A. glycinis alone

. These parasitic wasps are insect s whose immature stages (larvae and pupae) live as parasites that eventually kill their hosts (typically other insects).

Most of the e xisting soybean aphid management options (discussed below) are expensive, temporary, and/ or include non

- target impacts. For

these reasons, there is a need to iden tify and release a complement of effective, host

- specific biological control organism s against soybean aphid

in the continental United States

.



## II . Alternatives

This section will explain the two alte rnatives available to APHIS

– PPQ : no action (no issuance of permits) and issuance of permit s for environmental

release of A. rhamni in the continental United States

. Although APHIS ’

alternatives are limited to a decision o f whether to issue permit s for rele ase of A. rhamni

, we describe other methods currently used to control soybean aphid by soybean producers in the United States

. Use of t hese control

methods is not an APHIS decision

, and their use is li kely to continue

whether or not APHIS

- PPQ issued permit s for environmental release of A.

rhamni

.

APHIS

– PPQ considered a third alternative but will not analyze it further. Under this third alternative, APHIS

– PPQ would issue permits for the field

release of A. rhamni . The permits, however, would contain speci al provisions or requirements concerning release procedures or mitigating

measures , such as limited release s of A. rhamni in the United States

. There

are no issues raised i ndicat ing that special provisions or requirements are

necessary.



### A. No A ction

Un der the no action alternative, APHIS

– PPQ would not issue permit s for

the field release of A. rhamni for the control of soybean aphid

— t he

release of this biological control agent would not occur , and current

methods t o control soybean aphid in the United States will continue

. Use

of t hese methods is likely to continue even if APHIS

– PPQ issues permit s for release of A. rhamni

. Presently, control of soybean aphid in the United

States is limited to chemical control

, plant resistance , and biological

control me thods

.

Insecticide treatments are the most effective method to manage soybean aphids in North America. Foliar sprays of insecticides provide temporary suppression (7 to 14 days) of soybean aphid populations. I nsecticides 1. Chemical
C ontrol

2including esfenvalerate , dimetho ate, chlorpyrifos

, zeta

- cypermethrin

, methyl parathion

, permethrin

, and lambda cyhalothrin are or have been labeled for foliar application against soybean aphid (Ostlie, 2002 ). Soybean s eed treatments using the neonicotinoid insecticide s imidacloprid

and thiamethoxam for systemic control of soybean aphids also can be used (Pedersen and Lang, 2006 ; Magalhaes et al., 2009 )

.

Aphid

- resistant soybean plants were bred to support reduce d aphid populations. Host plant resistance works by inhibiting aphid growth and development and increasing plant tolerance to aphid feeding.

Aphelinus certus is a parasitoid native to Asia

, accidentally introduced i nto
the United States when the soybean aphid invaded (Heimpel et al., 2010)

. A

. certus has a broad host range (Heimpel et al., 2010; Frewin et al., 2010) , and was not considered for purposeful introduction against soybean aphid

. In the long term, introduction of another parasitoid is unlikely to

affect the abundance of A. ce rtus because these insects coexist in Asia.

Another Asian parasitoid, Binodoxys communis (Hymenoptera: Braconidae)

, was released in the United States beginning in 2007; unfortunately, it did not establish in the environment (Ragsdale et al., 2011). In 2 002, a previously introduced strain of the European parasitoid Aphelinus atriplicis was released against soybean aphid

. This strain was first released in the w estern United States against the Russian wheat aphid (Ho pper et al., 1998; Prokrym, 1998; Heraty et al., 2007)

, but it also

at tack s the soybean aphid (Wu et al., 2004) . Although this strain of Aphelinus atriplicis was released in nine Minnesota sites in 2002 (Heimpel et al., 2004) , it was not recovered since th at time.

Most recently

, APHIS

– PPQ approved the parasitoid Aphelinus glycinis for

environmental release into the United States ( Hopper, 2010; USDA

APHIS, 2012). This related species originated from X iyuan, Liaoning Province, Peoples Republic of China . The climate in this area closely

matches the climate in the northern Midwest United States where soybean aphid is a major pest. It is expect ed that A. glycinis will be come limited to this region , but may extend its range into Ontario and Quebec provinces in

Canada

. E stablishment is expected to occur primarily in soybean fields infested with soybean aphid during the growing season

.



### B

. Issue P ermit s for E nvironmental R elease of

**A. r**  **hamni**

**.**

Under this alternative, APHIS

– PPQ would issue permit s upon request and after evaluation of each application for the field release of A. rhamni for

the control of soybean aphid in the continental United States . Th ese permit s would contain no special provisions or requirements concerning release procedures or mitigating measures

. 3. Biologica l
Control 2. Host Plant
Resistance

3Insect Taxonomy

Order: Hymenoptera Family: Aphelini dae

Genus: Aphelinus Species: rhamni Woolley and Hopper

Common name : none

The species description of A. rh amni was published in 2012 (Hopper et al., 2012)

. Voucher specimens of A. rhamni are held at three locations. These include (1) the Beneficial Insect Introductions Research Unit, Agricultural

Research Service (ARS) , United States Department o f Agriculture, Newark, Delaware , (2) Texas A & M Unive rsity, Department of Entomology , and (3) the Smithsonian in the Systematic Entomology Laboratory, USDA

− ARS.

Aphelinus sp ecies are tiny (about 1 millimeter (mm) in length) , stingless parasitic wasps. T he immature st ages develop as internal parasit oids o f arthropods where , in this case, feeding of the wasp larva inside the host aphid eventually kills the host aphid

. Many species of Aphelinus have demonstrated their importance in the biological control of aphids ( Hemip tera: Aphididae).

Aphelinus rhamni females prefer second to fourth instar (an immature developmental stage) aphid hosts for oviposition (egg laying) , but will lay

eggs in all stages, including winged adults. Females use an egg laying organ called an ovip ositor to insert an egg into the aphid. The inserted egg hatches and the wasp larva feeds on the internal organs of the aphid host.

At 20 o C, the wasp s develop from egg to pupa in about 14 days; third instar

wasps kill their hosts leaving intact the aphid h ost external skeleton ( exoskeleton) hardened and black, in a process called mummification.

Adult A. rhamni wasps emerge from the mummified aphid about one week after pupation by chewing a hole through the host exoskeleton to escape. Adult wasps eat plant n ectar and honeydew (a sugary liquid secreted by aphids and other sap

- feed ing insects) . A dult females also feed on aphids to obtain nutrients for egg production by piercing or “stinging” the aphids with their ovipositor and drinking the hemolymph (insect bl ood) from the

wound , killing about two aphids per day by such feeding. Females emerge

as adults with five to 15 mature eggs, but produce more eggs throughout their lives, maturing about 10 eggs per day. In the laboratory, a dult wasps

live for two to three weeks, but they are unlikely to survive this long in the

field (Hopper, 2011)

.

Aphelinus species are weak fliers, searching primarily by walking (Fauvergue et al. , 1995; Mason and Hopper, 1997)

. Another related

pa rasitoid

, Aphelinus asychis, dispersed an average of 13 feet in one generation and 26 feet in three generations in fields with abundant hosts 1. **Aphelinus**
**rhamni**
Taxonomic

Information

2. Description

and Biology of **Aphelinis**
**rhamni**

4(Fauvergue and Hopper, 2009)

. Insects would have to disperse much further at the end of the growing season to track their hosts.

Aphelinus rhamni was collected from parasitized soybean aphid s on Rhamnus species in 2005 near Daxing ( Beijing, Hebei Province ) Peoples Republic of China

, and is not known to occur elsewhere (Hopper et al., 2012)

. The present distribution of soybean aphid in the United States matched fairly well with a climate

- based expected distribution of A.

rhamni (Dyme x Simulator Application software, version 2.01.025, (c) CSIRO, 2004). It is expect ed that A. rhamni may not establish in the full southern range of the soybean aphid because this pest is found further south than the climate match for Beijing. A phelinus rha mni is expected to

establish primarily in the northern half of the United States east of the Rocky Mountains (Hopper, 2011)

.

In the laboratory, an individual female A. rhamni wasp parasitiz es or kills by host feeding over 20 0 aphids during a lifetime o f three weeks, but long

lifetimes in the field are unlikely

. If females lived one week in t he field

, each is expected to p arasitize and host feed on 70 aphids, if aphids are sufficiently abundant (Hopper, 2011)

.



## III

. Affected Environment

A. Soybean Ap hid

The soybean aphid is a host alternating species, which means it must disperse between two different types of host plant s between summer and winter. Females reproduce asexually on soybean from June to September, producing winged and wingless forms . In September, winged mal es and parthenogenetic (a form of reproduction in which an unfertilized egg develops into a new individual) females migrate to their primary host plants, Rhamnus sp ecies , and the migratory females bear sexual, flightless females, which mate with the migratory males and lay overwintering eggs.

In the s pring, parthenogenetic females hatch from these eggs, and several generations of females reproduce asexually on Rhamnus species before winged aphids move to soybean plants

. Although soybean aphids are weak fliers, the rapid rate of spread of soybean aphid indicates a high rate of windborn dispersal. In the laboratory, soybean aphid has a high rate of population increase at 25 o C, with a steep decline to a negative rate when temperatures are h igher than 30 o C (McCornack et al., 2004) . This can explain why it has not moved further south in the U nited S tates where

temperatures are warmer

. At 25 o C, females can produce 73 offspring during their lifetime, with a generation time of 10 days. T he combi nation of high dispersal rate and high reproductive rate mean that soybean aphid can rapidly reach damaging levels over wide regions, if not properly managed (Hopper, 2011)

. 3

. Geographic

R ange of **Aphelinis**

**rhamni**

4 . Impact of **Aphelinus**
**rhamni**

on Soybean Aphid

5

### B . Areas A ffected by Soybean Aphid

The soybean aphid is native to E ast Asia, found in eastern Russia, China, Japan, and Southeast Asia (Vietnam, Thailand, Indonesia, Malaysia, and the Philippines). There are reports in Kenya, and it invaded Australia in

2000.

In North America, it is currently known from Ontario and Quebec in Canada, and from the following 2 9 U.S. states: Arkansas, Connecticut, Delaware, Georgia, Indiana, Iowa, Illinois, Kansas, Kentucky, Louisiana, Maine, Maryland, Massachusetts, Michigan, Minnesota, Mississippi,

Missour i, Nebraska, New Jersey, New York, North Dakota, Ohio, Oklahoma, Pennsylvania, South Dakota, Tennessee, Virginia, West Virginia, and Wisconsin (Ragsdale et al. , 20 11 ).

Soybean ( Glycine max ) is the summer host of soybean aphid while various buckthorn speci es ( Rhamnus sp ecies ) are the winter hosts. S everal native and a few introduced species of Rhamnus occur in the United States

.



### C

. Insects R elated to Soybean Aphid and **A. rhamni** 
in the United States

Information regarding insects taxonomically related t o soybean aphid is

included because closely related insect species have the greatest potential

for attack by A. rhamni

.

Of the 497 species of Aphis w ith distributions listed in Blackman and Eastop (2006), 119 are r eported from North America. Of these sp ecies found in North America, 33 are widely distributed throughout the world, and many of these are invasive exotic pests. Only 86 species of Aphis were described from collections in North America

, and consequently may be

considered native species. Fortuna tely, most native Aphis species are not closely related to soybean aphid ( Aphis glycines )

. The soybean aphid is closely related to the cotton aphid, Aphis gossypii , which is also native to Asia and a major agricultural pest, and to the buckthorn

- potato aph id, Aphis nasturtii , which shares its overwintering host ( Rhamnus sp ecies ). Aphis nasturtii is a pest of potatoes in North America , and believed to be

native to Asia (Hopper, 2011)

.

Thirty

- one species of Aphelinus are reported from North America

, howev er
, only about half of these species (14) were described from North America (listed in Appendix A with their authors and species complex membership, as well as reported distributions and hosts)

. These 14 Aphelinus species are reported exclusively or almost exclusively from North America, and likely are native.

The remaining Aphelinus species reported from North America were 2. Present

D istribution in

North America 1. Native and
Worldwide

Distribution

3

. Soybean Aphid

H osts

1

. Insects

Related

to Soybean Aphid

2

. I nsects
Related to **Aphelinus**

**rhamni**

6described elsewhere, are reported from one to several other continents, and therefore are unlikely to be native to North America. Se veral have been introduced for biological control of pest aphids (Hopper et al., 1998; Jackson et al., 197 1 ; van den Bosch et al., 1959)

. Three native Aphelinus species were reported from Aphis species, A. marlatti

, A. nigritis , and A. semiflavus , but only A. semiflavus parasitizes a species closely related to the soybean aphid, being reported from the cotton aphid in the M idwestern

United States

. T he native parasitoid species a ppear unlikely to switch to

attack the soybean aphid, because these types of switches have not been

observed (Lin and Ives, 2003; Rutledge et al., 2004; Kaiser et al., 2007; Costamagna and Landis, 2007; Noma and Brewer, 2008; Noma et al., 2010)

. Five n ative Aphelinus species ( A. lapisligni

, A. mali

, A. prociphili

, A. sanborniae

, and A. siphonophorae ) are in the mali complex to which A.

rhamni belongs . These species are more closely related to A. rhamni than

other native Aphelinus species , yet their hos ts are different from those parasitized by A. rhamni (see Appendix A)

. Therefore, A. rhamni is not expected to compete with native species of Aphelinus

.

Climate projections suggest both A. rhamni and A. glycinis are likely to

establish in the northern Mi dwest of the United States (Hopper, 2010; Hopper, 2011; Hopper, 2012). Establishment of A

. glycinis may preferentially occur on the soybean host, while A. rhamni is expected to

establish on the buckthorn hosts.



## I V. Environmental Consequences

A. No A ction

a . Effect of soybean aphid on host plants

Soybean aphids suck sap from plants. When aphid infestations are large,

infested leaves are wilted or curled. D irect feeding damage from aphids

may include plant stunting, reduced pod and seed counts, and yellowing of

leaves. Soybean aphids also t ransmit plant viruses such as alfalfa mosaic, soybean mosaic

, and bean yellow mosaic ; these diseases cause yield reduction in soybean s

.

For soybean growers in the north central United States, the soybean aphid is the first insect pest to consistently cause important losses over wide

areas (Ragsdale et al., 2004) , with yield decreases of up to 40 percent (Ragsdale et al., 2007)

. Invasion of the s oybean aphid led to dramatic changes in the pest management practices of soybean producers. In 2000,

less than one

- tenth of one percent of soybeans in the north central states

were treated with insecticides . B y 2006, over 13 percent of soybeans in

these states were treated with insecticides . This indicates soybean aphid

was r e sponsible for a 130

- fold increase in the use of insecticides (Ragsdale et al., 2011) . Control of the soybean aphid currently relies on 1

. Impact of Soybean Aphid on the
Environment

7insecticide use, and millions of acres of soybeans have been sprayed sin ce the invasion of this pest . This is a major disruption in the 70 million acres

of soybean production with in the country

.

The contin ued use of chemical control, plant resistance, and biological

control at current levels would result if the “no action ” alternative is

chosen

, and may continue even if permits are issued for environmental

release of A. rhamni

.

a. Chemical C ontrol

Foliar applications may not sufficiently control soybean aphids and prevent yield loss, especially if large quantities of so ybean aphids are surviving on lower leaves. Foliar sprays may pose a threat to bees and aphid natural enemies

, such as lady beetle larvae and predatory bugs. In a screening assay, both λ

- cyhalothrin and dimethoate were harmful to A.

certus (Frewin et al., 2012). This suggests chemical pest controls may limit the effectiveness of various parasitoid wasps released as potential biocontrol organisms.

Seed treatments o f systemic insecticide s such as imidacloprid do not last the entire growing season. In additio n, seed treatment insecticides are applied at the beginning of the growing season before it is known if treatment for soybean aphi d is even necessary. This creates a financial loss to growers if aphid populations would not have been high enough to trigger economic damage requiring pest treatments

.

b. Plant Resistance

Although plant resistance inhibit s aphid growth and development and

increas es plant tolerance to aphid feeding , p lant resistance in certain cases has been overcome by soybean aphids.

c. Biological Control

Two biological control organisms

, Aphelinus atriplicis and Binodoxys

communis were released against soybean aphid , h owever, neither organism e stablished in the environment and provide s cont rol of the soybean aphid. The related A. glyc inis recently was approved for

environmental release in the continental United States ( Hopper, 2010;

USDA

- APHIS, 2012) . T his organism is expected to establish primarily on the soybean host.

These impacts from the use of other control methods may have

e nvironmental consequences even with the implementat ion of the biological control alternative , d epending on the efficacy of A. rhamni to
reduce soybean aphid infestations in the continental United States

. 2. Impact from

the U se of

O ther C ontrol

Methods

8

### B. Issue P ermit s for E nvironmental R elease of **A.**
**rhamni**

a. Scientific Literature

Published literature on the host range of A

. rhamni includes only the information in the species description. In the field, Aphis glycines is the only known host , but i n labo ratory experiments, Aphelinus rhamni parasitizes A phis glycines

, closely related species in th is genus

, and rarely

R hopalosiphum padi L. and Schizaphis graminum (Hopper et al., 2012)

. Other species in the mali complex tend to have narrow host ranges; a

not able exception is A. mali itself which reportedly has a broad host range. On the other hand, literature reports of broad host ranges may reflect confounding effect s from cryptic species within this genus (Heraty et al., 2007).

b. Host S pecificity T estin g

Host specificity testing of A. rhamni in containment examined three ways that aphids were killed: oviposition leading to mummification, stinging the aphid to death without mummification, and host feeding (Hopper, 2011).

Parasitism o f 13 aphid species in five genera in two tribes on 10 plant species in nine families was measured (Table 1) . The se aphids cover ed a range of closely related aphids reported as hosts of the A. mali complex ( see Appendix A), of which A. rhamni is a member. Furthermore, the

ap hid

- plant combinations provided contrasts of aphids in the same versus different genera

, on the same versus different plants ( see Appendix B). Based on the aphid distributions in Blackman and Eastop (2006), most of these aphid species occur within the nati ve geographic range of A. rhamni

, so these aphids and this parasitoid are likely to have been in contact for at least 10,000 years. The exceptions are D iuraphis noxia , which is native to central to western Asia and Europe, and Aphis oestlundi and Aphis

mon ardae , which are native to North America. 1. Impact of

**A. rhamni**  on Non

- target
Insects

9Table 1

. Aphid and host plant species used to test Aphelinus

rhamni host specificity.

Aphid species Plant species Plant family Aphis glycines Glycine max Fabaceae Aphis gossypii Gossypium hirsutum Malvaceae Aphis gossypii Asclepias syriaca Apocynaceae Aphis monardae Monarda fistulosa Laminacea Aphis oestlundi Oenothera biennis Onagraceae Aphis nerii Asclepias syriaca Apocynaceae Aphis asclepiadis Asclepias syriaca Apocynaceae Aphis craccivora Vigna unguiculata Fabaceae Aphis rumicis Rumex altissimus Polygonaceae Rhopalosiphum maidis Hordeum vulgare Poaceae Rhopalosiphum padi Hordeum vulgare Poaceae Schizaphis graminum Hordeum vulgare Poaceae Diuraphis noxi a Hordeum vulgare Poaceae Myzus persicae Raphanus sativus Brassicaceae

Tests of parasitism in 24

- hour exposures of aphids to A. rhamni consisted

of A. rhamni females being placed in cages with 100 aphids of a single species on plant foliage. In th is study, A. rhamni rarely or never parasitized aphids outside of the genus Aphis. Within the genus Aphis, A

. rhamni produced mummified aphids on species closely related to the target aphid, A. glycines (e.g., A. gossypii, A. monardae, and A. oestlundi )

, b ut not on more distantly related Aphis species.

Direct observation of the b ehavior of A. rhamni and aphids determine d host use patterns . Continuous microscopic observation of each female parasitoid (and the aphids to which she was exposed) for 25 minute s under magnification showed A. glycines is the most suitable species for A.

rhamni development, by producing an adult parasitoid per oviposition . In most other species in the genus Aphis

, there were less than half as many A.

rhamni adults per oviposition produced

.

In summary, laboratory experiments showed oviposition and host feeding by A. rhamni was restricted almost completely to species in the genus Aphis closely related to the target. Furthermore, parasitoid stinging that

did not lead to mummificatio n or host feeding did not increase mortality of any aphid species. Handling times of aphids by A. rhamni were long, so

ant

- fended native aphids, such as A. monardae and A. oestlundi , should

not be at risk in the field. The target pest, soybean aphid, was t he most suitable aphid species for A. rhamni development (Hopper, 2011)

. See Appendix B for detail s on the host specificity study.

Additional studies determined if A. rhamni might be likely to shift to non

target aphid hosts under certain conditions. In t hese studies, it was found

10that stresses decreas ing the length of life of A. rhamni did not increase its acceptance of low quality aphid host species. Specificity appears to be

stable , and low

- quality, non

- target species should not have increased risk

of a ttack when A. rhamni is stressed from starvation or age.

Once a biological control agent such as A. rhamni is released into the

environment and become s established, there is a poss ibility it could move from the target insect ( soybean aphid ) to attack nontarget insects, such as

native aphid species. Native species that are closely related to the target species are the most likely to be attacked ( Louda et al., 2003 ). If other aphid sp ecies were to be attacked by A. rhamni , the resulting effects could be environmental impacts that may not be easily reversed. B iological control agents such as A. rhamni generally spread without intervention by

man. In principle, therefore, release of thes e parasitoids at even one site should be considered equivalent to release over the entire area in which potential hosts occur and in which the climate is suitable for reproduction

and survival.

In addition, these agents may not be successful in reducin g soybean aphid populations in the continental United States

. Approximately 12 percent of all parasitoid introductions have led to significant sustained control of the target pests, but the majority of introductions have failed to provide control of the pe st (Greathead and Greathead, 1992) either because

introduction did not lead to establishment or establishment did not lead to control (Lane et al., 1999).

Actual impacts on soybean aphid populations by A. rhamni will not be

known until after release and establishment occurs

. Actual impacts on the

establishment of A. glycinis , including potential competitive interactions

will not be known until after release and establishment of A. rhamni

occurs. The environmental consequences discussed under the no action alternative may occur even with the implementation of the action alternative, depending on the efficacy of A. rhamni to reduce soybean aphid in the continental United States

.

“Cumulative impacts are defined as the impact s on the env ironment which results from the incremental impact of the action when added to other past, present and reasonably foreseeable future actions regardless of what agencies or person undertakes such o ther actions” (40 CFR 1508.7).

To date, o ther soybean aphid biological control organisms released in the

United States have not successfully established. Therefore, competitive

interactions would not occur among any organisms previously released.

The release of A. glycinis is not expected to produce competitive

i nteractions with A. rhamni because habitat partitioning of the two parasitoids between soybean aphid s on the two alternate plant hosts (soybean and Rhamnus species) is anticipated ( K. Hopper, pers. comm. )

. 2. Uncertainties Regarding the
Environmental

R elease of

**A phelinus** 
**rhamni.**

Cumula tive

I mpacts 3. Cumulative Impacts

11The two species also have different predicted geog raphical distributions based on climate matching, with the distribution of A. glycinis extending

further north and A. rhamni further south (Hopper, 2010). A s discussed previously, A. rhamni is not expected to compete with native species of Aphelinus

.

T he non

- native parasitoid Aphelinus certus Yasnosh accidentally

established in North America (Heimpel et al., 2010, Frewin et al., 2010) . I t was found over a wide geographic range in the eastern United States and

Ca nada since 2005

. I n laboratory experimen t s, i t has a host range of at least four genera in two subfamilies of aphids, and on at least four host plant species ( Heimpel et al., 2010 ). The abundance of A. certus in the

United States is unlikely to be affecte d by introduction of A. rhamni

because th is parasitoid’s host range is narrower than for A. certus

. Aphelinus certus might interfere with establishment of A. rhamni , but the

coexistence of these parasitoid species in Asia suggests any interference is

likel y to be short

- lived

.

Release of A. rhamni is not expected to have any negative cumulative impacts in the continental United States because of its host specificity to soybean aphids

. Effective biological control of soybean aphid will have

beneficial effec ts for soybean growers in the United States , may result in a long

- term, non

- damaging method to assist in the control of soybean aphid

, and may reduce the use of insecticides on soybean.

Section 7 of the Endangered Species Act (E SA) and ESA’s implementing regulations require Federal agencies to ensure that their actions are not likely to jeopardize the continued existence of federally listed threatened

and endangered species

, or result in the destruction or adverse

modification of critical habitat.

APHIS has determined that

, based on the host specificity of A. rhamni

, there will be no effect on any listed insect species or designated critical

habitat in the continental United States

. In host specificity testing, A.

rhamni only att ack ed species of aphid s

. There are n o federally listed threatened or endangered insects belong to the aphid family

, Aph ididae (USFWS, 20 1 2 ). There are n o federally listed species are known to depend on or utilize soybean aphid

.



## V. Other Issues

Consistent with Executive Order (EO) 12898, “Federal Actions to Address Environmental Justice in Minority Populations and Low

- income Populations,” APHIS considered the potential for disproportionately high

and adverse human health or environ mental effects on any minority populations a nd low

- income populations. There are no adverse

environmental or human health effects anticipated from the field release of 4. Endangered Species Act

12A. rhamni and its release will not have disproportionate adverse effects to any minority or low

- income populations.

Consistent with EO 13045, “Protection of Children From Environmental Health Risks and Safety Risks,” APHIS considered the potential for disproportionately high and adverse environmental health and safety risks

to children. Th ere are no circumstances that would trigger the need for special environmental reviews involved in implementing the preferred alternative. Therefore, there are no disproportionate effects on children anticipated because of the field release of A. rhamni

.

EO 1317 5

, “ Consultation and Coo r dination with Indian Tribal

Govern m ents ” , w a s issued to ensure t h at there would be “ m eaningful

consultation and collaboration with tribal officials in the dev e lop m ent of Federal policies that have tribal i mpli c ations....” Cons istent w ith EO

131 7 5, A P H I S s e nt letters of notification a n d requests fo r com m ent a n d consultation on the pro p osed action to tribes in areas where the soybean aphid occurs. A P HIS will continue to consult and c o llaborate with Indian

tri b al officials to ensu re that they are wel l

- infor m ed and represent e d in policy a nd program decisions th a t m ay i m pact t heir agricultural interests,

in accorda n ce with EO 1 3175

.



## VI. Agencies, Organizations, and
Individuals Consulted

T his EA was prepared and reviewed by APHIS and ARS

. The addresses

of p articipating APHIS units and any applicable cooperators are provided.

U.S. Department of Agriculture Animal and Plant Health Inspection Service Policy and Program Development

Environmen tal and Risk Analysis Services 4700 River Road, Unit 149 Riverdale, MD 20737

U. S. Department of Agriculture Animal and Plant Health Inspection Service Plant Protection and Quarantine Registrations, Identification, Permits, and Plant Safeguarding

4700 Ri ver Road, Unit 133 Riverdale, MD 20737

U

. S

. Department of Agriculture Agricultural Research Service

Beneficial Insect Introductions Research Unit 501 South Chapel Street

13Newark, DE 19713

14

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22

### Appendix A. Aphelinus species described from North America and their species complexes, distributions, and reported hosts

Species Author Species complex Reported distribution Rep orted hosts

howardii Dalla Torre,

1898 abdominalis California (Howard, 1895) Acyrthosiphon pisum (Herting, 1972) ; Macrosiphum euphorbiae (Sullivan and van den Bosch, 1971) ; Illinoia liriodendri (Zuparko and Dahlsten, 1993) jucundus Gahan, 1924 abdominalis California (Gahan, 1924) ; Ne w York (Griswold, 1926) Acyrthosiphon malvae (Griswold, 1926) ; Acyrthosiphon pseudodirhodum (Griswold, 1927) ; Aulacorthum solani (Herting, 1972) ; Macrosiphum euphorbiae (Gahan, 1924) ; Myzus persicae (Griswold, 1927) semiflavus Howard, 1908 asychis Colorado (Howard, 1908) ; Indiana (Hartley, 1922) ; Kansas, Oklahoma (Kelly, 1917) ; Minnesota, New

Mexico (Webster and Phillips, 1912) ; New York (Griswold, 1927) ; Ohio (Girault, 1917) Acyrthosiphon pisum

, Aphis gossypii

, Cerusaphis viburnicola

, Rhopalosiphum maidis

, Macr osiphoniella sanborni

, Macrosiphum

avenae (Hartley, 1922) ; Aphis rumicis, Brevicoryne brassicae, Lipaphis pseudobrassicae, Sitobion avenae (Peck, 1963) ; Chaitophorus

vim inalis

, Schizaphis graminum (Webster and Phillips, 1912) ; Myzus persicae (Howard, 1908) ; Myzaphis rosarum (Peck, 1963)

marlatti Ashmead,

1888 asychis Kansas (Ashmead, 1888b) ; Ontario ; Quebec (Peck, 1963) Aphis sp. (Ashmead, 1888b) ; Myzus persicae (Herting, 1972)

lapisligni Howard, 1917 mali Oregon (Howard, 1917) ; Idaho (Smith, 1923) ; Washington (Smith, 1923) Brachycaudus helichrysi (Peck, 1963) ; Nearctaphis bakeri (Howard, 1917)

mali Haldeman,

1851 mali Arkansas (Becker, 1918) ; Colorado (Gillette and Taylor, 19 08) ; District of Columbia, Illinois,

Missouri (Howard, 1881) ; Kansas (Lohrenz, 1911) ; Nebraska (Bruner, 1894) ; Pennsylvania (Cresson, 1862) Eriosoma americanum (Girault, 1909) ; Eriosoma crataegi (Howard, 1881) ; Eriosoma lanigerum (Haldeman, 1859)

prociphili Carver, 1980 mali Iowa (Carver, 1980) Prociphilus fraxinifolii (Carver, 1980)

sanborniae Gahan, 1924 mali Penns ylvania (Gahan, 1924) ; Ohio (Delong and Jones, 1926) Sanbornia juniperi (Gahan, 1924) ; Nasonovia houghtonensis (Delong

and Jones, 1926) siphonophorae A shmead,

1888 mali Florida (Ashmead, 1888a) ; New Jersey (Peck, 1963) Macrosiphum sp. (Ashmead, 1888a)

nigritus Howard, 1908 varipes South Carolina (Howard, 1908) ; Kansas, Minnesota (Webster and Phillips, 1912) ; Ne w Mexico (Webster, 1909) ; Oklahoma (Jackson et al., 1970) ;
Texas (Archer et al., 1974) Schizaphis graminum (Howard, 1908) ; Hysteroneura setariae (Webster and Phillips, 1912) ; Aphis helianthi (Rogers et al., 1972) ; Rhopalosiphum maidi (Jackson et al., 1970)

aureus Gahan, 1924 subgenus

Mesidia California (Gahan, 1924) Chaitophorus nigrae (Gahan, 1924)

automatus Girault, 1911 subgenus

Mesidia Illinois (Girault, 1911) ; Virginia (Girault, 1916) ; California (Zuparko and Dahlsten, 1995) Chaitophorus sp. (Girault, 1911) ; Hysteroneura setariae (Girault, 1916) ; Eucalli pterus tiliae (Zuparko and Dahlsten, 1995)

23gillettei Howard, 1914 subgenus

Mesidia Colorado (Howard, 1914) Diuraphis tritici (Howard, 1914)

perpallidus Gahan, 1924 subgenus

Mesidia Iowa (Ga han, 1924) ; Alabama (Edelson and Estes, 1987) ; California (Peck, 1963) ; Florida (Mizell and

Sc hiffhauer, 1990) ; Texas (Watterson and Stone, 1982) Chromaphis juglandicola (Pe ck, 1963) ; Melanocallis fumipennellus

, Monellia costalis (Peck, 1963) ; Monellia caryella (Bueno and Stone, 1983) ; Monelliopsis pecanis (Edelson and Estes, 1987)

24

## Appendix B

. Host specificity tests for **Aphelin u**  **s rhamni**  (Hopper 2011; 2012)

.

Parasitism in 24

- hour exposure of aphids on plant. I ndivid ual female parasitoids were exposed

to one of 12 aphid species. One to five day old females were used that had been with males and aphids since emergence and thus had the opportunity to mate, host

- feed, and oviposit. F emales

were isolated from males and ap hids for 24 hours before using them in experiments to insure that

the females had a full egg load. E ach female parasitoid was placed in a cage (10 centimeters (cm) diameter by 22 cm tall) enclosing the foliage of a potted plant of the appropriate species w ith 100 aphids of mixed instars of a single species. Female parasitoids were removed after 24 hours and were used only once.

Because these parasitoids can parasitize a maximum of 20 aphids in 24 hours, this abundance of aphids and period of exposure allow ed parasitoids to use their full egg complement. Furthermore, the density of aphids, amount of plant material, and cage size meant that parasitoids were not limited by search rate. Paras i tism scoring occurred after larval parasitoids

killed and mummified their hosts. Therefore, a combination of acceptance of hosts for oviposition and suitability of hosts for parasitoid development were measured

.

Female parasitoids had the choice to oviposit or not to oviposit in a particular host species. This is frequen tly the choice parasitoids make in the field. The purpose of this study was to measure parasitism in an environment that appears to harbor only one aphid species on only one plant species and where female parasitoids re

- encounter this combination repeated ly starting with a full egg complement.

Ten days after exposure of aphids to parasitoids, any mummified aphids were collected and held for adult parasitoid emergence. After the adults emerged, the number of mummified aphids and

the number and sex of adul t parasitoids were recorded

. Eight to 32 females on each of 12 aphid species were tested

.

After 24 hours exposure, the number of aphids mummified by A. rhamni varied with aphid species ( F = 13.1; df = 12, 213; P < 0.0001; Fig. 3). Female parasitoids rarely or never parasitized aphids outside the genus Aphis ; within the genus Aphis , female parasitoids produced mummified aphids on species phylogenetically close to A. glycines , e.g. A. gossypii

, A.

monardae , and A. oestlundi , but not on aphids that are more ph ylogenetically distant, e.g. A. nerii

and A. rumicis . The low numbers of mummified aphids for A. nerii may have resulted from their host plant, A. syriaca, which is known to produce toxic secondary chemicals which aphids could sequester for their defense ( Mooney et al., 2008, Warashina and Noro, 2000). This effect is supported by many fewer mummies for A. gossypii on A. syriaca than on G. hirsutum (Fig. 3).

The proportion of mummies from which parasitoid adults emerged was generally high (0.93±0.01) but va ried among species ( F = 4.6; df = 8, 67; P = 0.0002) because A. oestlundi had a lower adult emergence rate (0.64±0.01) than the other aphids. Sex ratio of parasitoid progeny

25did not differ from equality (0.52±0.02) and did not differ among aphid species



### .

Direct observations of behavior in e xperiments on the stresses of starvation and age, egg load, and experience affect ing acceptance of low versus high quality host species for oviposition by Aphelinus rhamni found f emale parasitoids may behave different ly after encountering low versus high quality hosts for longer periods. Starved Aphelinus rhamni females persisted in ovipositing more in high

- quality aphid species than in low quality aphid species

. Females oviposited about

twice as often in Aphis glyci nes than in A. craccivora or R. padi ( F = 5.8; df = 2,128; P = 0.004).

Starved and fed females laid about the same numbers of eggs (1.0 ± 0.2 versus 1.1 ± 0.2, respectively), and there was no interaction between nutrition and aphid species in their effect on oviposition. Aphid species and egg load did interact in their effect on oviposition ( F = 5.9; df = 3,124; P = 0.0009). Honey

- fed parasitoids exposed to A. glycines oviposited more when they carried more eggs, but oviposition in the other aphid species d id not increase with egg load,

and increased only slightly in water

- fed females exposed to A. glycines . This pattern is the opposite of what one would expect if female parasitoids became choosier as eggs became more limiting. The higher oviposition in A. g lycines did not result from stinging this species more often. Indeed, female parasitoids stung A. craccivora more often than A. glycines , which was stung at the about the same rate as R. padi (6.3±0.4, 4.1±0.3, and 3.7±0.3 aphids, respectively; F

26= 17.3; d f = 2,156; P < 0.0001), although A. glycines was oviposited in twice as often as the other aphid species. The lack of correlation between stinging and oviposition, probably results

because Aphelinus females usually do not directly contact aphids before ovi positor insertion, and thus are may not assess host quality until aphids are stung. On encountering a potential host, Aphelinus rhamni females stop at about half their body length away from the aphid. Without touching the aphid, females sway from side to s ide several times with antennae extended (an approach). Stinging did not vary with nutrition, egg load, nor did these factors interact with aphid species in its effect on numbers of aphids stung.

Effect of age on oviposition in low versus high quality hos ts. Ten

- day old Aphelinus

rhamni females persisted in ovipositing more in a high quality aphid species than in a low quality aphid species. Females oviposited over twice as often in A. glycines than in R. padi ( F = 13.0; df = 1,76; P = 0.0006). Young and o ld females laid about the same number of eggs (1.1 ± 0.2 versus 0.9 ± 0.2 eggs, respectively), and there was no interaction between parasitoid age and aphid species.

As in the nutrition experiment, there was a strong interaction between aphid species and egg load in their effect on oviposition ( F = 6.6; df = 3,125; P = 0.0002). Young parasitoids exposed to A. glycines oviposited more when they carried more eggs, but oviposition did not increase significantly with egg load in parasitoids exposed to R. padi or in older parasitoids exposed to A. glycines . As in the nutrition experiment, the preference for high quality hosts persisted in the face of low versus high egg load. At the start of observations, young females carried about 3 more eggs than older female s (9.5 ± 0.9 versus 6.2 ± 0.6, respectively; F = 453.3; df = 2,77; P < 0.0001), and by the end of observations, both young and old females still had eggs they had not oviposited (8.0 ± 0.8 and 5.3±0.6, respectively).

The higher oviposition in A. glycines was partly a result of stinging this aphid species more

often. Unlike the starvation experiment, where A. glycines and R. padi where stung equally often, female parasitoids in this experiment stung A. glycines slightly more often than R. padi

.

Effect of age on oviposition in low versus high quality hosts. Ten

- day old Aphelinus rhamni females persisted in ovipositing more in a high quality aphid species than in a low quality aphid species. Females oviposited over more than twice as often in A. glycines tha n in R. padi ( F = 13.0; df = 1,76; P = 0.0006). Young and old females laid about the same number of eggs (1.1± 0.2 versus 0.9 ± 0.2 eggs, respectively), and there was no interaction between parasitoid

age and aphid species.

As in the nutrition experiment, there was a strong interaction between aphid species and egg load in their effect on oviposition ( F = 6.6; df = 3,125; P = 0.0002). Young parasitoids exposed to A. glycines oviposited more when they carried more eggs, but oviposition did not increase signi ficantly with egg load in parasitoids exposed to R. padi or in older parasitoids exposed to A. glycines . As in the nutrition experiment, the preference for high quality hosts persisted in the face of low versus high egg load. At the start of observations, young females carried about 3 more eggs tha n older females (9.5 ± 0.9 versus 6.2 ± 0.6, respectively; F = 453.3; df = 2,77; P < 0.0001), and by the end of observations, both young and old females still

27had eggs they had not oviposited (8.0 ± 0.8 and 5.3±0. 6, respectively). The higher oviposition in

A. glycines was partly a result of stinging this aphid species more often.

Unlike the starvation experiment, where A. glycines and R. padi where stung equally often, female parasitoids in this experiment stung A. glycines slightly more often than R. padi (5.6±0.4 versus 4.4±0.3, respectively; F = 5.3; df = 1,77; P = 0.02).However, this difference was only 1.3 fold more stings in A. glycines than R. padi , whereas oviposition was 2.5 fold greater in A. glycines th an in R. padi . Stinging did not vary with age, egg load, nor did these factors interact with aphid species in its effect on numbers of aphids stung. The frequency of host feeding did not vary with age, but parasitoids fed on A. glycines more frequently tha n on R. padi (32 percent

versus 6 percent of parasitoids fed on each species; likelihood ratio χ 2 = 8.2; n = 74; P = 0.003),

and this difference was similar to that for honey

- fed parasitoids in the nutrition experiment of the same age. Host feeding did not vary with egg load for either aphid species, age, or their

c ombinations.

Dif ferences in aphid defense cannot explain differences in oviposition between species because A. glycines defended itself as much as R. padi (2.3 ± 0.3 versus 2.1±0.3 defensive behaviors per approach), but received over twice as many eggs. Amount of aphid de fense behavior did not vary with aphid species, age, or their interaction.

Both young and old Aphelinus rhamni females produced more progeny on high quality host species than on low quality host species when exposed to the aphids for 24 hours

.

Conclusion s. Stresses that decreased life expectancy did not increase acceptance of low quality host species by Aphelinus rhamni . Egg load was also lower in stressed females, which resorbed eggs in response to starvation and aging, so changes in life expectancy and egg load may have counterbalanced one another. However, the decrease in egg load was small relative to the decrease in life expectancy, particularly for starved females, which carried about three

- quarters
of the egg load of honey

- fed females, but would die the day following the assays.

Furthermore, although oviposition increased with egg load for females exposed to A. glycines

, oviposition did not vary with egg load for females exposed to low quality aphid species. The outcome of this pattern was that fema les became less choosy as egg load decreased, rather than becoming choosier , as theory predicts. Besides having lower life expectancy, older females were kept without aphids as they aged so they experienced a more aphid

- poor environment than did

young para sitoids. It was expected this would make parasitism of low quality aphids by older females more frequent, yet it did not. These results demonstrate that A. rhamni specificity is behaviorally stable rather than dynamic, and low

- quality non

- target species sh ould not have

increased risk of attack when parasitoids are stressed from starvation or age or when they accumulate eggs.

28