United States 

Department of 

Agriculture 

Forest 

Service 

Northern 

Region 

200 East Broadway 

P.O. Box 7669 

Missoula, MT 59807 

Numbered Report 07-07 September 2007

COLONIZATION OF HAWAIIAN ACACIA KOA

SEEDLINGS WITH FUSARIUM SPECIES 

R.L. James1, A. Yeh2, and N.S. Dudley2

1

USDA Forest Service, Coeur d’Alene, ID, 2Hawaii 

Agriculture Research Center, Aiea, HI

ABSTRACT 

Acacia koa seedlings were evaluated from plantings and commercial nurseries for colonization by 

Fusarium spp., particularly F. oxysporum, the putative cause of koa wilt/dieback disease in Hawaii. We 

sampled 162 seedlings, 90% of which were from nurseries on the islands of Hawaii, Maui, Kauai, and 

Oahu. Fusarium oxysporum was detected on about 78% of sampled seedlings. It was most prevalent on 

roots; more than 50% of all sampled root pieces were colonized by this species. Ten additional Fusarium

species were isolated from seedlings, but all colonized plant tissues at much lower levels. More than 70% 

of all samples from seedlings yielded Fusarium spp. The potential importance of this high level of 

colonization is discussed. 

INTRODUCTION 

One of the most important tree species in Hawaii 

is Acacia koa A. Gray, which is a fast growing 

native (Dudley 2002). There is much recent 

interest in increasing koa plantings because of its 

high value and the availability of previous 

agriculture land for potential plantations. One of 

the major limiting factors in increased 

production of koa is an important wilt/dieback 

disease that seems to be increasing in 

importance. The primary cause of this disease is 

believed to be the soil-borne vascular wilt 

pathogen Fusarium oxysporum f.sp. koae

(Anderson and others 2002; Gardner 1980), 

although weather and site factors undoubtedly 

play important roles in disease severity 

(Anderson and others 2002). 

Disease symptoms can occur on plants of all 

ages from young seedlings to large, old-growth 
2

trees (Anderson and others 2002; Gardner 1980; 

James and others 2007). Seedlings tend to wilt 

and die more rapidly than older trees (James 

2004a). Previous work (Gardner 1980) indicated 

that F. oxysporum may contaminate and be 

disseminated on koa seeds. Another major 

concern is the possibility of the pathogen 

spreading on infected nursery seedling stock. 

This could potentially spread the pathogen to 

new areas. Therefore, we conducted a survey to 

evaluate the extent and distribution of F. 

oxysporum and other Fusarium species on some 

recently-planted seedlings as well as on koa 

seedling nursery stock. 

MATERIALS AND METHODS 

Planted koa seedlings exhibiting wilt symptoms 

were occasionally collected during field 

examinations of diseased areas. Seedlings grown 

in 20 commercial nurseries on the four major 

Hawaiian Islands (Hawaii, Maui, Kauai, 

Oahu)(Table 1) were also examined for any 

indication of disease or reduced vigor, i.e., 

wilting, foliar chlorosis or necrosis, and dieback. 

We tried to sample the poorest looking nursery 

seedlings such as those not growing well or with 

some foliar discoloration. However, in many 

cases most of our sampled seedlings appeared 

healthy. The number of seedlings sampled varied 

among the nurseries. Ninety percent of all 

sampled seedlings came from commercial 

nurseries. More than half of all sampled 

seedlings were from the Big Island (Table 1). 

Selected seedlings were extracted from soil or 

their containers, kept refrigerated, their roots 

thoroughly washed to remove soil or growing 

media, and shipped to the laboratory for analysis. 

Roots of all seedlings were sampled for 

Fusarium colonization; stems and/or branches 

were sampled from only about 38% of the 

seedlings. Roots and stem/branch sections were 

dissected into pieces about 5 mm in length. 

Randomly-selected pieces were surface sterilized 

in 0.525% aqueous sodium hypochlorite (10% 

bleach solution), rinsed in sterile water, and 

placed on a selective agar medium for Fusarium

and closely-related fungi (Komada 1975). Plates 

were incubated under diurnal cycles of cool, 

fluorescent light at about 24o

C for 7-14 days. 

Selected emerging fungi were transferred to 

potato dextrose and carnation leaf agar (Fisher 

and others 1982) for identification using the 

taxonomy of Nelson and others 1983. 

Percentages of sampled pieces colonized by 

particular Fusarium species were calculated. 

RESULTS AND DISCUSSION 

Eleven different Fusarium species were isolated 

from seedlings (Table 2). The most commonlyisolated species by far was F. oxysporum; it was 

found on more than half of the sampled root 

pieces and almost a fourth of the stem or branch 

pieces. This species was isolated from about 

78% of all sampled seedlings; it was prevalent 

on seedlings both in plantings and within 

nurseries. We found F. oxysporum commonly on 

the roots of many seedlings that displayed little 

or no disease symptoms. All sampled roots 

appeared healthy with no indications of necrosis 

or discoloration. Vascular discoloration was 

noted within stems of some, but not all, seedlings 

from which F. oxysporum was isolated. 

Although F. oxysporum was common on most 

sampled seedlings, we do not know the 

capability of such isolates to elicit disease on 

koa. We plan to evaluate pathogenic potential of 

some of these isolates in future controlled 

greenhouse inoculation tests. However, at the 

present time we don’t know what proportions of 

the F. oxysporum population colonizing 

seedlings are potentially pathogenic. It is 

possible that all or most of the isolates are either 

saprophytic or non-pathogenic endophytes 

(Gordon and Martyn 1997; Kistler 1997). Since 

standard wilt symptoms were not usually found 

in sampled nurseries, we suspect that the 

majority of the isolates we obtained are non 

pathogens. On the other hand, the planted 

seedlings we sampled exhibited wilt or dieback 

symptoms and we would expect a greater 

proportion the fungal population from these 

seedlings to be pathogenic. 
3

We have found F. oxysporum commonly 

colonizing roots of large koa trees (James and 

others 2007). Roots of large trees also appear 

healthy, even though they may be extensively 

colonized. Isolates of F. oxysporum capable of 

inducing wilt symptoms initially infect cortical 

root cells and rapidly grow into vascular 

systems, where spores may be systemically 

distributed throughout infected plants (James 

2004b; James and Dumroese 2007; Nelson and 

others 1981). We found that F. oxysporum was 

systemically distributed in those seedlings in 

which roots, stems and branches were sampled. 

We are unsure about potential inoculum sources 

of F. oxysporum in koa nurseries. High levels of 

this species have not been routinely detected on 

sampled koa seeds (James 2004a; James and 

others 2006). Plastic and styrofoam containers 

reused for several seedling crops may be 

important sources of pathogen inoculum (James 

2004b; James and Dumroese 2007). Most of the 

containers used in Hawaiian nurseries were 

plastic tubes of varying sizes. Previous work 

(Dumroese and others 1993, 2002) indicated that 

Fusarium inoculum can remain viable on such 

containers for prolonged periods; sterilization 

with hot water is usually required to remove 

inoculum. Another potential source of F. 

oxysporum inoculum is the growing media used 

by nursery growers, although commercial peatbased media is usually not contaminated by 

potentially-pathogenic Fusarium (James 1985). 

We have not tested the media and containers 

used by Hawaiian nursery growers for potential 

pathogens. Some media contained rather large 

pieces of wood and volcanic rock and it is 

possible that this material may potentially harbor 

Fusarium inoculum. 

Fusarium oxysporum is usually soil-borne and 

many of the sampled nurseries are located in 

areas where wind-blown soil contamination 

occurs. Therefore, blowing soil may be an 

important source of inoculum within nurseries, 

although this has not yet been tested. 

The other most common Fusarium species 

isolated from koa seedlings were F. solani

(Mart.) Appel & Wollenw., F. semitectum Berk. 

& Rav. and F. subglutinans (Wollenw. & 

Reinking). Fusarium solani is a common soilborne fungus capable of causing several types of 

plant diseases, including root decay (James 

2004b; Nelson and others 1981); we have found 

it most commonly within the interior portions of 

large roots, stems and branches of koa trees 

exhibiting wilt/dieback symptoms (Daehler and 

Dudley 2002; James and others 2007). Fusarium 

solani is often associated with activity of black 

twig borers (Xylosandrus compactus Eichhoff 

(Coleoptera: Scolytidae), which commonly infest 

declining trees (Daehler and Dudley 2002). 

Both F. semitectum and F. subglutinans are 

commonly isolated from Acacia koa seed (James 

2004a; James and others 2006). These species 

have also been detected on larger trees with 

disease symptoms, although at relatively low 

levels (James and others 2007). We suspect that 

some seedlings become infected from 

contaminated seeds following germination. 

Although both Fusarium species may be plant 

pathogens (Logreico and others 2002; Onyike 

and Nelson 1992; Satou and others 2001), their 

abilities to elicit diseases on koa seedlings have 

not been adequately tested. Limited tests 

indicated that both F. subglutinans and F. 

semitectum have pathogenic potential on young 

koa seedlings under controlled greenhouse 

inoculation conditions (Dudley and others 2007). 

However, much further testing will be necessary 

before we can make definitive conclusions 

regarding the pathogenic potential of these two 

Fusarium species. 

The other seven Fusarium species isolated from 

koa seedlings were found at very low levels 

(Table 2). We suspect that few, if any, of these 

are capable of causing koa diseases. Most of 

them may be non-pathogenic endophytes that do 

not adversely affect their hosts. 

In conclusion, we confirmed that F. oxysporum

commonly colonizes Acacia koa seedlings from 
4

both plantings and commercial nurseries. 

However, nursery growers need not be 

concerned unless the majority of isolates within 

their nurseries are pathogenic. Determining 

pathogenic potential of F. oxysporum

populations usually requires expensive, timeconsuming inoculation tests. However, in some 

cases specific molecular genetic markers have 

been developed to differentiate pathogenic from 

non-pathogenic isolates (Bao and others 2002; 

Lori and others 2004; Roncero and others 2003). 

With such markers, characterization of fungal 

populations may be determined much more 

quickly and at less cost. Using such techniques 

to characterize Fusarium populations associated 

with koa seedlings will greatly improve our 

understanding of the potential impacts of these 

fungi on nursery crops. 

LITERATURE CITED 

Anderson RC, Gardner DE, Daehler CC, Meinze 

FC. 2002. Dieback of Acacia koa in Hawaii: 

ecological and pathological characteristics of 

affected stands. Forest Ecololgy and 

Management 162:273-286. 

Bao JR, Fravel DR, O’Neill NR, Lazarovits G, 

van Berkum P. 2002. Genetic analysis of 

pathogenic and nonpathogenic Fusarium 

oxysporum from tomato plants. Canadian 

Journal of Botany.80:271-279. 

Daehler CC, Dudley NS. 2002. Impact of the 

black twig borer, an introduced insect pest, on 

Acacia koa in the Hawaiian Islands. 

Micronesica Supplement 6: 35-53. 

Dudley NS. 2002. 2-Year Growth Performance 

of selected Acacia koa progeny at Maunawili, 

Oahu. Hawaii Agriculture Research Center 

Report (Unpublished). 14p. 

Dudley NS, James RL, Sniezko RA, Yeh A. 

2007. Pathogenicity of four Fusarium species 

on Acacia koa seedlings. USDA Forest 

Service, Northern Region, Forest Health 

Protection. Report 07-04. 9pp. 

Dumroese RK, James RL, Wenny DL. 1993. 

Sodium metabisulfite reduces fungal inoculum 

in containers used for conifer nursery crops. 

Tree Planters' Notes 44(4):161-165. 

Dumroese RK, James RL, Wenny DL. 2002. Hot 

water and copper coatings in reused containers 

decrease inoculum of Fusarium and 

Cylindrocarpon and increase Douglas fir 

seedling growth. HortScience 37:943-947. 

Fisher NL, Burgess LW, Toussoun TA, Nelson 

PE. 1982. Carnation leaves as a substrate and 

for preserving cultures of Fusarium species. 

Phytopathology 72:151-153. 

Gardner DE. 1980. Acacia koa seedling wilt 

caused by Fusarium oxysporum. Phytopathology 70:594-597. 

Gordon TR, Martyn RD . 1997. The evolutionary 

biology of Fusarium oxysporum. Annual 

Review of Phytopathology 35:111-128. 

James RL. 1985. Diseases associated with 

containerized seedling soil mixes. Tree 

Planters' Notes. 36(2):3-5. 

James RL. 2004a. Fusarium colonization of 

seeds, seedpods, and diseased seedlings of 

Acacia koa from Hawaii. USDA Forest 

Service, Northern Region, Forest Health 

Protection. Nursery Disease Notes No. 159. 

22p. 

James RL. 2004b. The role of Fusarium species 

in the etiology of conifer seedling diseases in 

forest nurseries. In: Proceedings of the Hawaii 

International Conference on Sciences, January, 

2004, Honolulu, Hawaii. 17p. 
5

James RL, Dudley NS, Yeh Y. 2006. 

Investigating koa wilt in Hawaii: Examining 

Acacia koa seeds and seedpods for Fusarium 

species. Native Plants 7(3):315-323. 

James RL, Dudley NS, A Yeh Y. 2007. 

Colonization of diseased Acacia koa trees with 

Fusarium species. USDA Forest Service, 

Northern Region, Forest Health Protection. 

Report 07- (In press). 

James RL, Dumroese RK. 2007. Evolution of 

our knowledge of Fusarium diseases in forest 

nurseries. In: J.R. Sutherland and Z. 

Prochazkova (eds.). Proceedings of the Sixth 

Meeting of IUFRO Working Party 7.03.04 

(Diseases and Insects in Forest Nurseries). 

Communicationes Instituti Forestalis 

Bohemicae 23:53-66. 

Kistler HC. 1997. Genetic diversity in the plantpathogenic fungus Fusarium oxysporum. 

Phytopathology 87:474-479. 

Komada H. 1975. Development of a selective 

medium for quantitative isolation of Fusarium 

oxysporum from natural soil. Review of Plant 

Protection Research (Japan) 8:114-125. 

Logrieco A, Mule G, Moretti A, Bottalico A. 

2002. Toxigenic Fusarium species and 

mycotoxins associated with maize ear rot in 

Europe. European Journal of Plant Pathology 

108:598-609. 

Lori G, Edel-Hermann V, Gautheron N, 

Alabouvette C. 2004. Genetic diversity of 

pathogenic and nonpathogenic populations of 

Fusarium oxysporum isolated from carnation 

fields in Argentina. Phytopathology 94:661-

668. 

Nelson PE, Toussoun TA, Cook RJ. 1981. 

Fusarium Diseases, Biology, and Taxonomy. 

University Park (PA): The Pennsylvania State 

University Press. 457p. 

Nelson PE, Toussoun TA, Marasas WFO . 1983. 

Fusarium species: an illustrated manual for 

identification. The Pennsylvania State 

University Press. University Park. 193p. 

Onyike NBN, Nelson PE. 1992. Fusarium

species associated with sorghum grain from 

Nigeria, Lesotho, and Zimbabwe. Mycologia 

84:452-458. 

Roncero MIG, Hera C, Ruiz-Rubio M, GarciaMaceira FI, Madrid MP, Caracuel Z, Calero F, 

Delgado-Jarana J, Roldan-Rodriguez R, 

Martinez-Rocha AL, Velasco C, Roa J, MartinUrdiroz M, Cordoba D, Di Pietro A. 2003. 

Fusarium as a model for studying virulence in 

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Molecular Plant Pathology 62:87-98. 

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6

Table 1. Locations where Acacia koa seedlings were sampled for colonization by Fusarium spp. 

Island [County] Number of Nurseries 

Sampled1

Percent of Seedlings 

Sampled 

Hawaii [Big Island] 7 57 

Maui 6 28 

Kauai 4 12 

Oahu 3 3 

1

90% 

Table 2. Colonization of Acacia koa Seedlings with Fusarium spp. 

Number of Roots Stems/Branches All Samples 

Seedlings Sampled 158 62 162 

Pieces Sampled 1892 533 2425 

Fusarium spp. Percent Colonization 

oxysporum

1

 56.1 21.2 48.4 

solani 9.6 8.1 9.2 

semitectum 7.9 6.0 7.5 

subglutinans 4.3 4.1 4.3 

equiseti 1.4 3.6 1.9 

proliferatum 1.2 0.9 1.1 

avenaceum 0.05 3.6 0.8 

graminearum 0.8 0.2 0.7 

acuminatum 0.4 0.4 0.4 

sporotrichioides 0.4 0.2 0.4 

anthophilum 0 0.9 0.2 

All Fusarium 77.1 46.1 70.3 

No Fungi 0.2 36.0 8.1 

1

77.8%of all sampled seedlings were infected by F. oxysporum. 

R.L. James is Plant Pathologist, USDA Forest Service, Forest Health Protection, 3815 Schreiber Way, 

Coeur d’Alene, ID 83815; rjames@fs.fed.us. A. Yeh is Plant Propagation Specialist, Hawaii Agriculture 

Research Center, 942 W. Kawailani Street, Hilo, HI 96778; ayeh@hawaii.rr.com. N.S. Dudley is 

Forester; Hawaii Agriculture Research Center, 99-193 Aiea Heights Drive, Aiea, HI 96701; 

NDudley@harc-haspa.com.