United States 

Department of 

Agriculture 

Forest 

Service 

Northern 

Region 

200 East Broadway 

P.O. Box 7669 

Missoula, MT 59807 

Numbered Report 07-06 September 2007

COLONIZATION OF DISEASED ACACIA KOA

TREES WITH FUSARIUM SPECIES 

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

1

USDA Forest Service, Coeur d’ Alene, ID 

2

Hawaii Agriculture Research Center, Aiea, HI 

ABSTRACT 

Fifty-two Acacia koa trees with wilt/dieback disease symptoms in Hawaii were sampled for associated 

Fusarium spp. within their roots, stems and branches. Fusarium oxysporum was isolated from nearly 35% 

of all sampled roots and was especially common on healthy-appearing fine feeder roots. Fusarium solani 

was isolated most frequently from stem and branch tissues, but also occurred on more than 10% of 

sampled roots. A total of fourteen Fusarium spp. were detected within diseased trees; we suspect that 

most of these were probably non-pathogenic fungal endophytes. We found that F. oxysporum commonly 

colonized diseased trees, especially roots. Coupled with our results from pathogenicity testing, we believe 

that this fungal species is probably important in koa wilt/dieback disease etiology. 

INTRODUCTION 

Acacia koa A. Gray is one of the most important 

native forest trees in Hawaii. Koa wood is highly 

prized for several important products, including 

surf boards, paneling, furniture, and miscellaneous carvings. This tree species grows very 

quickly under Hawaiian forest conditions and 

high-valued trees can be produced in relatively 

short rotations (Dudley 2002). 

An important disease resulting in wilt and 

dieback symptoms was first described on koa in 

1980 (Gardner 1980). Etiological studies 

determined that the primary cause of the disease 

was root-infecting strains of Fusarium 

oxysporum Schlecht. (Anderson and others 2002; 

Gardner 1980). Isolates capable of eliciting 

disease symptoms were placed within the taxon 
2

F. oxysporum f.sp. koae. Subsequent work 

evaluated ecological factors important in disease 

occurrence, spread and intensity (Anderson and 

others 2002). In addition, one study (Daehler and 

Dudley 2002) found that the black twig borer 

[Xylosandrus compactus Eichhoff (Coleoptera: 

Scolytidae)] often infested declining trees and 

was commonly associated with colonization of 

stem and branch wood by another Fusarium

species [F. solani (Mart.) Appel & Wollenw.]. 

Although Koch’s postulates have been 

completed several times, confirming pathogenicity of F. oxysporum on koa, controversy 

still remains regarding the actual importance of 

this fungus in disease etiology. We have 

completed several greenhouse pathogenicity tests 

and found that certain isolates of F. oxysporum

are highly virulent on susceptible koa seedlots 

(Dudley and others 2007a), although the 

proportion of isolates from populations 

associated with diseased trees that are capable of 

being aggressive pathogens is relatively low. 

Although some reports indicated that F. 

oxysporum may infest koa seeds (Gardner 1980; 

Anderson and others 2002), we have only 

detected this species at very low levels on or 

within seeds (James 2004; James and others 

2006). However, several other Fusarium spp. 

have routinely been isolated from koa seeds 

(James 2004; James and others 2006). 

An important part of our investigations 

concerning koa wilt/dieback disease included 

disease-impact surveys on the four major 

Hawaiian Islands (Oahu, Kauai, Maui, and 

Hawaii). The goal of these surveys was to 

determine distribution and severity of the disease 

throughout the natural and planted range of koa. 

As an important part of these surveys, isolations 

were made from diseased trees to determine 

occurrence and extent of colonization by F. 

oxysporum and other Fusarium spp. that may 

influence disease etiology. This report 

summarizes our findings regarding colonization 

of diseased koa trees by Fusarium. 

MATERIALS AND METHODS 

Koa trees exhibiting external symptoms of wilt 

and/or dieback disease were located primarily 

via roadside surveys or from information 

provided by landowners. Fifty-two trees were 

selected based on external indications of disease, 

including foliar necrosis, dieback, and wilting. 

Selected trees were also examined for definitive 

green-black staining within the center of stems, 

branches and large roots, which we have found 

commonly associated with the disease. Wood 

samples from symptomatic trees were collected 

from selected branches, portions of the main 

stem, and from root systems. Emphasis was 

placed on collecting root samples, particularly 

fine feeder roots, since this is likely the point of 

infection by wilt-inducing organisms. Collected 

samples were kept cool within ice chests and 

subsequently washed thoroughly to remove soil 

and shipped to the laboratory for analysis. 

In the laboratory, samples were divided into five 

categories (branches, stems, fine roots, 

secondary roots, and tertiary roots); the latter two 

categories were selected based on branching 

pattern and size of roots. All samples were 

thoroughly rinsed again under running tap water. 

Small pieces of wood (approximately 5 mm2

) 

were aseptically extracted from samples, 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). Fine roots were dissected into 

pieces about 5 mm long and randomly-selected 

for placement on the selective medium following 

surface sterilization. Plates were incubated under 

diurnal cycles of cool, fluorescent light at about 

24o

C for 7-14 days. Selected emerging fungi 

were single-spored and transferred to potato 

dextrose and carnation leaf agar (Fisher and 

others 1982) for identification of associated 

Fusarium spp. using the taxonomy of Nelson and 

others 1981. Percent colonization of wood 

samples by different Fusarium spp. was 

calculated. 
3

RESULTS 

Twelve different Fusarium species were isolated 

from roots of trees displaying wilt/dieback 

symptoms (Table 1). By far the most common 

Fusarium species isolated from roots was F. 

oxysporum. This species was obtained from 

nearly half of the fine roots sampled, but was 

less common within larger secondary and tertiary 

roots. The second most commonly-isolated 

Fusarium species from roots was F. solani, 

which was found on slightly more than 11% of 

sampled root pieces. The ten other Fusarium

spp. were isolated infrequently. Overall, 

Fusarium spp. were obtained from about 2/3 of 

the fine root pieces and nearly half of all sampled 

root pieces (Table 1). 

Fusarium spp. colonized stem and branch tissues 

at lower levels than roots (Table 2), although a 

wide diversity of species were isolated. The most 

commonly isolated Fusarium species was F. 

solani, followed by F. oxysporum and F. 

proliferatum (Matsushima) Nirenberg. Fusarium

spp. were about equally distributed throughout 

stem and branch tissues, i.e., about the same 

colonization levels were detected within tissues 

near the center (usually adjacent to the greenblack staining) and tissues just inside the 

cambium (Table 2). Slightly less than a third of 

all sampled wood pieces from stems and 

branches were colonized with Fusarium spp. We 

also found that over 40% of all sampled wood 

pieces were not colonized by any detectable 

fungi. 

Collated levels of root, stem, and branch 

colonization by Fusarium spp. within sampled 

diseased trees are summarized in Table 3. 

Fourteen different Fusarium spp. were detected 

within the 52 sampled trees exhibiting 

wilt/dieback symptoms. Although most species 

were isolated infrequently, F. oxysporum and F. 

solani were fairly common. A total of 42% of all 

sampled wood pieces from diseased trees were 

colonized by Fusarium spp. 

DISCUSSION 

We found that Fusarium spp. are common 

residents of koa trees exhibiting wilt/dieback 

disease symptoms in Hawaii. Many different 

Fusarium spp. were detected, but the most 

common were F. oxysporum within roots and F. 

solani within stems and branches. We suspect 

that many of the other Fusarium spp. colonized 

wood tissues as non-pathogenic fungal 

endophytes and they probably do not adversely 

affect their hosts. Some of these species, such as 

F. subglutinans (Wollenw. & Reinking) and F. 

semitectum Berk. & Rav., are commonly found 

on koa seeds (James 2004; James and others 

2006) and may have colonized seedlings 

following seed germination. Several of the others 

may have been introduced in other ways, such as 

on feeding insects and/or direct infection of 

tissues from either soil- or wind-borne inoculum. 

Fusarium oxysporum was concentrated within 

koa root systems. Fungal isolates from this 

species complex were especially common on 

fine roots. We did not find necrosis or other 

disease symptoms on colonized roots. Based on 

pathogenicity tests conducted thus far, only a 

small percentage of F. oxysporum isolates from 

diseased trees were highly virulent on inoculated 

seedlings from susceptible seedlots (Dudley and 

others 2007a, 2007b). Unfortunately, highlyvirulent isolates cannot be identified on the basis 

of morphological characteristics (Gordon and 

Martyn 1997; Kistler 1997). In some cases, they 

may be differentiated using special molecular 

markers (Bao and others 2002; Lori and others 

2004; Roncero and others 2003). However, their 

ability to exhibit aggressiveness and elicit 

disease must be confirmed via inoculation tests. 

Preliminary molecular genetic analyses of F. 

oxysporum populations associated with diseased 

koa trees indicated that populations are quite 

genetically diverse and that highly-virulent 

isolates may be clustered together, indicating 

genetic similarity (Yang and others 2007). 

Strictly non-pathogenic F. oxysporum isolates 

may likewise be clustered together within 
4

phylogenetic trees. We are in the process of 

relating genetic clustering of selected F. 

oxysporum isolates with their ability to cause 

disease on koa seedlings. When this work is 

completed, we will have a clearer idea of the 

relationships between genetics and pathogenicity 

within F. oxysporum populations associated with 

diseased koa in Hawaii. Since strains of fungi 

within the F. oxysporum species complex vary in 

their ability to cause plant diseases (Gordon and 

Martyn 1997; Kistler 1997), it is no surprise that 

only a portion of the population is pathogenic. If 

molecular markers can be developed which can 

quickly differentiate highly-virulent strains, we 

will be in a much better position to quantify 

pathogenic strains in nature. 

We also found fairly high levels of F. solani

within diseased koa trees, especially in stem and 

branch tissues. This collaborates previous 

findings where this species commonly colonized 

the interior of koa trees infested with black twig 

borers (Daehler and Dudley 2002). We have only 

evaluated a few isolates of F. solani for their 

potential to cause wilt disease on koa seedlings 

(Dudley and others 2007b). It appears that some 

strains of this fungus may be moderately virulent 

on koa, at least under greenhouse inoculation 

conditions. It is possible that F. solani may 

contribute to disease severity by increasing plant 

stresses, thus making trees more susceptible to 

virulent, wilt-inducing isolates strains of F. 

oxysporum. However, much more work is 

needed to evaluate possible synergistic roles of 

different Fusarium spp. in disease severity. 

This work increases the evidence that F. 

oxysporum is probably important in the etiology 

of wilt/dieback disease of Acacia koa trees. 

Strains of this fungus are usually present at 

relatively high levels within diseased trees, 

especially within root systems. Selected strains 

have exhibited high virulence in greenhouse 

inoculation tests on seedlings (Dudley and others 

2007a). We have not yet systematically sampled 

koa trees without disease symptoms to compare 

levels of Fusarium colonization of non-diseased 

with diseased trees. Such work would improve 

our understanding of the effects of Fusarium 

colonization on production of disease. 

ACKNOWLEDGEMENTS 

We acknowledge the USDA Forest Service, 

Forest Health Protection, Pacific Southwest 

Region and the Hawaii Agriculture Research 

Center for financial assistance and the assistance 

of J.B. Friday (University of Hawaii Cooperative 

Extension Service) for collecting some of the 

samples. 

LITERATURE CITED 

Anderson RC, Gardner DE, Daehler CC, 

Meinzer 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, Yeh A. 2007a. 

Comparative virulence of Hawaiian Fusarium 

oxysporum isolates on Acacia koa seedlings. 

USDA Forest Service, Northern Region, Forest 

Health Protection. Report 07- (In preparation). 
5

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

2007b. Pathogenicity of four Fusarium species 

on Acacia koa seedlings. USDA Forest 

Service, Northern Region, Forest Health 

Protection. Report 07-04.10p. 

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. 2004. 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., Dudley NS, Yeh A. 2006. 

Investigating koa wilt in Hawaii: Examining 

Acacia koa seeds and seedpods for Fusarium 

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

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. 

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, Marasas WFO . 1983. 

Fusarium species: an illustrated manual for 

identification. The Pennsylvania State 

University Press. University Park. 193p. 

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 

soilborne plant pathogens. Physiological and 

Molecular Plant Pathology 62:87-98. 

Yang B, James R, Uchida J, Zhong S. 2007. 

Molecular characterization of Fusarium 

oxysporum isolates from koa (Acacia koa) in 

Hawaii. Phytopathology 97:S127. 
6

Table 1. Root colonization of Acacia koa trees exhibiting wilt/dieback symptoms with Fusarium spp. 

Fine Roots Secondary Roots1 Tertiary Roots2 Number of Trees All Roots 

Sampled 46 33 18 18 

Pieces Sampled 763 600 430 1793 

Fusarium spp. Percent Colonization 

oxysporum 44.4 29.7 23.7 34.5 

solani 16.2 8.3 6.5 11.3 

graminearum 2.1 0 0.9 1.1 

semitectum 0.6 1.0 1.2 0.9 

lateritium 0.6 0.5 2.1 0.9 

sambucinum 0.9 0.5 1.2 0.8 

avenaceum 1.6 0 0 0.7 

equiseti 1.2 0 0 0.5 

subglutinans 0.8 0.3 0 0.4 

proliferatum 0.9 0 0 0.4 

acuminatum 0.8 0 0 0.3 

sporotrichioides 0.1 0 0.5 0.2 

All Fusarium 66.6 38.0 34.9 49.4 

No Fungi 0 19.5 22.6 11.9 

1

Roots adjoining fine feeder roots 

2

Larger roots (usually a few cm in diameter) connected to secondary roots 
7

Table 2. Stem and branch colonization of Acacia koa trees exhibiting wilt/dieback symptoms with 

Fusarium spp. 

Inner1 Outer2 Number of Trees All Samples 

43 43 43 

Pieces Sampled 596 523 1119 

Fusarium spp. Percent Colonization 

solani 16.1 17.2 16.6 

oxysporum 8.0 7.6 7.9 

proliferatum 4.0 4.2 4.1 

avenaceum 0.2 1.1 0.6 

graminearum 0.5 0.2 0.4 

subglutinans 0.3 0 0.2 

acuminatum 0.3 0.2 0.2 

sambucinum 0 0.4 0.2 

chlamydosporum 0.2 0 0.1 

semitectum 0.3 0 0.1 

decemcelluare 0 0.2 0.1 

sporotrichioides 0 0.2 0.1 

All Fusarium 29.5 30.8 30.1 

No Fungi 41.1 45.1 43.0 

1

Within the center of stems and branches, usually adjacent to a central core of green-black staining 

2

Adjacent to and inside the cambium near the bark 
8

Table 3. Colonization of Acacia koa trees exhibiting wilt/dieback symptoms with Fusarium spp. 

Number of Trees Sampled 52 

Number of Wood Pieces Sampled 29121

Fusarium Species Percent Colonization 

oxysporum 24.3 

solani 13.5 

proliferatum 1.8 

graminearum 0.8 

avenaceum 0.6 

semitectum 0.6 

sambucinum 0.6 

lateritium 0.6 

subglutinans 0.3 

acuminatum 0.3 

equiseti 0.3 

sporotrichioides 0.1 

chlamydosporum 0.03 

decemcelluare 0.03 

All Fusarium Species 42.0 

No Fungi 23.9 

1

Includes pieces from roots, stems and branches 

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

Coeur d’Alene, ID 83815; rjames@fs.fed.us. N.S. Dudley is Forester; Hawaii Agriculture Research 

Center, 99-193 Aiea Heights Drive, Aiea, HI 96701; NDudley@harc-haspa.com. A. Yeh is Plant 

Propagation Specialist, Hawaii Agriculture Research Center, 942 W. Kawailani Street, Hilo, HI 96778; 

ayeh@hawaii.rr.com.