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Department of Plant Pathology, Pennsylvania State University, University Park, Pennsylvania 16802
Melanie L. Lewis Ivey
Department of Plant Pathology, Ohio State University, Ohio Agricultural Research and Development Center, Wooster, Ohio 44691
Georgina Hakiza
Coffee Research Institute (CORI), P.O.Box 185, Mukono-Kituza, Uganda
Jean H. Juba
Department of Plant Pathology, Pennsylvania State University, University Park, Pennsylvania 16802
Sally A. Miller
Department of Plant Pathology, Ohio State University, Ohio Agricultural Research and Development Center, Wooster, Ohio 44691
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ABSTRACT |
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Tracheomycosis or coffee wilt has emerged as a major disease of robusta coffee in Uganda in the past 10 years. Coffee wilt historically has been associated with Fusarium xylarioides Steyaert (teleomorph Gibberella xylarioides Heim and Sacc.), a species that has been classified as a member of Fusarium section Lateritium. We investigated the molecular phylogenetics of fusarial coffee wilt isolates by generating partial DNA sequences from two protein coding regions, translation elongation factor 1-
and beta-tubulin, in 36 isolates previously identified as F. xylarioides and related fusaria from coffee and other woody hosts, as well as from 12 isolates associated with a current coffee wilt outbreak in Uganda. These isolates fell into two morphologically and phylogenetically distinct groups. The first group was found to represent previously unidentified members of the Gibberella fujikuroi species complex (GFC), a clade that replaces the artificial Fusarium section Liseola. This group of isolates fit the original description of F. xylarioides, thus connecting it to the GFC. The second group, which was diverse in its morphology and DNA sequences, comprised four distinct lineages related to Fusarium lateritium. Our finding of unrelated species associated with coffee wilt disease has important implications regarding its epidemiology, etiology and control.
Key words: coffee wilt, Fusarium lateritium, Gibberella fujikuroi, section Liseola
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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listed this species as "insufficiently documented," and stated that isolates from diseased coffee plants from Ethiopia were similar to a "female" strain described by Booth (1971), but most fusariologists (Booth 1971, Gerlach and Nirenberg 1982, Gordon 1952) recognized the species and placed it in Fusarium section Lateritium. However, because section Lateritium was found to be nonmonophyletic in this study, we have elected to use "Lateritium clade" in reference to those taxa that appear to be monophyletic in this study. So-called "male" and "female" isolates described by Booth (1971) were thought to exhibit dimorphism in conidial and colony characters, with "male" isolates producing thin, elongated, 5–7-septate sporodochial conidia (often referred to as macroconidia) typical of Lateritium clade fusaria, and "female" isolates producing shorter, highly curved, 0–3-septate conidia. In diseased trees, the sexual stage Gibberella xylarioides Heim and Sacc. forms readily in the cracks of stem bark from the collar region of dying trees. Booth (1971) noted that G. xylarioides perithecia form in culture when opposing mating-types are brought together, thus inferring that the fungus is heterothallic. The existence of "male" and "female" strains was questioned by Gerlach and Nirenberg (1982), whose description of F. xylarioides included only the "female" conidial morphology, as did the original description of this species (Steyaert 1948).
Members of the Lateritium clade have been associated with diseases of coffee. F. stilboides Wollenweber causes bark and fruit rots of citrus and coffee (Gerlach and Nirenberg 1982). This species was considered a synonym of F. lateritium by a number of authors (Bilai 1955, Nelson et al 1983, Snyder and Hansen 1945). F. lateritium var. longum also has been associated with rot diseases of citrus and coffee. This taxon was considered a synonym of F. stilboides by some authors (Bilai 1955, Booth 1971) and of F. lateritium by others (Gordon 1952, Snyder and Hansen 1945, Subramanian 1971), while Gerlach and Nirenberg (1982) maintained it as a unique taxon. Producing long, thin sporodochial conidia that are often five or more septate, these species possess a morphology more typical of F. lateritium and its presumed relatives than F. xylarioides as described by Steyaert (1948) and Gerlach and Nirenberg (1982). Gerlach (1978) proposed that "male" isolates were mutants or variants of F. stilboides.
The Gibberella fujikuroi species complex (GFC) is a monophyletic and diverse group of approximately 50 phylogenetic species, many of which remain unnamed (O’Donnell et al 1998a, O’Donnell et al 2000). This complex is divided into three subclades, often referred to as African, American and Asian, based on the putative geographic origin of most of the species within them. The GFC approximates Fusarium section Liseola as defined by a number of authors, except it contains a number of taxa that produce chlamydospores. However, as defined morphologically, section Liseola is paraphyletic because it excluded chlamydospore-producing taxa. The GFC includes a number of diverse plant pathogens, including fusaria causing ear rot of corn (F. verticillioides, F. proliferatum, F. subglutinans), pitch canker of pine (F. circinatum), mango malformation (F. mangiferae, F. sterilihyphosum (Britz et al 2002)) and the gibberellin-induced bakanae disease of rice (F. fujikuroi), as well as numerous species that produce mycotoxins such as fumonisins and monilformin (Marasas et al 2001). A number of species recently have been recognized as members of the African clade of the GFC based on phylogenetic evidence (O’Donnell et al 1998a), including F. udum, which was placed in section Elegans by some authors (Wollenweber and Reinking 1935, Subramanian 1971, Gerlach and Nirenberg 1982) and in section Lateritium by others (Booth 1971). F. udum causes vascular wilt diseases of woody hosts, particularly Cajanus cajan (pigeon pea). It produces conidia with strongly curved or hooked apical cells, along with longer, up to 5-septate conidia, from densely and irregularly branched conidiophores. The apparently heterothallic sexual stage of F. udum, G. indica B Rai & RS Upadhyay has been observed on the roots and collars of wilted pigeon pea plants (Rai and Upadhyay 1982). In addition, F. denticulatum, originally identified as F. lateritium from sweet potato, also is nested within the African clade of the GFC. In his description of G. xylarioides Heim noted similarities between the F. xylarioides anamorph and the strongly falciform conidia of ‘F. moniliforme var. anthophilum’ (now recognized as F. anthophilum) as illustrated by Wollenweber and Reinking (1935) and thus concluded that F. xylarioides belonged in Section Liseola (Heim 1950). This observation received little recognition perhaps in part due to the described production of chlamydospores by F. xylarioides, which would preclude its inclusion in section Liseola. In fact a number of diverse fusaria are known to produce highly curved conidia (e.g., F. inflexum, F. avenaceum var. volutum, insect-associated species such as F. larvarum and members of the GFC such as F. succisae and F. udum), and later descriptions of F. anthophilum do not depict the highly curved conidia illustrated in Wollenweber’s and Reinking’s (1935) monograph (Gerlach and Nirenberg 1982, Nelson et al 1983).
Two endemic species of coffee are cultivated in eastern Africa (Simpson and Ogorzaly 2001): Coffea canephora Pierre (robusta) and C. arabica L. (arabica), with robusta accounting for up to 90% of production in Uganda. Before the appearance of coffee wilt in 1993 only minor diseases were associated with robusta in Uganda (Hakiza and Mwebesa 1997). Coffee wilt can attack all stages of growth, from seedlings to mature plants, and infected plants show 100% mortality. Symptoms include wilting, defoliation and blue-black streaks in the wood and under the bark (Flood 1996, Guillemat 1946, Pochet 1988, Waller and Brayford 1990). On a multistemmed coffee plant, the external symptoms occur sequentially until all stems/branches are killed. Coffee berries on affected plants ripen prematurely and dry up but remain attached to the branches. Brown sunken lesions at the end of the stalk bearing berries also may be observed. Although pruned plants or stumps may sprout new suckers that appear healthy, the plants do not recover.
We used molecular phylogenetics to investigate the identity of fusaria associated with coffee wilt. Toward this end we determined partial DNA sequences from two protein coding regions, translation elongation factor 1- (tef) and beta tubulin (benA), from 36 putative Lateritium clade fusaria previously identified as F. xylarioides, F. stilboides or F. lateritium, associated with coffee or other woody hosts, and analyzed them phylogenetically.
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MATERIALS AND METHODS |
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). For the Ugandan isolates, stem and/or branch tissue showing typical coffee wilt symptoms were collected from farmers’ fields in nine districts where robusta is a major cash crop. Isolation of the pathogen was initiated within 7 d of sample collection. Samples were cut into 15–20 cm lengths and placed in envelopes for transport to the laboratory. Samples (20 mm) from the margins of infected tissue were split lengthwise, surface sterilized in 2% (v/v) Jik (3.5 % sodium hypochlorite; Reckitt Benckiser East Africa Ltd., Nairobi, Kenya) for 3 min, rinsed three times in sterile distilled water, placed onto 2% tap water agar and incubated at 25 ± 2 C under fluorescent light 4 d. Cultures were purified by transferring 1 cm2 plugs from the growing edges to potato-dextrose agar (PDA) and then purified by single-spore isolation. Other isolates were obtained from the Fusarium Research Center (FRC) Culture Collection at Pennsylvania State University.
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). Isolates include those recently collected from an outbreak of coffee wilt on robusta in Uganda, as well as the ex-type culture of G. xylarioides (CBS 258.52 = NRRL 25486) and other isolates in the FRC collection that previously were identified as F. lateritium, F. stilboides or F. xylarioides from coffee, citrus and in one case soil. All cultures were derived from single conidia and were stored on synthetic low-nutrient agar (SNA) (Nirenberg 1976) slants at 4 C or lyophilized as described by Nelson et al (1983). Isolates from Uganda were deposited in the FRC collection under accession numbers L-388–397, L-399 and L-400. To study the morphological characteristics of the Ugandan isolates, they were grown on potato-dextrose agar slants (PDA; Difco Laboratories, Detroit, Michigan) and SNA and carnation leaf agar (CLA) plates (Fisher et al 1982), under black and white fluorescent lights with a 12 h photoperiod.
DNA extraction, PCR and nucleotide sequencing.— –
For DNA extraction, mycelium was transferred from PDA to 250 mL Erlenmeyer flasks containing potato-dextrose broth (Difco Laboratories, Detroit, Michigan). After 5 d growth at 25 ± 2 C without shaking, mycelium was harvested and DNA was extracted from the mycelium using a DNeasy Plant Minikit (Qiagen, Valencia, California). Polymerase chain reaction (PCR) amplification of the beta-tubulin gene region and sequencing of the 1000 bp amplicon was performed using primers bena-T1 and bena-T22 (O’Donnell and Cigelnik 1997). An approximately 690 bp portion of the translation elongation factor 1- gene also was amplified and sequenced using primers ef1 and ef2 (O’Donnell et al 1998b). PCR for both primer sets was performed in a PTC-100 Programmable thermocycler (MJ Research Inc., Waltham, Massachusetts) using these conditions: 1x PCR buffer, 2 mM MgCl, 0.2 mM dNTP, 0.2 µM of each primer, 0.01 U Taq polymerase and 50 ng template DNA. These PCR parameters were used: 2 min at 94 C, followed by 35 cycles of 1 min at 94 C, 1 min at 53 C and 1 min at 72 C, followed by 5 min at 72 C. DNA sequencing was performed in both directions, using the same primers as used in PCR, except that the reverse primer bena-T2 was used instead of bena-T22 (O’Donnell and Cigelnik 1997). Sequences were generated using an Applied Biosystems Prism BigDye sequencing kit according to manufacturer’s instructions, except that 8 µL volumes were used, and analyzed on an Applied Bio-systems 3730 DNA Analyzer (PE Applied Biosystems, Foster City, California). Sequences were edited manually and deposited in GenBank under accession numbers AY707101–AY707173.
DNA sequence and phylogenetic analysis.— –
BLAST (Altschul et al 1990) was used to perform similarity searches comparing fusarial coffee wilt sequences with those in the GenBank database and local sequence databases, as described in Geiser et al (2004). Based on BLAST results, sequences were aligned by eye into files containing DNA sequences representing the phylogenetic breadth of the GFC (O’Donnell et al 2000), or to a new alignment of Lateritium clade sequences. Where identical sequences were found among isolates, a single representative was included in the phylogenetic analysis, and then its haplotype mates were added back to the inferred phylogenetic tree at the same position. Alignments were deposited in TreeBASE under accession number SN 1981. Maximum likelihood (ML) analyses were performed using the PAUP* phylogenetics package (beta versions 4.0b9–11:(Swofford 2003)). ModelTest version 3.0.6 (Posada and Crandall 1998) was used in conjunction with PAUP* to determine appropriate evolutionary models for ML analysis, which identified these models for the five different datasets analyzed phylogenetically: Lateritum clade tef (TrN model (Tamura and Nei 1993) with gamma shape parameter [G] set at 0.3229); Lateritum clade benA (HKY model (Hasegawa et al 1985) with transition/transversion ratio (TRatio) set at 2.5363 and G = 0.5481); GFC tef (TrN model with G = 0.3671); GFC benA (HKY model with TRatio = 2.4578 and G = 0.4240); and GFC combined (TrN model with proportion of invariable sites [I] set at 0.3221 and G = 0.7292). All models used observed base frequencies. For maximum likelihood analyses, alignment gaps were considered missing sites and heuristic searches were performed using random sequence addition and TBR branch swapping. Bootstrapping was performed using maximum parsimony as the criterion, using random sequence addition, and MAXtrees set at 10 000.
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RESULTS |
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and Steyaert (1948), and of the "female" strain of F. xylarioides described by Booth (1971). Colonies of isolates L-388–397, L-399 and L-400 from Uganda were examined morphologically on PDA, SNA and CLA. Colonies on PDA and SNA were pale to colorless after 3 d and pale orange 4–14 d later depending on the isolate. The orange pigmentation was more intense in the center of the colony and faded to pale orange to white at the growing margin. Colony appearance on the reverse was the same. Orange pionnotes were produced in concentric rings by Day 4. On PDA and SNA, hyaline mycelium was sparse and oppressed. On SNA, 0 –3-septate conidia, always curved in the apical cell and often in the basal cell (FIG. 1), were formed from irregularly branched conidiophores in sporodochia or pionnotes. On CLA, isolates formed abundant protoperithecia and sporodochia on carnation leaf pieces, and abundant, usually 0 –1-septate conidia with an exaggerated curve were produced on the agar surface, with the tips of the apical and basal cells sometimes nearly touching each other (FIG. 2). Few aerial mycelia and no chlamydospores were observed after 21 d. Additional isolates from African coffee, L-96, L-102, L-125, L-126 and L-128, were similar morphologically in terms of conidial morphology and protoperithecium production but showed differences in colony color on PDA, ranging from pale, off-white shades to orange.
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). DNA sequence analysis.— – BLAST searches using partial translation elongation factor 1-alpha (tef) sequences showed that 18/36 isolates, comprising the ex-type of G. xylarioides CBS 258.52, the 12 isolates from Uganda (L-388–397, L-399–400) and isolates L-96, L-102, L-125, L-126, and L-128, appeared to be part of the Gibberella fujikuroi species complex. These isolates have the F. xylarioides "female" strain morphology. Two tef alleles were observed among these isolates that differed at 17/690 sites (2.5%). Partial beta-tubulin (benA) sequences of these isolates, however, were identical and also showed a clear grouping with the GFC.
Phylogenetics of isolates from coffee associated with the GFC.— –
tef and benA sequences from GFC-associated coffee wilt isolates were added to DNA sequence alignments representing the known phylogenetic breadth of the GFC (O’Donnell et al 1998a, O’Donnell et al 2000) and subjected to phylogenetic analysis using maximum likelihood as the criterion. Heuristic searches identified a single tree (FIG. 7A). The two tef alleles grouped in the African clade of the GFC, in a 98% bootstrap-supported clade with F. udum, F. phyllophilum and an undescribed Fusarium sp. NRRL 26064 from sorghum seed. However they were not inferred to be one another’s closest relatives, given that the tef allele from the ex-type and isolate L-102 were more closely related to an undescribed Fusarium sp. NRRL 26064 than to the tef allele from other coffee wilt isolates. As is often the case with the tef locus alone, the African clade sensu O’Donnell (1998a) was not inferred to be monophyletic.
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). This tree also indicated a close phylogenetic relationship among these coffee wilt isolates and F. udum, F. phyllophilum and Fusarium sp. NRRL 26064, with 100% bootstrap support.
Because no strongly supported discordances were observed between the two gene trees, the data were combined and subjected to a phylogenetic analysis. This analysis yielded a single tree (FIG. 8), grouping all GFC coffee wilt isolates with F. udum, F. phyllophilum and the undescribed Fusarium sp. NRRL 26064 with 100% bootstrap support. Monophyly of the GFC coffee wilt isolates was inferred based on the combined data, albeit with weak (72%) bootstrap support. The beta-tubulin and combined trees both showed the three major biogeographic clades proposed by O’Donnell (1998a) to be monophyletic, including the African clade (FIGS. 7, 8).
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Maximum likelihood analysis of the tef and benA datasets for the 18 Lateritium clade-associated coffee isolates, with isolates of F. oxysporum and F. inflexum used as outgroup sequences, yielded trees with nearly identical topologies and four strongly supported groups (FIG. 9). These groups did not correlate well with the previous species identifications. The first group (I) consisted of coffee isolates from around the world, including western, southern and eastern Africa, Brazil, New Guinea and New Caledonia. These isolates had been identified previously either as F. lateritium, F. stilboides or F. xylarioides. The next two groups (IIA and IIB) were closely related and consisted of citrus isolates from New Zealand, Philippines and New Caledonia, and Philippines soil and coffee, previously identified either as F. lateritium or ‘F. lateritium/stilboides’. The fourth distinct group (III) consisted of two isolates originally identified as F. lateritium that had nearly identical sequences from coffee in Papua, New Guinea. The tef allele shared by these two isolates was more similar to that F. lateritium reference isolate L-55 than those of other coffee isolates, but they differed from L-55 at ~2.3% of the tef sites.
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Isolates in Groups IIA and IIB also produced a red pigment on PDA, with pinkish aerial mycelium. Few aerial conidia were produced. Included in Group IIA was isolate L-405 (= NRRL 25485 = BBA 63887) from citrus in New Zealand. This isolate was described as a member of F. stilboides by Gerlach and Nirenberg (1982), and our observations of Groups IIA and IIB were consistent with their description of this species.
Isolates in Group III produced an orange pigment on PDA and some with aerial mycelium. Zero to two-septate aerial conidia were produced abundantly from a mixture of mono- and polyphialides, in addition to long, thin sporodochial conidia typical of F. lateritium.
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DISCUSSION |
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with strong support in both genes, individually and in combination. Both genes also indicated F. udum, F. phyllophilum and an undescribed Fusarium sp. NRRL 26064 were their closest relatives, with consistently high bootstrap support. Included in this group was the ex-type of G. xylarioides (CBS 258.52 = NRRL 25486), the assumed sexual stage of this fungus. Morphologically these isolates matched the original description of F. xylarioides (Steyaert 1948) as well as that of the so-called "female" strains of this species described by Booth (1971), producing abundant 0–3-septate, highly curved conidia. On CLA, these isolates produced abundant protoperithecia, suggesting the possibility of an active sexual stage, consistent with F. xylarioides whose Gibberella sexual state is often observed in the field. We conclude that F. xylarioides Steyaert is a member of the GFC. This is consistent with the placement of F. xylarioides in section Liseola (Heim 1950).
One of the closest relatives of F. xylarioides, F. udum, is also an unusual member of the GFC. It too produces abundant 0–3-septate conidia that often are curved, although it also produces larger, 4–5-septate conidia. It also produces densely and irregularly branched conidiophores that are reminiscent of F. xylarioides (Gerlach and Nirenberg 1982). This species was assigned to GFC based on molecular phylogenetics (O’Donnell et al 1998a). In previous treatments, F. udum was placed in sections Eupionnotes (Wollenweber and Reinking 1935), Lateritium (Booth 1971) and Elegans (Gerlach and Nirenberg 1982). Like F. xylarioides, F.udum also causes a vascular wilt of a woody host, Cajanus cajan (pigeon pea or red gram), and has been observed to produce its sexual stage in the field (Rai and Upadhyay 1982). Although GFC comprises a diverse assemblage of plant pathogens, these two species are the only members of this complex associated with true vascular wilt diseases. Fusarial vascular wilt diseases are most commonly associated with the F. oxysporum complex, which approximates Fusarium section Elegans. Traditional sections Elegans and Liseola were distinguished by morphological characters that included the presence of chlamydospores (never observed in section Liseola) and the production of polyphialides (never observed in section Elegans). Molecular phylogenetic analyses have demonstrated that both of these characters are likely plesiomorphic in the F. oxysporum and G. fujikuroi species complexes (Geiser et al 2001, O’Donnell et al 1998a), as now appears to be the case with the ability to cause true vascular wilt diseases.
Isolates identified as F. xylarioides all had identical benA sequences, but two distinct tef alleles were observed. The two tef alleles were 2.5% divergent and were not inferred as monophyletic (FIG. 7A). This finding leaves open the distinct possibility that F. xylarioides as discussed here might comprise multiple related cryptic species that the benA locus failed to resolve. The two tef alleles observed were associated with geography, with L-102 and the G. xylarioides ex-type isolate CBS 258.52 possessing one allele and both coming from western Africa (Guinea and Ivory Coast respectively) and the remaining isolates with the alternate allele coming from countries in eastern Africa (Ethiopia, Uganda). Additional studies are under way to explore species boundaries in the group, using morphology, mating, and multilocus DNA sequence data.
O’Donnell et al (1998a) defined the "African" clade of the GFC based on the large number of species in the clade with African hosts, positing an African origin for this diverse group. There are exceptions to this trend, particularly F. verticillioides, which is known mostly in association with maize (American in origin) and is cosmopolitan in its distribution. All isolates of authentic F. xylarioides analyzed in this study were African in origin, and robusta and arabica coffee varieties are native to Africa (Simpson and Ogorzaly 1995), overall suggesting a common biogeographic origin for F. xylarioides and its only known host.
Coffee-associated fusaria related to Fusarium lateritium.— –
The remaining 18 isolates studied appeared to be diverse, but all showed a phylogenetic connection to the Lateritium clade. The majority (12/18) of isolates fell into one major clade, Clade I (FIG. 9A, B). These isolates did not exhibit morphological and molecular phylogenetic characteristics that allow a strong connection to a described species beyond F. lateritium broadly defined, and might represent a previously unnamed taxon. However all these geographically diverse isolates came from coffee, either from berries, bark or vascular tissue, and had been identified previously as F. lateritium, F. stilboides or F. xylarioides.
Isolate L-96 (= MRC 1853), an authentic member of F. xylarioides as delimited here, was derived from a single macroconidium from a culture of BBA 62458, an isolate considered in the description of F. xylarioides contained in Gerlach and Nirenberg (1982). Derived from a single spore from the same culture at the same time was isolate L-95 (= MRC 1845). Isolates L-95 and L-96 are distinct morphologically, with the latter showing characteristics of the "female" morphology described by Booth (1971). They also are distinct phylogenetically, with L-95 being a member of Clade I of the Lateritium clade and L-96 being a member of the GFC.
Clades IIA and IIB in the Lateritium clade include isolates from citrus and coffee in New Caledonia, New Zealand and the Philippines. Isolate L-405 (= BBA 63887, NRRL 25485, CBS 746.79) in Clade IIA was described by Gerlach and Nirenberg (1982) as a member of F. stilboides var. stilboides, which is known to cause bark and fruit rot diseases of citrus and coffee in the tropics and subtropics (Booth 1971, Gerlach and Nirenberg 1982). These isolates were variable morphologically but generally fit the description of F. stilboides (Gerlach and Nirenberg 1982). Relationships of the five isolates comprising Clades IIA and IIB were identical within the tef and benA gene trees. However, because these clades differed at up to 4.7% of their nucleotide sites over both genes, we suspect that they comprise F. stilboides and one or more cryptic species.
Clade III was represented by two nearly identical isolates, L-110 and L-112, both isolated from perithecia on coffee twigs in Papua New Guinea. These isolates produced long, often six or more septate sporodochial conidia reminiscent of F. lateritium. They showed a strongly supported connection to F. lateritium isolate L-55 in their tef alleles (98% bootstrap support). However these two isolates differed from L-55 by 2.6%, suggesting these two groups are possibly not conspecific.
Unlike the well characterized GFC (O’Donnell et al 1998a, O’Donnell et al 2000), no comprehensive multilocus molecular phylogenetic studies have been performed on the Lateritium clade. The results here suggest that there might be a number of cryptic species within this group, warranting additional scrutiny of isolates from hosts other than coffee and additional substrates.
Sexual stages and associated dimorphism.— –
Previous reports of sexual dimorphism in F. xylarioides appear to be a matter of mistaken identity. So-called "female" isolates clearly correspond to F. xylarioides, while "male" isolates correspond to members of the Lateritium clade. Booth’s (1971) description of the so-called "male" isolate of F. xylarioides included long, thin, 5–7 or more septate sporodochial conidia with a beaked basal cell, a characteristic we observed in all of the Lateritium clade studied from coffee. Gerlach (1978) stated that the "male" strain corresponded to a mutant or variety of F. stilboides. Different degrees of male and female sexual tendency frequently are observed in isolates of Fusarium species (Leslie and Klein 1996), but this is not known to correspond to dimorphism in morphological characteristics. Confusion about sexual dimorphism may have stemmed from the study of mixed cultures, as appears to have occurred where cultures of L-95 (GFC) and L-96 (Lateritium clade) apparently were derived from a single isolate. This might be the result of frequent co-occurrence of these very different fusaria on coffee plants, which in turn might lead to frequent co-isolation.
Are G. xylarioides and F. xylarioides the same species?— –
In addition to reported sexual dimorphism, a frequently observed sexual stage, Gibberella xylarioides, has been assigned to F. xylarioides. Previous evidence suggested that G. xylarioides indeed is the teleomorph of F. xylarioides. Gerlach and Nirenberg (1982) noted that isolates derived from single ascospores corresponded consistently with their concept of F. xylarioides and the so-called female morphology. In this study, the ex-type culture of G. xylarioides was identical in sequence to L-102 (= BBA 62721 = CBS 749.79), an isolate with clear F. xylarioides morphology, further suggesting a connection. However we have not gained access to type material associated with F. xylarioides and can only note similarity between the observed morphology of F. xylarioides as delimited here and the illustrations in its original description (Steyaert 1948). The discovery of two divergent tef alleles that might track with Western versus Eastern African origins, suggesting a possible cryptic species boundary, leaves open the possibility that the two names refer to two different species. The type of F. xylarioides was taken from a diseased coffee plant sent from Bangui, Central African Republic, and Steyaert inferred a connection between this collection and the fungus causing coffee wilt disease, in what was then French Equatorial Africa as well as the border region between Congo and Sudan (Steyaert 1948). Because no connection can be made between the type of F. xylarioides and these two potential cryptic species, we prefer to maintain the anamorph-teleomorph connection between G. xylarioides and F. xylarioides based on previous and current morphological observations.
Fusaria from coffee and coffee wilt.— –
The question is left open as to the differential roles of F. xylarioides and true Lateritium clade fusaria in coffee wilt. It appears that both groups can be isolated from coffee wilt-inflicted plants. Isolates of the true F. xylarioides from Uganda cause coffee wilt when inoculated onto coffee plants, and the same fungus was cultured successfully from the diseased plants (Lewis Ivey et al 2003). Future studies should focus on the differential roles of Lateritium clade and GFC fusaria in this disease, both alone and in co-infection. Lateritium clade fusaria are known widely as wound pathogens of woody hosts, causing a wide variety of diseases (Gerlach and Nirenberg 1982). Members of this clade might be secondary invaders or opportunists on dead or dying plant parts that have been stricken with coffee wilt by infection with F. xylarioides. In addition the recognition of F. xylarioides as a member of the GFC might aid research on this pathogen, opening up the potential for knowledge transfer from its genetically well studied relatives such as F. verticillioides (Kroken et al 2003, Schoch et al 2003).
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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1 Corresponding author. E-mail: dgeiser{at}psu.edu
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LITERATURE CITED |
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