J. C. Correll

Characterization of diversity in Colletotrichum acutatum sensu lato by sequence analysis of two gene introns, mtDNA and intron RFLPs, and mating compatibility

A diverse collection of isolates identified as Colletotrichum acutatum, including a range of fruit-rot and foliar pathogens, was examined for mtDNA RFLPs and RFLPs and sequence variation of a 900-bp intron of the glutamine synthetase (GS) gene and a 200-bp intron of the glyceraldehyde-3-phosphate dehydrogenase (GPDH) gene. RFLPs of mtDNA, RFLPs of the 900-bp GS intron and sequence analysis of each intron identified the same seven distinct molecular groups, or clades, within C. acutatum sensu lato. Sequence analysis produced highly concordant tree topologies with definitive phylogenetic relationships within and between the clades. The clades might represent phylogenetically distinct species within C. acutatum sensu lato. Mating tests also were conducted to assess sexual compatibility with tester isolates known to outcross to form the teleomorph Glomerella acutata. Mating compatibility was identified within one clade, C, and between two phylogenetically distinct clades, C and J4. The C clade represented isolates from a wide range of hosts and geographic origins. J4 clade contained isolates from Australia or New Zealand recovered from fruit rot and pine seedlings with terminal crook disease. That isolates in two phylogenetically distinct clades were capable of mating suggests that genetic isolation occurred before reproductive isolation. No other isolates were sexually compatible with the mating testers, which also were in groups C and J4. Certain clades identified by mtDNA and intron analysis (D1, J3 and J6) appeared to represent relatively host-limited populations. Other clades (C1, F1 and J4) contained isolates from a wide range of hosts. Isolates described as C. acutatum f. sp. pineum were clearly polyphyletic.

One fungus, one name

In this letter, we advocate recognizing the genus Fusarium as the sole name for a group that includes virtually all Fusarium species of importance in plant pathology, mycotoxicology, medicine, and basic research. This phylogenetically guided circumscription will free scientists from any obligation to use other genus names, including teleomorphs, for species nested within this clade, and preserve the application of the name Fusarium in the way it has been used for almost a century. Due to recent changes in the International Code of Nomenclature for algae, fungi, and plants, this is an urgent matter that requires community attention. The alternative is to break the longstanding concept of Fusarium into nine or more genera, and remove important taxa such as those in the F. solani species complex from the genus, a move we believe is unnecessary. Here we present taxonomic and nomenclatural proposals that will preserve established research connections and facilitate communication within and between research communities, and at the same time support strong scientific principles and good taxonomic practice.

One fungus, one name: defining the genus Fusarium in a scientifically robust way that preserves longstanding use.

In this letter, we advocate recognizing the genus Fusarium as the sole name for a group that includes virtually all Fusarium species of importance in plant pathology, mycotoxicology, medicine, and basic research. This phylogenetically guided circumscription will free scientists from any obligation to use other genus names, including teleomorphs, for species nested within this clade, and preserve the application of the name Fusarium in the way it has been used for almost a century. Due to recent changes in the International Code of Nomenclature for algae, fungi, and plants, this is an urgent matter that requires community attention. The alternative is to break the longstanding concept of Fusarium into nine or more genera, and remove important taxa such as those in the F. solani species complex from the genus, a move we believe is unnecessary. Here we present taxonomic and nomenclatural proposals that will preserve established research connections and facilitate communication within and between research communities, and at the same time support strong scientific principles and good taxonomic practice.

Mapping Quantitative Trait Loci Responsible for Resistance to Sheath Blight in Rice

Rice sheath blight (ShB), caused by the soilborne pathogen Rhizoctonia solani, annually causes severe losses in yield and quality in many rice production areas worldwide. Jasmine 85 is an indica cultivar that has proven to have a high level of resistance to this pathogen. The objective of this study was to determine the ability of controlled environment inoculation assays to detect ShB resistance quantitative trait loci (QTLs) in a cross derived from the susceptible cv. Lemont and the resistant cv. Jasmine 85. The disease reactions of 250 F(5) recombinant inbred lines (RILs) were measured on the seedlings inoculated using microchamber and mist-chamber assays under greenhouse conditions. In total, 10 ShB-QTLs were identified on chromosomes 1, 2, 3, 5, 6, and 9 using these two methods. The microchamber method identified four of five new ShB-QTLs, one on each of chromosomes 1, 3, 5, and 6. Both microchamber and mist-chamber methods identified two ShB-QTLs, qShB1 and qShB9-2. Four of the ShB-QTLs or ShB-QTL regions identified on chromosomes 2, 3, and 9 were previously reported in the literature. The major ShB-QTL qShB9-2, which cosegregated with simple sequence repeat (SSR) marker RM245 on chromosome 9, contributed to 24.3 and 27.2% of total phenotypic variation in ShB using microchamber and mistchamber assays, respectively. qShB9-2, a plant-stage-independent QTL, was also verified in nine haplotypes of 10 resistant Lemont/Jasmine 85 RILs using haplotype analysis. These results suggest that multiple ShB-QTLs are involved in ShB resistance and that microchamber and mist-chamber methods are effective for detecting plant-stage-independent QTLs. Furthermore, two SSR markers, RM215 and RM245, are robust markers and can be used in marker-assisted breeding programs to improve ShB resistance.

Characterization of Glomerella acutata, the teleomorph of Colletotrichum acutatum

The association between the anamorph Colletotrichum acutatum and a new teleomorph, Glom- erella acutata, was documented. Fertile perithecia were produced in a series of crosses between self-ster- ile monoconidial strains of C. acutatum, demonstrat- ing a previously undescribed teleomorph directly as- sociated with the anamorph originally described byJ. H. Simmonds in 1965. Strains capable of crossing in- cluded the C. acutatum type specimen, originally iso- lated from papaya (Carica papaya) in Australia by Simmonds, and other archived and contemporary strains from several hosts from the United States, Aus- tralia and New Zealand. The resulting teleomorph, Glomerella acutata, was described and illustrated as a new species which can be distinguished from other taxa by a combination of features of the anamorph, the teleomorph, and mitochondrial and nuclear DNA. Perithecia of G. acutata were black-brown, pre- dominantly ampulliform, 125 x 312 ILm (mean 183 ILm) diam, ostiolate and periphysate. Asci were nar- rowly clavate and bore up to eight oblong to ellip- soidal ascospores which measured 8.5-(15.7)-25.1 X 3.1-(5.8)-8.1 ,um.

Characterization of Pyricularia grisea in the United States using independent genetic and molecular markers.

ABSTRACT A total of 540 isolates of Pyricularia grisea from rice in the United States were examined for vegetative compatibility, MGR586 DNA fingerprint diversity, and mating type based on hybridization with the mat1-1 and mat1-2 sexual mating type alleles. The collections contained both archived and contemporary field isolates representative of the known MGR586 lineages and races that occur throughout the United States. Complementary nitrate nonutilizing (nit) or sulfate nonutilizing (sul) mutants were used to assess vegetative compatibility in P. grisea. There was a complete correspondence between vegetative compatibility groups (VCGs), MGR586 lineage, and mating type among 527 contemporary isolates (collected between 1991 and 1997) from Arkansas, Louisiana, Missouri, Mississippi, and Texas; all isolates in MGR586 lineages A, B, C, and D belonged to VCGs US-01, US-02, US-03, and US-04, respectively. In addition, all isolates tested in VCGs US-01 and US-04 had the mat1-1 mating type allele whereas those in VCGs US-02 and US-03 had the mat1-2 allele. The strict association of independent markers during this sample period was consistent with a strictly asexual mode of reproduction. However, examination of archived isolates collected in the 1970s and 1980s and contemporary isolates revealed an incongruent relationship between the independent markers. MGR586 C and E isolates were vegetatively compatible which indicated that multiple robust MGR586 delineated lineages could be nested within certain VCGs. Although isolates in lineages C and E were vegetatively compatible, they were of opposite mating type. Several hypotheses, including recombination, could account for the incongruence between the various markers. Among the eight MGR586 lineages (A through H) that occur in the United States, all isolates in lineages A, D, E, G, and H had the mat1-1 allele, whereas isolates in lineages B, C, and F had the mat1-2 allele. Nit mutants can be recovered relatively easy from P. grisea and should allow large numbers of individuals within a population to be assessed for vegetative compatibility. VCGs may prove to be an effective multilocus marker in P. grisea. Thus, VCGs should be a useful means for characterizing genetic structure in populations of the rice blast fungus worldwide, provide a useful genetic framework to assist in interpreting molecular population data, and may provide insight into potential sexual or asexual recombination events.

Systematic Numbering of Vegetative Compatibility Groups in the Plant Pathogenic Fungus Fusarium oxysporum

First author: Plant Pathology Department, University of Florida, Gainesville 32611-0680; second author: Laboratoire de Recherches sur la Flore Pathogene du Sol, INRA, 17 rue Sully, B.V. 154

Diversification and evolution of the avirulence gene AVR-Pita1 in field isolates of Magnaporthe oryzae

Rice blast disease is the single most destructive plant disease that threatens stable rice production worldwide. Race-specific resistance to the rice blast pathogen has not been durable and the mechanism by which the resistance is overcome remains largely unknown. Here we report the molecular mechanisms of diversification and the instability of the avirulence gene AVR-Pita1 in field strains of Magnaporthe oryzae interacting with the host resistance gene Pi-ta and triggering race-specific resistance. Two-base-pair insertions resulting in frame-shift mutations and partial and complete deletions of AVR-Pita1 were identified in virulent isolates. Moreover, a total of 38 AVR-Pita1 haplotypes encoding 27 AVR-Pita1 variants were identified among 151 avirulent isolates. Most DNA sequence variation was found to occur in the exon region resulting in amino acid substitution. These findings demonstrate that AVR-Pita1 is under positive selection and mutations of AVR-Pita1 are responsible for defeating race-specific resistance in nature.

Clarification of the Etiology of Glomerella Leaf Spot and Bitter Rot of Apple Caused by Colletotrichum spp. Based on Morphology and Genetic, Molecular, and Pathogenicity Tests

ABSTRACT Morphological characteristics and vegetative compatibility groups (VCGs) of 486 isolates of Glomerella cingulata, Colletotrichum gloeosporioides, and C. acutatum collected from apple leaves with Glomerella leaf spot (GLS) symptoms and fruit with bitter rot symptoms in the United States and Brazil were studied. From this collection, 155 isolates of G. cingulata (93 from fruit, 61 from leaves, and 1 from buds), 42 isolates of C. gloeosporioides from fruit, and 14 isolates of C. acutatum (10 from fruit and 4 from leaves) were studied using mitochondrial (mt)DNA restriction fragment length polymorphism (RFLP) haplotypes. A subset of 24 isolates was studied by examining the sequence of a 200-bp intron of the glyceraldehyde 3-phosphate dehydrogenase (GDPH) nuclear gene. In addition, 98 isolates were tested for pathogenicity on leaves of cvs. Gala and Golden Delicious in the greenhouse, and 24 isolates were tested for pathogenicity on fruit of cv. Gala in growth chambers. In total, 238 and 225 isolates of G. cingulata were separated into four distinct morphological types and six VCGs, respectively. Five morphological types and six VCGs were identified among 74 and 36 isolates of C. gloeosporioides, respectively. Three morphological types and four VCGs were identified among 74 and 23 isolates of C. acutatum, respectively. Seven different mtDNA RFLP haplotypes were observed within isolates of G. cingulata, two within isolates of C. gloeosporioides, and two within isolates of C. acutatum. Phylogenetic trees, inferred based on maximum likelihood and maximum parsimony methods using the intron sequence, produced similar topologies. Each species was separated into distinct groups. All isolates tested were pathogenic on fruit, though only isolates with specific VCGs and haplotypes were pathogenic to leaves. Vegetative compatibility was a better tool than molecular characters for distinguishing isolates of G. cingulata pathogenic on both leaves and fruit from the ones pathogenic only on fruit. Isolates of G. cingulata capable of causing both GLS and bitter rot were included in haplotypes and groups based on the sequence analysis of the 200-bp intron that also included isolates capable of causing bitter rot only. Additionally, isolates of G. cingulata from the United States and Brazil which cause GLS were included in different haplotypes and sequence analysis groups. Therefore, one hypothesis is that isolates of G. cingulata from the United States capable of causing both GLS on foliage and bitter rot on fruit may have arisen independently of Brazilian isolates of G. cingulata capable of causing both GLS and bitter rot, and the two groups of isolates may represent distinct populations.

Vegetative compatibility and virulence of the spinach anthracnose pathogen, Colletotrichum dematium

We characterized 215 isolates of Colletotrichum dematium collected from spinach in Arkansas, California, New Jersey, Oklahoma, Texas, and Ontario, Canada, for vegetative compatibility (using nitrate-nonutilizing mutants) and lesion type (primary or secondary). Isolates recovered from spinach leaves not previously damaged were identified as primary anthracnose isolates, whereas those recovered from leaves with white rust lesions (caused by Albugo occidentalis) were identified as secondary anthracnose isolates. Conidial size and colony color on acidified lima bean agar were recorded