Evaluation of potential biocontrol agents against Claviceps africana in vitro and in vivo

Undiluted culture filtrates of two commercial products of Trichoderma spp., Trichopel and Trichoflow, and two isolates of Penicillium citrinum completely inhibited the conidial germination of macroconidia of Claviceps africana , the cause of ergot or sugary disease of sorghum ( Sorghum bicolor ) in vitro . Similarly, Pseudomonas aeruginosa and Burkholderia cepacia completely inhibited macroconidial germination, with the former being more effective at high dilutions. In contrast, these bacterial isolates failed to inhibit infection in vivo in glasshouse tests with ergot-inoculated sorghum, but all fungal biocontrol agents (including an isolate of Epicoccum nigrum ) reduced the severity of disease (percentage of infected spikelets per panicle), in some cases completely inhibiting the development of ergot. In a second glasshouse trial, optimum control was achieved when the biocontrol agents were applied 3–7 days before inoculation with conidia of C. africana .     

Genetic diversity of Claviceps africana on sorghum and Hyparrhenia

RAPD markers were used to survey genetic variability among 140 isolates of Claviceps africana collected from Southern Africa, India, Thailand, Australia and the Americas in 1992–2002. Amplified fragment length polymorphisms were determined for a subset of the isolates. Both markers gave similar results in phenetic analysis of genetic distances between haplotypes of different geographical origin. In the Americas, a single RAPD haplotype was found throughout the various countries. The Eastern lineage consisted of two close haplotypes (one from India, the other from Thailand and Australia). Among five specialized isolates of C. africana from the alternative hosts ( Hyparrhenia spp.), three haplotypes were found. Eleven private alleles distinguished the Hyparrhenia population from that on sorghum. rDNA sequences of sorghum and Hyparrhenia isolates differed in three positions. The African sorghum population of C. africana consisted of 10, mostly closely related haplotypes. Low genotypic diversity ( H _E = 0·0337) and the fact that most of the variation originated from between populations ( G _ST = 0·866) suggested founder effects following recent invasion. In Southern Africa, no significant differentiation was found among six populations. Therefore the data were pooled and tested for prevalence of clonal or sexual reproduction. The presence of the over-represented, widespread RAPD haplotype A; gametic disequilibrium (37% loci detected by exact tests); index of association ( I _A) significantly >0; and the high proportion of compatible loci (in the clone-corrected and total data sets found to be 94 and 99%, respectively) support the hypothesis of clonality as the predominant means of reproduction.     

Ovary colonization by Claviceps africana is related to ergot resistance in male-sterile sorghum lines

Ergot, caused by Claviceps africana , has emerged as a serious threat to sorghum hybrid seed production worldwide. In the absence of gene-for-gene-based qualitative resistance in commercial cultivars, varieties with high pollen production that can escape ergot infection are preferred. Recent demonstration of differences in ergot susceptibility among male-sterile lines has indicated the presence of partial resistance. Using chitin-specific fluorescin-isothiocyanate-conjugated wheat germ agglutin and callose-specific aniline blue, this study investigated the process of sorghum ovary colonization by C. africana . Conidia germinated within 24 h after inoculation (a.i.); the pathogen was established in the ovary by 79 h a.i., and at least half of the ovary was converted into sphacelial tissue by 120 h a.i. Changes in fungal cell wall chitin content and strategic callose deposition in the host tissue were associated with penetration and invasion of the ovary. The rate of ovary colonization differed in three male-sterile lines that also differed in ergot susceptibility. This work demonstrates a possible histological basis for partial resistance in male-sterile sorghum lines that could lay the foundation for variety improvement through further breeding and selection.     

Windborne spread of ergot disease (Claviceps africana) in sorghum A-lines in Zimbabwe

Ergot disease spread rapidly in Zimbabwe amongst replicated plots of male-sterile sorghum A-lines, from a group of centrally situated and precociously inoculated plants. Prominent secondary conidiation by the pathogen, Claviceps africana , on the surface of exuded honeydew provided airborne spores which were trapped in a Burkard continuous spore trap and showed diurnal peaks of concentration in air close to the primary source of inoculum. The rate of disease spread ( r =0·2; range 0·14–0·58) closely matched that recorded for other plant pathogens such as Phytophthora infestans and Puccinia graminis tritici , and it is concluded that the characteristic secondary conidia of C. africana were the principal epidemiological agents within the experimental area. Ergot spread by windborne secondary conidia has significant epidemiological and economic implications for sorghum hybrid breeding in southern Africa.     

Sorghum ergot can develop without local Claviceps africana inoculum from nearby infected plants

Batches of glasshouse-grown flowering sorghum plants were placed in circular plots for 24 h at two field sites in southeast Queensland, Australia on 38 occasions in 2003 and 2004, to trap aerial inoculum of Claviceps africana. Plants were located 20–200 m from the centre of the plots. Batches of sorghum plants with secondary conidia of C. africana on inoculated spikelets were placed at the centre of each plot on some dates as a local point source of inoculum. Plants exposed to field inoculum were returned to a glasshouse, incubated at near-100% relative humidity for 48 h and then at ambient relative humidity for another week before counting infected spikelets to estimate pathogen dispersal. Three times as many spikelets became infected when inoculum was present within 200 m of trap plants, but infected spikelets did not decline with increasing distance from local source within the 200 m. Spikelets also became infected on all 10 dates when plants were exposed without a local source of infected plants, indicating that infection can occur from conidia surviving in the atmosphere. In 2005, when trap plants were placed at 14 locations along a 280 km route, infected spikelets diminished with increasing distance from sorghum paddocks and infection was sporadic for distances over 1 km. Multiple regression analysis showed significant influence of moisture related weather variables on inoculum dispersal. Results suggest that sanitation measures can help reduce ergot severity at the local level, but sustainable management will require better understanding of long-distance dispersal of C. africana inoculum.     

Genetic Relatedness of African and United States Populations of Cercospora zeae-maydis

Two taxonomically identical but genetically distinct sibling species, designated groups I and II, of Cercospora zeae-maydis cause gray leaf spot of maize in the United States. Isolates of the gray leaf spot pathogen from Africa were compared with isolates from the United States by amplified fragment length polymorphism (AFLP) analysis and restriction digests of internal transcribed spacer (ITS) regions and 5.8S ribosomal DNA (rDNA), as well as by morphological and cultural characteristics. The isolates from Africa were morphologically indistinguishable from the U.S. isolates in both groups, but like isolates of group II, they grew more slowly and failed to produce detectable amounts of cercosporin in culture. Analysis of restriction fragments from the ITS and rDNA regions digested with five endonucleases indicated that all of the African isolates shared the profile of the C. zeae-maydis group II population from the eastern United States and, thus, are distinct from the group I population, which is more prevalent in the United States and other parts of the world. Cluster analysis of 85 AFLP loci confirmed that the African and U.S. group II populations were conspecific (greater than 97% average similarity) with limited variability. Among all group II isolates, only 8 of 57 AFLP loci were polymorphic, and none was specific to either population. Thus, although gray leaf spot was reported in the United States several decades prior to the first record in Africa, the relative age of the two populations on their respective continents could not be ascertained with confidence. The absence of C. zeae-maydis group I in our samples from four countries in the major maize-producing region of Africa as well as the greater AFLP haplotype diversity found in the African group II population, however, suggest that Africa was the source of C. zeae-maydis group II in the United States. The overall paucity of AFLP variation in this sibling species further suggests that its origin is recent or that the ancestral population experienced a severe bottleneck prior to secondary migration.     

Genetic diversity and pathotype determination of Colletotrichum sublineolum isolates causing anthracnose in sorghum

Amplified fragment length polymorphism (AFLP) based genetic diversity was analyzed for 232 Colletotrichum sublineolum isolates collected between 2002 and 2004 from three geographically distinct regions of Texas, and from Arkansas, Georgia, and Puerto Rico. Results revealed significant levels of polymorphism (59%) among the isolates. Even so, genetic similarity between isolates was high, ranging from 0.78 to 1.00. Clustering of similar isolates did not correlate with either geographic origin or year of collection. Pathotypes of 20 of the isolates were determined using 14 sorghum lines previously used in Brazil and the United States and 4 from Sudan. Seventeen new pathotypes were established from the 18 isolates that gave uniform and consistent reactions on all host differentials over 2?years of greenhouse testing. Differentials BTx378 and QL3 were resistant to all isolates while BTx623 and TAM428 were universally susceptible both years. Each of these lines had shown differential responses in prior studies indicating that the pathogen population has sufficient diversity to adapt rapidly to changes in resistant host lines deployed. When the 2-step pathotype classification scheme was used, the 18 isolates examined in this study were placed in four pathotype groups (A, C, D and G), which would further then be separated into ten distinct pathotypes. Common sets of differentials and a standardized nomenclature will allow for comparison to be made among pathotypes of C. sublineolum detected from different regions and also could help direct planting of appropriate sorghum lines and aid in the development of more durable forms of resistance.     

Sibling species of cercospora associated with gray leaf spot of maize

ABSTRACT Monoconidial isolates of the fungus causing gray leaf spot of maize were obtained from diseased leaves collected throughout the United States and analyzed for genetic variability at 111 amplified fragment length polymorphism (AFLP) loci. Cluster analysis revealed two very distinct groups of Cercospora zeae-maydis isolates. Both groups were found to be relatively uniform internally with an average genetic similarity among isolates of approximately 93 and 94%, respectively. The groups were separated from each other by a genetic distance of approximately 80%, a distance greater than that separating each group from the sorghum pathogen, C. sorghi (67 to 70%). Characteristics and dimensions of conidia and conid-iophores produced on infected plants or nutrient media were unreliable criteria for taxonomic differentiation of isolates composing the two groups of C. zeae-maydis. Nucleotide sequences of 5.8S ribosomal DNA (rDNA) and the internal transcribed spacer (ITS) regions were identical within each group but different between the two groups and different from C. sorghi. Restriction fragment length polymorphisms generated by digestion of the 5.8S rDNA and ITS regions with TaqI readily distinguished each group and C. sorghi. Isolates in one group were generally distributed throughout maize-producing regions of the United States; isolates in the other group were localized in the eastern third of the country. Both types were present in the same fields at some locations. The genetic distance based on AFLP profiles and different ITS nucleotide sequences between the two morphologically indistinguishable groups indicate that they are sibling species. Although it is unlikely that breeding for resistance to gray leaf spot will be confounded by local or regional variation in the pathogen, a vigilant approach is warranted, because two pathogenic species exist with unknown abilities to evolve new pathotypes.     

Worldwide Phylogeography of the Citrus Brown Spot Pathogen, Alternaria alternata

ABSTRACT Sixty-five isolates of Alternaria alternata were sampled from brown spot lesions on tangerines and mandarins (Citrus reticulata) and tangerine x grapefruit (C. reticulata x C. paradisi) hybrids in the United States, Colombia, Australia, Turkey, South Africa, and Israel to investigate the worldwide phylogeography of the fungus. Genetic variation was scored at 15 putative random amplified polymorphic DNA (RAPD) loci and 465 bp of an endo-polygalacturonase (endo-PG) gene was sequenced for each isolate. Cluster analysis of RAPD genotypes revealed significant differentiation between United State and Colombia isolates and Turkey, South Africa, Israel, and Australia isolates. Sequencing of endo-PG revealed 21 variable sites when the outgroup A. gaisen (AK-toxin-producing pathogen of Japanese pear) was included and 13 variable sites among the sampled isolates. Nucleotide substitutions at 10 of 13 variable sites represented silent mutations when endo-PG was translated in frame. Eight distinct endo-PG haplotypes were found among the sampled isolates and estimation of a phylogeny with endo-PG sequence data revealed three clades, each with strong bootstrap support. The most basal clade (clade 1) was inferred based on its similarity to the outgroup A. gaisen and consisted exclusively of pathogenic isolates from the United States and Colombia. Clade 2 consisted of pathogenic and nonpathogenic isolates from the United States, Australia, South Africa, and Israel and clade 3 contained pathogenic and nonpathogenic isolates from Australia, South Africa, Israel, and Turkey. Quantitative estimates of virulence (disease incidence) were obtained for isolates from the United States, Colombia, South Africa, Israel, and Turkey by spray inoculating detached citrus leaves and counting the number of lesions 24 h after inoculation. Large differences in virulence were detected among isolates within each location and isolates from the United States were significantly more virulent than isolates from other locations. Several isolates from Colombia, South Africa, Israel, and Turkey had low virulence and 8% of all isolates were nonpathogenic. All but one of the nonpathogenic isolates were found in clade 2 of the endo-PG phylogeny, which also included the most highly virulent isolates sampled.     

Genetic Characterization of Cercospora sorghi from Cultivated and Wild Sorghum and Its Relationship to Other Cercospora Fungi

ABSTRACT Genetic variability and population structure of Cercospora sorghi from wild and cultivated sorghum were investigated to gain insight into their potential impact on epidemics of gray leaf spot of sorghum in Africa. Population structure was examined using data derived from amplified fragment length polymorphism (AFLP) of C. sorghi by Nei's test for population differentiation, G(ST), and analysis of molecular variation (AMOVA). Two ecological populations of C. sorghi in Uganda were devoid of population structure (G(ST) = 0.03, small ef, CyrillicF(ST) = 0.01, P = 0.291). AMOVA revealed that genetic variability was due mainly to variations within (99%) rather than between (0.35%) populations, and Nei's genetic distance between the two populations was 0.014. Phenetic analysis based on AFLP data and polymerase chain reaction-restriction fragment length polymorphism analyses of the internal transcribed spacer regions of rDNA and mitochondrial small subunit rDNA separated Cercospora cereal pathogens from dicot pathogens but did not differentiate among C. sorghi isolates from wild and cultivated sorghum. Our results indicate that Ugandan populations of C. sorghi compose one epidemiological unit and suggest that wild sorghum, while not affecting genetic variability of the pathogen population, provides an alternative host for generating additional inoculum.     

Virulence and Molecular Diversity in Cochliobolus sativus

ABSTRACT Spot blotch, caused by the fungal pathogen Cochliobolus sativus, is an important disease of barley in many production areas of the world. To assess genetic diversity in this pathogen, a worldwide collection of C. sativus isolates was evaluated for virulence on barley and DNA polymorphism. Three pathotypes (0, 1, and 2) were identified among the 22 isolates tested in this study and the 36 isolates characterized previously on three barley differentials (ND5883, Bowman, and NDB112) that differ in their resistance to C. sativus. Pathotype 2, which exhibits high virulence on cv. Bowman, was only found in North Dakota, whereas the other two pathotypes occurred in many other regions of the world. Genetic diversity of the 58 C. sativus isolates, together with isolates of three related pathogenic Cochliobolus spp. (C. heterostrophus, C. carbonum, and C. victoriae) was analyzed using amplified fragment length polymorphism (AFLP) markers. A total of 577 polymorphic AFLP markers were recorded among the 70 isolates of the four Cochliobolus spp. using eight primer combinations. Cluster analysis revealed distinct groups corresponding to the four different species, except in one case where race 0 of C. carbonum was placed in an outgroup that may belong to a different species. In C. sativus, 95 polymorphic AFLP markers were detected with the eight primer pairs used, and each isolate exhibited a unique AFLP pattern. Allelic diversity in the pathotype 2 group was lower (0.10) than in the pathotype 0 (0.23) and pathotype 1 (0.15) groups, indicating that pathotype 2 may have arisen more recently. Cluster analysis did not reveal a close correlation between pathotypes and AFLP groups, although two AFLP markers unique to pathotype 2 isolates were identified. This low correlation suggests that genetic exchange may have occurred through parasexual recombination in the fungal population. Some isolates collected from different regions of the world were clustered into the same AFLP group, suggesting that migration of the fungal pathogen around these regions has occurred.     

A new Fusarium lineage within the Gibberella fujikuroi species complex is the main causal agent of mango malformation disease in Brazil

Mango malformation is a serious disease in tropical and subtropical areas of the world and has been attributed to various Fusarium spp., including F. mangiferae , F. proliferatum , F. sacchari , F. sterilihyphosum and F. subglutinans . Isolates of Fusarium associated with mango malformation from Brazil, Egypt, India, South Africa and the United States were evaluated through amplified fragment length polymorphisms (AFLPs) and partial DNA sequences of the genes encoding β-tubulin ( tub2 ) and translation elongation factor 1-α ( tef1 ). These techniques were used to delimit species and to estimate the genetic and phylogenetic relatedness of the isolates. In the AFLP analysis, most of the Brazilian isolates formed a unique cluster. Additionally, one small cluster was formed by isolates of F. sterilihyphosum from Brazil and South Africa, and another by isolates of F. mangiferae from Egypt, India, South Africa and the United States. In the phylogenetic analysis, most of the Brazilian isolates represented a new phylogenetic lineage in the Gibberella fujikuroi species complex, where they formed a sister clade to F. sterilihyphosum. Representatives of both clades were pathogenic to mango (cv. Tommy Atkins) and Koch's postulates were completed for isolates belonging to the new lineage and to F. sterilihyphosum . Thus, most of the mango malformation disease in Brazil is due to a distinct phylogenetic lineage of Fusarium , and to a lesser extent by F. sterilihyphosum. The new phylogenetic lineage identified in this study, together with F. mangiferae and F. sterilihyphosum , are the only known taxa of Fusarium proven to be capable of causing mango malformation.     

Molecular characterization of Endothia gyrosa isolates from Eucalyptus in South Africa and Australia

Endothia gyrosa is a canker pathogen best known as the causal agent of pin oak blight in North America, and causes cankers on other woody hosts such as Castanea spp. and Liquidambar spp. In South Africa, Australia and Tasmania, a fungus identified as E. gyrosa has been recorded on Eucalyptus spp. Some morphological differences exist between the North American fungus and the isolates from Eucalyptus . Phylogenetic relationships between E. gyrosa from North America and E. gyrosa from South Africa and Australia, as well as that of the related fungi Cryphonectria parasitica and C. cubensis , were studied using PCR-based restriction fragment length polymorphism (RFLP) and sequences of the internal transcribed spacer (ITS) region of the rRNA operon. Endothia gyrosa isolates from South Africa produced the same RFLP banding patterns as those from Australia, which differed markedly from North American isolates of E. gyrosa . In a phylogram based on the DNA sequences, the Australian and South African isolates of E. gyrosa resided in a single, well resolved clade, distinct from North American isolates. Isolates of C. parasitica grouped in the same clade as the South African and Australian isolates of E. gyrosa , but C. cubensis was distantly related to them. The molecular data suggest that the E. gyrosa isolates from South Africa and Australia represent a distinct taxon, and probably belong to the genus Cryphonectria .     

Support for a stepwise mutation model for pathogen evolution in Australasian Puccinia striiformis f.sp. tritici by use of molecular markers

Since its initial detection in Australia in 1979, wheat yellow (stripe) rust ( Puccinia striiformis f.sp. tritici ) has evolved in Australia and New Zealand into more than 20 pathotypes with assorted virulence characteristics. This evolution is believed to have occurred in a stepwise fashion from an original single pathotype, with no subsequent new introductions. A combination of random amplified polymorphic DNA (RAPDs) and amplified fragment length polymorphisms (AFLPs) was used to examine the level of molecular variation in Australian and New Zealand isolates, and to compare this with variation amongst other isolates of P. striiformis . Using 60 RAPD primers on seven Australian isolates representing seven different pathotypes collected between 1979 and 1991, more than 300 potentially polymorphic loci were analysed and no polymorphisms were detected. Using the same primers on two UK isolates, 3% of loci showed a polymorphism. A similar level of polymorphism was found between UK isolates using AFLP primers, and between 5 and 15% of fragments were polymorphic between an isolate from the UK, an isolate from Denmark, and one from Colombia. However, no AFLP polymorphisms were found amongst 14 Australian and New Zealand isolates tested, at over 100 potentially polymorphic loci. The lack of molecular variation in the Australian and New Zealand collection is consistent with the stepwise mutation theory of pathotype evolution from a single introduction.     

Genetic Diversity and the Presence of Two Distinct Groups in Ophiostoma clavigerum Associated with Dendroctonus ponderosae in British Columbia and the Northern Rocky Mountains

ABSTRACT The sapstaining fungal pathogen Ophiostoma clavigerum is associated with the mountain pine beetle (Dendroctonus ponderosae), which is currently the most destructive forest pest in North America. The genetic diversity of O. clavigerum populations collected from five sites in Canada and two sites in the United States was estimated with amplified fragment length polymorphism (AFLP) analysis. Genomic DNA from 170 O. clavigerum isolates was digested with EcoRI and PstI and amplified with six primer sets. A total of 469 AFLP markers consisting of 243 monomorphic and 226 polymorphic loci were scored. The overall genetic diversity of the O. clavigerum population was low (Hs = 0.0531) and the differentiation of the seven O. clavigerum populations was moderate (Phi = 0.143). Genetic distances among the populations were not significantly correlated with geographic distance (r = 0.3235, P = 0.074). Two genetically distinct groups in the O. clavigerum populations were shown by unique AFLP profiles and the unweighted pair group method with arithmetic averages. Further work to characterize biological differences between the two groups will be needed to confirm whether cryptic species are present in the O. clavigerum population.     

Emergence of a novel population of Puccinia striiformis f. sp. tritici in Eastern United States

The geographic range of stripe rust of wheat, caused by Puccinia striiformis f. sp. tritici, has increased dramatically since 2000 in the United States. Yield losses to the disease have been most severe in the eastern United States, where measurable yield loss had been rare prior to 2000. The objective of this study was to examine the phenotypic and genotypic variation among isolates of P. striiformis f. sp. tritici collected from populations in the eastern United States before and since 2000. Virulence phenotype and amplified fragment length polymorphism (AFLP) markers were used to examine 42 isolates collected between 1960 and 2004. In addition, the genetic structure of 59 isolates collected in 2005 using a hierarchical sampling strategy was examined. The data indicated that the contemporary isolates (collected since 2000) were very distinct from older isolates (collected before 2000) based on virulence and AFLP markers, and that the old population prevalent before 2000 may have been replaced by the contemporary population. The old and new populations appear to be genetically distinct and may represent an exotic introduction rather than a mutation in isolates of the old population.     

Characterization of fluorescent Pseudomonas spp. associated with roots and soil of two sorghum genotypes

Sorghum is used as bioenergy feedstock, animal feed, and food. Economical methods for disease prevention and control are valuable for producers. Fluorescent Pseudomonas spp. were isolated from sorghum roots and surrounding soil with the goal of finding isolates that significantly inhibited sorghum fungal pathogens. Fluorescent pseudomonads were collected from seedlings of sorghum cultivars RTx433 and Redlan and wheat cultivar Lewjain, grown in two soils. Lewjain is known to support growth of producers of the antibiotic, 2,4-diacetylphloroglucinol (2,4-DAPG). Isolates from all three plants were assessed for hydrogen cyanide (HCN) and extracellular protease production, and for a 2,4-DAPG gene, phlD. Both soil type and plant type affected HCN- and protease-production, but phlD was not affected. Subsets of phlD ⁺ isolates were chosen to determine phlD genotypes and to conduct in vitro inhibition assays against sorghum pathogens. Most isolates from sorghum and wheat were genotype D, previously associated with superior root colonization. phlD ⁺ sorghum isolates were co-cultured with five sorghum pathogens. One isolate from each sorghum line exhibited inhibition to all five pathogens but more Redlan isolates were inhibitory to the virulent pathogen, Fusarium thapsinum, than RTx433 isolates. Nearly all inhibitory isolates from either sorghum cultivar were from one soil type. This is consistent with what had been previously observed in field studies: that soil type played a significant role in determining characteristics of fluorescent Pseudomonas spp. isolated from roots or soil, but sorghum genotype also had a considerable effect.     

Characterization and distribution of mating type genes in the dothistroma needle blight pathogens

ABSTRACT Dothistroma septosporum and D. pini are the two causal agents of Dothistroma needle blight of Pinus spp. in natural forests and plantations. Degenerate primers amplified portions of mating type genes (MAT1-1-1 and MAT1-2) and chromosome walking was applied to obtain the full-length genes in both species. The mating-type-specific primers designed in this study could distinguish between the morphologically similar D. pini and D. septosporum and between the different mating types of these species. Screening of isolates from global collections of D. septosporum showed that only MAT2 isolates are present in Australian and New Zealand collections, where only the asexual form of the fungus has been found. In contrast, both mating types of D. septosporum were present in collections from Canada and Europe, where the sexual state is known. Intriguingly, collections from South Africa and the United Kingdom, where the sexual state of the fungus is unknown, included both mating types. In D. pini, for which no teleomorph is known, both mating types were present in collections from the United States. These results provided new insights into the biology and global distribution of two of the world's most important pine pathogens and should facilitate management of the diseases caused by these fungi.     

玉米大斑病菌有性杂交F1代菌株的生理小种鉴定和AFLP分析

玉米大斑病菌是异宗配合真菌，有性杂交有可能增强病菌的致病力，或形成新的致病小种，因此对该病菌有性杂交后代进行致病性测定和遗传多态性分析对控制该病菌的危害具有重要意义。对亲本菌株132、135和它们杂交产生的70个单子囊孢子F1代菌株进行了生理小种鉴定和AFLP（扩增性片段长度多态性）分析。生理小种鉴定结果表明，F1代菌株中与亲本菌株132（23N号小种）属于同一小种类型的占41．4％，与亲本菌株135（23号小种）相同的占20．0％，另外还出现了0、1、2、3、13、123、12N、13N和123N号小种，所占比例分别为2．9％、1．4％、2．9％、2．9％、4．3％、8．6％、1．4％、4．3％和10．0％，说明有性杂交可使后代菌株的致病性发生比较广泛的变异。AFLP分析表明，F1代菌株之间分子遗传相似系数在0．87～0．99之间，其中84．3％的F1代菌株与亲本菌株的遗传相似系数在0．878以上，但与亲本菌株132同源性较强的F1代菌株数目大约是与亲本菌株135的5倍，说明不同菌株具有不同的遗传传递能力。比较生理小种鉴定和AFLP分析结果，发现生理小种分化和AFLP分子遗传多态性间有一定的相关性，但不能完全对应，不存在遗传谱系就等于小种的简单对应关系。