Characterization of chickpea differentials for pathogenicity assay of ascochyta blight and identification of chickpea accessions resistant to Didymella rabiei

Forty-eight chickpea germplasm lines, including 22 differentials used in previous studies, were characterized for disease phenotypes following inoculation with six isolates of Didymella (anamorph Ascochyta ) rabiei , representing a wide spectrum of pathogenic variation. Representative isolates were also directly compared with six previously identified races on eight chickpea genotypes. Many of the chickpea differentials reacted similarly to inoculation with each isolate of D. rabiei , and several previously identified races caused similar levels of disease on the differentials. This indicates that the number of differentials can be reduced significantly without sacrificing accuracy in describing pathogenic variation of D. rabiei on chickpea. Pathogenic variation among samples of US isolates allowed classification of the isolates into two pathotypes. The distribution of disease phenotypes of the 48 germplasm lines was bimodal after inoculation with pathotype I isolates, whereas the distribution of disease phenotypes was continuous after inoculation with pathotype II isolates. Such distinct distribution patterns suggest that chickpea plants employ different resistance mechanisms to each pathotype and that the two pathotypes may have different genetic mechanisms controlling pathogenicity. The advantages of using the two-pathotype system in assaying pathogenicity of the pathogen and in studying resistance mechanisms of the host are discussed. Three chickpea accessions, PI 559361, PI 559363 and W6 22589, showed a high level of resistance to both pathotypes, and can be employed as resistance sources in chickpea breeding programmes for resistance to ascochyta blight.     

Genetic Diversity in South American Colletotrichum gloeosporioides Isolates from Stylosanthes guianensis, a Tropical Forage Legume

The degree of genetic diversity of 127 Colletotrichum gloeosporioides isolates from Stylosanthes guianensis genotypes in South America was measured at the molecular level by random amplified polymorphic DNA (RAPD) with nine arbitrary primers of 10 bases, and by restriction fragment length polymorphism (RFLP) with a non-LTR (long terminal repeats) retrotransposon DNA sequence. The RAPD products revealed scorable polymorphism among the isolates, and a total of 80 band positions were scored. Sixty-three of the 127 isolates were clustered into 13 distinct lineages usually correlating with geographic origin. Where isolates from various regions were clustered together, most had identical host genotype origin. The pathogen population sampled from Carimagua, Colombia, a long-time Stylosanthes breeding and selection site, with a savanna ecosystem, was highly diverse. A set of 12 S. guianensis genotype differentials was used to characterize pathogenic variability of 104 isolates and their virulence patterns were grouped into 57 pathotypes. However, when they were tested on four Australian differentials, they grouped into 11 pathotypes. As shown in previous studies, no strict correlations existed between genetic diversity measured by RAPD or RFLP, and pathotype defined by pathogenicity pattern on the differentials. Southern blot analysis of the 127 isolates revealed 23 hybridizing fragments, resulting in 41 fingerprint patterns among the 127 isolates. Relationships between RFLP and RAPD variables were examined using Spearman's Rank Correlation Coefficient, which showed that the two measures of genotypic variation are in agreement.     

Distribution and Biocontrol Potential of phlD(+) Pseudomonads in Corn and Soybean Fields

ABSTRACT The abundance and diversity of phlD(+) Pseudomonas spp. colonizing the rhizospheres of young, field-grown corn and soybean plants were assayed over a 3-year period. Populations of these bacteria were detected on the large majority of plants sampled in the state of Ohio, but colonization was greater on corn. Although significant variation in the incidence of rhizosphere colonization was observed from site to site and year to year on both crops, the magnitude of the variation was greatest for soybean. The D genotype was detected on plants collected from all 15 counties examined, and it represented the most abundant subpopulation on both crops. Additionally, six other genotypes (A, C, F, I, R, and S) were found to predominate in the rhizosphere of some plants. The most frequently observed of these were the A genotype and a newly discovered S genotype, both of which were found on corn and soybean roots obtained from multiple locations. Multiple isolates of the most abundant genotypes were recovered and characterized. The S genotype was found to be phylogenetically and phenotypically similar to the D genotype. In addition, the novel R genotype was found to be most similar to the A genotype. All of the isolates displayed significant capacities to inhibit the growth of an oomycete pathogen in vitro, but such phenotypes were highly dependent on media used. When tested against multiple oomycete pathogens isolated from soybean, the A genotype was significantly more inhibitory than the D genotype when incubated on 1/10x tryptic soy agar and 1/5x corn meal agar. Seed inoculation with different isolates of the A, D, and S genotypes indicated that significant root colonization, generally in excess of log 5 cells per gram of root, could be attained on both crops. Field trials of the A genotype isolate Wayne1R indicated the capacity of inoculant populations to supplement the activities of native populations so as to increase soybean stands and yields. The relevance of these findings to natural and augmentative biocontrol of root pathogens by these bacteria is discussed.     

Development of simple sequence repeat markers for the classification of the clubroot pathogen <Emphasis Type="Italic">Plasmodiophora brassicae</Emphasis>

Clubroot disease caused by Plasmodiophora brassicae is one of the most serious diseases in cruciferous crops. To classify isolates, we developed simple sequence repeat (SSR) markers for P. brassicae. Twenty-four Japanese isolates were used in this study: 12 isolates of an unknown pathotype from the Kyoto Prefecture, as well as 12 isolates of known pathotypes, including three single-spore lines. From the 12 isolates from Kyoto Prefecture, 11 were classified into either pathotype 2 (three isolates) or 4 (eight isolates). We designed 23 SSR markers based on the P. brassicae genome, of which 11 markers from intergenic regions showed polymorphisms in the 24 isolates. Many haploid isolates belonging to pathotypes 2 and 4 were monomorphic, and typical alleles were detected in some isolates not belonging to pathotype 4. Two bands were detected for eight SSR loci in five isolates, indicating that different genotypes were mixed in these isolates. We constructed a phylogram based on the 11 polymorphic SSRs. Pathotypes 2 and 4 formed a cluster, from which pathotypes 3 and 1 were successively placed. These results strongly suggest a close genetic relationship between isolates in pathotypes 2 and 4, consistent with our finding that isolates in these two pathotypes were found at one collection site. In combination with pathotype classification and other marker systems, the SSR markers can be used for more detailed analyses to improve the control of clubroot disease.     

Genetic and pathogenic diversity within Ascochyta rabiei(Pass.) Lab. populations in Pakistan causing blight of chickpea (Cicer arietinum L.)

Pathogenic and genetic diversity in Ascochyta rabiei populations in Pakistan were evaluated. Biological pathotyping of 130 A. rabiei isolates (obtained from hierarchically collected samples) was conducted on a set of three chickpea differentials, i.e. ILC 1929 (susceptible), ILC 482 (tolerant) and ILC 3279 (resistant), under controlled conditions. Disease severity data were recorded 12 days after inoculation. Statistical analysis grouped the isolates into three pathotype classes. Four isolates belonged to pathotype I (least aggressive), 79 isolates to pathotype II (medium aggressive) and 47 isolates to pathotype-III (highly aggressive).Genetic analysis was performed using RAPDs and oligonucleotide fingerprinting, where Hinf I-digested DNA was hybridized to the³²P-endlabeled oligonucleotide probes (CAA)₅, (GAA)₅, (GA)₈, (CA)₈and (GATA)₄. Dendrograms produced by cluster analysis discriminated 46 genotypes in the A. rabiei population of Pakistan. Genetic distances and relatedness between isolates were calculated. At a genetic distance of 0.3, genotypes were divided into six distinct genotype groups A, B, C, D, E and F containing 16, 11, 2, 5, 5 and 7 isolates, respectively. Most of the genotypes were area specific or predominated in certain areas but did not belong to a distinct pathotype, while most of the aggressive isolates (pathotype III) occurred in Northern Punjab and in the North Western Frontier Province.     

Trichothecene mycotoxin genotypes of Fusarium graminearum sensu stricto and Fusarium meridionale in wheat from southern Brazil

A total of 82 fungal isolates was obtained from wheat kernel samples affected by fusarium head blight collected from 20 locations in southern Brazil. Polymerase chain reaction (PCR) assays were used to characterize trichothecene mycotoxin genotypes [deoxynivalenol (DON), nivalenol (NIV) and two acetylated derivatives of DON]. To identify isolates that producing DON and NIV, portions of the Tri13 gene were amplified. To identify 3-acetyl-deoxynivalenol (3-ADON) and 15-acetyl-deoxynivalenol (15-ADON) genotypes, portions of Tri3 and Tri12 were amplified. Nearly all of the isolates studied (76/82) were of the DON/15-ADON genotype. Six of the isolates were of the NIV genotype. The DON/3-ADON genotype was not observed. Portions of three genes were sequenced from representative isolates of the NIV and DON/15-ADON genotypes and compared with sequences from curated reference isolates of Fusarium in GenBank. blast queries for individual gene sequences and pairwise comparisons of percentage identity and percentage divergence based on 1676 bp of concatenated DNA sequence suggested that the isolates representing the DON/15-ADON genotype were Fusarium graminearum sensu stricto and the isolates representing the NIV genotype were Fusarium meridionale . This is the first detailed report of trichothecene mycotoxin genotypes of F. graminearum and F. meridionale in Brazil.     

Assessment of biological and molecular variability between and within field isolates of Plasmodiophora brassicae

Plasmodiophora brassicae is an obligate biotroph that causes clubroot, one of the most damaging diseases of crucifers. Differential cultivars and random amplified polymorphic DNA markers were used to assess the extent of genetic diversity among nine single-gall populations of P. brassicae and 37 single-spore isolates (SSI) derived from four of those field samples. Isolates were classified into eight pathotypes, and each isolate was associated with a unique molecular genotype. Virulence and DNA polymorphisms were detected within and between field isolates, and among SSIs from different pathotypes, hosts and geographical origins. The relatively high level of genetic diversity among field isolates was similar to that among SSIs derived from a single-club field isolate. Molecular and pathogenicity-based classifications were not clearly correlated, but isolates belonging to pathotype P1 were clustered. Two RAPD markers were specific to pathotype P1. The finding that genetic differences can occur in P. brassicae field isolates will be an important consideration in resistance genetic studies and in choosing breeding strategies to develop durable clubroot resistance.     

The Population Structure of Phytophthora infestans from the Toluca Valley of Central Mexico Suggests Genetic Differentiation Between Populations from Cultivated Potato and Wild Solanum spp

ABSTRACT The population structure of Phytophthora infestans in the Toluca Valley of central Mexico was assessed using 170 isolates collected from cultivated potatoes and the native wild Solanum spp., S. demissum and S. xendinense. All isolates were analyzed for mitochondrial DNA (mtDNA) haplotype and amplified fragment length polymorphism (AFLP) multi-locus fingerprint genotype. Isolate samples were monomorphic for mtDNA haplotype because all isolates tested were of the Ia haplotype. A total of 158 multilocus AFLP genotypes were identified among the 170 P. infestans isolates included in this study. P. infestans populations sampled in the Toluca Valley in 1997 were highly variable and almost every single isolate represented a unique genotype based on the analysis of 165 AFLP marker loci. Populations of P. infestans collected from the commercial potato-growing region in the valley, the subsistence potato production area along the slopes of the Nevado de Toluca, and the native Solanum spp. on the forested slopes of the volcano showed a high degree of genetic diversity. The number of polymorphic loci varied from 20.0 to 62.4% for isolates collected from the field station and wild Solanum spp. On average, 81.8% (135) of the AFLP loci were polymorphic. Hetero-zygosity varied between 7.7 and 19.4%. Significant differentiation was found at the population level between strains originating from cultivated potatoes and wild Solanum spp. (P = 0.001 to 0.022). Private alleles were observed in individual isolates collected from all three populations, with numbers of unique dominant alleles varying from 9 to 16 for isolates collected from commercial potato crops and native Solanum spp., respectively. Four AFLP markers were exclusively found present in isolates collected from S. demissum. Indirect estimation of gene flow between populations indicated restricted gene flow between both P. infestans populations from cultivated potatoes and wild Solanum hosts. There was no evidence found for the presence of substructuring at the subpopulation (field) level. We hypothesize that population differentiation and genetic isolation of P. infestans in the Toluca Valley is driven by host-specific factors (i.e., R-genes) widely distributed in wild Solanum spp. and random genetic drift.     

Variation in aggressiveness is detected among Puccinia triticina isolates of the same pathotype and clonal lineage in the adult plant stage

Puccinia triticina reproduces asexually in France and thus individual genotype is the unit of selection. A strong link has been observed between genotype identities (as assessed by microsatellite markers) and pathotypes (pools of individuals with the same combination of qualitative virulence factors). Here, we tested whether differences in quantitative traits of aggressiveness could be detected within those clonal lineages by comparing isolates of identical pathotype and microsatellite profile. Pairs of isolates belonging to different pathotypes were compared for their latent period, lesion size and spore production capacity on adult plants under greenhouse conditions, with a high number of replicates. Isolates of the same pathotype showed remarkably similar values for the measured traits, except in three situations: differences were obtained within two pathotypes for latent period and within one pathotype for sporulation capacity. One of these differences was tested again and confirmed. This indicates that the average aggressiveness level of a leaf rust pathotype may increase without any change in its virulence factors or microsatellite profile.     

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.     

Selection for Pathogenicity to Strawberry in Populations of Colletotrichum gloeosporioides from Native Plants

ABSTRACT Colletotrichum gloeosporioides causes a serious crown rot of strawberry and some isolates from native plants are pathogenic to strawberry. C. gloeosporioides from lesions on wild grape and oak were sampled at two sites adjacent to commercial strawberry fields in Florida and two distant sites. Random amplified polymorphic DNA (RAPD) marker data and restriction enzyme digests of amplified rDNA were used to determine whether isolates were from the same C. gloeosporioides subgroup that infects strawberry. There were 17 to 24 native host isolates from each site that clustered with a group of strawberry crown isolates based on RAPD markers. Among strawberry isolates, there were two rDNA genotypes identified by restriction enzyme analysis. Both genotypes were present among native host isolates sampled from all four sites. There was some evidence that the different rDNA genotypes differentiated two closely related subpopulations, although the proportion of pathogenic isolates from native hosts among the two different genotypes was not different. The incidence of isolates pathogenic to strawberry was greater at sites close to strawberry fields relative to sites distant from strawberry fields for isolates with a BstUI(-)/MspI(+) rDNA genotype (44 versus 13%), a BstUI(+)/MspI(-) genotype (57 versus 16%), or when both genotypes were analyzed together (46 versus 15%). Based on these results, it appears that the C. gloeosporioides subgroup that causes crown rot on strawberry is widely distributed in Florida and that selection for pathogenicity on strawberry occurs in the area where this host is grown in abundance.     

Clonal Spread of Sclerotium cepivorum in Onion Production with Evidence of Past Recombination Events

Three field populations of Sclerotium cepivorum from Ontario and Que-bec in Canada, one field sample from the North Island of New Zealand, as well as isolates from other onion- and garlic-growing areas were classified in mycelial compatibility groups (MCGs) and further characterized by DNA fingerprints and DNA sequence polymorphisms at six genomic regions. MCGs in the population samples were genetically distinct clones. Members of a clone belonged to a unique MCG with an associated multi-locus genotype and DNA fingerprint. The structure in each field population was predominantly clonal based on the repeated sampling of genotypes over a wide geographic area. Only three clones were identified within each of the population samples. The hypothesis that these clones were introduced to the three agricultural production areas is supported by their recovery from additional locations. Some evidence for past recombination was detected when genotypes from the three population samples and from other locations were pooled.     

Characterization and Distribution of Two Races of Phialophora gregata in the North-Central United States

ABSTRACT Genetic variation and variation in aggressiveness in Phialophora gregata f. sp. sojae, the cause of brown stem rot of soybean, was characterized in a sample of 209 isolates from the north-central region. The isolates were collected from soybean plants without regard to symptoms from randomly selected soybean fields. Seven genotypes (A1, A2, A4, A5, A6, M1, and M2) were distinguished based on DNA fingerprinting with microsatellite probes (CAT)(5) and (CAC)(5), with only minor genetic variation within the A or M genotypes. Only the A1, A2, and M1 genotypes were represented by more than one isolate. The A genotypes dominated in the eastern Iowa, Illinois, and Ohio samples, whereas the M genotypes were dominant in samples from western Iowa, Minnesota, and Missouri. In growth chamber experiments, isolates segregated into two pathogenicity groups based on their aggressiveness toward soybean cvs. Kenwood and BSR101, which are relatively susceptible and resistant, respectively, to brown stem rot. In both root dip inoculation and inoculation by injecting spores into the stem near the ground line (stab inoculations), isolates of the A genotypes caused greater foliar symptoms and more vascular discoloration than isolates of the M genotypes on both cultivars of soybean. All isolates caused foliar symptoms in both cultivars and in three additional cultivars of soybean with resistance to brown stem rot. Greater differences between the A and M genotypes were seen in foliar symptoms than in the linear extent of xylem discoloration, and greater differences were seen in Kenwood than in BSR101. Inoculation of these genotypes into five cultivars of soybean with different resistance genes to brown stem rot showed a genotype x cultivar interaction. A similar distinction was found in an earlier study of the adzuki bean pathogen, P. gregata f. sp. adzukicola, and consistent with the nomenclature of that pathogen, the soybean pathogens are named the aggressive race (race A) and the mild race (race M) of P. gregata f. sp. sojae.     

Genetic and morphologic diversity of <Emphasis Type="Italic">Echinochloa crus-galli</Emphasis> populations from different origins

Echinochloa crus-galli (L.) P. Beauv. (barnyardgrass) is an annual weed that is native to Asia and found throughout the world. The broad ecological tolerance and competitive ability of E. crus-galli makes it the most important weed species in rice. Genetic studies of plants are becoming increasingly common because reliable information is necessary to better understand population dynamics, occurrence of herbicide resistance, and demographic data. Echinochloa crus-galli populations from 34 different locations in Turkey were compared with respect to morphological differences and genetic variation. For morphological variation, five seeds of each population were sown in pots and grown in a screenhouse using a randomized block design. Morphological parameters such as germination speed, flowering time, leaf area, plant height, spikelet length, above-ground biomass, root dry weight and number of seeds were measured. Distinct differences among populations with respect to hierarchical cluster analysis were observed. Genetic variations among populations were performed using random amplified polymorphic DNA (RAPD) markers. The seven RAPD primers amplified 55 bands whose molecular weight varied between 200 and 4000 bp. The percentage of polymorphic bands was 74.54%. Results showed high morphological and genetic variability among individual genotypes within geographic locations. Phenotypic and genetic variability among E. crus-galli populations would be influenced by agricultural practices, crop characteristics, geographic location and herbicide pressure. Differences between weed populations may affect response to chemical or biological control.     

Genetic diversity and pathogenicity of Sphaceloma rosarum (teleomorph Elsinoë rosarum) causing spot anthracnose on roses

Roses produced or grown in the field, as well as pot‐grown and cut roses, are attacked by different fungal pathogens causing leaf spot diseases. The incorrect identification and scoring of these pathogens and the lack of information about their genetic and pathotype diversity hamper resistance breeding. This is especially true for the hemibiotrophic ascomycete Sphaceloma rosarum, which is often confused with other fungi. Here for the first time, the genetic variability between isolates at both the molecular and morphological level is analysed. Eighty leaf spot samples were collected from different rose genotypes at five different locations, and 15 single conidial isolates established. All of the samples showed high morphological similarities to the reference isolate CBS 213.33 that was obtained from a public repository. By sequencing a part of the large subunit (LSU) of the 28S ribosomal RNA and phylogenetic analysis, high sequence similarities were shown to other Sphaceloma species for 13 of the isolates and the CBS reference. One of the isolates clustered with Septoria species and another clustered with Seimatosporium species. UPGMA clustering with 145 polymorphic AFLP markers resulted in five distinct groups in the majority rule consensus tree for the 14 S. rosarum isolates, including the CBS reference. Jaccard similarities ranged from 0·31 to 0·91. A detached leaf assay using a differential set of five rose genotypes led to the classification of the five tested isolates as five distinct pathotypes. Therefore, grouping depending on the avirulence gene diversity was clearly different from clustering using selectively neutral AFLP markers that were evenly distributed throughout the genome.     

Host resistance mediated inter-genotype competition and temporal variation in <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> f. sp. <Emphasis Type="Italic">vasinfectum</Emphasis>

Temporal variation in Fusarium oxysporum f. sp. vasinfectum (Fov) populations was determined by comparing the genetic diversity of pathogen isolates recovered from three consecutive cotton crops (2002, 2004 and 2006) in the Boggabilla area of New South Wales, Australia. A total of 288 isolates were collected, among which 25 distinct AFLP genotypes were identified. These genotypes were classified into two main groups corresponding to known vegetative compatibility groups (VCG)—01111 and 01112. The Fov populations were dominated by four genotypes (I-A, I-B, II-A, II-B) that accounted for 87.5% of the isolates. Significant temporal variation was observed in both sampled fields with 6.8% and 10.7% of total genetic variation being attributed to differences among collections in different years. Genetic diversity based on Nei’s gene diversity and the Shannon-Wiener index increased over time. Significant changes in the frequency of the dominant Fov genotypes were observed in one field, where genotype I-A declined from 84.8% to 40.0% over the study period (2002–2006), while genotype I-B increased from 7.6% to 35.4%. Strong inter-genotype competition was detected in glasshouse bioassays with 93.4% of symptomatic plants sampled from dual inoculation trials being infected by single genotypes. Competition was differentially mediated by cotton cultivars as the competitive ability of pathogen genotype I-B was enhanced on the resistant cultivar Sicot 189 relative to the susceptible cultivar Siokra 1–4. This suggests that host-mediated inter-genotype competition may play an important role in temporal variation in Fov populations in the field.     

Electrophoretic karyotype variation among pathotypes of Fusarium oxysporum f.sp. dianthi

Karyotype analysis by pulsed-field gel electrophoresis was applied to characterize isolates of Fusarium oxysporum f.sp. dianthi , the causal agent of Fusarium wilt on carnation. Eleven distinct chromosomal DNA patterns were detected among 38 pathogenic isolates, and the total genome size was estimated to range from 23·7 to 36·4 Mb. Except for isolates belonging to pathotypes 2 and 4 , all members of the same pathotype shared overlapping electrophoretic karyotypes. Karyotypes of isolates assigned to pathotypes 1 and 8 showed a high degree of similarity, in accordance with VCG and RFLP analysis. The same electrophoretic karyotype was also shared by members of pathotypes 2 and 5, thus confirming results obtained by both VCG and RFLP grouping, A single representative of pathotype 6, previously confined to the same VCG and RFLP group as pathotypes 2 and 5, had a slightly different chromosomal pattern. Isolates assigned to pathotype 4 showed four related karyotypes which partially differed in both the number and size of chromosomal bands. However, all strains assigned to this pathotype shared a basic profile of nine chromosomal bands, while two low-molecular-weight bands were present or absent. The findings are discussed with regard both to the suitability of race distinction in the case of the special form dianthi of F. oxysporum and to the use of karyotype analysis by PFGE as a tool for the study of the population genetics of this fungus.     

Comparison of genotyping by sequencing and microsatellite markers for unravelling population structure in the clonal fungus Verticillium dahliae

Microsatellite genotyping of a large sample of isolates of Verticillium dahliae from diverse locations recently identified seven distinct genotypic clusters. However, these clusters were not put in the context of phenotypes known to be correlated with clonal lineages in V. dahliae. The objective of this study was to compare clusters defined by microsatellite markers with clonal lineages defined by single‐nucleotide polymorphisms (SNPs) and vegetative compatibility groups (VCGs). Genotyping isolates known to belong to specific clonal lineages (based on SNPs) with microsatellite markers determined the correspondence of clusters and lineages. All but one cluster corresponded to a known clonal lineage, allowing analysis of correlations of phenotypes with microsatellite genotypes from other studies. As shown previously, most race 1 isolates are in lineage 2A, and most isolates with the defoliating pathotype are in lineage 1A. Phylogenetic incompatibility was used to test for recombination or homoplasy caused by hypervariable microsatellite loci; incompatibility was highly correlated with the number of alleles per locus, suggesting that homoplasy caused by parallel evolution of microsatellite alleles is the cause of incompatibility. Microsatellite genotyping of lineage 1A isolates from cotton and olive in Spain over a 29‐year period revealed remarkably little variation; these markers did not mutate enough to provide insight on the spatial and temporal expansion of this clone. Overall, this study showed that microsatellite genotyping can be used to identify clonal lineages in V. dahliae, which has predictive power for inferring phenotypes of phytopathological relevance such as race and pathotype.     

Genotypic variability and aggressiveness of Bipolaris oryzae in the Philippines

The genotypic diversity of a collection of 352 isolates of Bipolaris oryzae obtained from 11 locations in the Philippines was estimated. The isolates could be divided into 50 haplotypes based on variation in microsatellite DNA with a moderately high genotypic diversity value of 0.88. Thirty nine haplotypes were represented by three or fewer isolates, whereas 80 % of the isolates belonged to only eight haplotypes, each containing 10 to 88 isolates indicating the prevalence of clonality. AMOVA revealed that the greatest variation was associated with the brown spot isolates collected within provinces (50.81 %), among varieties within provinces (48.17 %) and within ecosystems (49.33 %). Intensive sampling from a single field showed that the population was mostly clonal with about 98 % of the isolates belonging to a single VNTR haplotype. However, isolates within this haplotype exhibited a continuous range of aggressiveness when inoculated onto susceptible rice variety IR72. Several types of lesions were observed in the field during sampling, but the isolates obtained from each type of lesion produced a range of different lesion types when inoculated onto leaves of IR72, indicating that the type of lesion observed in the field was not related to the genotype of the pathogen. These results show that rice fields across the Philippines may contain B. oryzae isolates with considerable genotypic diversity, but an individual field may have both clonal and unique genotypes.     

Genetic Diversity and Structure of the Apiosporina morbosa Populations on Prunus spp

ABSTRACT Populations of Apiosporina morbosa collected from 15 geographic locations in Canada and the United States and three host species, Prunus virginiana, P. pensylvanica, and P. padus, were evaluated using the sequence-related amplified polymorphism (SRAP) technique to determine their genetic diversity and population differentiation. Extensive diversity was detected in the A. morbosa populations, including 134 isolates from Canada and the United States, regardless of the origin of the population. The number of polymorphic loci varied from 6.9 to 82.8% in the geographic populations, and from 41.4 to 79.3% in the populations from four host genotypes based on 58 polymorphic fragments. In all, 44 to 100% of isolates in the geographic populations and 43.6 to 76.2% in populations from four host genotypes represented unique genotypes. Values of heterozygosity (H) varied from 2.8 to 28.3% in the geographic populations and 10.2 to 26.1% in the populations from four host genotypes. In general, the A. morbosa populations sampled from wild chokecherry showed a higher genetic diversity than those populations collected from other host species, whereas the populations isolated from cultivated chokecherry, P. virginiana 'Shubert Select', showed a reduction of genetic diversity compared with populations from wild P. virginiana. Significant population differentiation was found among both the geographic populations (P < 0.05) and populations from different host genotypes (P < 0.02). In the geographic populations, most of populations from cultivated and wild P. virginiana were closely clustered, and no population differentiation was detected except for the populations from Morris, Morden, and Winnipeg, Manitoba, Canada. Furthermore, the populations from P. virginiana in the same geographic locations had higher genetic identity and closer genetic distance to each other compared with those from different locations. Four populations from P. virginiana, P. pensylvanica, and P. padus, were significantly differentiated from each other (P < 0.02), except there was no differentiation between the Shubert Select and wild chokecherry populations (>P> = 0.334). Indirect estimation of gene flow showed that significant restricted gene flow existed between populations from different regions and host species. Gene flow rates (Nm) varied from <1 to 12.5, with higher gene flow rates among population pairs from the same host species (P = 1.000). The analysis of molecular variance revealed that a major genetic variance source came from the genetic variation among isolates within populations regardless of the origin and host genotype of the population. Although some locations had a limited number of isolates, the results of this study clearly showed that the genetic diversity and population differentiation of A. morbosa were closely associated with host genotypes and geographic locations, but mostly with the former.