Population genetic structure of four regional populations of the barley pathogen Pyrenophora teres f. maculata in Iran is characterized by high genetic diversity and sexual recombination

The leaf spot form of the barley disease net blotch, caused by the fungus Pyrenophora teres f. maculata (PTM), is an increasingly important foliar disease of barley. Studies of population genetic structure and reproductive mode are necessary to make predictions of the evolutionary potential of the pathogen. Sources of resistance to PTM have been found in Iranian landraces, which may have the potential to improve plant breeding efforts. However, little is known about the population genetic structure of this fungus in Iran. In this study, we analysed the frequency of the mating type genes to assess the potential for sexual mating of PTM collected from four provinces—Khuzestan, Hamadan, Golestan, and East Azerbaijan—and we investigated the population genetic structure using seven simple sequence repeat markers. High genotype diversity, linkage equilibrium, and equal ratios of mating types frequencies in the PTM populations at Khuzestan and Hamadan support the occurrence of sexual reproduction in these populations, while in Golestan and East Azerbaijan populations, significant gametic disequilibrium and relatively low genotype diversity suggest a higher incidence of clonality or different demographic histories. Unequal mating type frequencies in Golestan confirm a predominance of asexual reproduction. Finally, we found significant evidence for strong population structure with most of the genetic variation represented within regional populations (89%). Overall, our study provides evidence for high genetic variation in Iranian PTM populations, which may be the basis for rapid adaptive evolution in this pathosystem. This highlights the need for integrated efforts to control the disease.     

Population structure of the ash dieback pathogen,Hymenoscyphus fraxineus,in relation to its mode of arrival in the UK

The ash dieback fungus, Hymenoscyphus fraxineus, a destructive, alien pathogen of common ash (Fraxinus excelsior), has spread across Europe over the past 25 years and was first observed in the UK in 2012. To investigate the relationship of the pathogen's population structure to its mode of arrival, isolates were obtained from locations in England and Wales, either where established natural populations of ash had been infected by wind‐dispersed ascospores or where the fungus had been introduced on imported planting stock. Population structure was determined by tests for vegetative compatibility (VC), mating type and single‐nucleotide polymorphisms (SNPs). VC heterogeneity was high at all locations, with 96% of isolate pairings being incompatible. Frequencies of the MAT1‐1‐1 and MAT1‐2‐1 idiomorphs were approximately equal, consistent with H. fraxineus being an obligate outbreeder. Most SNP variation occurred within study location and there was little genetic differentiation between the two types of location in the UK, or between pathogen populations in the UK and continental Europe. There was modest differentiation between UK subpopulations, consistent with genetic variation between source populations in continental Europe. However, there was no evidence of strong founder effects, indicating that numerous individuals of H. fraxineus initiated infection at each location, regardless of the route of pathogen transmission. The ssRNA virus HfMV1 was present at moderate to high frequencies in all UK subpopulations. The results imply that management of an introduced plant pathogen requires action against its spread at the continental level involving coordinated efforts by European countries.     

Population structure of Sclerotinia sclerotiorum in crop and wild hosts in the UK

Sclerotinia sclerotiorum is an important pathogen of many crop plants which also infects wild hosts. The population structure of this fungus was studied for different crop plants and Ranunculus acris (meadow buttercup) in the UK using eight microsatellite markers and sequenced sections of the intergenic spacer (IGS) region of the rRNA gene and the elongation factor 1‐alpha (EF) gene. A total of 228 microsatellite haplotypes were identified within 384 isolates from 12 S. sclerotiorum populations sampled in England and Wales. One microsatellite haplotype was generally found at high frequency in each population and was distributed widely across different hosts, locations and years. Fourteen IGS and five EF haplotypes were found in the 12 populations, with six IGS haplotypes and one EF haplotype exclusive to buttercup. Analysis of published sequences for S. sclerotiorum populations from the USA, Canada, New Zealand and Norway showed that three of the IGS haplotypes and one EF haplotype were widely distributed, while eight IGS haplotypes were only found in the UK. Although common microsatellite and IGS/EF haplotypes were found on different hosts in the UK, there was evidence of differentiation, particularly for one isolated population on buttercup. However, overall there was no consistent differentiation of S. sclerotiorum populations from buttercup and crop hosts. Sclerotinia sclerotiorum therefore has a multiclonal population structure in the UK and the wide distribution of one microsatellite haplotype suggests spatial mixing at a national scale. The related species S. subarctica was also identified in one buttercup population.     

Spatial genetic structure and dispersal of the cacao pathogen Moniliophthora perniciosa in the Brazilian Amazon

Moniliophthora perniciosa is the causal agent of witches’ broom in Theobroma cacao (cacao). Three biotypes of M. perniciosa are recognized, differing in host specificity, with two causing symptoms on cacao or Solanaceae species (C‐ and S‐biotypes), and the third found growing endophytically on lianas (L‐biotype). The objectives of this study were to clarify the genetic relationship between the three biotypes, and to identify those regions in the Brazilian Amazon with the greatest genetic diversity for the C‐biotype. Phylogenetic reconstruction based on the rRNA ITS regions showed that the C‐ and S‐biotypes formed a well‐supported clade separated from the L‐biotype. Analysis of 131 isolates genotyped at 11 microsatellite loci found that S‐ and especially L‐biotypes showed a higher genetic diversity. A significant spatial genetic structure was detected for the C‐biotype populations in Amazonia for up to 137 km, suggesting ‘isolation by distance’ mode of dispersal. However, in regions containing extensive cacao plantings, C‐biotype populations were essentially ‘clonal’, as evidenced by high frequency of repeated multilocus genotypes. Among the Amazonian C‐biotype populations, Acre and West Amazon displayed the largest genotypic diversity and might be part of the centre of diversity of the fungus. The pathogen dispersal may have followed the direction of river flow downstream from Acre, Rondônia and West Amazon eastward to the rest of the Amazon valley, where cacao is not endemic. The Bahia population exhibited the lowest genotypic diversity, but high allele richness, suggesting multiple invasions, with origin assigned to Rondônia and West Amazon, possibly through isolates from the Lower Amazon population.     

Dominance of a single clonal lineage in the Phytophthora infestans population from northern Shaanxi,China revealed by genetic and phenotypic diversity analysis

Late blight caused by Phytophthora infestans is the most serious disease of potato worldwide. To understand the P. infestans population structure in northern Shaanxi, an emerging potato production region in China, 125 single‐lesion isolates were randomly collected from farmers' fields in 2009 and characterized phenotypically and genotypically. A mating type assay showed that 94 isolates were A1 mating type. Virulence determination of selected isolates on a set of differential potato lines containing R1 to R11, respectively, showed the presence of two pathotypes, of which the pathotype lacking avirulence genes Avr3, Avr4 and Avr10 was dominant. Isolates lacking all avirulence factors Avr1 to Avr11 were detected but at lower frequency (13·6%). Analysis for mtDNA haplotype showed all 61 examined isolates were IIa. A total of seven multilocus genotypes were distinguished among 125 isolates, as determined with seven polymorphic microsatellite markers. The genotype SG‐1 was dominant in the population with a frequency of 75·2% and was present throughout the region. Analysis of the phenotypic and genotypic structures of P. infestans populations indicated strict clonal reproduction of the pathogen and suggested that sexual reproduction probably does not occur. Potential implications for disease management are discussed.     

Population genetic diversity and structure of the pecan scab pathogen,Venturia effusa,on cv. Desirable and native seedlings,and the impact of marker number

Scab (Venturia effusa) is the major cause of economic loss in pecan in the south-eastern USA. We explored population genetic diversity and structure among orchards of cv. Desirable and native seedlings, and within-orchard variability among trees of all cultivars sampled. We compared the ability of 30, 15 and 7 previously developed microsatellites to characterize the population genetic diversity and structure of V. effusa. Analyses of molecular variance (AMOVA) provided little evidence of structure dependent on cultivar, but there was some evidence of structure between orchards of a cultivar based on distance. Individual orchard AMOVA showed that three of 11 orchards had between-tree population structure. Among six populations from cv. Desirable, a Mantel test showed that geographic distance was related to the pairwise genetic divergence (R² = 0.84). Among 11 orchards of various cultivars there was little difference in diversity using 30, 15 or 7 markers, or population structure based on AMOVA. Some minor differences in population structure were seen based on discriminant analysis of principal components, or dendrograms. Thus, depending on the objectives, future studies may use as few as 15 or 7 markers without losing ability to discern population genetic diversity or structure. More populations exhibited linkage disequilibrium when using 15 or 30 markers compared to when using seven markers. Knowledge of population genetics of V. effusa in relation to host genotype is needed to understand pathogen population interactions and gene flow, knowledge that will help underpin future breeding efforts to develop durable resistance in this long-lived orchard tree.     

Insights into the genetic diversity and evolution of Little cherry virus 1

Little cherry virus 1 (LChV‐1), a member of the recently proposed genus Velarivirus, is a sweet cherry pathogen that has been recently reported to infect other Prunus species and is associated with various plant disorders. In this work the incidence of the virus on its putative hosts and possible mechanisms driving its evolution were investigated. Due to problems encountered with LChV‐1 detection, a new nested RT‐PCR assay was developed and applied. The virus was found to be prevalent in cherry plantations in Greece and only occasionally detected in other Prunus species. Sequences corresponding to the partial RNA‐dependent RNA polymerase (RdRp), heat‐shock protein homologue (HSP70h) and coat protein (CP) genes were determined from Greek LChV‐1 isolates originating from different hosts; these were analysed, along with published homologous genomic regions from other isolates. Phylogenetic analysis of the three genes revealed the segregation of four evolutionary distinct groups showing no host or geography‐based clustering. Mean genetic distances among the four groups were high with the CP region showing the highest divergence, although intragroup variability levels were low. Nevertheless, estimations of the mean ratio of nonsynonymous substitutions per synonymous site to synonymous substitutions per synonymous site (dN/dS) for the partial RdRp, HSP70h and CP indicated that these genomic regions are under negative selection pressure. Interestingly, a recombination event was identified at the 3′ end of RdRp on a Greek virus isolate, thus highlighting the role of this mechanism in the evolutionary history of LChV‐1.     

Genetic diversity and pathogenicity of Monilinia polystroma – the new pathogen of cherries

Brown rot caused by fungi belonging to the genus Monilinia is one of the major limiting factors of sour and sweet cherry production. Up to now, three species, M. fructigena, M. laxa and M. fructicola, have been identified as causal agents of brown rot on cherries worldwide. From 2010 to 2013, during the monitoring of cherry orchards in different areas of Poland, a fourth species, M. polystroma, was isolated from brown rot symptoms on sour and sweet cherry fruits. To the best of the authors’ knowledge, this is the first time M. polystroma has been reported as the causal agent of brown rot on cherries. The genetic diversity of M. polystroma isolates from cherries and other hosts was analysed using PCR MP, ISSR and RAPD techniques and showed its clear distinctness from other Monilinia spp. tested. The cluster analysis of fingerprinting data revealed a high similarity of M. polystroma isolates from Poland and their close relationship with the reference strain from Japan, indicating that this species is a recently introduced pathogen. The highest genetic distance between the examined isolates and the highest number of different genotypes was observed in an ISSR assay. Detailed genetic diversity characteristics revealed that M. polystroma isolates from cherries did not create a distinct group but were intermingled with M. polystroma isolates from other hosts. The results of the pathogenicity test conducted on different fruit species indicated a lack of host specificity for M. polystroma isolates.     

Population structure and migration of the witches’ broom pathogen Moniliophthora perniciosa from cacao and cultivated and wild solanaceous hosts in southeastern Brazil

Moniliophthora perniciosa, causal agent of witches’ broom disease in cacao plantations in South America and the Caribbean Islands, has co‐evolved with its host cacao, but the pathogen has also emerged in many solanaceous hosts in Brazil, including economically important food crops and wild species. This study was carried out to: (i) determine the existence of host subpopulations of M. perniciosa in Brazil; (ii) estimate gene and genotypic diversity of M. perniciosa host subpopulations infecting solanaceous hosts in southeastern Bahia and Minas Gerais states, Brazil; and (iii) estimate the amount and directionality of historical migration of M. perniciosa subpopulations. Up to 203 M. perniciosa isolates collected from solanaceous hosts with symptoms from Bahia and Minas Gerais states in Brazil and from Theobroma spp. (cacao) and Herrania spp. were characterized with 11 microsatellite markers. Factorial correspondence analyses, minimum‐spanning network and Bayesian clustering revealed genetic clusters associated with their host of origin. Significant subpopulation differentiation was evident (Φ_ST = 0.30, P ≤ 0.05) among M. perniciosa host subpopulations. Most of the multilocus microsatellite genotypes (MLMGs) were host‐specific, with few MLMGs shared among subpopulations. Pairwise comparisons among M. perniciosa host subpopulations were significant, except between jurubeba (Solanum paniculatum) and cultivated solanaceous subpopulations. The combined analyses rejected the null hypothesis that M. perniciosa in Brazil is a single genetic population not structured by host. These findings support a scenario of introduction and subsequent adaptation to solanaceous hosts that should be taken into consideration to improve mitigation and management of M. perniciosa.     

Host switching between native and non‐native trees in a population of the canker pathogen Chrysoporthe cubensis from Colombia

The purpose of this study was to test the hypothesis that Chrysoporthe cubensis on native trees in South America could be the source of the pathogen that causes severe stem cankers and often mortality in commercially propagated Eucalyptus trees. This was done by investigating populations originating from two adjacent Eucalyptus (Myrtaceae) plantations in Colombia, and wild Miconia rubiginosa trees (Melastomataceae) growing alongside these stands. Polymorphic microsatellite markers were used to quantify allele sizes in 20 and 39 isolates from the two Eucalyptus stands and 32 isolates from adjacent M. rubiginosa trees. Gene and genotypic diversities were calculated from these data, and population differentiation and assignment tests were performed to ascertain whether the populations were genetically different. Results showed that there were no differences between any of the populations using these techniques, and that they can be treated as a single population. Therefore, the results support the hypothesis that host switching has occurred in C. cubensis in Colombia.     

Genetic diversity and population structure analysis of isolates of the rice false smut pathogen Ustilaginoidea virens in India

Genetic diversity assessment and population structure analysis are essential for characterization of pathogens and their isolates. Markers are essential tools for exploring genetic variation among the isolates. False smut of rice caused by Ustilaginoidea virens, formerly Villosiclava virens, is a major emerging disease of rice in India. A high level of variability is observed at the field level, but no information is available from India on genetic diversity and population structure. This is the first report of genetic diversity and population structure of U. virens from India that included 63 isolates distributed across the vast geographical area of eastern and north-eastern India (18.9 to 26.7°N and 82.6 to 94.2°E). Seventeen RAPDs and 14 SSRs were identified as polymorphic and a total of 140 alleles were detected across the populations. The average number of alleles per locus for each primer was 4.5. All the isolates were grouped into two major clusters, with partial geographical segregation that was supported by principal coordinate analysis. Mantel test suggested genetic distance within the isolates increased with increasing geographical distance. Analysis of molecular variation showed more genetic variation within populations and less among populations. This outcome will help in understanding genetic diversity of U. virens from eastern and north-eastern India and in planning effective management strategies.     

西藏小麦条锈菌群体结构与遗传多样性

为查明西藏小麦条锈菌Puccinia striiformis f. sp. tritici群体结构和遗传多样性,采用中国鉴别寄主和近等基因系鉴别寄主,以及竞争性等位基因特异性PCR-单核苷酸多态性（kompetitive al-lele specific PCR-single nucleotide polymorphism,KASP-SNP）分子标记对2017年采自西藏的150个小麦条锈菌菌系分别进行表型分析和基因型分析。表型分析结果显示,中国鉴别寄主将150个菌系区分为 12 个已知小种、6 个已知致病类型和 13 个未知致病类型,所有菌系均不能侵染中四和Triticum spelta album鉴别寄主。近等基因系鉴别寄主将150个菌系区分为88个毒性类型,这些毒性类型均不侵染携带抗性基因Yr5、Yr10或Yr15的品种。基因型分析结果显示,26对引物将150个菌系划分为73个基因型,表明西藏小麦条锈菌群体基因型丰富。基因流分析结果表明,波密县与洛扎县小麦条锈菌亚群体之间的基因流N_m最高,达5.86,米林县西部与波密县、洛扎县、巴宜县、米林县东部条锈菌亚群体之间的N_m较低,分别为0.25、0.34、0.42和0.67,表明西藏不同地区条锈菌群体之间基因交流强度差异较大。说明西藏作为我国小麦条锈病的独立流行区,条锈菌群体毒性结构复杂,遗传多样性高。     

Microsatellite and mating type markers reveal unexpected patterns of genetic diversity in the pine root‐infecting fungus Grosmannia alacris

Grosmannia alacris is a fungus commonly associated with root‐infesting bark beetles occurring on Pinus spp. The fungus has been recorded in South Africa, the USA, France, Portugal and Spain and importantly, has been associated with pine root diseases in South Africa and the USA. Nothing is known regarding the population genetics or origin of G. alacris, although its association with root‐infesting beetles native to Europe suggests that it is an invasive alien in South Africa. In this study, microsatellite markers together with newly developed mating type markers were used to characterize a total of 170 isolates of G. alacris from South Africa and the USA. The results showed that the genotypic diversity of the South African population of G. alacris was very high when compared to the USA populations. Two mating types were also present in South African isolates and the MAT1‐1/MAT1‐2 ratio did not differ from 1:1 (χ² = 1·39, P = 0·24). This suggests that sexual reproduction most probably occurs in the fungus in South Africa, although a sexual state has never been seen in nature. In contrast, the large collection of USA isolates harboured only a single mating type. The results suggest that multiple introductions, followed by random mating, have influenced the population structure in South Africa. In contrast, limited introductions of probably a single mating type (MAT1‐2) may best explain the clonality of USA populations.     

Genetic analysis of the population structure of the rice false smut fungus,Villosiclava virens,in China using microsatellite markers mined from a genome assembly

Studies on population genetics of Villosiclava virens are limited because of the lack of polymorphic markers. Based on a draft genome sequence of isolate HWD‐2 produced in this study, 20 of 403 potential simple sequence repeats (SSR) loci showed polymorphisms in preliminary screening using eight diverse V. virens isolates. Among polymorphic loci, most of them with tetra‐ to hexanucleotide unit motifs showed higher levels of polymorphism than loci with smaller motifs. After testing with 20 polymorphic SSR markers, the 87 isolates of V. virens from eight populations in China showed a high level of genetic diversity, with each as a unique haplotype. This differs from some previous findings showing little to no genetic variation based on random amplified polymorphic DNA and amplified fragment length polymorphism analyses. Among eight populations from major rice production areas of China, the population from Guangxi province in south China showed the highest levels of polymorphism, which led to the speculation that it might be closer to the centre of origin of this pathogen. The northern, central and eastern populations (Jilin, Liaoning, Hubei, Hunan, Jiangxi and Zhejiang), when considered together as a group, showed significant molecular variation compared to the southern populations (Fujian and Guangxi) (Φ_PT = 0·043, P = 0·037). A significant relationship (Mantel test, P = 0·027) but with low correlation (R² = 0·23) was also found between geographic distance and genetic distance. The 20 polymorphic SSR primer pairs designed in this study provide a tool for further research on the population diversity of this emerging fungal pathogen of rice.     

Occurrence and diversity of Tomato spotted wilt virus isolates breaking the Tsw resistance gene of Capsicum chinense in Yunnan,southwest China

Widely used resistant peppers (Capsicum spp.) bearing the Tsw locus triggered the rapid emergence of resistance‐breaking (RB) isolates of Tomato spotted wilt virus (TSWV) around the world. However, although TSWV‐induced diseases have rapidly increased in Yunnan, southwest China, in recent years, no information is available about the diversity of TSWV isolates in this region. In this study, the occurrence of natural TSWV RB variants among isolates collected in Yunnan is reported. Initially, a TSWV isolate from asparagus lettuce (TSWV‐LE) was collected in Yunnan in 2012. Surprisingly, this isolate of TSWV induced systemic necrosis on pepper carrying the Tsw resistance gene. Novel TSWV isolates, collected in 2015, included a tomato isolate (TSWV‐YN18) and a tobacco isolate (TSWV‐YN53) that also overcame Tsw‐mediated resistance. TSWV‐YN18 induced systemic ringspots, whereas TSWV‐YN53 caused systemic chlorotic mottling. Variations in the TSWV nonstructural (NSs) protein are the key determinants associated with Tsw resistance‐breaking isolates. It was found that TSWV‐LE NSs retained the hypersensitive response (HR) induction, whereas TSWV‐YN18 and TSWV‐YN53 NSs were unable to induce HR. However, the NSs of all three RB isolates suppressed RNA silencing. Sequence analysis of the NSs revealed that RB isolates of Yunnan have no amino acid mutation sites common to other previously reported RB isolates. However, two amino acids (F74 and K272) on TSWV‐LE NSs make it distinct from TSWV‐YN18 and TSWV‐YN53. The occurrence of different RB isolates and the failure of Tsw‐mediated resistance control pose serious threats to domestic pepper crops in southwest China.     

Genetic and phenotypic diversity of Mediterranean populations of the olive knot pathogen,Pseudomonas savastanoi pv. savastanoi

Pseudomonas savastanoi pv. savastanoi (Psav) is a member of P. syringae sensu lato, and causes olive knot disease, a disease first reported over 2000 years ago. Analysing 124 isolates of Psav from 15 countries by rep‐PCR, the population genetic structure of Psav was investigated. A total of 113 distinct fingerprints were detected. Cluster analysis revealed the existence of two clusters and four subclusters. These clusters were associated with the geographic origin of isolates, which in turn correspond to historic human migration events and trade routes across the Mediterranean Sea. In contrast, multilocus sequence typing (MLST) of 2788 bp of the gapA, gltA, gyrB and rpoD genes found only one variable site among 77 representative isolates. Virulence variation was observed within the Psav population, with the most virulent strains generating knots that had a weight that was 10‐fold greater than those generated by the least virulent strains. Taken together, these data suggest that today's Psav population is the result of clonal expansion of a single strain, that moderate migration of the pathogen occurred between countries, and that changes in virulence arose during its evolution.     

基于mtCOI基因序列的我国云南省和缅甸、柬埔寨草地贪夜蛾种群遗传多样性分析

为制定有效的草地贪夜蛾防控措施,分别自缅甸、柬埔寨和我国云南省采集4、2和8个种群共542个草地贪夜蛾样品,基于mtCOI基因序列分析这14个种群的遗传多样性指数、遗传分化系数及基因流。结果表明,缅甸草地贪夜蛾种群的单倍型多样性指数和平均核苷酸差异数分别介于0.273～0.396和4.643～6.727之间,高于我国云南省的草地贪夜蛾种群,分别介于0.047～0.214和0.791～3.636之间;除ZT种群、LS种群和TO种群外,其它11个种群的Fu’s F均为显著正值,表明缅甸、柬埔寨和我国云南省的草地贪夜蛾种群未经历种群扩张事件;在14个种群中,LS种群与其它种群分化较明显,TK种群、MY种群、CP种群、MS种群和KY种群有效迁入个数和有效迁出个数之和较高,分别为699.41、682.50、855.76、684.56和701.31,推测这5个种群在草地贪夜蛾基因交流中起着类似"中转站"的作用。     

Development,characterization and application of genomic SSR markers for the oat stem rust pathogen Puccinia graminis f. sp. avenae

Oat stem rust, caused by Puccinia graminis f. sp. avenae (Pga), is one of the most severe diseases of oats worldwide. Population studies are scarce for this pathogen, mainly due to the lack of polymorphic molecular markers suitable for genetic analysis. In this study, an Australian Pga isolate was sequenced, the abundance of simple sequence repeats (SSRs) was determined and PCR‐based polymorphic markers suitable for genetic diversity analysis were developed. The amplification of 194 primer pairs was initially assessed using a set of 12 isolates of different cereal rust species and their formae speciales. A high frequency of cross‐species amplification was observed for most markers; however, 36 SSRs were diagnostic for P. graminis only. A subset of 19 genome‐derived SSRs were deemed useful for genetic diversity analysis of Pga and were assessed on 66 Pga isolates from Australia, Brazil and Sweden. Brazilian and Australian isolates were characterized by one and two predominant clonal lineages, respectively. In contrast, the Swedish isolates, previously shown to undergo sexual recombination, were highly diverse (nine distinct genotypes out of 10 isolates) and divided into two subpopulations. The genome‐derived SSR markers developed in this study were well suited to the population studies undertaken, and have diagnostic capabilities that should aid in the identification of unknown rust pathogen species.     

The origin and genetic diversity of the causal agent of Asian soybean rust,Phakopsora pachyrhizi,in South America

A sequence‐based approach was used to investigate molecular genetic variations in Phakopsora pachyrhizi, an obligate biotrophic pathogen that causes Asian soybean rust. In Argentina, the samples came from uredinium‐bearing leaves taken from 11 soybean fields; in Brazil, the samples comprised urediniospores from leaves of 10 soybean genotypes that had been grown in three experimental stations during two growing seasons. PCR‐based cloning techniques were used to generate DNA sequences for two gene regions and alignments were supplemented with data from GenBank. A total of 575 sequences for the internal transcribed spacer region (18 ribotypes) and 160 partial sequences for a housekeeping gene encoding ADP‐ribosylation factor (10 haplotypes) were obtained. Ribotype accumulation curves predicted that about 20 bacterial clones would recover 5–6 ribotypes (c. 70–80% of the total molecular variation) per locality. The samples from the three experimental stations in Brazil displayed most (14 out of 16) ribotypes found worldwide; the lack of genetic structure and differentiation at a diverse geographic scale suggests that both local and distant sources provide airborne inoculum during disease establishment. Soybean genotypes with resistance genes for the Asian soybean rust did not decrease the molecular genetic variation of fungal populations.     

Characterization,genetic diversity and distribution of Xanthomonas campestris pv. campestris races causing black rot disease in cruciferous crops of India

Black rot, caused by Xanthomonas campestris pv. campestris (Xcc) is a disease of crucifer crops. The objective of this study was to characterize races of Xcc, their distribution and genetic diversity in India. Two hundred and seventeen isolates of bacteria were obtained from 12 different black rot‐infected crucifer crops from 19 states of India; these were identified as Xcc based on morphology, hrpF gene and 16S rRNA gene based molecular markers and pathogenicity tests. Characterization of races was performed by using a set of seven differential crucifer hosts, comprising two cultivars of turnip (Brassica rapa var. rapa) and cultivars of Indian mustard (B. juncea), Ethiopian mustard (B. carinata), rapeseed mustard (B. napus), cauliflower (B. oleracea) and Savoy cabbage (B. oleracea var. sabauda). Races 1, 4 and 6 of Xcc were identified and, among these races, race 1 followed by race 4 dominated most of the states of India. Genetic diversity of the Indian isolates of Xcc was analysed using repetitive sequence‐based PCR (rep‐PCR) including primers for REP (repetitive extragenic palindromic), ERIC (enterobacterial repetitive intergenic consensus) and BOX (amplifying with BOX A1 R primer) repetitive elements. This method of fingerprinting grouped the isolates into 56 different DNA types (clusters) with a 75% similarity coefficient. Among these clusters, DNA types 22 and 53 contained two different races 1 and 4, whereas DNA type 12 contained races 1, 4 and 6. However, no clear relationship was observed between fingerprints and races, hosts or geographical origin.