SSR Variability of Stem Rust Pathogen on Spring Bread Wheat in Russia.

Wheat stem rust, caused by Puccinia graminis f. sp. tritici, which used to be a harmful disease of winter wheat in the southern part of Russia, has been largely affecting the yield of spring bread wheat in the territories of the temperate climate zone since 2009. In total, 222 P. graminis f. sp. tritici isolates were obtained from samples of susceptible cultivars of spring bread wheat in Central and Volga regions and Omsk and Novosibirsk provinces in 2019. Genotyping of the isolates was carried out at 16 simple-sequence repeat (SSR) loci. Number of alleles, proportion of heterozygotes, and deviation from Hardy-Weinberg equilibrium were determined at each SSR locus. Based on genetic variability of SSR genotypes, it was shown that the P. graminis f. sp. tritici population is subdivided into two large clusters in the territory of the Russian temperate climate zone: the "European" population (the Central region) and the "Asian" one (the Volga region and two main wheat provinces of Western Siberia). Both of the P. graminis f. sp. tritici populations are characterized by a mixed mode of reproduction (sexual and clonal) but different sources of inoculum seem to shape a genotype structure within them. A group of P. graminis f. sp. tritici genotypes with high variability, the inbreeding coefficient closed to zero, and low observed heterozygosity was revealed among samples from Omsk. Moreover, two singular SSR genotypes identified among the Asian samples of P. graminis f. sp. tritici isolates should attract special attention in the monitoring of stem rust in order to disclose unexpected rapid changes of the pathogen in the corresponding regions and to prevent disease outbreak.

Virulence Survey of Puccinia striiformis in Israel Revealed Considerable Changes in the Pathogen Population During the Period 2001 to 2019.

A total of 353 urediniospore isolates of Puccinia striiformis f. sp. tritici (Pst) collected in Israel during 2001 to 2019 were analyzed. Pst pathogenicity was studied with a set of 20 differentials (17 Avocet and 3 other lines). Three periods were compared: 2001 to 2007, 2009 to 2016, and 2017 to 2019. No virulence to Yr5 or Yr15 was detected. Virulence frequencies on Yr4, Yr10, Yr24, and YrSp genes rose to the moderate level (0.28 to 0.44) in 2017 to 2019. Virulence frequencies to Yr2 and Yr9 decreased. One Pst phenotype was identified in all three periods, but its frequency drastically decreased from 0.74 in 2001 to 2016 to 0.21 in 2017 to 2019. The most probable scenario of emergence of wheat yellow rust in Israel is wind dissemination of Pst urediniospores from the Horn of Africa. Variability of the Pst population increased amid considerable evolution with two major transformations in 2009 and 2017. The first modification can be attributed to changes in wheat genetic background in Israel due to deployment of new cultivars resistant to yellow rust since 2004. The second shift in 2017 can be primarily explained by intensive deployment of wheat cultivars resistant to the stem rust race Ug99 in the 2010s in the Horn of Africa. This led to changing genetic backgrounds of the cultivated wheats in the donor region and development and long-distance spread of new Pst phenotypes to Israel. Two singular multivirulent Pst phenotypes were identified in 2019, one of them being closely related to the aggressive Warrior race. Such phenotypes may potentially defeat existing resistances.

Analysis of Host-Specific Differentiation of Puccinia striiformis in the South and North-West of the European Part of Russia.

Yellow (stripe) rust, caused by Puccinia striiformis Westend. (Pst), is a major disease of cereals worldwide. We studied Pst virulence phenotypes on Triticum aestivum, Triticum durum, and triticale in three geographically distant regions of the European part of Russia (Dagestan and Krasnodar in North Caucasus, and Northwest) with different climate and environmental conditions. Based on the set of twenty differential lines, a relatively high level of population diversity was determined with 67 different Pst pathotypes identified among 141 isolates. Only seven pathotypes were shared by at least two hosts or occurred in the different regions. No significant differentiation was found between regional Pst collections of pathotypes either from T. aestivum or from T. durum. A set of Pst pathotypes from triticale was subdivided into two groups. One of them was indistinguishable from most durum and common wheat pathotypes, whereas the second group differed greatly from all other pathotypes. All sampled Pst isolates were avirulent on lines with Yr5, Yr10, Yr15, and Yr24 genes. Significant variation in virulence frequency among all Pst collections was observed on lines containing Yr1, Yr3, Yr17, Yr27, and YrSp genes and cvs Strubes Dickkopf, Carstens V, and Nord Desprez. Relationships between Russian regional collections of Pst from wheat did not conform to those for P. triticina.

Race Characterization and Molecular Genotyping of Puccinia triticina Populations from Durum Wheat in Russia.

Variability of the Russian population of Puccinia triticina from durum wheat was studied with virulence and simple sequence repeat (SSR) markers. The pathogen was sampled during 2017 to 2019 in all regions with sizable durum wheat (Triticum durum) growing areas from winter (North Caucasus) and spring (Middle Volga, Ural, and West Siberia) wheat. A total of 474 isolates were tested on a set of 20 Lr-gene lines. Molecular genotypes for 105 selected isolates were determined at 11 SSR loci. Variable virulence/avirulence reaction was observed only on three Lr-gene lines, whereas just five SSR loci were polymorphic with two alleles at each. Seven different virulence phenotypes and 11 SSR genotypes were found among 474 and 105 isolates, respectively, indicating a very low variability of the pathogen. One virulence phenotype and three SSR genotypes occurred in all Russian regions. However, two phenotypes were specific to the European regions of Russia (North Caucasus and Middle Volga), while another two were found only in the Asian part of Russia (Ural and West Siberia). Significant differentiation between six populations of P. triticina from durum wheat in the Asian and European (mainly North Caucasus) regions was also shown with numerous metrics and approaches for data with and without clone correction. Relationships among the regional populations of P. triticina from durum wheat established with virulence phenotypes significantly associated with those for SSR genotypes and was similar to the relationships among the regional populations of the pathogen from common wheat.

Severe limitations of the FEve metric of functional evenness and some alternative metrics.

The metric of functional evenness FEve is an example of how approaches to conceptualizing and measuring functional variability may go astray. This index has several critical conceptual and practical drawbacks: Different values of the FEve index for the same community can be obtained if the species have unequal species abundances; this result is highly likely if most of the traits are categorical.Very minor differences in even one pairwise distance can result in very different values of FEve.FEve uses only a fraction of the information contained in the matrix of species distances. Counterintuitively, this can cause very similar FEve scores for communities with substantially different patterns of species dispersal in trait space.FEve is a valid metric only if all species have exactly the same abundances. However, the meaning of FEve in such an instance is unclear as the purpose of the metric is to measure the variability of abundances in trait space. We recommend not using the FEve metric in studies of functional variability. Given the wide usage of FEve index over the last decade, the validity of the conclusions based on those estimates is in question. Instead, we suggest three alternative metrics that combine variability in species distances in trait space with abundance in various ways. More broadly, we recommend that researchers think about which community properties (e.g., trait distances of a focus species to the nearest neighbor or all other species, variability of pairwise interactions between species) they want to measure and pick from among the appropriate metrics.

Stem Endophytic Mycobiota in Wild and Domesticated Wheat: Structural Differences and Hidden Resources for Wheat Improvement.

Towards the identification of entophytic fungal taxa with potential for crop improvement, we characterized and compared fungal endophyte communities (FECs) from domesticated bread wheat and two wheat ancestors, Aegilopssharonensis and Triticumdicoccoides. Data generated by next generation sequencing identified a total of 1666 taxa. The FECs in the three plant species contained high proportions of random taxa with low abundance. At plant species level, the majority of abundant taxa were common to all host plants, and the collective FECs of each of the three plant species had similar diversity. However, FECs from the wild plants in specific sites were more diverse and had greater richness than wheat FECs from corresponding specific fields. The wild plants also had higher numbers of differentially abundant fungal taxa than wheat, with Alternaria infectoria being the most abundant species in wild plants and Candida sake the most abundant in wheat. Network analysis on co-occurrence association revealed a small number of taxa with a relatively high number of co-occurrence associations, which might be important in community assembly. Our results show that the actual endophytic cargo in cultivated wheat plants is limited relative to wild plants, and highlight putative functional and hub fungal taxa with potential for wheat improvement.

Significant host- and environment-dependent differentiation among highly sporadic fungal endophyte communities in cereal crops-related wild grasses.

Endophytic fungi compose a significant part of plant microbiomes. However, while a small number of fungal taxa have proven beneficial impact, the vast majority of fungal endophytes remain uncharacterized, and the drivers of fungal endophyte community (FEC) assembly are not well understood. Here, we analysed FECs in three cereal crops-related wild grasses - Avena sterilis, Hordeum spontaneum and Aegilops peregrina - that grow in mixed populations in natural habitats. Taxa in Ascomycota class Dothideomycetes, particularly the genera Alternaria and Cladosporium, were the most abundant and prevalent across all populations, but there was also high incidence of basidiomyceteous yeasts of the class Tremellomycetes. The fungal community was shaped to large extent by stochastic processes, as indicated by high level of variation even between individuals from local populations of the same plant species, and confirmed by the neutral community model and Raup-Crick index. Nevertheless, we still found strong determinism in FEC assembly with both incidence and abundance data sets. Substantial differences in community composition across host species and locations were revealed. Our research demonstrated that assembly of FECs is affected by stochastic as well as deterministic processes and suggests strong effects of environment heterogeneity and plant species on community composition. In addition, a small number of taxa had high incidence and abundance in all of the 15 populations. These taxa represent an important part of the core FEC and might be of general functional importance.

Measuring similarity between gene interaction profiles.

BACKGROUND: Gene and protein interaction data are often represented as interaction networks, where nodes stand for genes or gene products and each edge stands for a relationship between a pair of gene nodes. Commonly, that relationship within a pair is specified by high similarity between profiles (vectors) of experimentally defined interactions of each of the two genes with all other genes in the genome; only gene pairs that interact with similar sets of genes are linked by an edge in the network. The tight groups of genes/gene products that work together in a cell can be discovered by the analysis of those complex networks. RESULTS: We show that the choice of the similarity measure between pairs of gene vectors impacts the properties of networks and of gene modules detected within them. We re-analyzed well-studied data on yeast genetic interactions, constructed four genetic networks using four different similarity measures, and detected gene modules in each network using the same algorithm. The four networks induced different numbers of putative functional gene modules, and each similarity measure induced some unique modules. In an example of a putative functional connection suggested by comparing genetic interaction vectors, we predict a link between SUN-domain proteins and protein glycosylation in the endoplasmic reticulum. CONCLUSIONS: The discovery of molecular modules in genetic networks is sensitive to the way of measuring similarity between profiles of gene interactions in a cell. In the absence of a formal way to choose the "best" measure, it is advisable to explore the measures with different mathematical properties, which may identify different sets of connections between genes.

Dissimilarity of individual microsatellite profiles under different mutation models: Empirical approach.

Microsatellites (simple sequence repeats, SSRs) still remain popular molecular markers for studying neutral genetic variation. Two alternative models outline how new microsatellite alleles evolve. Infinite alleles model (IAM) assumes that all possible alleles are equally likely to result from a mutation, while stepwise mutation model (SMM) describes microsatellite evolution as stepwise adding or subtracting single repeat units. Genetic relationships between individuals can be analyzed in higher precision when assuming the SMM scenario with allele size differences as a proxy of genetic distance. If population structure is not predetermined in advance, an empirical data analysis usually includes (a) estimating proximity between individual SSR profiles with a selected dissimilarity measure and (b) determining putative genetic structure of a given set of individuals using methods of clustering and/or ordination for the obtained dissimilarity matrix. We developed new dissimilarity indices between SSR profiles of haploid, diploid, or polyploid organisms assuming different mutation models and compared the performance of these indices for determining genetic structure with population data and with simulations. More specifically, we compared SMM with a constant or variable mutation rate at different SSR loci to IAM using data from natural populations of a freshwater bryozoan Cristatella mucedo (diploid), wheat leaf rust Puccinia triticina (dikaryon), and wheat powdery mildew Blumeria graminis (monokaryon). We show that inferences about population genetic structure are sensitive to the assumed mutation model. With simulations, we found that Bruvo's distance performs generally poorly, while the new metrics are capturing the differences in the genetic structure of the populations.

Cost of resistance to trematodes in freshwater snail populations with low clonal diversity.

BACKGROUND: The persistence of high genetic variability in natural populations garners considerable interest among ecologists and evolutionary biologists. One proposed hypothesis for the maintenance of high levels of genetic diversity relies on frequency-dependent selection imposed by parasites on host populations (Red Queen hypothesis). A complementary hypothesis suggests that a trade-off between fitness costs associated with tolerance to stress factors and fitness costs associated with resistance to parasites is responsible for the maintenance of host genetic diversity. RESULTS: The present study investigated whether host resistance to parasites is traded off with tolerance to environmental stress factors (high/low temperatures, high salinity), by comparing populations of the freshwater snail Melanoides tuberculata with low vs. high clonal diversity. Since polyclonal populations were found to be more parasitized than populations with low clonal diversity, we expected them to be tolerant to environmental stress factors. We found that clonal diversity explained most of the variation in snail survival under high temperature, thereby suggesting that tolerance to high temperatures of clonally diverse populations is higher than that of populations with low clonal diversity. CONCLUSIONS: Our results suggest that resistance to parasites may come at a cost of reduced tolerance to certain environmental stress factors.

The components of biodiversity, with a particular focus on phylogenetic information.

We present a framework for biodiversity metrics that organizes the growing panoply of metrics. Our framework distinguishes metrics based on the type of information-abundance, phylogeny, function-and two common properties-magnitude and variability. Our new metrics of phylogenetic diversity are based on a partition of the total branch lengths of a cladogram into the proportional share of each species, including: a measure of divergence which standardizes the amount of evolutionary divergence by species richness and time depth of the cladogram; a measure of regularity which is maximal when the tree is perfectly symmetrical so that all species have the same proportional branch lengths; a measure that combines information on the magnitude and variability of abundance with phylogenetic variability, and a measure of phylogenetically weighted effective mean abundance; and indicate how those metrics can be decomposed into α and ß components. We illustrate the utility of these new metrics using empirical data on the bat fauna of Manu, Peru. Divergence was greatest in lowland rainforest and at the transition between cloud and elfin forests, and least in upper elfin forests and in cloud forests. In contrast, regularity was greatest in lowland rainforest, dipping to its smallest values in mid-elevation cloud forests, and then increasing in high elevation elfin forests. These patterns indicate that the first species to drop out with increasing elevation are ones that are closely related to other species in the metacommunity. Measures of the effective number of phylogenetically independent or distinct species decreased very rapidly with elevation, and ß-diversity was larger. In contrast, a comparison of feeding guilds shows a different effect of phylogenetic patterning. Along the elevational gradient, each guild generally loses some species from each clade-rather than entire clades-explaining the maintenance of functional diversity as phylogenetic diversity decreases.

Diversity of fungal endophytes in recent and ancient wheat ancestors Triticum dicoccoides and Aegilops sharonensis.

Endophytes have profound impacts on plants, including beneficial effects on agriculturally important traits. We hypothesized that endophytes in wild plants include beneficial endophytes that are absent or underrepresented in domesticated crops. In this work, we studied the structure of endophyte communities in wheat-related grasses, Triticum dicoccoides and Aegilops sharonensis, and compared it to an endophyte community from wheat (T. aeastivum). Endophytes were isolated by cultivation and by cultivation-independent methods. In total, 514 intergenic spacer region sequences from single cultures were analyzed. Categorization at 97% sequence similarity resulted in 67 operational taxonomic units (OTUs) that were evenly distributed between the different plant species. A narrow core community of Alternaria spp. was found in all samples, but each plant species also contained a significant portion of unique endophytes. The cultivation-independent analysis identified a larger number of OTUs than the cultivation method, half of which were singletons or doubletons. For OTUs with a relative abundance >0.5%, similar numbers were obtained by both methods. Collectively, our data show that wild grass relatives of wheat contain a wealth of taxonomically diverse fungal endophytes that are not found in modern wheat, some of which belong to taxa with known beneficial effects.

Differentiation Among Blumeria graminis f. sp. tritici Isolates Originating from Wild Versus Domesticated Triticum Species in Israel.

Israel and its vicinity constitute a center of diversity of domesticated wheat species (Triticum aestivum and T. durum) and their sympatrically growing wild relatives, including wild emmer wheat (T. dicoccoides). We investigated differentiation within the forma specialis of their obligate powdery mildew pathogen, Blumeria graminis f. sp. tritici. A total of 61 B. graminis f. sp. tritici isolates were collected from the three host species in four geographic regions of Israel. Genetic relatedness of the isolates was characterized using both virulence patterns on 38 wheat lines (including 21 resistance gene differentials) and presumptively neutral molecular markers (simple-sequence repeats and single-nucleotide polymorphisms). All isolates were virulent on at least some genotypes of all three wheat species tested. All assays divided the B. graminis f. sp. tritici collection into two distinct groups, those from domesticated hosts and those from wild emmer wheat. One-way migration was detected from the domestic wheat B. graminis f. sp. tritici population to the wild emmer B. graminis f. sp. tritici population at a rate of five to six migrants per generation. This gene flow may help explain the overlap between the distinct domestic and wild B. graminis f. sp. tritici groups. Overall, B. graminis f. sp. tritici is significantly differentiated into wild-emmer and domesticated-wheat populations, although the results do not support the existence of a separate f. sp. dicocci.

Structure and Migration in U.S. Blumeria graminis f. sp. tritici Populations.

While wheat powdery mildew occurs throughout the south-central and eastern United States, epidemics are especially damaging in the Mid-Atlantic states. The structure of the U.S. Blumeria graminis f. sp. tritici population was assessed based on a sample of 238 single-spored isolates. The isolates were collected from 16 locations in 12 states (18 site-years) as chasmothecial samples in 2003 or 2005, or as conidial samples in 2007 or 2010. DNA was evaluated using nine single nucleotide polymorphism (SNP) markers in four housekeeping genes, and 10 simple sequence repeat (SSR) markers. The SSR markers were variably polymorphic, with allele numbers ranging from 3 to 39 per locus. Genotypic diversity was high (210 haplotypes) and in eight of the site-years, every isolate had a different SSR genotype. SNP haplotypic diversity was lower; although 15 haplotypes were identified, the majority of isolates possessed one of two haplotypes. The chasmothecial samples showed no evidence of linkage disequilibrium (P = 0.36), while the conidial samples did (P = 0.001), but the two groups had nearly identical mean levels of genetic diversity, which was moderate. There was a weakly positive relationship between genetic distance and geographic distance (R(2) = 0.25, P = 0.001), indicating modest isolation by distance. Most locations in the Mid-Atlantic and Great Lakes regions clustered together genetically, while Southeast locations formed a distinct but adjacent cluster; all of these were genetically separated from Southern Plains locations and an intermediate location in Kentucky. One-way migration was detected at a rate of approximately five individuals per generation from populations west of the Appalachian Mountains to those to the east, despite the fact that the Atlantic states experience more frequent and damaging wheat mildew epidemics. Overall, the evidence argues for a large-scale mosaic of overlapping populations that re-establish themselves from local sources, rather than continental-scale extinction and re-establishment, and a low rate of long-distance dispersal roughly from west to east, consistent with prevailing wind directions.

Afternoon Ascospore Release in Claviceps purpurea Optimizes Perennial Ryegrass Infection.

In Kentucky bluegrass (Poa pratensis), Claviceps purpurea, the causal agent of ergot, typically releases ascospores during the early-morning hours, between about midnight and 10:00 a.m., corresponding to time of flowering, when the unfertilized ovaries are most susceptible to infection. During aeromycology studies of C. purpurea in perennial ryegrass (Lolium perenne) in northeastern Oregon during 2008 to 2010 and 2013, a strain of C. purpurea was found that released ascospores in the afternoon, coinciding with flowering in perennial ryegrass. Under controlled environmental conditions, sclerotia from perennial ryegrass and Kentucky bluegrass released spores in the afternoon and morning, respectively, consistent with timing of spore release under field conditions. Internal transcribed spacer (ITS) sequences of single sclerotial isolates from Kentucky bluegrass and perennial ryegrass were consistent with C. purpurea, although minor variations in ITS sequences among isolates were noted. Differences between Kentucky bluegrass and perennial ryegrass isolates were observed in random amplified polymorphic DNA. Evidence is provided for adaptation of C. purpurea to perennial ryegrass by means of delayed spore release that coincides with afternoon flowering in perennial ryegrass.

Comprehensive Evaluation of Virulence and Resistance Data: A New Analysis Tool.

The functioning and features of the new software package VAT (Virulence Analysis Tool) are introduced. VAT provides a range of methods for the analysis of plant pathosystems. The techniques are applicable to other binary data sets that are organized in large two-way tables, e.g., molecular marker data. The main features are data entry, descriptive tools, and inference statistics by resampling. About 50 well-established or newly developed indices allow a detailed diversity analysis of sexually and asexually reproducing populations. VAT facilitates a comprehensive, effective, and logically consistent evaluation and presentation of virulence and resistance data. A translation option simplifies the comparison of results from differently coded pathotypes. The software package comes with a detailed manual and is freely available on the internet at tau.ac.il/lifesci/departments/plant_s/members/kosman/VAT.html and at va-tipp.de .

Conceptual analysis of methods applied to assessment of diversity within and distance between populations with asexual or mixed mode of reproduction.

Measures of diversity within populations, and distance between populations, are compared for organisms with an asexual or mixed mode of reproduction. Examples are drawn from studies of plant pathogenic fungi based on binary traits including presence/absence of DNA bands or virulence/avirulence to differential hosts. Commonly used measures of population diversity or genetic distance consider either genotype frequencies or allele frequencies. Kosman's diversity and distance measures are the most suitable for populations with an asexual or mixed mode of reproduction, because by considering genetic patterns of all individuals they take into account not just the genotype frequencies but also the genetic similarities between genotypes in the populations. The Kosman distance and diversity measures for populations can be calculated using different measures of dissimilarity between individuals (the simple mismatch, Jaccard and Dice coefficients of dissimilarity). Kosman's distances based on the simple mismatch and Jaccard dissimilarities are metrics. Comparisons of diversity indices for hypothetical examples as well as for actual data sets are presented to demonstrate that inferences from diversity analysis of populations can be driven by techniques of diversity and distance assessments and not only data driven.

Virulence and Diversity of Blumeria graminis f. sp. hordei in Israel and in the Czech Republic.

Three hundred and nine isolates were obtained from three natural populations of Blumeria graminis f. sp. hordei occurring on wild barley (Hordeum vulgare subsp. spontaneum) at two locations in Israel during 1997 and 1999. Their virulence frequency was determined on 32 differential lines. No isolate was virulent on the differential lines possessing the genes Mla13, mlo, Mlf1, and Mli, and conversely no isolate was avirulent on the differential lines possessing the genes MlRu2, MlLa, Mlh, Mla8, Mla25, and Mlj. The frequencies of isolates overcoming the genes Mlg, Mla7, and Mla27 were 0 to 16% at individual locations; frequencies of isolates overcoming the genes Mla9, Mla17, and Mla18 ranged from 37 to 78%, and frequencies of virulences to genes Mla1, Mla3, Mla6, Mlp1, Mlat, Mla12, Mlra, Mlk1, Mla19, Mla20, Mla26, Mla28, Mla29, Mla30, Mla32, and mlt1 were 79 to 99%. Based on examination of 376 isolates collected in the same years from the Czech Republic, these populations differed greatly from the Israeli ones. The Czech populations showed greater diversity of virulence and lower mean virulence complexity than the Israeli populations. Diversity in the Israeli populations differed also among clusters of niches at the same location.