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We used a population genomics approach to test the hypothesis of clonal expansion of a highly fit genotype in populations of Verticillium dahliae. This fungal pathogen has a broad host range and can be dispersed in contaminated seed or other plant material. It has a highly clonal population structure, with several lineages having nearly worldwide distributions in agricultural crops. Isolates in lineage 1A are highly virulent and cause defoliation in cotton, okra, and olive (denoted 1A/D), whereas those in other lineages cause wilting but not defoliation (ND). We tested whether the highly virulent lineage 1A/D could have spread from the southwestern United States to the Mediterranean basin, as predicted from historical records. We found 187 single-nucleotide polymorphisms (SNPs), determined by genotyping by sequencing, among 91 isolates of lineage 1A/D and 5 isolates in the closely related lineage 1B/ND. Neighbor-joining and maximum-likelihood analyses on the 187 SNPs showed a clear divergence between 1A/D and 1B/ND haplotypes. Data for only 77 SNPs were obtained for all 96 isolates (no missing data); lineages 1A/D and 1B/ND differed by 27 of these 77 SNPs, confirming a clear divergence between the two lineages. No evidence of recombination was detected within or between these two lineages. Phylogenetic and genealogical analyses resulted in five distinct subclades of 1A/D isolates that correlated closely with geographic origins in the Mediterranean basin, consistent with the hypothesis that the D pathotype was introduced at least five times in independent founder events into this region from a relatively diverse source population. The inferred ancestral haplotype was found in two isolates sampled before 1983 from the southwestern United States, which is consistent with historical records that 1A/D originated in North America. The five subclades coalesce with the ancestral haplotype at the same time, consistent with a hypothesis of rapid population expansion in the source population during the emergence of 1A/D as a severe pathogen of cotton in the United States. [ABSTRACT FROM AUTHOR]

Most asexual species of fungi have either lost sexuality recently, or they experience recombination by cryptic sexual reproduction. Verticillium dahliae is a plant-pathogenic, ascomycete fungus with no known sexual stage, even though related genera have well-described sexual reproduction. V. dahliae reproduces mitotically and its population structure is highly clonal. However, previously described discrepancies in phylogenetic relationships among clonal lineages may be explained more parsimoniously by recombination than mutation; therefore, we looked for evidence of recombination within and between clonal lineages. Genotyping by sequencing was performed on 141 V. dahliae isolates from diverse geographic and host origins, resulting in 26,748 single-nucleotide polymorphisms (SNPs). We found a strongly clonal population structure with the same lineages as described previously by vegetative compatibility groups (VCGs) and molecular markers. We detected 443 recombination events, evenly distributed throughout the genome. Most recombination events detected were between clonal lineages, with relatively few recombinant haplotypes detected within lineages. The only three isolates with mating type MAT1-1 had recombinant SNP haplotypes; all other isolates had mating type MAT1-2. We found homologs of eight meiosis-specific genes in the V. dahliae genome, all with conserved or partially conserved protein domains. The extent of recombination and molecular signs of sex in (mating-type and meiosis-specific genes) suggest that V. dahliae clonal lineages arose by recombination, even though the current population structure is markedly clonal. Moreover, the detection of new lineages may be evidence that sexual reproduction has occurred recently and may potentially occur under some circumstances. We speculate that the current clonal population structure, despite the sexual origin of lineages, has arisen, in part, as a consequence of agriculture and selection for adaptation to agricultural cropping systems. [ABSTRACT FROM AUTHOR]

Background: The grape powdery mildew fungus, Erysiphe necator, was introduced into Europe more than 160 years ago and is now distributed everywhere that grapes are grown. To understand the invasion history of this pathogen we investigated the evolutionary relationships between introduced populations of Europe, Australia and the western United States (US) and populations in the eastern US, where E. necator is thought to be native. Additionally, we tested the hypothesis that populations of E. necator in the eastern US are structured based on geography and Vitis host species. Results: We sequenced three nuclear gene regions covering 1803 nucleotides from 146 isolates of E. necator collected from the eastern US, Europe, Australia, and the western US. Phylogeographic analyses show that the two genetic groups in Europe represent two separate introductions and that the genetic groups may be derived from eastern US ancestors. Populations from the western US and Europe share haplotypes, suggesting that the western US population was introduced from Europe. Populations in Australia are derived from European populations. Haplotype richness and nucleotide diversity were significantly greater in the eastern US populations than in the introduced populations. Populations within the eastern US are geographically differentiated; however, no structure was detected with respect to host habitat (i.e., wild or cultivated). Populations from muscadine grapes, V. rotundifolia, are genetically distinct from populations from other Vitis host species, yet no differentiation was detected among populations from other Vitis species. Conclusions: Multilocus sequencing analysis of the grape powdery mildew fungus is consistent with the hypothesis that populations in Europe, Australia and the western US are derived from two separate introductions and their ancestors were likely from native populations in the eastern US. The invasion history of E. necator follows a pattern consistent with plant-mediated dispersal, however, more exhaustive sampling is required to make more precise conclusions as to origin. E. necator shows no genetic structure across Vitis host species, except with respect to V. rotundifolia. [ABSTRACT FROM AUTHOR]

Abstract: Evidence for parasexuality in natural populations of haploid fungi requires the demonstration of diploids or heterokaryons and recombinant genotypes in the absence of sex. We studied clonal populations of the chestnut blight fungus, Cryphonectria parasitica, in southeastern Europe and found evidence of parasexuality in two locations. In Osoj, Macedonia, we found one isolate (Os05-66) that had two alleles at six codominant loci, giving a haplotype that was a composite of two clones in this population. Six single-conidial isolates from Os05-66 had two alleles at some loci, suggesting partial diploidy or aneuploidy, and we found four recombinant haplotypes among single-conidial isolates from hyphal-tip isolates of the same isolate. In Teano, Italy, we found two heterokaryon isolates that were partial composites of two dominant clones. Single-conidial isolates from hyphal-tip isolates had recombinant haplotypes. These results provide evidence that is consistent with the hypothesis of parasexuality in C. parasitica in Europe, similar to an earlier report in a natural population in the USA. [Copyright &y& Elsevier]

Abstract: This study focuses on a pair of fungal species, Moelleriella libera and M. raciborskii (Ascomycota: Clavicipitaceae) from the neotropics and paleotropics, respectively, that are phenotypically nearly indistinguishable. Molecular analyses based on DNA sequences from RNA polymerase II subunit 2 (RPB2), translation elongation factor 1-α (EF1-α) and β-tubulin genes confirm that they are recently derived sister species. Speciation appears to have followed an historical transoceanic dispersal event. Models of population structure and migration from TCS, IM, and coalescent-based analyses suggest there is little gene flow between the two species. The direction of dispersal, investigated using the progression rule and coalescent-based gene genealogies, was likely from the New World to the Old World. [Copyright &y& Elsevier]

Expanding populations are often less genetically diverse at their margins than at the centre of a species’ range. Established, older populations of the chestnut blight fungus, Cryphonectria parasitica, are more variable for vegetative compatibility (vc) types than in expanding populations in southeastern Europe where C. parasitica has colonized relatively recently. To test whether vc types represent clones, we genotyped 373 isolates of C. parasitica from southern Italy, Romania, Bulgaria, Macedonia, Greece and Turkey using 11 sequence-characterized amplified region (SCAR) markers. Ten SCAR loci and six vegetative incompatibility ( vic) loci were polymorphic in these samples. These populations are clonal by all criteria tested: (i) among 373 isolates, we found only eight multilocus haplotypes, and the same haplotypes were found in multiple countries, sometimes separated in time by as much as 12 years; (ii) the number of haplotypes observed was significantly less than expected under random mating; (iii) populations are in linkage disequilibrium; (iv) the two sets of independent markers, SCARs and vc types, are highly correlated; and (v) sexual structures of C. parasitica were found only in Bulgaria and Romania. One mating type ( MAT-1) was found in 98% of the isolates sampled. In contrast, a population in northern Italy, in the central part of the range in Europe, had 12 multilocus haplotypes among 19 isolates. The spread of a few clones could be the result either of founder effect and restricted migration, or these clones have greater fitness than others and spread because they are better adapted to conditions in southeastern Europe. [ABSTRACT FROM AUTHOR]

A marked-isolate, release-recapture experiment was conducted to assess the relative contributions of seed-transmitted (released isolates) versus all other inocula to foliar and grain populations of Phaeosphaeria nodorum in winter wheat rotated with nonsusceptible crops in New York and Georgia, United States. Seed infected with two distinct groups of marked isolates of P. nodorum containing rare alleles (identified by amplified fragment length polymorphisms [AFLPs]) and balanced for mating type were planted in experimental field plots in two locations in each state. Recapture was done by isolating P. nodorum from leaves showing necrotic lesions at spring tillering and flowering stages, and mature grains from spikes showing glume blotch. Isolates from these samples were genotyped by AFLPs and categorized as released or nonreleased to infer sources of inoculum. Both infected seed and other sources of the pathogen contributed significant primary inocula to populations recovered from leaves and harvested grain. Seed-transmitted genotypes accounted for a total of 57% of all isolates recovered from inoculated plots, with a range of 15 to 90% of the populations of P. nodorum collected over the season in individual, inoculated plots at the four locations. Plants in the noninoculated control plots also became diseased and 95% or more of the isolates recovered from these plots were nonreleased genotypes. Although other potential sources of P. nodorum within and adjacent to experimental plots were not ruled out, nonreleased genotypes likely were derived from immigrant ascospores potentially from sources at a considerable distance from the plots. Our results suggest that, although reduction of seedborne inoculum of P. nodorum may delay foliar epidemics, this strategy by itself is unlikely to result in high levels of control in eastern North America because of the additional contribution from alternative sources of inoculum. [ABSTRACT FROM AUTHOR]

We found high diversity of vegetative compatibility (vc) types in native populations of the chestnut blight fungus, Cryphonectria parasitica, in Japan and China; almost every isolate was in a unique vc type. In Japan we found 71 vc types in a sample of 79 isolates pooled from six populations. Within two populations in China, all isolates (n = 28 and 11) had unique vc types; we found 15 vc types among 25 isolates in a third Chinese population where multiple isolates were collected from some trees. None of the isolates from China and only three isolates in the 71 vc types from Japan were compatible with any of 64 vc type testers from Europe, which have known vegetative incompatibility genotypes. To our knowledge this is the first report of vc type diversity for C. parasitica in Japan or of any comparisons of vc types between Asia and Europe. The most significant result of this survey is the identification fungal isolates for expanding knowledge of the genetics of vegetative incompatibility. [ABSTRACT FROM AUTHOR]

Abstract: To find markers linked to vegetative incompatibility (vic) genes in the chestnut blight fungus, Cryphonectria parasitica, we constructed a preliminary linkage map. In general, this map is characterized by low levels of polymorphism, as evident from the more than 24 linkage groups observed, compared to seven expected from electrophoretic karyotyping. Nonetheless, we found markers closely linked to two vic genes (vic1 and vic2) making them candidates for positional cloning. Two markers were found to be linked to vic2: one cosegregated with vic2, i.e., it is 0.0 cM from vic2, the other was at a distance of 4.5cM; a single marker was found 4.0cM from vic1. The closest markers linked to three other vic genes (vic4, vic6, and vic7) were >15cM away; additional markers are needed before efficient positional cloning of these three vic genes can be realized. In contrast to the low levels of polymorphism observed across most of the C. parasitica genome, the linkage group containing the MAT locus appears to harbor an extremely high level of RAPD heterogeneity and reduced recombination. Markers within this highly heterogeneous region are in linkage disequilibrium in some natural populations; however, recombination is clearly evident between this region and the MAT locus. [Copyright &y& Elsevier]

Population genetic and epidemiological studies have resulted in different hypotheses about the predominant source of primary inoculum in the Phaeosphaeria nodorum-wheat pathosystem (i.e., sexually derived, windborne ascospores versus asexual or seedborne inoculum). We examined the genetic structure of seedborne populations of P. nodorum as a further step toward evaluating the hypothesis that seedborne inoculum is an important contributor to foliar epidemics in New York's rotational wheat fields. In all, 330 seedborne isolates from seven field populations were genotyped at 155 amplified fragment length polymorphism loci. Seed-borne populations possessed high levels of genotypic diversity, with virtually every isolate (326/330) having a unique haplotype. As in previous population genetic studies of P. nodorum, we found low levels of gametic disequilibrium, although we could reject the null hypothesis of random mating with the index of association test for two populations. Thus, genotypically diverse and seemingly panmictic populations of P. nodorum that have been observed in wheat foliage could be derived from seedborne primary inoculum. Although sexual reproduction and recombination may contribute to the diversity of foliar populations of P. nodorum, population genetic data do not rule out seed as a source of primary inoculum. Further experimentation will be needed to determine definitively the relative importance of windborne ascospores and seedborne asexual inoculum in epidemics of Stagonospora nodorum blotch in New York. [ABSTRACT FROM AUTHOR]

Most hypovirulence in the chestnut blight fungus, Cryphonectria parasitica, is associated with infection by fungal viruses in the family Hypoviridae. Hypovirulence has controlled chestnut blight well in some locations in Europe and in Michigan in the United States. In contrast, with few exceptions, biological control has failed almost completely in eastern North America. Therapeutic treatment of individual cankers is successful in most cases, but the success of hypovirulence at the population level depends on the natural spread of viruses. Characteristics of three interacting trophic levels (virus, fungus, and tree), plus the environment, determine the success or failure of hypovirulence. Vegetative incompatibility restricts virus transmission, but this factor alone is a poor predictor of biological control. Any factor reducing the rate of chestnut blight epidemics enhances hypovirus invasion. Overall, however, not enough is understood about the epidemiological dynamics of this system to determine the crucial factors regulating the establishment of hypovirulence in chestnut forests. [ABSTRACT FROM AUTHOR]

Plant pathogens often exhibit variation in virulence, the ability to cause disease on host plants with specific resistance, evident from the diversity of races observed within pathogen species. The evolution of races in asexual fungal pathogens has been hypothesized to occur in a stepwise fashion, in which mutations to virulence accumulate sequentially in clonal lineages, resulting in races capable of overcoming multiple host plant resistance genes or multiple resistant cultivars. In this study, we demonstrate a simple stepwise pattern of race evolution in Fusarium oxysporum f. sp. ciceris, the fungus that causes Fusarium wilt of chickpeas. The inferred intraspecific phylogeny of races in this fungus, based on DNA fingerprinting with repetitive sequences, shows that each of the eight races forms a monophyletic lineage. By mapping virulence to each differential cultivar (used for defining races) onto the inferred phylogeny, we show that virulence has been acquired in a simple stepwise pattern, with few parallel gains or losses. Such a clear pattern of stepwise evolution of races, to our knowledge, has not been demonstrated previously for other pathogens based on analyses of field populations. We speculate that in other systems the stepwise pattern is obscured by parallel gains or losses of virulence caused by higher mutation rates and selection by widespread deployment of resistant cultivars. Although chickpea cultivars resistant to Fusarium wilt are available, their deployment has not been extensive and the stepwise acquisition of virulence is still clearly evident. [ABSTRACT FROM AUTHOR]

Using isolates collected over 2 decades, we determined the population structure and dynamics of the rice blast fungus, Magnaporthe grisea, in Korea at both the genotypic and phenotypic levels. Pathotype analysis on 6,315 isolates collected from 328 rice cultivars from 1981 to 2000 revealed the presence of a total of 91 pathotypes. Among these 91 pathotypes, nine dominated, comprising 76.5% of the isolates. The expected number of pathotypes (corrected for sample size) increased significantly during the course of this study. On average, six (ranging from 0 to 20) new commercial cultivars were introduced annually between 1981 and 1998. However, the overall cultivar diversity, estimated using the Shannon index, was low. Most of the new cultivars were not planted to a large area because the seven most common cultivars each year occupied over 70% of the rice-cultivated area. The frequencies of the nine dominant pathotypes from these seven cultivars were highly correlated with those from the entire set of cultivars. To understand genetic diversity within and between pathotypes, 176 isolates collected from 1984 to 1999 were randomly sampled and analyzed by DNA fingerprinting. High similarities were observed among isolates; overall similarities were greater than 63% in combined MGR586 and MAGGY DNA fingerprints. Unlike most other populations of M. grisea, DNA fingerprints showed no clear lineage structure. No groups were supported by bootstrap values greater than 10%. Furthermore, there was no significant correlation between DNA fingerprint similarities and pathotypes. Genetic similarity was significantly greater (P < 0.001) within years than between years, although the difference was small. Our data suggest that M. grisea populations in Korea have been mostly dominated by a single clonal lineage. We cannot conclude from these data that selection by the host population has been a major force in the evolution of M. grisea in Korea. [ABSTRACT FROM AUTHOR]

Although the genetic components of mating systems in fungi are well understood as laboratory phenomena, surprisingly little is known about their function in nature or about their role in determining mating patterns and population genetic structure. Our study of the mating system of the haploid ascomycete fungus, Cryphonectria parasitica, resulted in the following. (1) Laboratory crosses among 20 isolates, chosen randomly from North America and China, resolved into two incompatibility groups (occurring on both continents), confirming that C. parasitica has a diallelic, bipolar sexual self-incompatibility system, typical of other self-incompatible Ascomycetes, in which mating is only successful between isolates of opposite mating type. (2) PCR-based markers for mating-type alleles correlated perfectly with mating-type phenotypes of individual isolates. (3) Three genotypes, isolated from natural populations in Virginia and West Virginia, were inoculated onto chestnut trees in two sites in West Virginia and were confirmed to have self-fertilized and outcrossed in both sites. (4) Ten isolates, of a total of over 200 assayed, were confirmed to have self-fertilized in the laboratory, albeit at very low frequency. Five of these 10 isolates were ramets of a single genet, suggesting a genetic basis underlying the proclivity to self-fertilize in the laboratory. (5) Self-fertilization could not be induced in the laboratory with exudates (ostensibly containing pheromones) from isolates of opposite mating type. These results demonstrate that, a sexual self-incompatibility system notwithstanding, self-fertilization occurs under both laboratory and field conditions in C. parasitica. The disparity between observations of frequent selfing in nature and rare selfing in the laboratory suggests that the mating system is under ecological as well as genetic control. [ABSTRACT FROM AUTHOR]

Deals with a study which tested predictions about Cryphonectria parasitica. Vegetative compatibility type distribution in C. parasitica from Teano, Italy and populations in the United States; Allele frequencies, number of polymorphic loci and mean allelic diversities at six vic loci in populations of C. parasitica; Gene diversity analysis in European populations of C. parasitica.

Presents a study which identified the vegetative incompatibility in Cryphonectria parasitica in Europe, in relation to genetic aspects. What is vegetative of heterokaryon incompatibility; Method used to determine vic genotypes; Classification of C. parasitica.

Examines the relationship between recombination and the multilocus structure of fungal populations. Significance of recombination; Models of multilocus population structure; Inferences about random mating in fungal populations.

Consumption of food contaminated with aflatoxin, from crops infected by Aspergillus flavus, is associated with acute toxicosis, cancer, and stunted growth. Although such contamination is more common in the lower latitudes of the United States, it is unclear whether this pattern is associated with differences in the relative frequencies of aflatoxigenic individuals of A. flavus. To determine whether the frequency of the aflatoxin-producing ability of A. flavus increases as latitude decreases, we sampled 281 isolates from field soils in two north-south transects in the United States and tested them for aflatoxin production. We also genotyped 161 isolates using 10 microsatellite markers to assess population structure. Although the population density of A. flavus was highest at lower latitudes, there was no difference in the frequency of aflatoxigenic A. flavus isolates in relation to latitude. We found that the U.S. population of A. flavus is subdivided into two genetically differentiated subpopulations that are not associated with the chemotype or geographic origin of the isolates. The two populations differ markedly in allelic and genotypic diversity. The less diverse population is more abundant and may represent a clonal lineage derived from the more diverse population. Overall, increased aflatoxin contamination in lower latitudes may be explained partially by differences in the population density of A. flavus, not genetic population structure. [ABSTRACT FROM AUTHOR]

We are entering a new era in plant pathology in which whole-genome sequences of many individuals of a pathogen species are becoming readily available. Population genomics aims to discover genetic mechanisms underlying phenotypes associated with adaptive traits such as pathogenicity, virulence, fungicide resistance, and host specialization, as genome sequences or large numbers of single nucleotide polymorphisms become readily available from multiple individuals of the same species. This emerging field encompasses detailed genetic analyses of natural populations, comparative genomic analyses of closely related species, identification of genes under selection, and linkage analyses involving association studies in natural populations or segregating populations resulting from crosses. The era of pathogen population genomics will provide new opportunities and challenges, requiring new computational and analytical tools. This review focuses on conceptual and methodological issues as well as the approaches to answering questions in population genomics. The major steps start with defining relevant biological and evolutionary questions, followed by sampling, genotyping, and phenotyping, and ending in analytical methods and interpretations. We provide examples of recent applications of population genomics to fungal and oomycete plant pathogens. [ABSTRACT FROM AUTHOR]

For the last three decades, evolutionary biologists have sought to understand which factors modulate the evolution of parasite virulence. Although theory has identified several of these modulators, their effect has seldom been analysed experimentally. We investigated the role of two such major factors—the mode of transmission, and host adaptation in response to parasite evolution—in the evolution of virulence of the plant virus Cucumber mosaic virus (CMV) in its natural host Arabidopsis thaliana. To do so, we serially passaged three CMV strains under strict vertical and strict horizontal transmission, alternating both modes of transmission. We quantified seed (vertical) transmission rate, virus accumulation, effect on plant growth and virulence of evolved and non-evolved viruses in the original plants and in plants derived after five passages of vertical transmission. Our results indicated that vertical passaging led to adaptation of the virus to greater vertical transmission, which was associated with reductions of virus accumulation and virulence. On the other hand, horizontal serial passages did not significantly modify virus accumulation and virulence. The observed increases in CMV seed transmission, and reductions in virus accumulation and virulence in vertically passaged viruses were due also to reciprocal host adaptation during vertical passages, which additionally reduced virulence and multiplication of vertically passaged viruses. This result is consistent with plant-virus co-evolution. Host adaptation to vertically passaged viruses was traded-off against reduced resistance to the non-evolved viruses. Thus, we provide evidence of the key role that the interplay between mode of transmission and host-parasite co-evolution has in determining the evolution of virulence. [ABSTRACT FROM AUTHOR]

Random mating and recombination in heterothallic ascomycetes should result in high genotypic diversity, 1:1 mating-type ratios, and random associations of alleles, or linkage equilibrium, at different loci. To test for random mating in populations of the grape powdery mildew fungus Erysiphe necator, we sampled isolates from vineyards of Vitis vinifera in Burdett, NY (NY09) and Winchester, VA (VA09) at the end of the epidemic in fall 2009. We also sampled isolates from the same Winchester, VA vineyard in spring 2010 at the onset of the next epidemic. Isolates were genotyped for mating type and 11 microsatellite markers. In the spring sample, which originated from ascospore infections, nearly every isolate had a unique genotype. In contrast, fall populations were less diverse. In all, 9 of 45 total genotypes in VA09 were represented by two or more isolates; 3 of 40 total genotypes in NY09 were represented by two or more isolates, with 1 genotype represented by 20 isolates. After clone correction, mating-type ratios in the three populations did not deviate from 1:1. However, even with clone correction, we detected significant linkage disequilibrium (LD) in all populations. Mantel tests detected positive correlations between genetic and physical distances within vineyards. Spatial autocorrelation showed aggregations up to 42 and 3 m in VA09 and NY09, respectively. Spatial autocorrelation most likely results from short dispersal distances. Overall, these results suggest that spatial genetic aggregation and clonal genotypes that arise during the asexual phase of the epidemic contribute to persistent LD even though populations undergo sexual reproduction annually. [ABSTRACT FROM AUTHOR]

Eastern North America is considered the center of diversity for many Vitis spp. and for the grape powdery mildew pathogen, Erysiphe necator. However, little is known about populations of E. necator from wild Vitis spp. We determined the phenotypic variation in pathogenicity and aggressiveness of E. necator among isolates from wild and domesticate Vitis spp. from diverse geographic regions in the eastern United States. To test pathogenicity, we inoculated 38 E. necator isolates on three wild Vitis spp., two commercially grown hybrids and the European wine grape, Vitis vinifera. V. rotundifolia (muscadine grape) was the only host species on which complete host specialization was evident: it was only susceptible to isolates collected from V. rotundifolia. All isolates, regardless of source host, were pathogenic on the other Vitis spp. We found no differences in components of aggressiveness latent period and lesion size among isolates from different source hosts when inoculated on V. vinifera, which is highly susceptible to powdery mildew. However significant variation was evident among isolates on the more resistant V. labruscana 'Niagara'. Isolates from the wild species V aestivalis were the most aggressive whereas isolates from V. vinifera were not more aggressive than isolate from other source hosts. Greater aggressiveness was also detected among isolates from the southeastern United States compared with isolates from the northeastern United States. [ABSTRACT FROM AUTHOR]

Six vegetative incompatibility (vic) loci have been identified in Cryphonectria parasitica based on barrage formation during mycelial interactions. We used hygromycin B- and benomyl-resistance as forcing markers in C. parasitica strains to test whether heteroallelism at each vic locus prevents heterokaryon formation following mycelial interactions. Paired strains that had allelic differences at any of viol, 2, 3, 6 or 7 but not vic4 displayed heterokaryon incompatibility function, as recognized by slow growth or aberrant morphology. While clearly forming barrages in mycelial interactions, paired strains with different alleles at vic4 formed stable heterokaryons. With examples from other fungi, this inconsistency at vic4 suggests that barrage formation and heterokaryon incompatibility are not different manifestations of the same process. Rather, the evidence indicates that heterokaryon incompatibility represents a component of a vegetative incompatibility system that may also use cell-surface or extracellular factors to trigger programmed cell death to modulate nonself recognition in fungi. [ABSTRACT FROM AUTHOR]

Erysiphe necator overwinters as ascospores in cleistothecia and mycelium in dormant buds of grapevines. Shoots developing from infected buds early in the growing season are covered with dense mycelium and are known as "flag shoots". Combining epidemiological and genetic analyses, the objective of this study was to analyze the spatial and genetic structure of a flag shoot subpopulation of E. necator as a way to assess the contribution of flag shoots as primary inoculum, and to determine if flag shoot subpopulations are clonal with only one mating type. One vineyard in Tuscany, Italy was surveyed intensively for flag shoots for 8 years; isolations of E. necator were made from flag shoots for 5 years. We observed distinct disease foci developing around flag shoots early in epidemics, demonstrating a steep dispersal gradient of conidia and the importance of flag shoots as primary inoculum sources. Flag shoots were spatially aggregated within and between years, most likely as a result of short-distance dispersal of conidia from flags early in the season when dormant buds for the next year's shoots are formed and are susceptible to infection. The two mating types were found in 1:1 ratios in this flag shoot subpopulation. Genotypic diversity, based on inter-simple sequence repeat markers, was high in all years with only two haplotypes occurring twice, and subpopulations were genetically differentiated between years. Similarities between haplotypes were not spatially autocorrelated. One multilocus analysis of population structure is consistent with the hypothesis of random mating but another is not. These results are not consistent with expectations for a strictly clonal or strictly randomly mating flag shoot subpopulation. Instead, the hypothesis that the flag shoot subpopulation of E. necator may reproduce clonally and sexually needs further testing. [ABSTRACT FROM AUTHOR]

Selfing in the chestnut blight fungus, Cryphonectria parasitica, occurs by two different genetic mechanisms. Most self-fertile isolates of C. parasitica are heterokaryotic for mating type, and the progeny from selfing segregate for mating type. Further, we resolved mating-type (MAT) heterokaryons into homokaryons of both mating types by isolating uninucleate asexual spores (conidia). However, because ascospore progeny, with rare exceptions, are not MAT heterokaryons, C. parasitica must lack a regular mechanism to maintain heterokaryosis by selfing. We hypothesize that heterokaryon formation may occur either because of recurrent biparental inbreeding, or by mating-type switching, possibly one involving some kind of parasexual process. The second mechanism found for selfing in C. parasitica occurred less frequently. Three single-conidial isolates (MAT-1 and MAT-2) selfed and produced progeny that did not segregate for mating type. It is currently not known if meiosis occurs during ascospore formation by this mechanism.Index descriptors: Cryphonectria parasitica, Heterokaryosis, MAT, Mating type, Self-fertility, Self-incompatibility, Self-sterility [Copyright &y& Elsevier]

Fungi produce a wealth of pharmacologically bioactive secondary metabolites (SMs) from biosynthetic gene clusters (BGCs). It is common practice for drug discovery efforts to treat species' secondary metabolomes as being well represented by a single or a small number of representative genomes. However, this approach misses the possibility that intraspecific population dynamics, such as adaptation to environmental conditions or local microbiomes, may harbor novel BGCs that contribute to the overall niche breadth of species. Using 94 isolates of Aspergillus flavus, a cosmopolitan model fungus, sampled from seven states in the United States, we dereplicate 7,821 BGCs into 92 unique BGCs. We find that more than 25% of pangenomic BGCs show population-specific patterns of presence/absence or protein divergence. Population-specific BGCs make up most of the accessory-genome BGCs, suggesting that different ecological forces that maintain accessory genomes may be partially mediated by population-specific differences in secondary metabolism. We use ultra-high-performance high-resolution mass spectrometry to confirm that these genetic differences in BGCs also result in chemotypic differences in SM production in different populations, which could mediate ecological interactions and be acted on by selection. Thus, our results suggest a paradigm shift that previously unrealized population-level reservoirs of SM diversity may be of significant evolutionary, ecological, and pharmacological importance. Last, we find that several population-specific BGCs from A. flavus are present in Aspergillus parasiticus and Aspergillus minisclerotigenes and discuss how the microevolutionary patterns we uncover inform macroevolutionary inferences and help to align fungal secondary metabolism with existing evolutionary theory. [ABSTRACT FROM AUTHOR]

Verticillium dahliae is a soilborne fungal pathogen affecting many economically important crops that can also infect weeds and rotational crops with no apparent disease symptoms. The main research goal was to the hypothesis that V dahMae populations recovered from a symptomatic rotational crops and weed species are evolutionarily and genetically distinct from symptomatic hosts. We collected W dahliae isolates from symptomatic and asymptomatic hosts growing in fields with histories of Verticillium wilt of potato in Israel and Pennsylvania (United States), and used genotyping-by-sequencing to analyze the evolutionary history and genetic differentiation between populations of different hosts. phylogeny inferred from 26,934 single-nucleotide polymorphisms SNPs) in 126 W dahliae isolates displayed a highly clonal structure correlated with vegetative compatibility groups, and isolates grouped in lineages 2A, 28824,4A, and 48, with 77% of the isolates in lineage 4B. The lineages identified in this study were differentiated by host of origin; we found 2A, 213824, and 4A only in symptomatic hosts but isolates from asymptomatic hosts (weeds, oat, and sorghum) grouped exclusively within lineage 4B, and were genetically indistinguishable from 4B isolates sampled from symptomatic hosts (potato, eggplant, and avocado). Using coalescent analysis of 158 SNPs of lineage 48, we inferred a genealogy with clades that correlated with geographic origin. In contrast, isolates from asymptomatic and symptomatic hosts shared some of the same haplotypes and were not differentiated. We conclude that asymptomatic weeds and rotational hosts may be potential reservoirs for W dahliae populations of lineage 48, which are pathogenic to many cultivated hosts. [ABSTRACT FROM AUTHOR]

Cryphonectria parasitica is the causal agent of chestnut blight, a fungal disease that almost entirely eliminated mature American chestnut from North America over a 50-year period. Here, we formally report the genome of C. parasitica EP155 using a Sanger shotgun sequencing approach. After finishing and integration with simple-sequence repeat markers, the assembly was 43.8 Mb in 26 scaffolds (L50 = 5; N50 = 4.0Mb). Eight chromosomes are predicted: five scaffolds have two telomeres and six scaffolds have one telomere sequence. In total, 11,609 gene models were predicted, of which 85% show similarities to other proteins. This genome resource has already increased the utility of a fundamental plant pathogen experimental system through new understanding of the fungal vegetative incompatibility system, with significant implications for enhancing mycovirus-based biological control. [ABSTRACT FROM AUTHOR]

Verticillium dahliae is a plant pathogenic fungus that reproduces asexually and its population structure is highly clonal. In the present study, 78 E dahliae isolates from Iran were genotyped for mating type, single nucleotide polymorphisms (SNPs), and microsatellites to assign them to clonal lineages and to determine population genetic structure in Iran. The mating type of all isolates was MAT1-2. Based on neighbor-joining analysis and minimum spanning networks constructed from SNPs and microsatellite genotypes, respectively, all but four isolates were assigned to lineage 2B824; four isolates were assigned to lineage 4B. The inferred coalescent genealogy of isolates in lineage 2B824 showed a clear divergence into two clades that corresponded to geographic origin and host. Haplotypes of cotton and pistachio isolates sampled from central Iran were in one clade, and those of isolates from Prunus spp. sampled from northwestern Iran were in the other. The strong divergence in haplotypes between the two clades suggests that there were at least two separate introductions of lineage 2B824 to different pails of Iran. Given the history of cotton and pistachio cultivation and Verticillium wilt in Iran, these results are consistent with the hypothesis that cotton was historically a likely source inoculum causing Verticillium wilt in pistachio. [ABSTRACT FROM AUTHOR]

The role of microbial secondary metabolites in the ecology of the organisms that produce them remains poorly understood. Variation in aflatoxin production by Aspergillus flavus is maintained by balancing selection, but the ecological function and impact on fungal fitness of this compound are unknown. We hypothesize that balancing selection for aflatoxin production in A. flavus is driven by interaction with insects. To test this,we competed naturally occurring aflatoxigenic and non-aflatoxigenic fungal isolates against Drosophila larvae on medium containing 0-1750 ppb aflatoxin, using quantitative PCR to quantify A. flavus DNA as a proxy for fungal fitness. The addition of aflatoxin across this range resulted in a 26-fold increase in fungal fitness. With no added toxin, aflatoxigenic isolates caused higher mortality of Drosophila larvae and had slightly higher fitness than non-aflatoxigenic isolates. Additionally, aflatoxin production increased an average of 1.5-fold in the presence of a single larva and nearly threefold when the fungus was mechanically damaged. We argue that the role of aflatoxin in protection from fungivory is inextricably linked to its role in interference competition. Our results, to our knowledge, provide the first clear evidence of a fitness advantage conferred to A. flavus by aflatoxin when interacting with insects. [ABSTRACT FROM AUTHOR]

We studied the mechanisms of azole resistance in Erysiphe necator by quantifying the sensitivity to myclobutanil (EC50) in 65 isolates from the eastern United States and 12 from Chile. From each isolate, we sequenced the gene for sterol 14α-demethylase (CYP51), and measured the expression of CYP51 and homologs of four putative efflux transporter genes, which we identified in the E. necator transcriptome. Sequence variation in CYP51 was relatively low, with sequences of 40 U.S. isolates identical to the reference sequence. Nine U.S. isolates and five from Chile carried a previously identified A to T nucleotide substitution in position 495 (A495T), which results in an amino acid substitution in codon 136 (Y136F) and correlates with high levels of azole resistance. We also found a nucleotide substitution in position 1119 (A1119C) in 15 U.S. isolates, whose mean EC50 value was equivalent to that for the Y136F isolates. Isolates carrying mutation A1119C had significantly greater CYP5I expression, even though Al 119C does not affect the CYP51 amino acid sequence. Regression analysis showed no significant effects of the expression of efflux transporter genes on EC50. Both the Y136F mutation in CYP51 and increased CYP51 expression appear responsible for azole resistance in eastern U.S. populations of E. necator. [ABSTRACT FROM AUTHOR]

Race-specific resistance against powdery mildews is well documented in small grains but, in other crops such as grapevine, controlled analysis of host-pathogen interactions on resistant plants is uncommon. In the current study, we attempted to confirm powdery mildew resistance phenotypes through vineyard, greenhouse, and in vitro inoculations for test cross-mapping populations for two resistance sources: (i) a complex hybrid breeding line, 'Bloodworm 81-107-11', of at least Vitis rotundifolia, V. vinifera, V. berlandieri, V. rupestris, V. labrusca, and V. aestivalis background; and (ii) Vitis hybrid 'Tamiami' of V. aestivalis and V. vinifera origin. Statistical analysis of vineyard resistance data suggested the segregation of two and three race-specific resistance genes from the two sources, respectively. However, in each population, some resistant progeny were susceptible in greenhouse or in vitro screens, which suggested the presence of Erysiphe necator isolates virulent on progeny segregating for one or more resistance genes. Controlled inoculation of resistant and susceptible progeny with a diverse set of E. necator isolates clearly demonstrated the presence of fungal races differentially interacting with race-specific resistance genes, providing proof of race specificity in the grape powdery mildew pathosystem. Consistent with known race-specific resistance mechanisms, both resistance sources were characterized by programmed cell death of host epidermal cells under appressoria, which arrested or slowed hyphal growth; this response was also accompanied by collapse of conidia, germ tubes, appressoria, and secondary hyphae. The observation of prevalent isolates virulent on progeny with multiple race-specific resistance genes before resistance gene deployment has implications for grape breeding strategies. We suggest that grape breeders should characterize the mechanisms of resistance and pyramid multiple resistance genes with different mechanisms for improved durability. [ABSTRACT FROM AUTHOR]

SUMMARY Few plant pathogens have had a more profound effect on the evolution of disease management than Erysiphe necator, which causes grapevine powdery mildew. When the pathogen first spread from North America to England in 1845, and onwards to France in 1847, 'germ theory' was neither understood among the general populace nor even generally accepted within the scientific community. Louis Pasteur had only recently reported the microbial nature of fermentation, and it would be another 30 years before Robert Koch would publish his proofs of the microbial nature of certain animal diseases. However, within 6 years after the arrival of the pathogen, nearly 6 million grape growers in France were routinely applying sulphur to suppress powdery mildew on nearly 2.5 million hectares of vineyards (). The pathogen has remained a focus for disease management efforts ever since. Because of the worldwide importance of the crop and its susceptibility to the disease, and because conventional management with modern, organic fungicides has been compromised on several occasions since 1980 by the evolution of fungicide resistance, there has also been a renewed effort worldwide to explore the pathogen's biology and ecology, its genetics and molecular interactions with host plants, and to refine current and suggest new management strategies. These latter aspects are the subject of our review. Taxonomy: The most widely accepted classification follows. Family Erysiphaceae, Erysiphe necator Schw. [syn. Uncinula necator (Schw.) Burr., E. tuckeri Berk., U. americana Howe and U. spiralis Berk. & Curt; anamorph Oidium tuckeri Berk.]. Erysiphe necator var. ampelopsidis was found on Parthenocissus spp. in North America according to , although later studies revealed isolates whose host range spanned genera, making the application of this taxon somewhat imprecise (Gadoury and Pearson, 1991). The classification of the genera before 1980 was based on features of the mature ascocarp: (i) numbers of asci; and (ii) morphology of the appendages, in particular the appendage tips. The foregoing has been supplanted by phylogeny inferred from the internal transcribed spacer (ITS) of ribosomal DNA sequences (), which correlates with conidial ontogeny and morphology (). Host range: The pathogen is obligately parasitic on genera within the Vitaceae, including Vitis, Cissus, Parthenocissus and Ampelopsis (Pearson and Gadoury, 1992). The most economically important host is grapevine ( Vitis), particularly the European grape, V. vinifera, which is highly susceptible to powdery mildew. Disease symptoms and signs: In the strictest sense, macroscopically visible mildew colonies are signs of the pathogen rather than symptoms resulting from its infection, but, for convenience, we describe the symptoms and signs together as the collective appearance of colonized host tissues. All green tissues of the host may be infected. Ascospore colonies are most commonly found on the lower surface of the first-formed leaves near the bark of the vine, and may be accompanied by a similarly shaped chlorotic spot on the upper surface. Young colonies appear whitish and those that have not yet sporulated show a metallic sheen. They are roughly circular, ranging in size from a few millimetres to a centimetre or more in diameter, and can occur singly or in groups that coalesce to cover much of the leaf. Senescent colonies are greyish, and may bear cleistothecia in various stages of development. Dead epidermal cells often subtend the colonized area, as natural mortality in the mildew colony, the use of fungicides, mycoparasites or resistance responses in the leaf result in the deaths of segments of the mildew colony and infected epidermal cells. Severely affected leaves usually senesce, develop necrotic blotches and fall prematurely. Infection of stems initially produces symptoms similar to those on leaves, but colonies on shoots are eventually killed as periderm forms, producing a dark, web-like scar on the cane (Gadoury et al., 2011). Inflorescences and berries are most susceptible when young, and can become completely coated with whitish mildew. The growth of the berry epidermal tissue stops when severely infected, which may result in splitting as young fruit expand. Berries in a transitional stage between susceptible and resistant (generally between 3 and 4 weeks after anthesis) develop diffuse, nonsporulating mildew colonies only visible under magnification. Diffuse colonies die as berries continue to mature, leaving behind a network of necrotic epidermal cells (Gadoury et al., 2007). Survival over winter as mycelium in buds results in a distinctive foliar symptom. Shoots arising from these buds may be heavily coated with fungal growth, stark white in colour and stand out like white flags in the vine, resulting in the term 'flag shoots'. More commonly, colonization of a flag shoot is less extensive, and infection of a single leaf, or of leaves on one side of the shoot only, is observed (Gadoury et al., 2011). [ABSTRACT FROM AUTHOR]

Genetic nonself recognition systems such as vegetative incompatibility operate in many filamentous fungi to regulate hyphal fusion between genetically dissimilar individuals and to restrict the spread of virulence-attenuating mycoviruses that have potential for biological control of pathogenic fungi. We report here the use of a comparative genomics approach to identify seven candidate polymorphic genes associated with four vegetative incompatibility (vic) loci of the chestnut blight fungus Cryphonectria parasitica. Disruption of candidate alleles in one of two strains that were heteroallelic at vic2, vic6, or vic7 resulted in enhanced virus transmission, but did not prevent barrage formation associated with mycelial incompatibility. Detailed characterization of the vic6 locus revealed the involvement of nonallelic interactions between two tightly linked genes in barrage formation, heterokaryon formation, and asymmetric, gene-specific influences on virus transmission. The combined results establish molecular identities of genes associated with four C. parasitica vic loci and provide insights into how these recognition factors interact to trigger incompatibility and restrict virus transmission. [ABSTRACT FROM AUTHOR]

Asexual sporulation (conidiation) is coordinately regulated in the grape powdery mildew pathogen Erysiphe necator but nothing is known about its genetic regulation. We hypothesized that genes required for conidiation in other fungi would be upregulated at conidiophore initiation or full conidiation (relative to preconidiation vegetative growth and development of mature ascocarps), and that the obligate biotrophic lifestyle of E. necator would necessitate some novel gene regulation. cDNA amplified fragment length polymorphism analysis with 45 selective primer combinations produced ≈1,600 transcript-derived fragments (TDFs), of which 620 (39%) showed differential expression. TDF sequences were annotated using BLAST analysis of GenBank and of a reference transcriptome for E. necator developed by 454-FLX pyro-sequencing of a normalized cDNA library. One-fourth of the differentially expressed, annotated sequences had similarity to fungal genes of unknown function. The remaining genes had annotated function in metabolism, signaling, transcription, transport, and protein fate. As expected, a portion of orthologs known in other fungi to be involved in developmental regulation was upregulated immediately prior to or during conidiation; particularly noteworthy were several genes associated with the light-dependent VeA regulatory system. G-protein signaling (Pth11 and a kelch repeat), and nuclear transport (importin-β and Ran). This work represents the first investigation into differential gene expression during morphogenesis in E. necator and identifies candidate genes and hypotheses for characterization in powdery mildews. Our results indicate that, although control of conidiation in powdery mildews may share some basic elements with established systems, there are significant points of divergence as well, perhaps due, in part, to the obligate biotrophic lifestyle of powdery mildews. [ABSTRACT FROM AUTHOR]

Abstract: In ascomycetes, mating compatibility is regulated by the mating-type locus, MAT1. The objectives of this study were to identify and sequence genes at the MAT1 locus in the grape powdery mildew fungus, Erysiphe necator, to develop a PCR-based marker for determining mating type in E. necator, and to develop degenerate primers for amplification by PCR of conserved regions of mating-type idiomorphs in other powdery mildew fungi. We identified MAT1-2-1 of the MAT1-2 idiomorph in E. necator based on the homologous sequence in the genome of Blumeria graminis f. sp. hordei and we found MAT1-1-1 and MAT1-1-3 of the MAT1-1 idiomorph from transcriptome sequences of E. necator. We developed and applied a reliable PCR-based multiplex marker to confirm that genotype correlated with mating phenotype, which was determined by pairing with mating-type tester isolates. Additionally, we used the marker to genotype populations of E. necator from different Vitis spp. from throughout the USA. We found both mating types were present in all populations and mating-type ratios did not deviate from 1:1. The mating-type genes in E. necator are similar to those of other Leotiomycetes; however, the structure of the MAT1 locus in E. necator, like the MAT1-2 idiomorph of B. graminis, is markedly different from other ascomycetes in that it is greatly expanded and may contain a large amount of repetitive DNA. As a result, we were unable to amplify and sequence either idiomorph in its entirety. We designed degenerate primers that amplify conserved regions of MAT1-1 and MAT1-2 in E. necator, Podosphaera xanthii, Microsphaera syringae, and B. graminis, representing the major clades of the Erysiphales. These degenerate primers or sequences obtained in this study from these species can be used to identify and sequence MAT1 genes or design mating-type markers in other powdery mildew fungi as well. [Copyright &y& Elsevier]

Abstract: We surveyed native populations of the chestnut blight fungus, Cryphonectria parasitica, in Japan and China, and C. nitschkei, a sympatric species on chestnut trees in Japan, to learn more about the diversity of hypoviruses and other double-stranded (ds) RNA viruses. In a sample of 472 isolates of C. parasitica and 45 isolates of C. nitschkei from six prefectures in Japan, we found 27 containing one or more dsRNAs. Twelve isolates of C. parasitica and two isolates of C. nitschkei were infected with Cryphonectria hypovirus 1 (CHV-1); four of these 12 C. parasitica isolates also contained other dsRNAs that did not hybridize to CHV-1. In China, only one of 85 C. parasitica isolates was CHV-1-infected; no dsRNAs were detected in the other isolates from China. No other known hypoviruses were found in this study. However, we found two previously undescribed dsRNAs in Japan approximately 9kb in size that did not hybridize to each other or to any known dsRNAs from C. parasitica. We also found three additional groups of dsRNAs, one of which represents the genome of a new member of the virus family Chrysoviridae and was found only in C. nitschkei; the other two dsRNAs were found previously in isolates of C. parasitica from Japan or China. The most significant result of this survey is the discovery of novel dsRNAs that can be characterized in future research. [Copyright &y& Elsevier]

The overwintering mode of the grape powdery mildew fungus, Erysiphe necator (syn. Uncinula necator), as mycelium in dormant buds (resulting in symptoms known as flag shoots) or as ascospores in cleistothecia, affects the temporal dynamics of epidemics early in the growing season. We tested whether distinct genetic groups (I and III) identified previously in E. necator correlate to overwintering modes in two vineyards in Tuscany, Italy, to determine whether diagnostic genetic markers could be used to predict overwintering. Samples from one vineyard were collected from flag shoots; the other vineyard, 60 km away, had no flag shoots, and mildew colonies were assumed to be derived from ascospores. Genetic markers putatively diagnostic for groups I and III showed that both types were common in the flag shoot subpopulation. Both genetic types were found in the ascospore population, although group III was dominant. We did not find strong genetic differentiation between the two subpopulations based on inter-simple sequence repeat markers. Although there was significant (P < 0.001) genetic differentiation between these subpopulations in 1997 and when 1997 and 1998 subpopulations were pooled (θ = 0.214 and 0.150, respectively), no differentiation was evident between vineyards in 1998 (θ = 0.138, P = 0.872). Moreover, we did not observe distinct lineages corresponding to overwintering modes, as observed in previous studies. We could not determine if differentiation resulted from biological differences or restricted gene flow between the two vineyards. Our samples were taken from both subpopulations early in the epidemic, while previous studies confounded overwintering mode and sampling time. These results do not support a strong correlation between overwintering and genetic groups, highlighting the need to base population biology studies on sound biological and epidemiological knowledge. [ABSTRACT FROM AUTHOR]

Cryphonectria includes important tree pathogens as well as species believed to be saprophytes. Recent phylogenetic studies have concentrated on North American and southern hemisphere Cryphonectria spp., but little is known about Asian and European taxa. In this study we identify and differentiate among the species occurring in Europe, China, and Japan using morphological and phylogenetic comparisons among the Cryphonectria species. Phylogenetic comparisons were based on sequence data from the ribosomal ITS operon and two regions in the β-tubulin gene. Unknown Japanese and Chinese isolates showing different cultural features than those of Cryphonectria parasitica (Murrill) M.E. Barr from Japan and the USA, grouped with isolates of Cryphonectria nitschkei (G.H. Otth) M.E. Barr from Quercus spp. and Rhus javanica L. Isolates of Cryphonectria havanensis (Bruner) M.E. Barr from Quercus grosseserrata Blume, Castanopsis cuspidata Schottky, Pyrus sinensis Lindl., and Eucalyptus globulus Labill. also grouped in this phylogenetic clade. We propose that Cryphonectria nitschkei and the fungus that has been referred to as Cryphonectria havanensis in Japan should be treated as a single taxon. Phylogenetic and morphological data also suggest that there are two species currently representing Cryphonectria radicalis (Schwein.: Fr.) M.E. Barr in Europe. One of these species is similar to the type specimen of Cryphonectria radicalis, while the other species probably is new. [ABSTRACT FROM AUTHOR]

Genealogy-based methods were used to estimate migration of the fungal virus Cryphonectria hypovirus 1 between vegetative compatibility types of the host fungus, Cryphonectria parasitica, as a means of estimating horizontal transmission within two host populations. Vegetative incompatibility is a self/non-self recognition system that inhibits virus transmission under laboratory conditions but its effect on transmission in nature has not been clearly demonstrated. Recombination within and among different loci in the virus genome restricted the genealogical analyses to haplotypes with common mutation and recombinational histories. The existence of recombination necessitated that we also use genealogical approaches that can take advantage of both the mutation and recombinational histories of the sample. Virus migration between populations was significantly restricted. In contrast, estimates of migration between vegetative compatibility types were relatively high within populations despite previous evidence that transmission in the laboratory was restricted. The discordance between laboratory estimates and migration estimates from natural populations highlights the challenges in estimating pathogen transmission rates. Genealogical analyses inferred migration patterns throughout the entire coalescent history of one viral region in natural populations and not just recent patterns of migration or laboratory transmission. This application of genealogical analyses provides markedly stronger inferences on overall transmission rates than laboratory estimates do. [ABSTRACT FROM AUTHOR]

Summary Although Cryphonectria hypoviruses have been relatively successful as biological control agents of chestnut blight in Europe, their success in North America has been limited. Experimental releases of hypoviruses were made in 1978–82 at two sites in West Virginia forests with high densities of regenerating chestnut trees. Cryphonectria hypovirus 1 (CHV-1) from Europe, as well as American isolates of Cryphonectria parasitica containing CHV-3, were used for these releases. Although most trees died during the 5-year release period, it was not known if the hypoviruses persisted in the C. parasitica population at the two sites. When the experimental plots were revisited in 1994, few chestnut trees were found. The exception was one plot containing coppice sprouts that had grown from the root collars of the original trees. The authors intensively sampled C. parasitica from experimental plots and screened recovered isolates for double-stranded RNA (dsRNA). None of the isolates contained CHV-1; only six isolates contained CHV-3, all from the plots with the coppice sprouts. CHV-4, which occurs naturally in West Virginia forests and in two released isolates, hybridized to dsRNA from the isolates containing CHV-3, indicating mixed infections. CHV-4 also hybridized to dsRNA from other isolates sampled inside and outside the treated plots. In contrast to CHV-1 and CHV-3, however, CHV-4 has little effect on the growth or phenotype of C. parasitica . The limited persistence of CHV-1 and CHV-3 may have resulted when the C. parasitica population was reduced in size due to the failure of chestnut trees to resprout because of competition from other hardwood species. [ABSTRACT FROM AUTHOR]

Examines the genetic control of horizontal virus transmission in the chestnut blight fungus, Cryphonectria (C.) parasitica. Control of transmission by heteroallelism at vic loci, asymmetry, epistasis and host genetic background; Predictions from the logistic regression model; Control of virus transmission between individuals by vegetative incompatibility genes.

• Contamination with aflatoxin is correlated with broken outer layer of maize kernel. • Discoloration, but neither kernel breakage nor aflatoxin, is penalized by lower price. • Incentives to manage food safety losses in the informal maize market are absent. • Informing consumers that broken kernels are less safe could create such incentives. Like many other quality attributes, food safety tends to degrade after harvest, but unlike losses in quantity and many quality attributes, food safety losses are not readily observable by market actors. This implies the absence of incentives to address food safety losses specifically. To the extent that food safety is correlated with valued and observable food attributes, food safety losses may affect price indirectly, partially correcting this information failure. We analyze aflatoxin, a carcinogenic fungal contaminant, and visible quality data from 1500 maize samples and associated consumer surveys collected from clients of small-scale hammer mills in rural Kenya. We find that while losses in both food safety and observable quality increase with storage duration, only observable quality is rewarded by higher prices. There is no correlation between price and aflatoxin, implying an absence of market incentives to manage this aspect of quality loss. Further, of the two observable qualities on which data were collected, the one correlated most strongly with aflatoxin affects both prices and consumers' subjective quality assessment the least. Providing consumers with information about the correlation between visible grain attributes and contamination could allow consumers to direct contaminated grain to uses that minimize aflatoxin exposure and improve the incentives for provision of food safety in this market. [ABSTRACT FROM AUTHOR]