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In 1970, the Southern Corn Leaf Blight epidemic ravaged U.S. fields to great economic loss. The outbreak was caused by never-before-seen, supervirulent, Race T of the fungus Cochliobolus heterostrophus. The functional difference between Race T and O, the previously known, far less aggressive strain, is production of T-toxin, a host-selective polyketide. Supervirulence is associated with ~1 Mb of Race T-specific DNA; only a fraction encodes T-toxin biosynthetic genes (Tox1). Tox1 is genetically and physically complex, with unlinked loci (Tox1A, Tox1B) genetically inseparable from breakpoints of a Race O reciprocal translocation that generated hybrid Race T chromosomes. Previously, we identified 10 genes for T-toxin biosynthesis. Unfortunately, high-depth, short-read sequencing placed these genes on four small, unconnected scaffolds surrounded by repeated A+T rich sequence, concealing context. To sort out Tox1 topology and pinpoint the hypothetical Race O translocation breakpoints corresponding to Race T-specific insertions, we undertook PacBio long-read sequencing which revealed Tox1 gene arrangement and the breakpoints. Six Tox1A genes are arranged as three small islands in a Race T-specific sea (~634 kb) of repeats. Four Tox1B genes are linked, on a large loop of Race T-specific DNA (~210 kb). The race O breakpoints are short sequences of race O-specific DNA; corresponding positions in race T are large insertions of race T-specific, A+T rich DNA, often with similarity to transposable (predominantly Gypsy) elements. Nearby, are 'Voyager Starship' elements and DUF proteins. These elements may have facilitated Tox1 integration into progenitor Race O and promoted large scale recombination resulting in race T. IMPORTANCE In 1970 a corn disease epidemic ravaged fields in the United States to great economic loss. The outbreak was caused by a never-before seen, supervirulent strain of the fungal pathogen Cochliobolus heterostrophus. This was a plant disease epidemic, howev

Knowledge of pathogen biology and genetic diversity is a cornerstone of effective disease management, and accurateidentification of the pathogen is a foundation of pathogen biology. Species names provide an ideal framework for storageand retrieval of relevant information, a system that is contingent on a clear understanding of species boundaries andconsistent species identification. Verticillium, a genus of ascomycete fungi, contains important plant pathogens whosespecies boundaries have been ill defined. Using phylogenetic analyses, morphological investigations and comparisons toherbarium material and the literature, we established a taxonomic framework for Verticillium comprising ten species, five ofwhich are new to science. We used a collection of 74 isolates representing much of the diversity of Verticillium, andphylogenetic analyses based on the ribosomal internal transcribed spacer region (ITS), partial sequences of the proteincoding genes actin (ACT), elongation factor 1-alpha (EF), glyceraldehyde-3-phosphate dehydrogenase (GPD) and tryptophansynthase (TS). Combined analyses of the ACT, EF, GPD and TS datasets recognized two major groups within Verticillium, CladeFlavexudans and Clade Flavnonexudans, reflecting the respective production and absence of yellow hyphal pigments. CladeFlavexudans comprised V. albo-atrum and V. tricorpus as well as the new species V. zaregamsianum, V. isaacii and V.klebahnii, of which the latter two were morphologically indistinguishable from V. tricorpus but may differ in pathogenicity.Clade Flavnonexudans comprised V. nubilum, V. dahliae and V. longisporum, as well as the two new species V. alfalfae and V.nonalfalfae, which resembled the distantly related V. albo-atrum in morphology. Apart from the diploid hybrid V.longisporum, each of the ten species corresponded to a single clade in the phylogenetic tree comprising just one ex-typestrain, thereby establishing a direct link to a name tied to a herbarium specimen. A morphology-ba

Knowledge of pathogen biology and genetic diversity is a cornerstone of effective disease management, and accurateidentification of the pathogen is a foundation of pathogen biology. Species names provide an ideal framework for storageand retrieval of relevant information, a system that is contingent on a clear understanding of species boundaries andconsistent species identification. Verticillium, a genus of ascomycete fungi, contains important plant pathogens whosespecies boundaries have been ill defined. Using phylogenetic analyses, morphological investigations and comparisons toherbarium material and the literature, we established a taxonomic framework for Verticillium comprising ten species, five ofwhich are new to science. We used a collection of 74 isolates representing much of the diversity of Verticillium, andphylogenetic analyses based on the ribosomal internal transcribed spacer region (ITS), partial sequences of the proteincoding genes actin (ACT), elongation factor 1-alpha (EF), glyceraldehyde-3-phosphate dehydrogenase (GPD) and tryptophansynthase (TS). Combined analyses of the ACT, EF, GPD and TS datasets recognized two major groups within Verticillium, CladeFlavexudans and Clade Flavnonexudans, reflecting the respective production and absence of yellow hyphal pigments. CladeFlavexudans comprised V. albo-atrum and V. tricorpus as well as the new species V. zaregamsianum, V. isaacii and V.klebahnii, of which the latter two were morphologically indistinguishable from V. tricorpus but may differ in pathogenicity.Clade Flavnonexudans comprised V. nubilum, V. dahliae and V. longisporum, as well as the two new species V. alfalfae and V.nonalfalfae, which resembled the distantly related V. albo-atrum in morphology. Apart from the diploid hybrid V.longisporum, each of the ten species corresponded to a single clade in the phylogenetic tree comprising just one ex-typestrain, thereby establishing a direct link to a name tied to a herbarium specimen. A morphology-ba

BackgroundSelf/nonself discrimination is an essential feature for pathogen recognition and graft rejection and is a ubiquitous phenomenon in many organisms. Filamentous fungi, such as Neurospora crassa, provide a model for analyses of population genetics/evolution of self/nonself recognition loci due to their haploid nature, small genomes and excellent genetic/genomic resources. In N. crassa, nonself discrimination during vegetative growth is determined by 11 heterokaryon incompatibility (het) loci. Cell fusion between strains that differ in allelic specificity at any of these het loci triggers a rapid programmed cell death response.Methodology/principal findingsIn this study, we evaluated the evolution, population genetics and selective mechanisms operating at a nonself recognition complex consisting of two closely linked loci, het-c (NCU03493) and pin-c (NCU03494). The genomic position of pin-c next to het-c is unique to Neurospora/Sordaria species, and originated by gene duplication after divergence from other species within the Sordariaceae. The het-c pin-c alleles in N. crassa are in severe linkage disequilibrium and consist of three haplotypes, het-c1/pin-c1, het-c2/pin-c2 and het-c3/pin-c3, which are equally frequent in population samples and exhibit trans-species polymorphisms. The absence of recombinant haplotypes is correlated with divergence of the het-c/pin-c intergenic sequence. Tests for positive and balancing selection at het-c and pin-c support the conclusion that both of these loci are under non-neutral balancing selection; other regions of both genes appear to be under positive selection. Our data show that the het-c2/pin-c2 haplotype emerged by a recombination event between the het-c1/pin-c1 and het-c3/pin-c3 approximately 3-12 million years ago.Conclusions/significanceThese results support models by which loci that confer nonself discrimination form by the association of polymorphic genes with genes containing HET domains. Distinct allele classes

Anaplasma phagocytophilum is a tick-transmitted bacterial pathogen of humans and other animals, and is an obligate intracellular parasite. Throughout the course of infection, hosts acquire temporary resistance to granulocytic anaplasmosis as they develop immunity specific for the major antigen, major surface protein 2 (Msp2). However, the bacterium then utilizes a novel recombination mechanism shuffling functional pseudogenes sequentially into an expression cassette with conserved 5' and 3' ends, bypassing host immunity. Approximately 100 pseudogenes are present in the only fully sequenced human-origin HZ genome, representing the possibility for almost unlimited antigenic diversity. In the present study, we identified a select group of 20% of the A. phagocytophilum HZ msp2 pseudogenes that have matched preferentially to human, canine, and equine expression cassettes. Pseudogenes cluster predominantly in one spatial run limited to a single genomic island in less than 50% of the genome but phylogenetically related pseudogenes are neither necessarily located in close proximity on the genome nor share similar percent identity with expression cassettes. Pseudogenes near the expression cassette (and the origin) are more likely to be expressed than those farther away. Taken together, these findings suggest that there may be natural selection pressure to retain pseudogenes in one cluster near the putative origin of replication, even though global recombination shuffles pseudogenes around the genome, separating pseudogenes that share genetic origins as well as those with similar identities.