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Summary: Many pathogens evolved compartmentalized genomes with conserved core and variable accessory regions (ARs) that carry effector genes mediating virulence. The fungal plant pathogen Fusarium oxysporum has such ARs, often spanning entire chromosomes. The presence of specific ARs influences the host range, and horizontal transfer of ARs can modify the pathogenicity of the receiving strain. However, how these ARs evolve in strains that infect the same host remains largely unknown.We defined the pan‐genome of 69 diverse F. oxysporum strains that cause Fusarium wilt of banana, a significant constraint to global banana production, and analyzed the diversity and evolution of the ARs.Accessory regions in F. oxysporum strains infecting the same banana cultivar are highly diverse, and we could not identify any shared genomic regions and in planta‐induced effectors. We demonstrate that segmental duplications drive the evolution of ARs. Furthermore, we show that recent segmental duplications specifically in accessory chromosomes cause the expansion of ARs in F. oxysporum.Taken together, we conclude that extensive recent duplications drive the evolution of ARs in F. oxysporum, which contribute to the evolution of virulence. [ABSTRACT FROM AUTHOR]

This special issue presents new research of evolutionary processes in microbes associated with our near environment including human-associated microbes and microbes associated with food production. The study organisms include domesticated microorganisms (fungi and bacteria), microorganisms thriving in anthropic environments, host-associated microbiota, crop pathogens and animal pathogens. The study points to the importance of microbe-microbe interactions in shaping disease development and fungal metabolite production, an observation of great relevance in the context of disease treatment and crop protection. DOMESTICATED MICROORGANISMS AND MICROORGANISMS THRIVING IN ANTHROPIC ENVIRONMENTS The yeast I Brettanomyces bruxellensis i has adapted to many different anthropic environments, for example, wine, beer, bioethanol production and kombucha. [Extracted from the article]

Microorganisms are famous for adapting quickly to new environments. However, most evidence for rapid microbial adaptation comes from laboratory experiments or domesticated environments, and it is unclear how rates of adaptation scale from human‐influenced environments to the great diversity of wild microorganisms. We examined potential monthly‐scale selective pressures in the model forest yeast Saccharomyces paradoxus. Contrary to expectations of seasonal adaptation, the S. paradoxus population was stable over four seasons in the face of abiotic and biotic environmental changes. While the S. paradoxus population was diverse, including 41 unique genotypes among 192 sampled isolates, there was no correlation between S. paradoxus genotypes and seasonal environments. Consistent with observations from other S. paradoxus populations, the forest population was highly clonal and inbred. This lack of recombination, paired with population stability, implies that selection is not acting on the forest S. paradoxus population on a seasonal timescale. Saccharomyces paradoxus may instead have evolved generalism or phenotypic plasticity with regard to seasonal environmental changes long ago. Similarly, while the forest population included diversity among phenotypes related to intraspecific interference competition, there was no evidence for active coevolution among these phenotypes. At least ten percent of the forest S. paradoxus individuals produced "killer toxins," which kill sensitive Saccharomyces cells, but the presence of a toxin‐producing isolate did not predict resistance to the toxin among nearby isolates. How forest yeasts acclimate to changing environments remains an open question, and future studies should investigate the physiological responses that allow microbial cells to cope with environmental fluctuations in their native habitats. [ABSTRACT FROM AUTHOR]

Summary: Plants are colonized by microbial communities that have diverse implications for plant development and health. The establishment of a stable plant–bacteria interaction depends on a continuous coexistence over generations. Transmission via the seed is considered as the main route for vertical inheritance of plant‐associated bacteria. Nonetheless, the ecological principles that govern the plant colonization by seed endophytes remain understudied. Here we quantify the contribution of arrival time and colonization history to bacterial colonization of the wheat root. Establishing a common seed endophyte, Pantoea agglomerans, and wheat as a model system enabled us to document bacterial colonization of the plant roots during the early stages of germination. Using our system, we estimate the carrying capacity of the wheat roots as 108 cells g−1, which is robust among individual plants and over time. Competitions in planta reveal a significant advantage of early incoming colonizers over late‐incoming colonizers. Priming for the wheat environment had little effect on the colonizer success. Our experiments thus provide empirical data on the root colonization dynamics of a seed endophyte. The persistence of seed endophyte bacteria with the plant population over generations may contribute to the stable transmission that is one route for the evolution of a stable host‐associated lifestyle. [ABSTRACT FROM AUTHOR]

Cercospora beticola is a hemibiotrophic fungus that causes cercospora leaf spot disease of sugar beet (Beta vulgaris). After an initial symptomless biotrophic phase of colonization, necrotic lesions appear on host leaves as the fungus switches to a necrotrophic lifestyle. The phytotoxic secondary metabolite cercosporin has been shown to facilitate fungal virulence for several Cercospora spp. However, because cercosporin production and subsequent cercosporin‐initiated formation of reactive oxygen species is light‐dependent, cell death evocation by this toxin is only fully ensured during a period of light. Here, we report the discovery of the effector protein CbNip1 secreted by C. beticola that causes enhanced necrosis in the absence of light and, therefore, may complement light‐dependent necrosis formation by cercosporin. Infiltration of CbNip1 protein into sugar beet leaves revealed that darkness is essential for full CbNip1‐triggered necrosis, as light exposure delayed CbNip1‐triggered host cell death. Gene expression analysis during host infection shows that CbNip1 expression is correlated with symptom development in planta. Targeted gene replacement of CbNip1 leads to a significant reduction in virulence, indicating the importance of CbNip1 during colonization. Analysis of 89 C. beticola genomes revealed that CbNip1 resides in a region that recently underwent a selective sweep, suggesting selection pressure exists to maintain a beneficial variant of the gene. Taken together, CbNip1 is a crucial effector during the C. beticola–sugar beet disease process. [ABSTRACT FROM AUTHOR]

Homing endonucleases (HE) are enzymes capable of cutting DNA at highly specific target sequences, the repair of the generated double‐strand break resulting in the insertion of the HE‐encoding gene ("homing" mechanism). HEs are present in all three domains of life and viruses; in eukaryotes, they are mostly found in the genomes of mitochondria and chloroplasts, as well as nuclear ribosomal RNAs. We here report the case of a HE that accidentally integrated into a telomeric region of the nuclear genome of the fungal maize pathogen Ustilago maydis. We show that the gene has a mitochondrial origin, but its original copy is absent from the U. maydis mitochondrial genome, suggesting a subsequent loss or a horizontal transfer from a different species. The telomeric HE underwent mutations in its active site and lost its original start codon. A potential other start codon was retained downstream, but we did not detect any significant transcription of the newly created open reading frame, suggesting that the inserted gene is not functional. Besides, the insertion site is located in a putative RecQ helicase gene, truncating the C‐terminal domain of the protein. The truncated helicase is expressed during infection of the host, together with other homologous telomeric helicases. This unusual mutational event altered two genes: The integrated HE gene subsequently lost its homing activity, while its insertion created a truncated version of an existing gene, possibly altering its function. As the insertion is absent in other field isolates, suggesting that it is recent, the U. maydis 521 reference strain offers a snapshot of this singular mutational event. [ABSTRACT FROM AUTHOR]

Cercospora leaf spot, caused by the fungal pathogen Cercospora beticola, is the most destructive foliar disease of sugar beet worldwide. This review discusses C. beticola genetics, genomics, and biology and summarizes our current understanding of the molecular interactions that occur between C. beticola and its sugar beet host. We highlight the known virulence arsenal of C. beticola as well as its ability to overcome currently used disease management strategies. Finally, we discuss future prospects for the study and management of C. beticola infections in the context of newly employed molecular tools to uncover additional information regarding the biology of this pathogen. Taxonomy: Cercospora beticola Sacc.; Kingdom Fungi, Phylum Ascomycota, Class Dothideomycetes, Order Capnodiales, Family Mycosphaerellaceae, Genus Cercospora. Host range: Well‐known pathogen of sugar beet (Beta vulgaris subsp. vulgaris) and most species of the Beta genus. Reported as pathogenic on other members of the Chenopodiaceae (e.g., lamb's quarters, spinach) as well as members of the Acanthaceae (e.g., bear's breeches), Apiaceae (e.g., Apium), Asteraceae (e.g., chrysanthemum, lettuce, safflower), Brassicaceae (e.g., wild mustard), Malvaceae (e.g., Malva), Plumbaginaceae (e.g., Limonium), and Polygonaceae (e.g., broad‐leaved dock) families. Disease symptoms: Leaves infected with C. beticola exhibit circular lesions that are coloured tan to grey in the centre and are often delimited by tan‐brown to reddish‐purple rings. As disease progresses, spots can coalesce to form larger necrotic areas, causing severely infected leaves to wither and die. At the centre of these spots are black spore‐bearing structures (pseudostromata). Older leaves often show symptoms first and younger leaves become infected as the disease progresses. Management: Application of a mixture of fungicides with different modes of action is currently performed although elevated resistance has been documented in most employed fungicide classes. Breeding for high‐yielding cultivars with improved host resistance is an ongoing effort and prudent cultural practices, such as crop rotation, weed host management, and cultivation to reduce infested residue levels, are widely used to manage disease. Useful website: https://www.ncbi.nlm.nih.gov/genome/11237?genome_assembly_id=352037 [ABSTRACT FROM AUTHOR]

Summary: Access to greater genomic resolution through new sequencing technologies is transforming the field of plant pathology. As scientists embrace these new methods, some overarching patterns and observations come into focus. Evolutionary genomic studies are used to determine not only the origins of pathogen lineages and geographic patterns of genetic diversity, but also to discern how natural selection structures genetic variation across the genome. With greater and greater resolution, we can now pinpoint the targets of selection on a large scale. At multiple levels, crypsis and convergent evolution are evident. Host jumps and shifts may be more pervasive than once believed, and hybridization and horizontal gene transfer (HGT) likely play important roles in the emergence of genetic novelty. [ABSTRACT FROM AUTHOR]

Saccharomyces yeasts are emerging as model organisms for ecology and evolution, and researchers need environmental Saccharomyces isolates to test ecological and evolutionary hypotheses. However, methods for isolating Saccharomyces from nature have not been standardized, and isolation methods may influence the genotypes and phenotypes of studied strains. We compared the effectiveness and potential biases of an established enrichment culturing method against a newly developed direct plating method for isolating forest floor Saccharomyces spp. In a European forest, enrichment culturing was both less successful at isolating Saccharomyces paradoxus per sample collected and less labour intensive per isolated S. paradoxus colony than direct isolation. The two methods sampled similar S. paradoxus diversity: The number of unique genotypes sampled (i.e., genotypic diversity) per S. paradoxus isolate and average growth rates of S. paradoxus isolates did not differ between the two methods, and growth rate variances (i.e., phenotypic diversity) only differed in one of three tested environments. However, enrichment culturing did detect rare Saccharomyces cerevisiae in the forest habitat and also found two S. paradoxus isolates with outlier phenotypes. Our results validate the historically common method of using enrichment culturing to isolate representative collections of environmental Saccharomyces. We recommend that researchers choose a Saccharomyces sampling method based on resources available for sampling and isolate screening. Researchers interested in discovering new Saccharomyces phenotypes or rare Saccharomyces species from natural environments may also have more success using enrichment culturing. We include step‐by‐step sampling protocols in the supplemental materials. [ABSTRACT FROM AUTHOR]

Many filamentous plant pathogens exhibit high levels of genomic variability, yet the impact of this variation on host–pathogen interactions is largely unknown. We have addressed host specialization in the wheat pathogen Zymoseptoria tritici. Our study builds on comparative analyses of infection and gene expression phenotypes of three isolates and reveals the extent to which genomic variation translates into phenotypic variation. The isolates exhibit genetic and genomic variation but are similarly virulent. By combining confocal microscopy, disease monitoring, staining of ROS, and comparative transcriptome analyses, we conducted a detailed comparison of the infection processes of these isolates in a susceptible wheat cultivar. We characterized four core infection stages: establishment, biotrophic growth, lifestyle transition, and necrotrophic growth and asexual reproduction that are shared by the three isolates. However, we demonstrate differentiated temporal and spatial infection development and significant differences in the expression profiles of the three isolates during the infection stages. More than 20% of the genes were differentially expressed and these genes were located significantly closer to transposable elements, suggesting an impact of epigenetic regulation. Further, differentially expressed genes were enriched in effector candidates suggesting that isolate‐specific strategies for manipulating host defenses are present in Z. tritici. We demonstrate that individuals of a host‐specialized pathogen have highly differentiated infection programs characterized by flexible infection development and functional redundancy. This illustrates how high genetic diversity in pathogen populations results in highly differentiated infection phenotypes, which fact needs to be acknowledged to understand host–pathogen interactions and pathogen evolution. In our study, we address host specialization of the fungal plant pathogen Zymoseptoria tritici to wheat. We find an unexpected extent of genetic and phenotypic variation among three isolates that produce the same amount of disease on susceptible hosts. Our study reveals that host specialization in this organism entails a highly flexible infection program which may be important for rapid adaptation of this important crop pathogen. [ABSTRACT FROM AUTHOR]

Abstract: Host specialization is a key process in ecological divergence and speciation of plant‐associated fungi. The underlying determinants of host specialization are generally poorly understood, especially in endophytes, which constitute one of the most abundant components of the plant microbiome. We addressed the genetic basis of host specialization in two sympatric subspecies of grass‐endophytic fungi from the Epichloë typhina complex: subsp. typhina and clarkii. The life cycle of these fungi entails unrestricted dispersal of gametes and sexual reproduction before infection of a new host, implying that the host imposes a selective barrier on viability of the progeny. We aimed to detect genes under divergent selection between subspecies, experiencing restricted gene flow due to adaptation to different hosts. Using pooled whole‐genome sequencing data, we combined FST and DXY population statistics in genome scans and detected 57 outlier genes showing strong differentiation between the two subspecies. Genomewide analyses of nucleotide diversity (π), Tajima's D and dN/dS ratios indicated that these genes have evolved under positive selection. Genes encoding secreted proteins were enriched among the genes showing evidence of positive selection, suggesting that molecular plant–fungus interactions are strong drivers of endophyte divergence. We focused on five genes encoding secreted proteins, which were further sequenced in 28 additional isolates collected across Europe to assess genetic variation in a larger sample size. Signature of positive selection in these isolates and putative identification of pathogenic function supports our findings that these genes represent strong candidates for host specialization determinants in Epichloë endophytes. Our results highlight the role of secreted proteins as key determinants of host specialization. [ABSTRACT FROM AUTHOR]

The host-specific toxin ToxA produced by the wheat pathogens Pyrenophora tritici-repentis and Phaeosphaeria nodorum interacts with the product of the dominant plant gene Tsn1 to induce necrosis. The ToxA gene is thought to have been acquired by Py. tritici-repentis from Ph. nodorum through a recent horizontal gene transfer event. PCR and sequence analysis indicate that the level of ToxA variation, including gene deletion, in Ph. nodorum ( SnToxA) is significantly higher than in Py. tritici-repentis ( PtrToxA). We PCR-screened 788 isolates of Ph. nodorum originating from eight geographical regions to infer the pattern of SnToxA deletions. The frequency of deletions differed significantly among populations, ranging from 0% (Australia) to 98% (China). Sequence analysis of the SnToxA gene in 123 Ph. nodorum isolates revealed 13 distinct haplotypes. The distribution and diversity of haplotypes varied significantly among populations. The majority of SnToxA mutations were non-synonymous resulting in changes at the protein level. We applied different models of selection to infer the mode of evolution operating at the ToxA locus. Evidence for positive diversifying selection supports the hypothesis that evolution of the ToxA locus is driven by selection imposed by the host. The distribution of SnToxA alleles and deletions may reflect the distribution of different Tsn1 alleles in the corresponding host populations. [ABSTRACT FROM AUTHOR]

The global migration patterns of the fungal wheat pathogen Phaeosphaeria nodorum were analysed using 12 microsatellite loci. Analysis of 693 isolates from nine populations indicated that the population structure of P. nodorum is characterized by high levels of genetic diversity and a low degree of subdivision between continents. To determine whether genetic similarity of populations was a result of recent divergence or extensive gene flow, the microsatellite data were analysed using an isolation-with-migration model. We found that the continental P. nodorum populations diverged recently, but that enough migration occurred to reduce population differentiation. The migration patterns of the pathogen indicate that immigrants originated mainly from populations in Europe, China and North America. [ABSTRACT FROM AUTHOR]

A nested multiplex PCR (polymerase chain reaction) approach was used for multilocus genotyping of arbuscular mycorrhizal fungal populations. This method allowed us to amplify multiple loci fromGlomussingle spores in a single PCR amplification. Variable introns in the two protein coding genesGmFOX2andGmTOR2were applied as codominant genetic markers together with the LSU rDNA. Genetic structure ofGlomusspp. populations from an organically and a conventionally cultured field were compared by hierarchical sampling of spores from four plots in each field. Multilocus genotypes were characterized by SSCP (single stranded conformation polymorphism) and sequencing. All spore genotypes were unique suggesting that no recombination was taking place in the populations. There were no overall differences in the distribution of genotypes in the two fields and identical genotypes could be sampled from both fields. Analysis of gene diversity indicated thatGlomuspopulations are subdivided between plots within each field. There were however, no subdivision between the fields. [ABSTRACT FROM AUTHOR]

Arbuscular mycorrhizal fungi (AMF) form a mutualistic symbiosis with plant roots and are found in most ecosystems. In this study the community structure of AMF in a clade of the genus Glomus was examined in undisturbed costal grassland using LSU rDNA sequences amplified from roots of Hieracium pilosella. Roots were sampled from May to November along eight 30-m transects, 30–120 m apart. Phylogenetic analysis of the sequences revealed 11 phylogenetic clusters within the clade of Glomus. The phylogenetic clusters were patchily distributed within the area; time had no influence on the distribution pattern. The dominant cluster covered up to 10 m along the transect, whereas other clusters formed what can be interpreted as small individual mycelia. Four of the phylogenetic clusters included known species; the other clusters, including the dominant sequence types, were unknown. The dominant phylogenetic cluster enclosed nine haplotypes, and analyses of genetic diversity of this phylogenetic cluster showed that the total diversity could be found within single root fragments, suggesting that the multiple sequences were derived from a single individual. [ABSTRACT FROM AUTHOR]

Eleven polymorphic microsatellite loci and one minisatellite locus originating from expressed sequence tag (EST) libraries of Phaeosphaeria (syn. Stagonospora) nodorum were isolated and characterized. The satellite markers were used to genotype isolates from field populations collected in China, North America and South Africa. The number of alleles per locus ranged from two to 15. Genotype diversity ranged from 87.5 to 95.3 and gene diversity from 0.1 to 0.8. The variable levels of polymorphism within and among populations of P. nodorum renders these 12 satellite loci ideal markers for population genetic analysis of P. nodorum. [ABSTRACT FROM AUTHOR]