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Many filamentous ascomycetes develop three-dimensional fruiting bodies for production and dispersal of sexual spores. Fruiting bodies are among the most complex structures differentiated by ascomycetes; however, the molecular mechanisms underlying this process are insufficiently understood. Previous comparative transcriptomics analyses of fruiting body development in different ascomycetes suggested that there might be a core set of genes that are transcriptionally regulated in a similar manner across species. Conserved patterns of gene expression can be indicative of functional relevance, and therefore such a set of genes might constitute promising candidates for functional analyses. In this study, we have sequenced the genome of the Pezizomycete Ascodesmis nigricans, and performed comparative transcriptomics of developing fruiting bodies of this fungus, the Pezizomycete Pyronema confluens, and the Sordariomycete Sordaria macrospora With only 27 Mb, the A. nigricans genome is the smallest Pezizomycete genome sequenced to date. Comparative transcriptomics indicated that gene expression patterns in developing fruiting bodies of the three species are more similar to each other than to nonsexual hyphae of the same species. An analysis of 83 genes that are upregulated only during fruiting body development in all three species revealed 23 genes encoding proteins with predicted roles in vesicle transport, the endomembrane system, or transport across membranes, and 13 genes encoding proteins with predicted roles in chromatin organization or the regulation of gene expression. Among four genes chosen for functional analysis by deletion in S. macrospora, three were shown to be involved in fruiting body formation, including two predicted chromatin modifier genes.

Several Phyllosticta species are known as pathogens of Citrus spp., and are responsible for various disease symptoms including leaf and fruit spots. One of the most important species is P. citricarpa, which causes a foliar and fruit disease called citrus black spot. The Phyllosticta species occurring on citrus can most effectively be distinguished from P. citricarpa by means of multilocus DNA sequence data. Recent studies also demonstrated P. citricarpa to be heterothallic, and reported successful mating in the laboratory. Since the domestication of citrus, different clones of P. citricarpa have escaped Asia to other continents via trade routes, with obvious disease management consequences. This pathogen profile represents a comprehensive literature review of this pathogen and allied taxa associated with citrus, focusing on identification, distribution, genomics, epidemiology and disease management. This review also considers the knowledge emerging from seven genomes of Phyllosticta spp., demonstrating unknown aspects of these species, including their mating behaviour.TaxonomyPhyllosticta citricarpa (McAlpine) Aa, 1973. Kingdom Fungi, Phylum Ascomycota, Class Dothideomycetes, Order Botryosphaeriales, Family Phyllostictaceae, Genus Phyllosticta, Species citricarpa.Host rangeConfirmed on more than 12 Citrus species, Phyllosticta citricarpa has only been found on plant species in the Rutaceae.Disease symptomsP. citricarpa causes diverse symptoms such as hard spot, virulent spot, false melanose and freckle spot on fruit, and necrotic lesions on leaves and twigs.Useful websitesDOE Joint Genome Institute MycoCosm portals for the Phyllosticta capitalensis (https://genome.jgi.doe.gov/Phycap1), P. citriasiana (https://genome.jgi.doe.gov/Phycit1), P. citribraziliensis (https://genome.jgi.doe.gov/Phcit1), P. citrichinaensis (https://genome.jgi.doe.gov/Phcitr1), P. citricarpa (https://genome.jgi.doe.gov/Phycitr1, https://genome.jgi.doe.gov/Phycpc1), P. paracitricarpa (https://genome.jgi.doe.gov/Phy27169) genomes. All available Phyllosticta genomes on MycoCosm can be viewed at https://genome.jgi.doe.gov/Phyllosticta.

Carrillo, J.D., Rugman-Jones, PF., Husein, D., Stajich, J.E., Kasson, M.T., Carrillo, D., Stouthamer, R., and Eskalen, A. 2019. Members of the Euwallacea fornicatus species complex exhibit promiscuous mutualism with ambrosia fungi in Taiwan. A number of ambrosia beetles have come to prominence in recent years because of the damage they inflict on a variety of trees within invaded habitats across the globe. Ambrosia beetles rely on symbiotic microorganisms, mainly fungi, as a dedicated food source and carry those microorganisms around with them within specialized organs termed mycangia. Investigation of members of the Euwallacea fornicatus species complex and their fungal symbionts in Taiwan revealed promiscuous symbioses with ambrosial Fusaria clade (AFC) members, Graphium spp., and Paracremonium spp. based on co-phylogenetic analyses. For AFC members, a novel diagnostic PCR assay targeting mating type genes MAT1-1-1 and MAT1-2-1 was developed and validated by amplicon size and sequencing. Mating type screening of AFC members revealed the isolates screened are all heterothallic (self-sterile), with both MAT types represented and recovered from fungi vectored by E. fornicatus (tea shot hole borer), E. kuroshio (Kuroshio shot hole borer), and E. whitfordiodendrus (polyphagous shot hole borer) in Taiwan. Members of the Euwallacea fornicatus species complex and the variety of ambrosia fungi they utilize further confirms that their relationship with these fungi are more likely promiscuous in native areas, as opposed to strictly obligate to a specific combination of fungi as observed in invaded areas.

Phytophthora colocasiae is an important pathogen that causes great economic losses in taro production in tropical and subtropical regions, especially in Japan. Understanding the genetic variations in P. colocasiae populations and their transmission patterns in Japan is essential for effective disease control. Here, the genetic diversity of 358 P. colocasiae isolates, including 348 from Japan, 7 from China, and 3 from Indonesia, was assessed using 11 simple sequence repeat (SSR) primer pairs with high polymorphism. The phylogenetic tree of the SSR locus showed that the isolates from Japan could be divided into 14 groups, with group A being the dominant group. Among foreign isolates, only six from mainland China were similar to those from Japan and clustered in groups B and E. Analysis of molecular variance (AMOVA), principal components analysis (PCA), and cluster analysis (K = 3) results revealed a moderate level of genetic diversity, mainly within individuals. Populations showed high heterozygosity, a lack of regional differentiation, and frequent gene flow. Analysis of mating types and ploidy levels revealed that A2 and self-fertile (SF) A2 types and tetraploids were dominant across populations. Explanations and hypotheses for the results can provide more effective strategies for disease management of taro leaf blight. [ABSTRACT FROM AUTHOR]

Summary: The Closterium peracerosum–strigosum–littorale complex (Closterium, Zygnematophyceae) has an isogamous mating system. Members of the Zygnematophyceae are the closest relatives to extant land plants and are distantly related to chlorophytic models, for which a genetic basis of mating type (MT) determination has been reported. We thus investigated MT determination in Closterium.We sequenced genomes representing the two MTs, mt+ and mt−, in Closterium and identified CpMinus1, a gene linked to the mt− phenotype. We analyzed its function using reverse genetics methods.CpMinus1 encodes a divergent RWP‐RK domain‐containing‐like transcription factor and is specifically expressed during gamete differentiation. Introduction of CpMinus1 into an mt+ strain was sufficient to convert it to a phenotypically mt− strain, while CpMinus1‐knockout mt− strains were phenotypically mt+.We propose that CpMinus1 is the major MT determinant that acts by evoking the mt− phenotype and suppressing the mt+ phenotype in heterothallic Closterium. CpMinus1 likely evolved independently in the Zygnematophyceae lineage, which lost an egg–sperm anisogamous mating system. mt− specific regions possibly constitute an MT locus flanked by common sequences that undergo some recombination. [ABSTRACT FROM AUTHOR]

Potato (Solanum tuberosum L.) and tomato (S. lycopersicum L.) are the most economically important vegetable crops in Egypt and worldwide. The winter crop in Egypt is particularly prone to late blight caused by Phytophthora infestans. A total of 152 P. infestans isolates were isolated from the 2013, 2014, 2016 and 2018 winter crops with 82 isolates from potato, 69 from tomato and one isolate from eggplant (S. melongena L.). All isolates belonged to the A1 mating type with no evidence of A2 or self-fertile strains. The majority of isolates (53%) were sensitive to metalaxyl, 32% were intermediate and 15% were resistant. Variation in aggressiveness between three P. infestans isolates EG-005 (13_A2) and EG-276 (23_A1) from potato, and EG-237 (23_A1) from eggplant was determined on tuber slices and leaflets of 10 potato cultivars. The eggplant isolate EG-237 showed higher sporulation capacity compared with the other tested isolates and was able to infect potato (Lady Rosetta cv) and tomato (Super Strain B cv). The simple sequence repeat (SSR) genotyping data showed that in contrast to our previous work (3-year period 2010-12) in which the proportion of 13_A2 lineage was 35%, all isolates belonged to the 23_A1 lineage. There was no evidence for the existence of the A2 mating type or 13_A2 lineage even in the destroyed field crops of some cultivars (Cara, Bellini and Valor) that had been reported as resistant to 23_A1. The data have been submitted into the Euroblight database to allow temporal and spatial genetic diversity to be examined in comparison with other regional P. infestans populations. The AVR2 and AVR2-like RXLR effector genes were amplified and sequenced. In the avirulent AVR2 gene, only one heterozygous SNP was detected at position 31 in the N terminus in six isolates out of eleven, whereas two heterozygous SNPs were detected at position 29 in the N-terminus and ninety-two in the C- terminus of the AVR2-like gene. This suggests that changes in the previously reported virulence profile of 23_A1 are not related to commercial cultivars carrying the R2 gene. In addition, this is the first report of P. infestans on eggplant in Egypt. [ABSTRACT FROM AUTHOR]

In Bangladesh, the third largest producer of potatoes in Asia, late blight, caused by Phytophthora infestans, is the major constraint to production. Nevertheless, there is a lack of published information on the pathogen population. A collection of 69 isolates obtained from samples of infected potato foliage collected in 2018–2019 varied significantly in their aggressiveness to detached potato leaflets and tuber slices of cv. Diamant. On leaflets, most isolates were highly aggressive, colonizing more than half the surface in 7 days. On tuber slices, while some isolates were highly aggressive, those collected in 2019 were less so. There was a significant effect of the locations from which the isolates were obtained on aggressiveness to both leaflets and tuber slices, but aggressiveness to leaflets and to tuber slices was not significantly correlated. All isolates were A2 mating type and either intermediate (38%) or resistant (62%) in sensitivity to the fungicide metalaxyl. A subset of 24 isolates all had mitochondrial DNA haplotype Ia. Samples of P. infestans DNA (124) from infected potato foliage obtained from Bangladesh (2014–2019) were genotyped by 12-plex SSR. The vast majority (95%) were assigned to EU_13_A2, which was present in every year in which samples were collected. Numerous EU_13_A2 variants were identified, many specific to Bangladesh. Six samples (from 2017 to 2019) had genotypes distinct from EU_13_A2 and with no close match to known European lineages. It is concluded that the current P. infestans population of Bangladesh is dominated by the aggressive lineage EU_13_A2, also dominant in India and Pakistan. The implications of this for control of late blight are discussed. [ABSTRACT FROM AUTHOR]

Phytophthora infestans is the oomycete responsible for late blight disease of Solanaceae that causes both yield and economic losses. With the aim of reducing plant wilt and high management costs mainly due to wide fungicide applications, alternative eco-sustainable control strategies are needed. RNA interference (RNAi) is a powerful tool for gene function studies that can be accomplished by constitutive transformation or transient expression such as virus-induced gene silencing (VIGS) experiments. VIGS makes use of viruses to deliver sequences homologous to a target gene fragment and trigger RNAi. Indeed, a P. infestans ortholog of plant loricrin-like protein (LLP), named PiLLP, has been silenced using the direct infection of a recombinant vector based on the plant virus tobacco mosaic virus (TMV-PiLLP-1056), aiming to reduce the oomycete sexual reproduction. For this purpose, the gene coding for the green fluorescent protein (GFP) present in the TMV-GFP-1056 vector has been replaced with an antisense construct obtained by fusion PCR of the PiLLP 5′-UTR and 3′-UTR sequences. Here, we show that RNAi can be expressed in the A1 mating type of P. infestans strain 96.9.5.1 by VIGS using the direct infection of TMV-PiLLP-1056. We provide evidence that the recombinant vector can enter, replicate, and persist in mycelia of P. infestans where it induces the partial downregulation of the PiLLP transcript. Compared with the wild-type, the PiLLP-silenced A1 mating type had slower colony growth and a diminished virulence in detached tomato leaflets. This seems to be the first evidence of a constitutive gene downregulation of P. infestans using a recombinant vector based on a plus-sense RNA plant virus. [ABSTRACT FROM AUTHOR]

Fusarium circinatum is an introduced fungal pathogen extended to the northern regions of Spain that causes Pine Pitch Canker (PPC) disease. In this work, we analyzed the pathogen's genetic diversity to study changes over time and space since the first outbreak occurred in Spain. Using six polymorphic SSR markers, 15 MLGs were identified in 66 isolates, and only three haplotypes were found with frequencies higher than one. In general, genotypic diversity was low and decreased shortly over time in the northwestern regions while maintained at País Vasco, where only one haplotype (MLG32) was detected 10 years. This population also included isolates of a single mating type (MAT-2) and VCGs identified in only two groups, while isolates from NW regions were of both mating types and VCGs represented in 11 groups. The existence of haplotype MLG32 maintained on time and widely distributed suggests its good adaptation to the environment and the host. Results showed that the pathogen in País Vasco remains clearly differentiated from other northwestern populations. This fact was supported with no evidence of migration among regions. Results are explained by the asexual reproduction, but also selfing at least to a lesser extent that leads to identification of two new haplotypes. [ABSTRACT FROM AUTHOR]

Throughout history, the yeast Saccharomyces cerevisiae has played a central role in human society due to its use in food production and more recently as a major industrial and model microorganism, because of the many genetic and genomic tools available to probe its biology. However, S. cerevisiae has proven difficult to engineer to expand the carbon sources it can utilize, the products it can make, and the harsh conditions it can tolerate in industrial applications. Other yeasts that could solve many of these problems remain difficult to manipulate genetically. Here, we engineered the thermotolerant yeast Kluyveromyces marxianus to create a new synthetic biology platform. Using CRISPR-Cas9 (clustered regularly interspaced short palindromic repeats with Cas9)-mediated genome editing, we show that wild isolates of K. marxianus can be made heterothallic for sexual crossing. By breeding two of these mating-type engineered K. marxianus strains, we combined three complex traits-thermotolerance, lipid production, and facile transformation with exogenous DNA-into a single host. The ability to cross K. marxianus strains with relative ease, together with CRISPR-Cas9 genome editing, should enable engineering of K. marxianus isolates with promising lipid production at temperatures far exceeding those of other fungi under development for industrial applications. These results establish K. marxianus as a synthetic biology platform comparable to S. cerevisiae, with naturally more robust traits that hold potential for the industrial production of renewable chemicals.IMPORTANCE The yeast Kluyveromyces marxianus grows at high temperatures and on a wide range of carbon sources, making it a promising host for industrial biotechnology to produce renewable chemicals from plant biomass feedstocks. However, major genetic engineering limitations have kept this yeast from replacing the commonly used yeast Saccharomyces cerevisiae in industrial applications. Here, we describe genetic tools for genome editing and breeding K. marxianus strains, which we use to create a new thermotolerant strain with promising fatty acid production. These results open the door to using K. marxianus as a versatile synthetic biology platform organism for industrial applications.

Verticillium dahliae is a soil-borne fungal pathogen responsible for vascular wilt diseases in more than 300 dicotyledonous species, including solanaceous vegetable crops. In this study, a collection of 89 Brazilian V. dahliae isolates was characterized by combing molecular information for mating type and physiological race determination, as well as via virulence bioassays employing a set of differentials. Based on the virulence assays, three isolates were classified as race 1, 76 were classified as race 2, whereas ten isolates did not cause any symptom on the tested cultivars." In race-specific detection, a total of six isolates were identified as race 1, 70 as race 2, and 13 isolates displayed no amplicon with any primer set employed. Therefore, V. dahliae race 2 isolates are currently ubiquitous across major Solanaceae-producing areas in Brazil. Both MAT idiomorphs were detected, but a larger number of the V. dahliae isolates displayed the MAT1–1 (82%) pattern in our molecular analyses. The simultaneous presence of both MAT idiomorphs opens the possibility of sporadic events of sexual reproduction among V. dahliae populations from Brazil, enabling the potential emergence of either new recombinant isolates with broader host ranges or novel physiological races. Our results clearly indicated the need to intensify the search for effective sources of resistance to V. dahliae race 2 in tomato breeding programs under Brazilian conditions. [ABSTRACT FROM AUTHOR]

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Blackleg (Phoma) disease, caused by the ascomycete fungi Plenodomus biglobosus and P. lingam, threatens oilseed rape (OSR; Brassica napus) crops internationally. In many parts of the world, both species co-occur, but in China only P. biglobosus has so far been reported. Plenodomus biglobosus reproduces asexually (pycnidiospores), but also sexually (pseudothecia-yielding ascospores), via a heterothallic mating system requiring MAT1-1 and MAT1-2 genotypes. However, the roles of airborne ascospore inoculum in driving blackleg disease outbreaks in China are less well understood compared to elsewhere in the world. This is despite the very different agronomic cropping practices in parts of China, in which paddy rice and OSR are often grown in rotation; OSR stubble is often submerged under water for long periods potentially affecting pseudothecial development. Here, we indirectly investigate the potential role of sexual reproduction by developing new polymerase chain reaction (PCR) -based mating-type diagnostics for P. biglobosus and subsequently screening an international collection of 59 European and 157 Chinese isolates. Overall, in both Europe and China, P. biglobosus mating types did not deviate from a 1:1 ratio, such as is generally thought to occur under frequency-dependent selection in sexually reproducing pathogen populations. Both mating types were balanced in all the individual European countries tested (Austria, France, Poland, UK). Conversely, in China, mating types were only balanced in the eastern region; in the northern and southwestern regions there were skewed ratios, more typical of predominantly asexual reproduction, towards MAT1-1 and MAT1-2, respectively. The implications of these findings and future research directions for improved understanding of P. biglobosus epidemiology on OSR, particularly in China, are considered. [ABSTRACT FROM AUTHOR]

In recent years, Brown spot disease of rice (BSR) has been observed on leaves and seeds of rice in all rice-growing areas of Burkina Faso. Bipolaris oryzae and Exserohilum rostratum are the main fungal species isolated from BSR infected tissues and they are frequently observed in the same field. However, we are lacking information on the genetic diversity and population structure of these fungi in Burkina Faso. The mode of reproduction is also unknown. The genetic diversity of isolates of B. oryzae (n=61) and E. rostratum (n=151), collected from major ricegrowing areas of Burkina Faso, was estimated using genotyping-by-sequencing (GBS). The mean values for nucleotide diversity (p) were 1.9 x10-4 for B. oryzae and 4.8 x10-4 for E. rostratum. There is no genetic differentiation between the geographical populations of each species. The analysis of molecular variance revealed that 89% and 94% of the genetic variances were within the populations of B. oryzae and E. rostratum, respectively. For each species, four genetic clusters were identified by two clustering methods (DAPC and sNMF). The distribution of these genetic groups was independent of the geographical origin of the isolates. Evidence of recombination was detected in the populations of B. oryzae and E. rostratum. For B. oryzae balanced mating type ratios were supporting sexual reproduction. For E. rostratum overrepresentation of MAT1-2 isolates (79%) suggested a predominant asexual reproduction. This study provides important information on the biology and genetics of the two major fungi causing brown spot disease of rice in Burkina Faso. [ABSTRACT FROM AUTHOR]

Lecanosticta acicola is a pine needle pathogen causing brown spot needle blight that results in premature needle shedding with considerable damage described in North America, Europe, and Asia. Microsatellite and mating type markers were used to study the population genetics, migration history, and reproduction mode of the pathogen, based on a collection of 650 isolates from 27 countries and 26 hosts across the range of L. acicola. The presence of L. acicola in Georgia was confirmed in this study. Migration analyses indicate there have been several introduction events from North America into Europe. However, some of the source populations still appear to remain unknown. The populations in Croatia and western Asia appear to originate from genetically similar populations in North America. Intercontinental movement of the pathogen was reflected in an identical haplotype occurring on two continents, in North America (Canada) and Europe (Germany). Several shared haplotypes between European populations further suggests more local pathogen movement between countries. Moreover, migration analyses indicate that the populations in northern Europe originate from more established populations in central Europe. Overall, the highest genetic diversity was observed in south‐eastern USA. In Europe, the highest diversity was observed in France, where the presence of both known pathogen lineages was recorded. Less than half of the observed populations contained mating types in equal proportions. Although there is evidence of some sexual reproduction taking place, the pathogen spreads predominantly asexually and through anthropogenic activity. [ABSTRACT FROM AUTHOR]

Aflatoxin contamination of corn is a major threat to the safe food and feed. The United States Federal Grain Inspection Service (FGIS) monitors commercial grain shipments for the presence of aflatoxin. A total of 146 Aspergillus flavus were isolated from 29 highly contaminated grain samples to characterize the visual phenotypes, aflatoxin-producing potential, and genotypes to explore the etiological cause of high aflatoxin contamination of US corn. Five of the isolates had reduced sensitivity (43–49% resistant) to the fungicide azoxystrobin, with the remainder all being over 50% resistant to azoxystrobin at the discriminating dose of 2.5 µg/mL. Only six isolates of the highly aflatoxigenic S morphotype were found, and 48 isolates were non-aflatoxigenic. Analysis of the mating type locus revealed 45% MAT 1-1 and 55% MAT 1-2. The A. flavus population originating from the highly aflatoxin contaminated grain samples was compared to a randomly selected subset of isolates originating from commercial corn samples with typical levels of aflatoxin contamination (average < 50 ppb). Use of simple sequence repeat (SSR) genotyping followed by principal component analysis (PCoA) revealed a similar pattern of genotypic distribution in the two populations, but greater diversity in the FGIS-derived population. The noticeable difference between the two populations was that genotypes identical to strain NRRL 21882, the active component of the aflatoxin biocontrol product Afla-Guard™, were ten times more common in the commercial corn population of A. flavus compared to the population from the high-aflatoxin corn samples. The other similarities between the two populations suggest that high aflatoxin concentrations in corn grain are generally the result of infection with common A. flavus genotypes. [ABSTRACT FROM AUTHOR]

Blue cheeses, including renowned mold-ripened varieties such as Roquefort (France), Gorgonzola (Italy), Stilton (UK), Danablue (Denmark), and Cabrales (Spain), owe their distinct blue-green color and unique flavor to the fungal species Penicillium roqueforti. In Turkey, traditional cheeses similar to blue cheeses, namely mold-ripened Tulum and Civil, employ production techniques distinct from their European counterparts. Notably, mold-ripening in Turkish cheeses is spontaneous and does not involve starter cultures. Despite P. roqueforti being recognized for its distinct genetic populations sourced from various blue cheeses and non-cheese origins globally, the characteristics of the P. roqueforti population within Turkish cheeses remain unexplored. This study aimed to unravel the genetic characteristics and population structure of P. roqueforti from Turkish mold-ripened cheeses. Analysis of mold-ripened Civil (n = 22) and Tulum (n = 8) samples revealed 66 P. roqueforti isolates (76.6 % of total fungal isolates). Subsequently, these isolates (n = 66) and those from previous studies (Tulum n = 53, Golot n = 1) were used to assess genetic characteristics and mating genotypes. All 120 isolates harbored horizontal transfer regions (Wallaby and CheesyTer) and predominantly possessed the MAT1-2 mating genotype, similar to global blue cheese populations. However, most lacked the mpaC deletion associated with such populations. Analysis of the population with three polymorphic microsatellite markers revealed 36 haplotypes (HTs). Some cheeses contained isolates with different HTs or opposite mating genotypes, aligning with spontaneous fungal growth. Tulum and Civil isolates exhibited similar population diversity without forming distinct subgroups. Phylogenetic analysis of 20 selected isolates showed 75 % aligning with global blue cheese isolates, while 25 % formed unique clades. Overall, Turkish P. roqueforti isolates share genetic similarities with global populations but exhibit unique characteristics, suggesting potential new clades deserving further investigation. This research illuminates the characteristics of P. roqueforti isolates from Turkish cheeses, contributing to the knowledge of the global intraspecific diversity of the P. roqueforti species. • All P. roqueforti isolates originating from Turkey harbor Wallaby and CheesyTer. • Some cheeses contain isolates with opposite mating types and multiple strains • Isolates from Tulum and Civil cheeses do not represent distinct sub-populations. • Phylogenetically, most isolates group with Non-Roquefort and Termignon isolates. [ABSTRACT FROM AUTHOR]

In an attempt to determine the mating type of different Sporothrix schenckii sensu stricto isolates that remained viable after a long period of preservation in a culture collection and to correlate them with the degree of virulence/pathogenicity, a PCR technique using primers designed for the sequences of MAT1-1-1 and MAT1-2-1 genes and a murine experimental model were used. The results showed that there was no correlation between the mating type and virulence among the isolates. Furthermore, different degrees of virulence/pathogenicity, ranging from high to low, were found among them based on different virulence parameters. It was assumed that the long period of preservation favored the changes, yielding the isolation of variants. Thus, we believe that new technologies for studies on factors can improve our knowledge of the pathogenesis of sporotrichosis.

The heterochromatin spreading reaction is a central contributor to the formation of gene-repressive structures, which are re-established with high positional precision, or fidelity, following replication. How the spreading reaction contributes to this fidelity is not clear. To resolve the origins of stable inheritance of repression, we probed the intrinsic character of spreading events in fission yeast using a system that quantitatively describes the spreading reaction in live single cells. We show that spreading triggered by noncoding RNA-nucleated elements is stochastic, multimodal, and fluctuates dynamically across time. This lack of stability correlates with high histone turnover. At the mating type locus, this unstable behavior is restrained by an accessory cis-acting element REIII, which represses histone turnover. Further, REIII safeguards epigenetic memory against environmental perturbations. Our results suggest that the most prevalent type of spreading, driven by noncoding RNA-nucleators, is epigenetically unstable and requires collaboration with accessory elements to achieve high fidelity.

The capacity of Candida albicans to switch reversibly between the white phenotype and the opaque phenotype is required for the fungus to mate. It also influences virulence during hematogenously disseminated candidiasis. We investigated the roles of the mating type loci (MTL) and white-opaque switching in the capacity of C. albicans to mate in the oropharynx and cause oropharyngeal candidiasis (OPC). When immunosuppressed mice were orally infected with mating-competent opaque a/a and α/α cells either alone or mixed with white cells, no detectable mating occurred, indicating that the mating frequency was less than 1.6 × 10-6 Opaque cells were also highly attenuated in virulence; they either were cleared from the oropharynx or switched to the white phenotype during OPC. Although there were strain-to-strain differences in the virulence of white cells, they were consistently more virulent than opaque cells. In vitro studies indicated that relative to white cells, opaque cells had decreased capacity to invade and damage oral epithelial cells. The reduced invasion of at least one opaque strain was due to reduced surface expression of the Als3 invasin and inability to activate the epidermal growth factor receptor, which is required to stimulate the epithelial cell endocytic machinery. These results suggest that mating is a rare event during OPC because opaque cells have reduced capacity to invade and damage the epithelial cells of the oral mucosa.

Phylogenomic approaches have the potential to improve confidence about the inter-relationships of species in the order Mucorales within the fungal tree of life. Rhizopus species are especially important as plant and animal pathogens and bioindustrial fermenters for food and metabolite production. A dataset of 192 orthologous genes was used to construct a phylogenetic tree of 21 Rhizopus strains, classified into four species isolated from habitats of industrial, medical and environmental importance. The phylogeny indicates that the genus Rhizopus consists of three major clades, with R. microsporus as the basal species and the sister lineage to R. stolonifer and two closely related species R. arrhizus and R. delemar A comparative analysis of the mating type locus across Rhizopus reveals that its structure is flexible even between different species in the same genus, but shows similarities between Rhizopus and other mucoralean fungi. The topology of single-gene phylogenies built for two genes involved in mating is similar to the phylogenomic tree. Comparison of the total length of the genome assemblies showed that genome size varies by as much as threefold within a species and is driven by changes in transposable element copy numbers and genome duplications.

The shaping of individual cells requires a tight coordination of cell mechanics and growth. However, it is unclear how information about the mechanical state of the wall is relayed to the molecular processes building it, thereby enabling the coordination of cell wall expansion and assembly during morphogenesis. Combining theoretical and experimental approaches, we show that a mechanical feedback coordinating cell wall assembly and expansion is essential to sustain mating projection growth in budding yeast (Saccharomyces cerevisiae). Our theoretical results indicate that the mechanical feedback provided by the Cell Wall Integrity pathway, with cell wall stress sensors Wsc1 and Mid2 increasingly activating membrane-localized cell wall synthases Fks1/2 upon faster cell wall expansion, stabilizes mating projection growth without affecting cell shape. Experimental perturbation of the osmotic pressure and cell wall mechanics, as well as compromising the mechanical feedback through genetic deletion of the stress sensors, leads to cellular phenotypes that support the theoretical predictions. Our results indicate that while the existence of mechanical feedback is essential to stabilize mating projection growth, the shape and size of the cell are insensitive to the feedback.

The pathogenic fungus Candida albicans can undergo phenotypic switching between two heritable states: white and opaque. This phenotypic plasticity facilitates its colonization in distinct host niches. The master regulator WOR1 is exclusively expressed in opaque phase cells. Positive feedback regulation by Wor1 on the WOR1 promoter is essential for opaque formation, however the underlying mechanism of how Wor1 functions is not clear. Here, we use tandem affinity purification coupled with mass spectrometry to identify Wor1-interacting proteins. Tup1 and its associated complex proteins are found as the major factors associated with Wor1. Tup1 occupies the same regions of the WOR1 promoter as Wor1 preferentially in opaque cells. Loss of Tup1 is sufficient to induce the opaque phase, even in the absence of Wor1. This is the first such report of a bypass of Wor1 in opaque formation. These genetic analyses suggest that Tup1 is a key repressor of the opaque state, and Wor1 functions via alleviating Tup1 repression at the WOR1 promoter. Opaque cells convert to white en masse at 37°C. We show that this conversion occurs only in the presence of glycolytic carbon sources. The opaque state is stabilized when cells are cultured on non-glycolytic carbon sources, even in a MTLa/α background. We further show that temperature and carbon source affect opaque stability by altering the levels of Wor1 and Tup1 at the WOR1 promoter. We propose that Wor1 and Tup1 form the core regulatory circuit controlling the opaque transcriptional program. This model provides molecular insights on how C. albicans adapts to different host signals to undergo phenotypic switching for colonization in distinct host niches.

Lentinula edodes is a tetrapolar basidiomycete and its mating type is determined by two unlinked genetic loci, A and B. Theoretically, one dikaryotic strain could produce basidiospores with four different mating types in a 1:1:1:1 ratio. Previous studies have described the skewed segregation ratio of mating types among basidiospores of L. edodes. However, they were based only on morphological characteristics, such as clamp connection, to determine mating types. To clarify whether the segregation distortion of mating types is a general phenomenon in L. edodes, we analyzed the mating types of basidiospores obtained from three cultivars of L. edodes using recently developed DNA markers. We found that the skewed segregation of mating types was strain-specific, as reported previously. Among the three cultivars, one cultivar showed balanced segregation, while the other two displayed distorted segregation. We also examined the relationship between mating type and mycelial growth rate of monokaryons derived from each basidiospore. It was found that the monokaryotic mycelial growth rate was related to the A mating type but not to the B mating type. Therefore, homeodomain transcription factor genes that reside on the A locus or other genes linked to the A locus affect the growth rate of monokaryotic mycelia. Considering the importance of mating types in mushroom breeding, this study is informative for establishing an efficient breeding strategy as well as for understanding the mechanism of monokaryotic mycelial growth.

In recent years, Brown spot disease of rice (BSR) has been observed on leaves and seeds of rice in all rice-growing areas of Burkina Faso. Bipolaris oryzae and Exserohilum rostratum are the main fungal species isolated from BSR infected tissues and they are frequently observed in the same field. However, we are lacking information on the genetic diversity and population structure of these fungi in Burkina Faso. The mode of reproduction is also unknown. The genetic diversity of isolates of B. oryzae (n=61) and E. rostratum (n=151), collected from major rice-growing areas of Burkina Faso, was estimated using genotyping-by-sequencing (GBS). The mean values for nucleotide diversity (π) were 1.9 x10-4 for B. oryzae and 4.8 x10-4 for E. rostratum. There is no genetic differentiation between the geographical populations of each species. The analysis of molecular variance revealed that 89% and 94% of the genetic variances were within the populations of B. oryzae and E. rostratum, respectively. For each species, four genetic clusters were identified by two clustering methods (DAPC and sNMF). The distribution of these genetic groups was independent of the geographical origin of the isolates. Evidence of recombination was detected in the populations of B. oryzae and E. rostratum. For B. oryzae balanced mating type ratios were supporting sexual reproduction. For E. rostratum overrepresentation of MAT1-2 isolates (79%) suggested a predominant asexual reproduction. This study provides important information on the biology and genetics of the two major fungi causing brown spot disease of rice in Burkina Faso.

Keywords: gender studies; inclusive pedagogy; mating type; reproductive biology EN gender studies inclusive pedagogy mating type reproductive biology 1 3 3 07/19/23 20230701 NES 230701 Plant sex is complicated. Recognizing the value of distinguishing cultural "gender" from biological "sex" by avoiding the confusing phrase "plant gender" begs much larger questions about how other, more prevalent forms of gendered language limit the field. For instance, scholars interested in gender as a social and cultural construct adopted the phrase "sex assigned at birth" to refer to the effects of being raised with expectations linked to sex. [Extracted from the article]

Scab, caused by the plant-pathogenic fungus Venturia effusa, is a major disease of pecan in South America, resulting in loss of quantity and quality of nut yield. Characteristics of the populations of V. effusa in South America are unknown. We used microsatellites to describe the genetic diversity and population structure of V. effusa in South America, and determined the mating type status of the pathogen. The four hierarchically sampled orchard populations from Argentina (AR), Brazil (BRC and BRS), and Uruguay (UR) had moderate to high genotypic and gene diversity. There was evidence of population differentiation (Fst = 0.196) but the correlation between geographic distance and genetic distance was not statistically significant. Genetic differentiation was minimal between the UR, BRC, and BRS populations, and these populations were more clearly differentiated from the AR population. The MAT1-1 and MAT1-2 mating types occurred in all four orchards and their frequencies did not deviate from the 1:1 ratio expected under random mating; however, multilocus linkage equilibrium was rejected in three of the four populations. The population genetics of South American populations of V. effusa has many similarities to the population genetics of V. effusa previously described in the United States. Characterizing the populations genetics and reproductive systems of V. effusa are important to establish the evolutionary potential of the pathogen and, thus, its adaptability--and can provide a basis for informed approaches to utilizing available host resistance and determining phytosanitary needs. [ABSTRACT FROM AUTHOR]

In heterostylous, self-incompatible Turnera species, a member of the YUCCA gene family, YUC6, resides at the S-locus and has been hypothesized to determine the male mating type. YUCCA gene family members synthesize the auxin, indole-3-acetic acid, via a two-step process involving the TAA gene family. Consequently, it has been speculated that differences in auxin concentration in developing anthers are the biochemical basis underlying the male mating type. Here, we provide empirical evidence that supports this hypothesis. Using a transgenic knockdown approach, we show that YUC6 acts pleiotropically to control both the male physiological mating type and pollen size, but not the filament length dimorphism associated with heterostyly in Turnera. Using qPCR to assess YUC6 expression in different transgenic lines, we demonstrate that the level of YUC6 knockdown correlates with the degree of change observed in the male mating type. Further assessment of YUC6 expression through anther development, in the knockdown lines, suggests that the male mating type is irreversibly determined during a specific developmental window prior to microsporogenesis, which is consistent with the genetically sporophytic nature of this self-incompatibility system. These results represent the first gene controlling male mating type to be characterized in any species with heterostyly. [ABSTRACT FROM AUTHOR]

Long-term site-specific studies describing changes in the phenotypic variability of Phytophthora infestans populations allow quantitative predictions of pathogen spread and possible outbreaks of epidemics, and provide key input for regional resistance breeding programs. Late blight samples were collected from potato (Solanum tuberosum) breeding fields in Estonia during a twelve-year study period between 2001 and 2014. In total, 207 isolates were assessed for mating type and 235 isolates for metalaxyl resistance and 251 isolates for virulence factors. The frequency of mating types strongly fluctuated across the years, whereas the later period of 2010–2014 was dominated by the A2 mating. Despite fluctuations, both mating types were recorded in the same fields in most years, indicating sustained sexual reproduction of P. infestans with oospore production. Metalaxyl-resistant and intermediately resistant strains dominated in the first years of study, but with the progression of the study, metalaxyl-sensitive isolates became dominant, reaching up to 88%. Racial diversity, characterized by normalized Shannon diversity index decreased in time, varying from 1.00 in 2003 to 0.43 in 2013. The frequency of several virulence factors changed in a time-dependent manner, with R2 increasing and R6, R8, and R9 decreasing in time. Potato cultivar resistance background did not influence the frequency of P. infestans mating type, response to metalaxyl, and racial diversity. However, the diversity index decreased in time among isolates collected from resistant and susceptible cultivars, and remained at a high level in moderately resistant cultivars. These data demonstrate major time-dependent changes in racial diversity, fungicide resistance, and virulence factors in P. infestans, consistent with alterations in the control strategies and popularity of potato cultivars with different resistance. [ABSTRACT FROM AUTHOR]

The objective of this study was to investigate the role of the oomycete Phytophthora× cambivora in the decline affecting European beech (Fagus sylvatica) in the Nebrodi Regional Park (Sicily, southern Italy). In a survey of a beech forest stand in the heart of the park, Phytophthora× cambivora was the sole Phytophthora species recovered from the rhizosphere soil and fine roots of trees. Both A1 and A2 mating type isolates were found. Direct isolation from the stem bark of trees showing severe decline symptoms and bleeding stem cankers yielded exclusively P. gonapodyides, usually considered as an opportunistic pathogen. The mean inoculum density of P.× cambivora in the rhizosphere soil, as determined using the soil dilution plating method and expressed in terms of colony forming units (cfus) per gm of soil, the isolation frequency using leaf baiting, and the percentage of infected fibrous roots from 20 randomly selected beech trees with severe decline symptoms (50 to 100 foliage transparency classes) were 31.7 cfus, 80%, and 48.6%, respectively. These were significantly higher than the corresponding mean values of 20 asymptomatic or slightly declining trees, suggesting P.× cambivora is a major factor responsible for the decline in the surveyed stand. [ABSTRACT FROM AUTHOR]

Ascochyta fabae Speg. is a serious foliar fungal disease of faba bean and a constraint to production worldwide. This study investigated the phenotypic and genotypic diversity of the A. fabae pathogen population in southern Australia and the pathogenic variability of the population was examined on a differential set of faba bean cultivars. The host set was inoculated with 154 A. fabae isolates collected from 2015 to 2018 and a range of disease reactions from high to low aggressiveness was observed. Eighty percent of isolates collected from 2015 to 2018 were categorized as pathogenicity group (PG) PG- 2 (pathogenic on Farah) and were detected in every region in each year of collection. Four percent of isolates were non-pathogenic on Farah and designated as PG-1. A small group of isolates (16%) were pathogenic on the most resistant differential cultivars, PBA Samira or Nura, and these isolates were designated PG-3. Mating types of 311 isolates collected between 1991 and 2018 were determined and showed an equal ratio of MAT1--1 and MAT1--2 in the southern Australian population. The genetic diversity and population structure of 305 isolates were examined using DArTseq genotyping, and results suggest no association of genotype with any of the population descriptors viz.: collection year, region, host cultivar, mating type, or PG. A Genome-Wide Association Study (GWAS) was performed to assess genetic association with pathogenicity traits and a significant trait-associated genomic locus for disease in Farah AR and PBA Zahra, and PG was revealed. The high frequency of mating of A. fabae indicated by the wide distribution of the two mating types means changes to virulence genes would be quickly distributed to other genotypes. Continued monitoring of the A. fabae pathogen population through pathogenicity testing will be important to identify any increases in aggressiveness or emergence of novel PGs. GWAS and future genetic studies using biparental mating populations could be useful for identifying virulence genes responsible for the observed changes in pathogenicity. [ABSTRACT FROM AUTHOR]

Paramecium spp. are a genus of free-living protists that live mainly in freshwater environments. They are ciliates with high motility and phagocytosis and have been used to analyze cell motility and as a host model for pathogens. Besides such biological characteristics, apart from the usual morphological and genetic classification of species, the existence of taxonomies (such as syngens) and mating types related to Paramecium's unique reproduction is known. In this study, we attempted to develop a simple method to identify Paramecium strains, which are difficult to distinguish morphologically, using random amplified polymorphic DNA (RAPD) analysis. Consequently, we can observe strain-specific band patterns. We also confirm that the presence of endosymbiotic Chlorella cells affects the band pattern of P. bursaria. Furthermore, the results of the RAPD analysis using several P. caudatum strains with different syngens show that it is possible to detect a band specific to a certain syngen. By improving the reaction conditions and random primers, based on the results of this study, RAPD analysis can be applied to the identification of Paramecium strains and their syngen confirmation tests. [ABSTRACT FROM AUTHOR]

Powdery mildew is one of the most common and severe diseases of cucurbits, causing heavy yield losses in all growing areas when not successfully controlled. Two different fungal species, Podosphaera xanthii and Golovinomyces orontii, are generally recognized as causal agents. The results of monitoring carried out in 2016 and 2018 confirmed that P. xanthii is the exclusive pathogen causing cucurbit powdery mildew (CPM) in southern Italy. P. xanthii is a bipolar heterothallic fungus; a PCR‐based method for distinguishing MAT1‐1 and MAT1‐2 idiomorphs was applied for assessing mating type distribution in fungal populations present on cucurbits in different sites. The idiomorph MAT1‐2 was prevalent over the MAT1‐1 in 2016, whereas they were approximately in a 1:1 ratio in 2018; this finding corroborated the hypothesis that the MAT1‐1 idiomorph was more recently introduced in the area. Cyflufenamid‐resistant isolates were widespread in commercial greenhouses and field plantings even though use of this fungicide had been drastically reduced by the farmers 1 year before the monitoring due to the effectiveness losses observed in CPM control. Occurrence of cyflufenamid resistance and its impact on efficacy were evaluated in a field trial comparing different fungicide spray schedules. Cyflufenamid‐resistant isolates were detected even at the first appearance of symptoms on leaves, increasing over time. Isolates resistant to cyflufenamid showed a resistance factor as high as 900. Generally, P. xanthii was better controlled when cyflufenamid was used in integrated strategies rather than in spray schedules based on the exclusive use of the fungicide. [ABSTRACT FROM AUTHOR]

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ABSTRACT Transposable elements (TEs) play a pivotal role in shaping diversity in eukaryotic genomes. The covered smut pathogen on barley, Ustilago hordei, encountered a recent genome expansion. Using long reads, we assembled genomes of 6 U. hordei strains and 3 sister species, to study this genome expansion. We found that larger genome sizes can mainly be attributed to a higher genome fraction of long terminal repeat retrotransposons (LTR-RTs). In the studied smut genomes, LTR-RTs fractions are the largest in U. hordei and are positively correlated with the mating-type locus sizes, which is up to ~560 kb in U. hordei. Furthermore, LTR-RTs were found to be associated with higher nucleotide substitution levels, as these occur in specific genome regions of smut species with a recent LTR-RT proliferation. Moreover, genes in genome regions with higher nucleotide substitution levels generally reside closer to LTR-RTs than other genome regions. Genome regions with many nucleotide substitutions encountered an especially high fraction of CG substitutions, which is not observed for LTR-RT sequences. The high nucleotide substitution levels particularly accelerate the evolution of secretome genes, as their more accessory nature results in substitutions that often lead to amino acid alterations. IMPORTANCE Genomic alteration can be generated through various means, in which transposable elements (TEs) can play a pivotal role. Their mobility causes mutagenesis in itself and can disrupt the function of the sequences they insert into. They also impact genome evolution as their repetitive nature facilitates nonhomologous recombination. Furthermore, TEs have been linked to specific epigenetic genome organizations. We report a recent TE proliferation in the genome of the barley covered smut fungus, Ustilago hordei. This proliferation is associated with a distinct nucleotide substitution regime that has a higher rate and a higher fraction of CG substitutions. This different regime shapes the evolution of genes in subjected genome regions. We hypothesize that TEs may influence the error-rate of DNA polymerase in a hitherto unknown fashion.

Ascochyta fabae Speg. is a serious foliar fungal disease of faba bean and a constraint to production worldwide. This study investigated the phenotypic and genotypic diversity of the A. fabae pathogen population in southern Australia and the pathogenic variability of the population was examined on a differential set of faba bean cultivars. The host set was inoculated with 154 A. fabae isolates collected from 2015 to 2018 and a range of disease reactions from high to low aggressiveness was observed. Eighty percent of isolates collected from 2015 to 2018 were categorized as pathogenicity group (PG) PG-2 (pathogenic on Farah) and were detected in every region in each year of collection. Four percent of isolates were non-pathogenic on Farah and designated as PG-1. A small group of isolates (16%) were pathogenic on the most resistant differential cultivars, PBA Samira or Nura, and these isolates were designated PG-3. Mating types of 311 isolates collected between 1991 and 2018 were determined and showed an equal ratio of MAT1–1 and MAT1–2 in the southern Australian population. The genetic diversity and population structure of 305 isolates were examined using DArTseq genotyping, and results suggest no association of genotype with any of the population descriptors viz.: collection year, region, host cultivar, mating type, or PG. A Genome-Wide Association Study (GWAS) was performed to assess genetic association with pathogenicity traits and a significant trait-associated genomic locus for disease in Farah AR and PBA Zahra, and PG was revealed. The high frequency of mating of A. fabae indicated by the wide distribution of the two mating types means changes to virulence genes would be quickly distributed to other genotypes. Continued monitoring of the A. fabae pathogen population through pathogenicity testing will be important to identify any increases in aggressiveness or emergence of novel PGs. GWAS and future genetic studies using biparental mating populations could be useful for identifying virulence genes responsible for the observed changes in pathogenicity.

The genus Calonectria includes many aggressive plant pathogens causing diseases on various agricultural crops as well as forestry and ornamental tree species. Some species have been accidentally introduced into new environments via international trade of putatively asymptomatic plant germplasm or contaminated soil, resulting in significant economic losses. This review provides an overview of the taxonomy, population biology, and pathology of Calonectria species, specifically emerging from contemporary studies that have relied on DNA‐based technologies. The growing importance of genomics in future research is highlighted. A life cycle is proposed for Calonectria species, aimed at improving our ability to manage diseases caused by these pathogens. [ABSTRACT FROM AUTHOR]

Mating-type distribution within a phylogenetic tree, heterokaryon compatibility, and subsequent diploid formation were studied in 24 Aspergillus niger sensu stricto strains. The genomes of the 24 strains were sequenced and analyzed revealing an average of 6.162.0 variants/kb between Aspergillus niger sensu stricto strains. The genome sequences were used together with available genome data to generate a phylogenetic tree revealing 3 distinct clades within Aspergillus niger sensu stricto. The phylogenetic tree revealed that both MAT1-1 and MAT1-2 mating types were present in each of the 3 clades. The phylogenetic differences were used to select for strains to analyze heterokaryon compatibility. Conidial color markers (fwnA and brnA) and auxotrophic markers (pyrG and nicB) were introduced via CRISPR/Cas9-based genome editing in a selection of strains. Twenty-three parasexual crosses using 11 different strains were performed. Only a single parasexual cross between genetically highly similar strains resulted in a successful formation of heterokaryotic mycelium and subsequent diploid formation, indicating widespread heterokaryon incompatibility as well as multiple active heterokaryon incompatibility systems between Aspergillus niger sensu stricto strains. The 2 vegetatively compatible strains were of 2 different mating types and a stable diploid was isolated from this heterokaryon. Sclerotium formation was induced on agar media containing Triton X-100; however, the sclerotia remained sterile and no ascospores were observed. Nevertheless, this is the first report of a diploid Aspergillus niger sensu stricto strain with 2 different mating types, which offers the unique possibility to screen for conditions that might lead to ascospore formation in A. niger. [ABSTRACT FROM AUTHOR]

Aims: To investigate the genetic diversity, population structure and mating‐type distribution among the eco‐distinct isolates of Magnaporthe oryzae from Karnataka, India. Methods and Results: A set of 38 isolates of M. oryzae associated with leaf blast disease of rice were collected from different rice ecosystems of Karnataka, India, and analysed for their diversity at actin, β‐tubulin, calmodulin, translation elongation factor 1‐α (TEF‐1‐α), and internal transcribed spacer (ITS) genes/region. The isolates were grouped into two clusters based on the multilocus sequence diversity, the majority being in cluster‐IA (n = 37), and only one isolate formed cluster‐IB. Population structure was analysed using 123 SNP data to understand the genetic relationship. Based on K = 2 and ancestry threshold of >70%, blast strains were classified into two subgroups (SG1 and SG2) whereas, based on K = 4 and ancestry threshold of >70%, blast strains were classified into four subgroups (SG1, SG2, SG3 and SG4). We have identified 13 haplotype groups where haplotype group 2 was predominant (n = 20) in the population. The Tajima's and Fu's Fs neutrality tests exhibited many rare alleles. Further, the mating‐type analysis was also performed using MAT1 gene‐specific primers to find the potentiality of sexual reproduction in different ecosystems. The majority of the isolates (54.5%) had MAT1‐2 idiomorph, whereas 45.5% of the isolates possessed MAT1‐1 idiomorph. Conclusions: The present study found the genetically homogenous population of M. oryzae by multilocus sequence analysis. Both mating types, MAT1‐1 and MAT1‐2, were found within the M. oryzae population of Karnataka. Significance and impact of study: The study on the population structure and sexual mating behaviour of M. oryzae is important in developing region‐specific blast‐resistant rice cultivars. This is the first report of MAT1 idiomorphs distribution in the M. oryzae population in any Southern state of India. [ABSTRACT FROM AUTHOR]

How ploidy is determined in organisms is an important issue in bioscience. Polyploidy is believed to be relevant to useful traits of domesticated plants and microorganisms. As such, polyploidy is central to many applications in biotechnology. However, studies of polyploidy are poorly advanced because no methodologies to construct desired polyploid have been developed for any organism. Herein we describe the development of a novel breeding technology, mat α 2- PBT, to generate polyploid strains of Saccharomyces cerevisiae. S. cerevisiae has two mating types, a and α, determined by MAT a and MAT α gene each of which consists of a 1 and a 2 and α1 and α2 cistrons. This novel technology exploits an interesting feature of a specific mutation, mat α 2-102 , in the MAT α 2 gene. Unlike the MAT α wild-type strain, which gives a non-mating phenotype when mated with MAT a cells, the mat α 2-102 strain confers an α mating-type to a -type strains when mated with a -type strains. We constructed plasmid with the cloned mat α 2-102 mutant gene. An a- type cells harboring this plasmid displayed an α mating-type and mated with a- type cells. Because the resultant hybrid displays an α mating-type, it can mate again with a -type cells. By repeating this procedure, we have constructed an isogenic series of haploid to tetraploid of S. cerevisiae. Although whether even higher polyploid than tetraploid can be constructed by using this technology remains to be determined in the future, we believe that it became possible for the first time with mat α 2 -PBT method to investigate whether higher polyploid than tetraploid can be constructed. [ABSTRACT FROM AUTHOR]

There is limited understanding of the genetic variability in Phytophthora infestans in the major potato cultivation region of north-western Russia, where potato is grown primarily by small households with limited chemical treatment of late blight. In this study, the mating type, sensitivity to metalaxyl, and genotype and population genetic diversity (based on 12 simple sequence repeat (SSR) markers) of 238 isolates of P. infestans from the Pskov region during the years 2010–2013 were characterized. The aim was to examine the population structure, phenotypic and genotypic diversity, and the prevalent reproductive mode of P. infestans, as well as the influence of the location, time, and agricultural management practices on the pathogen population. The frequency of the A2 mating was stable over the four seasons and ranged from 33 to 48% of the sampled population. Both mating types occurred simultaneously in 90% of studied fields, suggesting the presence of sexual reproduction and oospore production in P. infestans in the Pskov region. Metalaxyl-sensitive isolates prevailed in all four years (72%), however, significantly fewer sensitive isolates were found in samples from large-scale conventional fields. A total of 50 alleles were detected in the 141 P. infestans isolates analyzed for genetic diversity. Amongst the 83 SSR multilocus genotypes (MLGs) detected, 65% were unique and the number of MLGs varied between locations from 3 to 20. These results, together with the high genotypic diversity observed in all the locations and the lack of significance of linkage disequilibrium, suggest that sexual recombination is likely responsible for the unique MLGs and the high genetic diversity found in the Pskov region population, resembling those of north-eastern European populations. [ABSTRACT FROM AUTHOR]

Divergence of breeding system plays an important role in fungal speciation. Ectomycorrhizal fungi, however, pose a challenge for the study of reproductive biology because most cannot be mated under laboratory conditions. To overcome this barrier, we sequenced the draft genomes of the ectomycorrhizal sister species Rhizopogon vinicolor Smith and Zeller and R. vesiculosus Smith and Zeller (Basidiomycota, Boletales)-the first genomes available for Basidiomycota truffles-and characterized gene content and organization surrounding their mating type loci. Both species possess a pair of homeodomain transcription factor homologs at the mating type A-locus as well as pheromone receptor and pheromone precursor homologs at the mating type B-locus. Comparison of Rhizopogon genomes with genomes from Boletales, Agaricales, and Polyporales revealed synteny of the A-locus region within Boletales, but several genomic rearrangements across orders. Our findings suggest correlation between gene content at the B-locus region and breeding system in Boletales with tetrapolar species possessing more diverse gene content than bipolar species. Rhizopogon vinicolor possesses a greater number of B-locus pheromone receptor and precursor genes than R. vesiculosus, as well as a pair of isoprenyl cysteine methyltransferase genes flanking the B-locus compared to a single copy in R. vesiculosus Examination of dikaryotic single nucleotide polymorphisms within genomes revealed greater heterozygosity in R. vinicolor, consistent with increased rates of outcrossing. Both species possess the components of a heterothallic breeding system with R. vinicolor possessing a B-locus region structure consistent with tetrapolar Boletales and R. vesiculosus possessing a B-locus region structure intermediate between bipolar and tetrapolar Boletales.

Smut fungi are globally distributed plant pathogens that infect agriculturally important crop plants such as maize or potato. To date, molecular studies on plant responses to smut fungi are challenging due to the genetic complexity of their host plants. Therefore, we set out to investigate the known smut fungus of Brassicaceae hosts, Thecaphora thlaspeos. T. thlaspeos infects different Brassicaceae plant species throughout Europe, including the perennial model plant Arabis alpina. In contrast to characterized smut fungi, mature and dry T. thlaspeos teliospores germinated only in the presence of a plant signal. An infectious filament emerges from the teliospore, which can proliferate as haploid filamentous cultures. Haploid filaments from opposite mating types mate, similar to sporidia of the model smut fungus Ustilago maydis. Consistently, the a and b mating locus genes are conserved. Infectious filaments can penetrate roots and aerial tissues of host plants, causing systemic colonization along the vasculature. Notably, we could show that T. thlaspeos also infects Arabidopsis thaliana. Exploiting the genetic resources of A. thaliana and Arabis alpina will allow us to characterize plant responses to smut infection in a comparative manner and, thereby, characterize factors for endophytic growth as well as smut fungi virulence in dicot plants.

Due to their economic relevance, the study of plant pathogen interactions is of importance. However, elucidating these interactions and their underlying molecular mechanisms remains challenging since both host and pathogen need to be fully genetically accessible organisms. Here we present milestones in the establishment of a new biotrophic model pathosystem: Ustilago bromivora and Brachypodium sp. We provide a complete toolset, including an annotated fungal genome and methods for genetic manipulation of the fungus and its host plant. This toolset will enable researchers to easily study biotrophic interactions at the molecular level on both the pathogen and the host side. Moreover, our research on the fungal life cycle revealed a mating type bias phenomenon. U. bromivora harbors a haplo-lethal allele that is linked to one mating type region. As a result, the identified mating type bias strongly promotes inbreeding, which we consider to be a potential speciation driver.

Saccharomycopsis species are natural organic sulphur auxotrophs. Their genomes do not encode genes for the uptake and assimilation of sulphate and thus these species cannot grow on media lacking e.g. methionine. Due to the similarity between sulphate and selenate, uptake and assimilation of selenate occurs through the same pathway starting from sulphate transporters encoded by the homologs of the SUL1 and SUL2 genes in S. cerevisiae. Lack of these transporters renders Saccharomycopsis species resistant to selenate levels that are toxic to other microorganisms. We used this feature to enrich environmental samples for Saccharomycopsis species. This led to the isolation of S. schoenii , S. lassenensis and a hitherto undescribed Saccharomycopsis species with limited by-catch of other yeasts, mainly belonging to Metschnikowia and Hanseniaspora. We performed growth and predation assays to characterize the potential of these new isolates as predacious yeasts. Most Saccharomycopsis species are temperature sensitive and cannot grow at 37°C; with the exception of S. lassenensis strains. Predation assays with S. schoenii and S. cerevisiae as prey indicated that predation was enhanced at 20°C compared to 30°C. We crossed an American isolate of S. schoenii with our German isolate using marker directed breeding. Viable progeny indicated that both strains are interfertile and belong to the same biological species. S. lassenensis is heterothallic, while S. schoenii and the new Saccharomycopsis isolate, for which we suggest the name S. geisenheimensis sp. nov., are homothallic. [ABSTRACT FROM AUTHOR]

Phytophthora colocasiae is an important pathogen that causes great economic losses in taro production in tropical and subtropical regions, especially in Japan. Understanding the genetic variations in P. colocasiae populations and their transmission patterns in Japan is essential for effective disease control. Here, the genetic diversity of 358 P. colocasiae isolates, including 348 from Japan, 7 from China, and 3 from Indonesia, was assessed using 11 simple sequence repeat (SSR) primer pairs with high polymorphism. The phylogenetic tree of the SSR locus showed that the isolates from Japan could be divided into 14 groups, with group A being the dominant group. Among foreign isolates, only six from mainland China were similar to those from Japan and clustered in groups B and E. Analysis of molecular variance (AMOVA), principal components analysis (PCA), and cluster analysis (K = 3) results revealed a moderate level of genetic diversity, mainly within individuals. Populations showed high heterozygosity, a lack of regional differentiation, and frequent gene flow. Analysis of mating types and ploidy levels revealed that A2 and self-fertile (SF) A2 types and tetraploids were dominant across populations. Explanations and hypotheses for the results can provide more effective strategies for disease management of taro leaf blight.

Phytophthora infestans is the oomycete responsible for late blight disease of Solanaceae that causes both yield and economic losses. With the aim of reducing plant wilt and high management costs mainly due to wide fungicide applications, alternative eco-sustainable control strategies are needed. RNA interference (RNAi) is a powerful tool for gene function studies that can be accomplished by constitutive transformation or transient expression such as virus-induced gene silencing (VIGS) experiments. VIGS makes use of viruses to deliver sequences homologous to a target gene fragment and trigger RNAi. Indeed, a P. infestans ortholog of plant loricrin-like protein (LLP), named PiLLP, has been silenced using the direct infection of a recombinant vector based on the plant virus tobacco mosaic virus (TMV-PiLLP-1056), aiming to reduce the oomycete sexual reproduction. For this purpose, the gene coding for the green fluorescent protein (GFP) present in the TMV-GFP-1056 vector has been replaced with an antisense construct obtained by fusion PCR of the PiLLP 5′-UTR and 3′-UTR sequences. Here, we show that RNAi can be expressed in the A1 mating type of P. infestans strain 96.9.5.1 by VIGS using the direct infection of TMV-PiLLP-1056. We provide evidence that the recombinant vector can enter, replicate, and persist in mycelia of P. infestans where it induces the partial downregulation of the PiLLP transcript. Compared with the wild-type, the PiLLP-silenced A1 mating type had slower colony growth and a diminished virulence in detached tomato leaflets. This seems to be the first evidence of a constitutive gene downregulation of P. infestans using a recombinant vector based on a plus-sense RNA plant virus.

Potato (Solanum tuberosum L.) and tomato (S. lycopersicum L.) are the most economically important vegetable crops in Egypt and worldwide. The winter crop in Egypt is particularly prone to late blight caused by Phytophthora infestans. A total of 152 P. infestans isolates were isolated from the 2013, 2014, 2016 and 2018 winter crops with 82 isolates from potato, 69 from tomato and one isolate from eggplant (S. melongena L.). All isolates belonged to the A1 mating type with no evidence of A2 or self-fertile strains. The majority of isolates (53%) were sensitive to metalaxyl, 32% were intermediate and 15% were resistant. Variation in aggressiveness between three P. infestans isolates EG-005 (13_A2) and EG-276 (23_A1) from potato, and EG-237 (23_A1) from eggplant was determined on tuber slices and leaflets of 10 potato cultivars. The eggplant isolate EG-237 showed higher sporulation capacity compared with the other tested isolates and was able to infect potato (Lady Rosetta cv) and tomato (Super Strain B cv). The simple sequence repeat (SSR) genotyping data showed that in contrast to our previous work (3-year period 2010–12) in which the proportion of 13_A2 lineage was 35%, all isolates belonged to the 23_A1 lineage. There was no evidence for the existence of the A2 mating type or 13_A2 lineage even in the destroyed field crops of some cultivars (Cara, Bellini and Valor) that had been reported as resistant to 23_A1. The data have been submitted into the Euroblight database to allow temporal and spatial genetic diversity to be examined in comparison with other regional P. infestans populations. The AVR2 and AVR2-like RXLR effector genes were amplified and sequenced. In the avirulent AVR2 gene, only one heterozygous SNP was detected at position 31 in the N terminus in six isolates out of eleven, whereas two heterozygous SNPs were detected at position 29 in the N-terminus and ninety-two in the C- terminus of the AVR2-like gene. This suggests that changes in the previously reported virulence profile of 23_A1 are not related to commercial cultivars carrying the R2 gene. In addition, this is the first report of P. infestans on eggplant in Egypt.