Maintenance of Sex-Related Genes and the Co-Occurrence of Both Mating Types in Verticillium dahliae

Verticillium dahliae is a cosmopolitan, soilborne fungus that causes a significant wilt disease on a wide variety of plant hosts including economically important crops, ornamentals, and timber species. Clonal expansion through asexual reproduction plays a vital role in recurring plant epidemics caused by this pathogen. The recent discovery of recombination between clonal lineages and preliminary investigations of the meiotic gene inventory of V. dahliae suggest that cryptic sex appears to be rare in this species. Here we expanded on previous findings on the sexual nature of V. dahliae. Only 1% of isolates in a global collection of 1120 phytopathogenic V. dahliae isolates contained the MAT1-1 idiomorph, whereas 99% contained MAT1-2. Nine unique multilocus microsatellite types comprised isolates of both mating types, eight of which were collected from the same substrate at the same time. Orthologs of 88 previously characterized sex-related genes from fungal model systems in the Ascoymycota were identified in the genome of V. dahliae, out of 93 genes investigated. Results of RT-PCR experiments using both mating types revealed that 10 arbitrarily chosen sex-related genes, including MAT1-1-1 and MAT1-2-1, were constitutively expressed in V. dahliae cultures grown under laboratory conditions. Ratios of non-synonymous (amino-acid altering) to synonymous (silent) substitutions in V. dahliae MAT1-1-1 and MAT1-2-1 sequences were indistinguishable from the ratios observed in the MAT genes of sexual fungi in the Pezizomycotina. Patterns consistent with strong purifying selection were also observed in 18 other arbitrarily chosen V. dahliae sex-related genes, relative to the patterns in orthologs from fungi with known sexual stages. This study builds upon recent findings from other laboratories and mounts further evidence for an ancestral or cryptic sexual stage in V. dahliae.大丽轮枝菌（Verticillium dahliae）是一种广布性土传真菌，可侵染多种植物寄主引发严重萎蔫病，寄主涵盖具有经济价值的作物、观赏植物及林木树种。无性繁殖介导的克隆扩增在该病原菌引发的周期性植物病害流行中发挥关键作用。近期研究发现克隆谱系间存在重组现象，且对大丽轮枝菌减数分裂基因库的初步分析提示，该物种中隐性有性生殖似乎较为罕见。本研究拓展了此前关于大丽轮枝菌有性生殖特性的研究结论。在全球收集的1120株植物病原性大丽轮枝菌菌株中，仅1%的菌株携带MAT1-1交配型基因盒（idiomorph），剩余99%均携带MAT1-2交配型基因盒。共鉴定出9种独特的多位点微卫星类型，其包含的菌株同时具备两种交配型，其中8种菌株采自同一时间的同一基质。在所检测的93个已在子囊菌门（Ascomycota）真菌模型系统中被表征的有性生殖相关基因中，大丽轮枝菌基因组内成功鉴定出88个对应的直系同源基因（orthologs）。针对两种交配型菌株的逆转录聚合酶链式反应（Reverse Transcription PCR, RT-PCR）实验结果显示，随机选取的10个有性生殖相关基因（包括MAT1-1-1与MAT1-2-1）在实验室培养条件下的大丽轮枝菌中均呈组成型表达。大丽轮枝菌MAT1-1-1与MAT1-2-1序列的非同义（氨基酸改变型）替换与同义（沉默型）替换比值，与盘菌亚门（Pezizomycotina）有性真菌的MAT基因比值无显著差异。相较于已知有性阶段真菌的直系同源基因，另外随机选取的18个大丽轮枝菌有性生殖相关基因也呈现出强烈纯化选择的特征模式。本研究基于其他实验室近期的研究成果，进一步为大丽轮枝菌存在祖先型或隐性有性生殖阶段提供了证据。