Table_1_Enlightening Gliotoxin Biological System in Agriculturally Relevant Trichoderma spp..xlsx

Gliotoxin (GT) is a dual fungal secondary metabolite (SM). It displays pleiotropic activities and possesses medicinal properties and biocontrol abilities but, unfortunately, has toxic properties in humans. Various Trichoderma species are used as fungal biological control agents (BCAs), as a sustainable alternative for crop protection worldwide. Among them is Trichoderma virens, a GT-producing fungus. Since no information was available on the genetically coded prerequisites for the production of GT in other Trichoderma spp., genome analyses were carried out in 10 Trichoderma spp. genomes. Moreover, a real-time PCR assay setup ad hoc and high-performance liquid chromatography (HPLC) analyses were employed to understand the GT-producing biological systems in T. virens GV29-8 (TvGv29-8) and Trichoderma afroharzianum T6776 (TaT6776), two relevant biocontrol fungi. The structure of the GT biosynthesis genes (GT-BG) is polymorphic, with two distinct types associated with the ability to produce GT. GliH, a key protein for GT synthesis, is absent in most of the Trichoderma GT biosynthetic pathways, which may be the reason for their inability to produce GT. The GT-BG are expressed in TvGv29-8 as expected, while they are silent in TaT6776. Interestingly, in the GT-non-producing TaT6776, only gliA (putative GT transporter) and gtmA (putative GT S-methyltransferase) were induced by exogenous GT, underlining the ability of this strain to reduce the deleterious effect of the toxin. This ability is confirmed by growth assays and by the detection of the bis-thiomethylated form of GT catalyzed by GtmA in the culture medium supplemented with GT. To the best of our knowledge, this is the first general description of the GT biological system in different Trichoderma spp. as far as the GT-BG content and organization is concerned and a preliminary insight into their functionality.

胶质霉素（Gliotoxin, GT）是一类双功能真菌次级代谢产物（secondary metabolite, SM），兼具多效活性，拥有药用价值与生防潜力，但同时对人体具有毒性。木霉菌属（Trichoderma）多个物种已被开发为真菌生防制剂（biological control agents, BCAs），是全球范围内作物保护的可持续替代方案。其中绿色木霉（Trichoderma virens）是一类可产GT的真菌。由于此前尚无关于其他木霉菌属物种合成GT的遗传编码先决条件的相关研究，本研究对10个木霉菌属物种的基因组展开了分析。此外，本研究搭建了定制化实时荧光定量PCR（real-time PCR）检测体系，并结合高效液相色谱（high-performance liquid chromatography, HPLC）分析，以探究两株典型生防真菌——绿色木霉GV29-8（TvGv29-8）与阿哈拉姆木霉T6776（Trichoderma afroharzianum T6776, TaT6776）的GT合成生物学系统。GT生物合成基因（GT biosynthesis genes, GT-BG）的结构具有多态性，存在两种与GT合成能力相关的不同类型。GT合成的关键蛋白GliH在多数木霉菌属的GT生物合成通路中缺失，这或许是多数木霉菌无法合成GT的原因。TvGv29-8中GT-BG的表达模式符合预期，而TaT6776中此类基因则处于沉默状态。有趣的是，在无法合成GT的TaT6776中，仅假定的GT转运蛋白编码基因gliA与假定的GT S-甲基转移酶编码基因gtmA可被外源GT诱导，这表明该菌株具备缓解该毒素毒害作用的能力。这一结论通过生长试验与在添加了GT的培养基中检测到GtmA催化生成的GT双硫甲基化产物得到了验证。据我们所知，本研究首次针对不同木霉菌属物种的GT生物合成基因的组成与组织形式展开了系统性描述，并对其功能进行了初步探究。