The adaptation of Fusarium culmorum to DMI fungicides is mediated by major transcriptome modifications in response to azole

Fusarium culmorum is a fungal pathogen that causes economically important diseases on a variety of crops. Fungicides can be applied to control this species with triazoles being the most efficient molecules. F. culmorum strains resistant to these molecules have been reported, but the underlying resistance mechanisms remain unknown. In this study, a tebuconazole-adapted F. culmorum strain was developed from the sensitive strain UK99 which had a level of fitness similar to its parental strain. The adapted strain showed cross-resistance to all demethylation inhibitors (DMIs), but not to other classes of fungicides tested. RNA-Seq analysis revealed that 4900 out of the 12,000 annotated genes of F. culmorum's genome had significantly different level of expression (q < 0.05) between the resistant strain and its parental strain after tebuconazole treatment. Among these changes, FcABC1 (FCUL_06717), a pleiotropic drug resistance transporter, had a 30-fold higher expression level upon tebuconazole treatment in the adapted strains as compared to the wild-type strain. The implication of this transporter in triazole resistance was subsequently confirmed in field strains harboring distinct levels of sensitivity to triazoles. FcABC1 is present in other species/genera, including F. graminearum in which it is known to be necessary for azole resistance. No difference in FcABC1 sequences, including the surrounding regions, were found when comparing the resistant strain to the wild-type strain. Fusarium culmorum is therefore capable to adapt to triazole pressure by overexpressing a drug resistance transporter when submitted to triazoles and the same mechanism is anticipated to occur in other species.

秆镰孢菌（Fusarium culmorum）是一类可在多种作物上引发具有重要经济影响病害的真菌病原菌。生产中可通过施用杀菌剂对其进行防控，其中三唑类（triazoles）药剂为目前防控该病原菌最有效的活性成分。已有研究报道了对三唑类药剂产生抗性的秆镰孢菌菌株，但此类抗性的内在分子机制至今仍未明确。本研究以对三唑类敏感的秆镰孢菌菌株UK99为亲本，通过驯化获得了一株适应戊唑醇（tebuconazole）的抗性菌株，该驯化菌株的适合度水平与亲本菌株基本一致。该抗性菌株对所有脱甲基化抑制剂（DMIs）均表现出交互抗性，但对本次试验中测试的其他类别杀菌剂无交互抗性。通过RNA测序（RNA-Seq）分析发现，在经戊唑醇处理后，抗性菌株与亲本菌株的12000个注释基因中，共有4900个基因的表达水平存在显著差异（q < 0.05）。其中，多效耐药转运蛋白（pleiotropic drug resistance transporter）编码基因FcABC1（FCUL_06717）在抗性菌株经戊唑醇处理后的表达量较野生型亲本菌株上调了30倍。后续通过对不同三唑类敏感性水平的田间菌株进行功能验证，确认了该转运蛋白在三唑类抗性形成中的关键作用。FcABC1同源基因广泛存在于其他物种类群中，包括禾谷镰孢菌（F. graminearum），该菌中FcABC1的同源基因已被证实与唑类杀菌剂抗性相关。对抗性菌株与野生型菌株的FcABC1基因及其侧翼序列进行比对后，未发现二者存在序列差异。综上，秆镰孢菌可通过上调药物抗性转运蛋白的表达水平，以适应三唑类药剂的选择压力，且该抗性机制预计在其他相关物种中同样存在。