BACKGROUNDSclerotium rolfsii is an extremely destructive phytopathogenic fungus that causes significant economic losses. Biocontrol strategies utilizing antagonistic microorganisms present a promising alternative for controlling plant pathogens. Bacillus megaterium L2 has been identified as a potential microbial biocontrol agent in our previous study; however, its efficacy in controlling pathogens has yet to meet current demands. This study aims to enhance the antifungal activity of strain L2 against S. rolfsii R‐67 through a two‐round mutagenesis strategy and to preliminarily investigate the mutagenesis mechanism of the high antifungal activity mutant.RESULTSWe obtained mutant Dr‐77 with the strongest antifungal activity against R‐67, and its cell‐free supernatant significantly reduced the infection potential of R‐67 to Amorphophallus konjac corms, which may be attributed to the antimicrobial compound phenylacetic acid (PAA), and PAA content in Dr‐77 (5.78 mg/mL) was 28.90 times higher than original strain L2. This compound exhibited strong antifungal ability against R‐67, with a half maximal effective concentration (EC50) value of 0.475 mg/mL, significantly inhibiting mycelial growth and destroying the ultrastructure of R‐67 at EC50 value. Notably, PAA also exhibited broad‐spectrum antifungal activity against six phytopathogens at EC50 value. Moreover, genome analysis revealed nine different gene mutations, including those involved in PAA biosynthesis, and the activities of prephenate dehydratase (PheA) and phenylacetaldehyde dehydrogenase (ALDH) in PAA biosynthesis pathway were significantly increased.CONCLUSIONThese results suggest that the elevated PAA content is a primary factor contributing to the enhanced antifungal activity of Dr‐77, and that this mutagenesis strategy offers valuable guidance for the breeding of functional microbial resources. © 2024 Society of Chemical Industry.

中文翻译：

---

常温和室温等离子体与核糖体工程技术相结合，以增强 Bacillus megateriumL2 对 Sclerotium rolfsii 的抗真菌活性


背景Clerotium rolfsii 是一种极具破坏性的植物病原真菌，会造成重大经济损失。利用拮抗微生物的生物防治策略为控制植物病原体提供了一种很有前途的替代方案。在我们之前的研究中，Bacillus megaterium L2 已被确定为一种潜在的微生物生物防治剂;然而，它在控制病原体方面的功效尚未满足当前的需求。本研究旨在通过两轮诱变策略增强菌株 L2 对 S. rolfsii R-67 的抗真菌活性，并初步研究高抗真菌活性突变体的诱变机制。结果我们获得了对 R-67 具有最强抗真菌活性的突变体 Dr-77，其无细胞上清液显著降低了 R-67 对魔芋球茎的感染可能性，这可能归因于抗菌化合物苯乙酸 （PAA），Dr-77 中 PAA 含量 （5.78 mg/mL） 是原始菌株 L2 的 28.90 倍。该化合物对 R-67 表现出较强的抗真菌能力，半数最大有效浓度 （EC50） 值为 0.475 mg/mL，在 EC50 值时显着抑制菌丝生长并破坏 R-67 的超微结构。值得注意的是，PAA 在 EC50 值下还表现出对 6 种植物病原体的广谱抗真菌活性。此外，基因组分析揭示了 9 种不同的基因突变，包括参与 PAA 生物合成的基因突变，并且 PAA 生物合成途径中 prephenate dehydratase （PheA） 和苯乙醛脱氢酶 （ALDH） 的活性显著增加。结论结果表明，PAA 含量升高是导致 Dr-77 抗真菌活性增强的主要因素，这种诱变策略为功能性微生物资源的培育提供了有价值的指导。© 2024 化工学会.