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香蕉细菌性软腐病菌tus基因对细菌生物被膜形成的影响

引用本文：蒋尚伯,孙大元,唐子晴,沈会芳,蒲小明,林壁润,杨祁云,张景欣.香蕉细菌性软腐病菌tus基因对细菌生物被膜形成的影响.植物保护学报,2023,50(5):1161-1175

DOI：10.13802/j.cnki.zwbhxb.2023.2023807

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作者	单位	E-mail
蒋尚伯	广东省农业科学院植物保护研究所, 广东省植物保护新技术重点实验室, 广州 510640
孙大元	广东省农业科学院植物保护研究所, 广东省植物保护新技术重点实验室, 广州 510640
唐子晴	广东省农业科学院植物保护研究所, 广东省植物保护新技术重点实验室, 广州 510640
沈会芳	广东省农业科学院植物保护研究所, 广东省植物保护新技术重点实验室, 广州 510640
蒲小明	广东省农业科学院植物保护研究所, 广东省植物保护新技术重点实验室, 广州 510640
林壁润	广东省农业科学院植物保护研究所, 广东省植物保护新技术重点实验室, 广州 510640
杨祁云	广东省农业科学院植物保护研究所, 广东省植物保护新技术重点实验室, 广州 510640	yangqiyun@gdaas.cn
张景欣	广东省农业科学院植物保护研究所, 广东省植物保护新技术重点实验室, 广州 510640	chougu@126.com

中文摘要:为解析硫转运蛋白（tRNA 2-thiouridine synthesizing protein，tus）基因对香蕉细菌性软腐病病原细菌——玉米迪基氏菌Dickeya zeae生物被膜形成的影响，采用同源重组方法构建6株tus基因的缺失突变体及回补菌株，分析代表性突变体的转录组差异，并测定不同突变体的运动能力、细胞壁降解酶分泌水平以及对烟草的致敏性和对寄主的毒力，解析突变体生物被膜表型发生变异的原因。结果显示，玉米迪基氏菌的6株tus基因缺失突变体ΔtusA、ΔtusB、ΔtusC、ΔtusD、ΔtusE和ΔmnmA均表现出生物被膜表型缺陷；ΔtusC突变体转录组比较分析表明，细菌生物被膜形成和运动性相关基因均属于细胞进程中的显著下调表达基因，且前者主要为VI型分泌系统（type VI secretion system，T6SS）基因，进一步分析发现3个位点的T6SS基因均出现显著下调表达趋势；6株tus基因缺失突变体的运动能力显著减弱，但是细胞壁降解酶分泌水平和对烟草的致敏性并未发生显著变化；接种野生型菌株和基因回补菌株后香蕉的病情指数为42.5~67.5，而接种突变体后香蕉的病情指数下降至5.0~17.5，突变体的毒力下降。表明tus基因在玉米迪基氏菌生物被膜形成过程中具有重要功能。

中文关键词:香蕉细菌性软腐病菌 tus基因转录组测序生物被膜运动性

Functional analysis of tus genes that affect the biofilm formation of banana soft rot pathogen Dickeya zeae

Author Name	Affiliation	E-mail
Jiang Shangbo	Guangdong Provincial Key Laboratory of High Technology for Plant Protection, Plant Protection Research Institute, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, Guangdong Province, China
Sun Dayuan	Guangdong Provincial Key Laboratory of High Technology for Plant Protection, Plant Protection Research Institute, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, Guangdong Province, China
Tang Ziqing	Guangdong Provincial Key Laboratory of High Technology for Plant Protection, Plant Protection Research Institute, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, Guangdong Province, China
Shen Huifang	Guangdong Provincial Key Laboratory of High Technology for Plant Protection, Plant Protection Research Institute, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, Guangdong Province, China
Pu Xiaoming	Guangdong Provincial Key Laboratory of High Technology for Plant Protection, Plant Protection Research Institute, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, Guangdong Province, China
Lin Birun	Guangdong Provincial Key Laboratory of High Technology for Plant Protection, Plant Protection Research Institute, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, Guangdong Province, China
Yang Qiyun	Guangdong Provincial Key Laboratory of High Technology for Plant Protection, Plant Protection Research Institute, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, Guangdong Province, China	yangqiyun@gdaas.cn
Zhang Jingxin	Guangdong Provincial Key Laboratory of High Technology for Plant Protection, Plant Protection Research Institute, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, Guangdong Province, China	chougu@126.com

Abstract:To reveal the effects of tRNA 2-thiouridine-synthesizing protein genes, tus, on biofilm formation of banana soft rot pathogen Dickeya zeae, the gene deletion mutants and their complementary strains were constructed for all of six tus genes by homologous recombination, and were used for exploring the mechanism of their biofilm phenotypic change. In combination with comparative transcriptome analysis of representative gene deletion mutant, six deletion mutants were further tested for bacterial motility, secretion of cell wall degradation enzymes, hypersensitive reaction in tobacco, and virulence on banana host. Six gene deletion mutants, ΔtusA, ΔtusB, ΔtusC, ΔtusD, ΔtusE and ΔmnmA, all showed obvious phenotypic defects on bacterial biofilm. Comparative transcriptome analysis between ΔtusC mutant and wild-type revealed that those genes related to bacterial biofilm formation and motility mainly consisted of the significantly down-regulated gene group within cellular process, while bacterial biofilm formation genes were mainly the type VI secretory system (T6SS) genes. Further analysis found that T6SS genes at three different genome loci were significantly down regulated. For the bacterial motility, these six gene mutants were significantly weakened at their bacterial motility, but there were no obvious changes in cell wall-degrading enzymes secretion and tobacco hypersensitivity. The disease indexes of banana seedlings inoculated with wild-type and gene complementary strains were 42.5-67.5, while those inoculated with six tus gene mutants decreased to 5.0-17.5. Thus, the tus genes play important roles in the formation of bacterial biofilm of this D. zeae pathogen.

keywords:banana bacterial soft rot pathogen tus gene RNA-seq biofilm motility

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