贝莱斯芽孢杆菌LF01基因组序列分析及其代谢产物的生防作用  

Genomic analysis of Bacillus velezensis LF01 strain and the biocontrol effect of its secondary metabolites

作　　者：张德锋[1,2] 高艳侠 可小丽 王亚军[1,2] 任燕 石存斌[1,2] ZHANG Defeng;GAO Yanxia;KE Xiaoli;WANG Yajun;REN Yan;SHI Cunbin(Key Laboratory of Fishery Drug Development,Ministry of Agriculture and Rural Affairs,Pearl River Fisheries Research Institute,Chinese Academy of Fishery Sciences,Guangzhou 510380,China;Key Laboratory of Aquatic Animal Immune Technology,Guangdong Province,Pearl River Fisheries Research Institute,Chinese Academy of Fishery Sciences,Guangzhou 510380,China)

机构地区：[1]中国水产科学研究院珠江水产研究所,农业农村部渔用药物创制重点实验室,广东广州510380 [2]中国水产科学研究院珠江水产研究所,广东省水产动物免疫技术重点实验室,广东广州510380

出　　处：《水产学报》2022年第2期196-206,共11页Journal of Fisheries of China

基　　金：国家重点研发计划(2019YFD0900102);国家现代农业产业技术体系(CARS-45);中国水产科学研究院中央级公益性科研院所基本科研业务费专项(2019XN-003);广东省促进经济发展专项(现代渔业发展用途)(粤农2019B8)。

摘　　要：为充分挖掘贝莱斯芽孢杆菌LF01菌株拮抗物质合成相关基因簇,实验对其进行了全基因组测序与分析,同时评价了LF01菌株拮抗物质的生物安全性及其生防作用效果。基于三代Nanopore测序平台对LF01菌株进行全基因组测序分析,并利用在线平均核苷酸同源性(ANI)和DNA-DNA杂交(DDH)分析,以及基因组遗传进化确定该菌株的分类学地位;通过antiSMASH在线分析LF01基因组中的拮抗物质合成基因簇。同时,基于CAZyme数据库分析LF01菌株对糖类物质的水解能力。LF01菌株拮抗物质通过腹腔注射的方式,检测其对尼罗罗非鱼和斑马鱼的致病性;拮抗物质饲喂尼罗罗非鱼后评价其生防作用效果。结果显示,LF01菌株基因组全长为3974023 bp,GC含量为46.56%,包含3843个编码基因。LF01菌株与贝莱斯芽孢杆菌的ANI和DDH分别≥97.66%和≥80.10%,同时基于SNP进化分析显示,该菌与贝莱斯芽孢杆菌聚为一支,表明LF01菌株在基因组分子水平上被鉴定为贝莱斯芽孢杆菌。LF01菌株包含了bacillaene、bacillibactin、杆菌霉素D(bacillomycin D)、溶杆菌素(bacilysin)、difficidin、泛革素(fengycin)、macrolactin H、plantazolicin和表面活性素(surfactin)等9个拮抗物质基因簇,约占基因组全长的8.83%。LF01菌株基因组中含有许多与纤维素、半纤维素、淀粉、几丁质、果胶、肽聚糖和葡聚糖降解有关的CAZy酶基因家族。LF01的拮抗物质对尼罗罗非鱼和斑马鱼具有较高的安全性,而且饲喂尼罗罗非鱼后能够显著提高其对无乳链球菌的抗病力。综上可知,LF01菌株含有大量拮抗物质合成基因簇,其拮抗物质具有较高的生物安全性,并且饲喂后能够显著提高尼罗罗非鱼的抗病力,表明LF01菌株拮抗物质在水产动物细菌性疾病的防控方面具有良好的开发应用前景。Bacillus velezensis is a novel species in the Bacillus genus which exhibits a broad spectrum of antimicrobial activities because it produces various secondary metabolites.The current research on B.velezensis mainly focuses on its use for the promotion of the growth of animals and plants,for antagonizing pathogens,and on investigating its gene cluster which plays significant roles in biological control,drug research and development,and food fermentation.The aims of this study are to explore the gene clusters related to antagonistic substances of B.velezensis LF01 strain based on the whole genome sequencing,and to evaluate the biological safety and biocontrol effect of the antagonistic substances of LF01 strain.In this study,the whole genome of LF01 strain was sequenced based on the third generation Nanopore sequencing platform,and its taxonomic status was identified based on ANI and DDH online analysis,and the genetic evolution at genome level.The gene clusters related to antagonistic substances of LF01 strain was analyzed based on antiSMASH software.The hydrolysis activity of LF01 strain to carbohydrate was analyzed based on CAZyme database.The toxicity of the antagonistic substances of LF01 strain to Oreochromis niloticus and Danio rerio was evaluated by intraperitoneal injection.The biocontrol effect of the antagonistic substances of LF01 strain to O.niloticus was evaluated by artificial infection after 5 d of feeding O.niloticus.The complete genome of LF01 strain had a total length of 3974023 bp and the GC content was 46.56%,and it contains 3843 coding genes.Compared to B.velezensis,the ANI and DDH values of LF01 strain were≥97.66%and≥80.10%,respectively.Furthermore,the genomic evolution analysis showed that LF01 strain was clustered with B.velezensis.These results indicated that LF01 strain was identified as B.velezensis at the genomic level.The LF01 strain contains nine gene clusters related to antagonistic substances,including bacillaene,bacillibactin,bacillomycin D,bacilysin,difficidin,fengycin,macrola

关 键 词：贝莱斯芽孢杆菌 基因组测序 拮抗物质 生防作用 

分 类 号：S942.3[农业科学—水产养殖]