河北链霉菌JL9001对番茄枯萎病的防治效果及全基因组序列分析

Biocontrol effect of Streptomyces hebeiensis JL9001 on tomato Fusarium wilt and whole genome sequence analysis

摘要:

目的 从太子参中分离出对番茄枯萎病具有较强生防潜力的河北链霉菌(Streptomyces hebeiensis) JL9001，解析其全基因组序列与功能注释信息，挖掘其次级代谢产物的基因信息，为番茄枯萎病的生物防治提供优良菌种资源及理论依据。方法 采用平板划线法研究不同培养基上菌落的形态，利用平板对峙法探究菌株JL9001对尖孢镰孢菌的拮抗效果；通过萃取发酵粗提物，运用微量稀释法探究其代谢产物对尖孢镰孢菌的活性；采用发酵液灌根法评估菌株JL9001对番茄枯萎病的防治效果。对菌株JL9001进行全基因组测序，并运用相关软件对测序数据进行种属鉴定、基因预测、功能注释以及次级代谢产物合成基因簇预测等。结果 菌株JL9001在SIM培养基上生长产孢效果最佳。拮抗试验表明，其对尖孢镰孢菌菌丝生长的抑制率达40.18%；在1 000 μg/mL浓度下萃取的代谢粗提物能够完全抑制尖孢镰孢菌。盆栽试验表明，浇灌JL9001发酵液后第13天对番茄枯萎病的抑制率达51.61%。菌株JL9001基因组序列总长为7 700 822 bp，G+C含量为71.46%，总基因数为6 589个；同时预测到27个次级代谢产物合成基因簇，其中包括萜烯、聚酮化合物以及铁载体等具有潜在抗菌活性的代谢产物。结论 本研究通过拮抗和盆栽试验表明，菌株JL9001可有效抑制番茄枯萎病的发生。对菌株JL9001基因组的构成和功能基因信息进行分析，为探究抗菌性天然产物的抑菌机制、分析其次级代谢产物合成基因簇以及评估链霉菌次生代谢产物的潜力奠定了基础。

Abstract:

Objective To isolate the Streptomyces hebeiensis strain JL9001 with significant biocontrol potential against tomato Fusarium wilt from Pseudostellaria heterophylla roots, elucidate the complete genome sequence and functional annotation of the strain, and extract genetic data pertaining to its secondary metabolites, thus offering a valuable microbial resource and a theoretical foundation for the biological management of tomato Fusarium wilt.Methods The colony morphology on various media was examined via the plate streaking technique. The antagonistic properties of strain JL9001 against Fusarium oxysporum were evaluated through the plate confrontation assay. The activities of metabolites (crude fermentation extract) against F. oxysporum were assessed via the microdilution method. The effectiveness of strain JL9001 in managing tomato Fusarium wilt was evaluated through root drenching with the fermentation broth. Whole genome sequencing of strain JL9001 was conducted, and the sequencing data were analyzed by appropriate software for species identification, gene prediction, functional annotation, and prediction of secondary metabolite biosynthesis gene clusters.Results Strain JL9001 demonstrated optimal spore production on the SIM medium. Antagonistic assays indicated that it inhibited the mycelial growth of F. oxysporum by 40.18%. Furthermore, the crude fermentation extract at a concentration of 1 000 μg/mL completely inhibited F. oxysporum. Pot trials revealed that irrigation with the fermentation broth of JL9001 resulted in a 51.61% reduction in tomato Fusarium wilt on day 13. The genome of strain JL9001 comprised 7 700 822 base pairs with the G+C content of 71.46%, encompassing 6 589 genes. Analysis predicted the presence of 27 biosynthetic gene clusters for secondary metabolites including terpenoids, polyketides, and siderophores, which may possess antimicrobial properties.Conclusion This study elucidates, through antagonistic and pot experiments, that strain JL9001 effectively mitigates the incidence of tomato Fusarium wilt. The analysis of the genomic composition and functional gene information of strain JL9001 provides a basis for exploring the antimicrobial mechanisms of natural products, examining secondary metabolite biosynthetic gene clusters, and assessing the potential of Streptomyces-derived secondary metabolites.