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Chinese Journal of Biological Control ›› 2023, Vol. 39 ›› Issue (4): 885-894.DOI: 10.16409/j.cnki.2095-039x.2023.02.035

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Antifungal Mechanism Analysis and Verification of Bacillus velezensis Ba-0321 Based on Whole Genome Sequencing

LI Xiaojie, QIU Rui, LIU Chang, YAO Chenxiao, BAI Jingke, CHEN Yuguo, LI Shujun   

  1. Tobacco Research Institute of Henan Academy of Agricultural Sciences/Key Laboratory for Green Preservation & Control of Tobacco Diseases and Pests in Huanghuai Growing Area, Xuchang 461000, China
  • Received:2022-11-03 Online:2023-08-25 Published:2023-08-25

Abstract: Bacillus velezensis Ba-0321, isolated from the rhizosphere soil of healthy tobacco plants, is a plant growth promoting rhizobacteria with strong antifungal activity. It has a good potential in biocontrol application. In the study, the inhibitory effects of strain Ba-0321 sterile filtrate on Fusarium oxysporum and Phytophthora nicotianae were determined by methods of co-culture. The whole genome of strain Ba-0321 was sequenced using the second-generation Illumina and the third-generation Nanopore platform, then analyzed for genome assembly, gene functional annotation, prediction of secondary metabolite synthetic gene clusters. The results showed that the sterile filtrate of strain Ba-0321 has inhibitory effects on both F. oxysporum and P. nicotianae. The culture medium containing 10% sterile filtrate has a higher inhibitory rate on the mycelial growth of the two pathogens, reaching 57.38% and 34.30%, respectively. The results of whole genome sequencing showed that the genome size of strain Ba-0321 was 4099109 bp, including 3897 coding genes, and the sequencing data are available in the GenBank database (accession No. CP101904). There were a large number of genes encoding enzymes, terpene, polyketone metabolic pathways, and genes involved in defense mechanisms have been annotated in the GO, COG and KEGG databases. Using anti-SMASH software, 13 secondary metabolite synthesis gene clusters were predicted encoding antibacterial substances such as Surfactin、Fengycin、difficidin、Bacillaene、Bacillibactin、Macrolactin H、Bacilysin. The existence of biosynthetic gene cluster for antibacterial substances and defence machainsm related genes in Ba-0321 strain genome were verified by PCR amplification and sequencing. The enzyme activity test results showed that the strain has resistant enzyme activities such as protease, chitinase, and cellulase.. This study provides a basis for analyzing the antimicrobial mechanism of strain Ba-0321 and mining antifungal related gene resources at the genomic level, which is of a great significance for research and application of the Ba-0321 strain in future.

Key words: Bacillus velezensis, whole genome, antifungal mechanism, functional annotation, functional verification

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