A biocontrol bacterium YN-2A with growth-promoting effect: isolation, identification, and genome sequencing

Abstract:

[Background] There are numerous microorganisms in the rhizosphere soil, many of which can antagonize pathogens and thus protect plants from diseases. [Objective] To isolate and identify a biocontrol bacterial strain capable of promoting plant growth and explore the antifungal mechanism from genomics and bioinformatics. [Methods] The dilution-plate coating method was employed to isolate a bacterial strain with antagonistic effects against Colletotrichum orbiculare. The strain was identified based on 16S rRNA gene sequencing and genome sequence alignment. The effects of the strain suspension (5×10⁸ CFU/mL) on tomato seeds and seedlings were determined. The antifungal spectrum of the strain was determined by the plate confrontation test. Furthermore, the third-generation (PacBio, Nanopore) sequencing was combined with the second-generation sequencing to analyze the genomic information and gene function annotation of the strain. [Results] A strain of biocontrol bacteria isolated from the rhizosphere soil of Schefflera heptaphylla was named YN-2A. Based on the alignment results of 16S rRNA gene and nucleotide sequence database (NT), the strain YN-2A was identified as Bacillus velezensis. Strain YN-2A was an aerobic Gram-positive bacterial strain with the ability to produce biofilms. Strain YN-2A suspension (5×10⁸ CFU/mL) promoted tomato seed germination and seedling growth and increased the leaf chlorophyll fluorescence parameters. Specifically, the suspension increased the germination rate at the time points of 72, 96 and 120 h by 1.45 times, 1.42 times and 1.09 times, respectively, and it increased the plant height of tomato seedlings by 5.90 cm. Among the fluorescence parameters, photochemical quenching (qP) exhibited the most significant increase of 1.27 times in the strain treatment group compared with the control group. The results of the plate confrontation test showed that strain YN-2A can antagonize a variety of fungal pathogens. The results of whole genome sequencing showed that the whole genome length of strain YN-2A was 4 046 066 bp, with the G+C content of 46% and 4 090 coding genes. The whole genome sequencing data was submitted to NCBI to obtain the GenBank accession number CP139086. Most genes were annotated to the amino acid and carbohydrate metabolic pathways. Particularly, 905 genes were annotated to reduced virulence in the pathogen host interactions (PHI) database, and 69 genes were annotated to the loss of pathogenicity database. It is thus hypothesized that the antifungal effect of YN-2A is attributed to the disease-resistant and virulence-reducing genes. [Conclusion] A biocontrol strain B.velezensis YN-2A was isolated and identified. The strain promoted tomato seed germination and seedling growth and demonstrated a broad antifungal spectrum. The whole genome information of strain YN-2A was obtained by sequencing, which provided reference information for deciphering the antifungal mechanism of strain YN-2A.

Key words:biocontrol bacteria;antagonistic effect;whole genome sequencing;gene function annotation