Complete genome sequence of Bacillus velezensis YYC, a bacterium isolated from the tomato rhizosphere

Introduction

Bacillus velezensis is an important member of plant growth-promoting rhizobacteria. This species was found to have multiple growth-promoting effects and to produce a variety of secondary metabolites with antibacterial activity (Chowdhury et al. 2015). B. velezensis can be widely isolated from diversified environments, such as plant rhizospheres, soil, rivers, human food, animal guts and seawater, and can easily be isolated and cultured (Ye et al. 2018). In our work, the B. velezensis YYC strain was isolated from the tomato rhizosphere in Heilongjiang Province, China. B. velezensis YYC increased tomato growth and induced systemic resistance against Ralstonia solanacearum (unpublished data). Strain YYC is a non-pathogenic bacterium. Genome sequencing of B. velezensis YYC will provide basic insight into the growth-promoting and biocontrol mechanism.

Data description

Genome sequencing, assembly and annotation

B. velezensis YYC strain was propagated in Luria-Bertani broth with shaking at 180 r/min overnight at 30°C. By alignments of the 16S ri-bosome RNA and housekeeping genes, it was identified as Bacillus velezensis. Bacterial genomic DNA extraction kit (Majorbio Bio-pharm Technology Co., Ltd., Shanghai, China) was used to extract genomic DNA. A TBS-380 fluorometer (Turner Bio Systems Inc., Sunnyvale, CA) was used to quantify the purified genomic DNA. PacBio RS II Single Molecule Real Time (SMRT) and Illumina sequencing platforms were used to sequence the genomic DNA. The sequencing yielded 170,436 reads, including 1,341,760,841 bp, with 337.7× sequence depth. A statistic of quality information was applied for quality trimming, by which the low-quality data could be removed to result in clean data. Using Unicycler (Version 0.4.7) (Wick et al. 2017), the reads were assembled into contigs. A complete genome was generated by inspecting and completing the last circular step. Finally, using the Illumina reads, error correction of the PacBio assembly results was performed.

The number of protein coding sequences (CDSs) in the B. velezensis YYC genome was predicted by Glimmer (version 3.02) (http://ccb.jhu.edu/software/glimmer/index.shtml) (Delcher et al. 2007) and GeneMarkS software (version 4.3) (Besemer et al. 2005). The transfer RNA (tRNA) gene was analyzed by tRNAscan-SE v2.0 software (Version 2.0) (http://trna.ucsc.edu/software) (Chan et al. 2019). Barrnap software (Version 0.8) (https://github.com/tseemann/barrnap) was utilized to predict ri-bosome RNA genes. By aligning reads with the Nonredundant (NR), Swiss-Prot, Kyoto Encyclopedia of Genes and Genomes (KEGG) (Kanehisa et al. 2016), Gene Ontology (GO) (Ashburner et al. 2000), Cluster of Orthologous Groups of proteins (COG) (Galperin et al. 2015) and protein families (Pfam) (Finn et al. 2014) databases, all genes were annotated. The bioactive secondary metabolites were predicted by antiSMASH software (Version 4.0.2) (Weber et al. 2015).

General genome features of B. velezensis YYC

Whole-genome sequencing showed that the B. velezensis YYC strain contained a genome size of approximately 3,973,236 bp, with an average G + C content of 46.52%. The Glimmer program predicted that the number of protein coding sequences (CDSs) was 4,034, and the average gene length was 877.29 bp. Furthermore, a total of 86 tRNA and 27 ri-bosome RNA genes were identified and analyzed in the genome. By aligning the genome to sequences from diverse databases, including the NR, Swiss-Prot, Pfam, COG, GO and KEGG databases, the numbers of identified genes were 4,034, 3,533, 3,337, 3,013, 2,668, and 2,163, respectively.

The KEGG database analysis showed a great number of two-component systems (113 genes) and ABC transporters (117 genes). Meanwhile, 69 genes were related to quorum sensing, which were important for cross-kingdom communication (Schikora et al. 2016).

Secondary metabolites related to biocontrol

AntiSMASH version 4.0.2 analysis identified 14 clusters of secondary metabolites (Fig. 1). Seven clusters of secondary metabolites were related to the synthesis of bacillaene, macrolactin, bacilysin, fengycin, difficidin, surfactin and bacillibactin. Some of these substances have antagonistic effects on bacteria, fungi, and viruses (Moldenhauer et al. 2007; Chowdhury et al.

2015; Wu et al. 2015; Zhang et al. 2018). Fengycin was found to induce resistance to plant diseases (Farzand et al. 2019).

Comparative genomics of diverse Bacillus strains

In addition, based on digital DNA–DNA hybridizations (dDDH), B. velezensis YYC was the most closely related with B. velezensis FZB42 (Table 1). And it shared 96.80% identity with the strain that was used as biofertilizer and biocontrol agent (B. velezensis FZB42). Compared with FZB42, lantipeptide biosynthesis gene cluster was special and only existed in the genome of B. velezensis YYC. It is engaged in the synthesis of locillomycin, which was related to hemolytic activity, swarming motility, biofilm formation, and colony morphology (Luo et al. 2019). In addition, four clusters encoding new metabolites with no reported description previously. Comparative analysis of secondary metabolite clusters of these strains were summarized for comparisons (Supplementary Table S1).

Table 1

Pairwise comparisons of *B. velezensis* YYC with type strain genomes
Subject strain	dDDH (d0, in %)	C.I. (d0, in %)	dDDH (d4, in %)	C.I. (d4, in %)	dDDH (d6, in %)	C.I. (d6, in %)	G + C content difference (in %)
Bacillus velezensis FZB42	96.8	[95.1–98.0]	90.8	[88.5–92.6]	97.5	[96.3–98.4]	0.05
Bacillus velezensis NRRL B-41580	91.8	[88.8–94.0]	85.4	[82.7–87.7]	93.3	[91.0–95.0]	0.21
Bacillus methylotrophicus KACC 13105	95	[92.7–96.6]	84.6	[81.9–87.0]	95.5	[93.7–96.8]	0.09
Bacillus siamensis KCTC 13613	89.1	[85.8–91.8]	56.9	[54.2–59.7]	85.4	[82.3–88.1]	0.19
Bacillus vanillea XY18	89	[85.6–91.7]	56.9	[54.2–59.7]	85.3	[82.2–88.0]	0.2
Bacillus amyloliquefaciens DSM 7	82.7	[78.8–86.0]	56	[53.2–58.7]	79.8	[76.3–82.8]	0.44
Bacillus nakamurai NRRL B-41091	73.4	[69.5–77.1]	31	[28.6–33.5]	61.2	[57.9–64.4]	1.26
Bacillus tequilensis NCTC13306	30.9	[27.5–34.5]	21.3	[19.0–23.7]	27.5	[24.6–30.6]	2.54
Bacillus spizizenii TU-B-10	33.6	[30.2–37.2]	21	[18.8–23.4]	29.2	[26.3–32.3]	2.7
Bacillus subtilis NCIB 3610	32.5	[29.1–36.0]	20.9	[18.7–23.3]	28.4	[25.5–31.5]	3.13
The dDDH values were provided along with their confidence intervals (C.I.).

Genes involved in promoting plant growth and triggering plant immunity

In addition to producing secondary metabolites with antifungal or antibacterial activity. B. velezensis YYC contains a various of genes implicated in biofilm formation and root colonization (Table 2). B. velezensis YYC contained the genes encoding acetolactate synthase (ilvH, ilvB), acetolactate decarboxylase (alsD), and butanediol dehydrogenase (butB), which have plant growth–promoting effects including stimulating root formation and increasing systemic disease resistance (He et al. 2013; Jayakumar et al. 2020). B. velezensis YYC also has the genes required for synthesis of 2,3-butanediol (alsD), the compound reported to trigger systemic resistance (He et al., 2013).

Table 2

Representative genes of *B. velezensis* YYC probably involved in plant bacterium interactions
Gene	Position	Protein	Description
spo0A	2432908 − 2432108	Sporulation transcription factor Spo0A	Biofilm formation
sinR	2466273–2466614	Transcriptional regulator	Biofilm formation
abrB	45929 − 45645	Transition state regulator Abh	Biofilm formation
resE	2260204–2261985	Sensor histidine kinase	Biofilm formation
lytS	2757501–2759282	Sensor histidine kinase	Biofilm formation
ycbA	248181–249473	Sensor histidine kinase	Biofilm formation
sacB	3926734–3928215	Levan sucrase	Root adhesion
efp	2452165–2452722	Elongation factor P	Essential for swarming motility
comP	3033195–3035495	Histidine kinase	Regulator of surfactin production
fliD	3414679–3416199	Flagellar capping protein	Elicitation of plant basal defence
flgK	3419847–3421364	Flagellar hook-associated protein FlgK	Elicitation of plant basal defence
xynB	1846654–1848069	1,4-beta-D-xylan xylohydrolase	Carbohydrate metabolism
lacR	1216343–1217104	Lactose phosphotransferase system repressor	Lactose metabolism
lacG	1214690–1216090	6-phospho-beta-galactosidase	Hydrolyzation of phospholactose
lacE	1212603–1214300	Phosphotransferase system	Cellobiose degradation
lacF	1214312–1214626	Phosphotransferase system	Cellobiose degradation
alsD	3488161–3488928	Acetolactate decarboxylase	Synthesis of 2,3-butanediol
pta	3639358–3640329	Phosphotransacetylase	Strongly upregulated by root exudate
ilvH	2697631–2698149	Acetolactate synthase	Promote plant growth
ilvB	2698146–2699963	Acetolactate synthase	Promote plant growth
alsD	3488161–3488928	Acetolactate decarboxylase	Promote plant growth
butB	629733–630773	Butanediol dehydrogenase	Promote plant growth

Nucleotide sequence accession number

The complete genome sequence of B. velezensis YYC was deposited in GenBank under accession number CP075055. (BioProject: PRJNA728388, BioSample: SAMN19079027).

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