Draft Genome Sequences of Four Plant Probiotic Bacillus Strains.

Here, we report the whole-genome sequences of four Bacillus strains that exhibit plant probiotic activities. Three of them are the type strains of Bacillus endophyticus, "Bacillus gaemokensis," and Bacillus trypoxylicola, and the other, Bacillus sp. strain KCTC 13219, should be reclassified into a species belonging to the genus Lysinibacillus.

GENOME ANNOUNCEMENT

Bacillus is one of the most extensively studied microorganisms among plant growth-promoting rhizobacteria (PGPR). Owing to the production of various antibiotics and the formation of stress-resistant endospores, Bacillus strains are more amenable to the formulation of commercial products and have been widely used as biocontrol agents (1). Induced systemic resistance (ISR) elicited by Bacillus strains has been recently recognized as one of the key mechanisms by which crops can protect themselves against phytopathogens with the aid of PGPR (2, 3).

The four Bacillus strains sequenced in this study, all purchased from the Korean Collection for Type Cultures (KCTC), were chosen based on the partial results of a systematic screening approach used to search for plant probiotic bacteria. The three type strains, Bacillus endophyticus KCTC 13922 (4), “Bacillus gaemokensis” KCTC 13318 (5), and Bacillus trypoxylicola KCTC 13244 (6), were shown to promote plant growth (Arabidopsis thaliana) by volatiles, whereas Bacillus sp. strain KCTC 13219 (=b04i-3) triggers ISR in A. thaliana against Pectobacterium carotovorum (our unpublished data). The type strains B. gaemokensis JCM 15801 and B. trypoxylicola NRBC 102646, obtained from the Japan Collection of Microorganisms (JCM) and the NITE Biological Resource Center (NBRC), respectively, were sequenced by other research groups and made available through accession numbers JOTM00000000 (7) and BCWA00000000 while this paper was being prepared.

Genome sequencing was carried out using the Illumina HiSeq 2000 platform at the National Instrument Center for Environmental Management at Seoul National University (Seoul, Republic of Korea). One hundred one-nucleotide paired reads produced from a library with a fragment size of ca. 500 bp were pretreated using Trimmomatic version 0.32 (8), and 600-Mb subsamples were randomly extracted from them. After k-mer-based error correction using SGA version 0.10.13 (9), de novo genome assembly was conducted using the A5-miseq pipeline (10). The assembled sequences were annotated using the RAST server (11) and Prokaryotic Genome Annotation Pipeline from the NCBI. Biosynthetic genes for secondary metabolites were predicted using antiSMASH 3.0 (12). The list of strains used in this study, the sequencing and assembly statistics, and the accession numbers are all provided in Table 1. Compared with the previously published records, the genome sequences of KCTC 13318 and KCTC 13244 obtained through this study were shown to be superior in terms of the assembly statistics.

TABLE 1 

Summary of genome sequencing results

Bacillus species	Draft coverage (×)a	Genome size (bp)	No. of contigs	G+C content (%)	No. of secondary metabolite biosynthesis gene clustersb	Accession no.
B. endophyticus KCTC 13922T	741	5,121,484	42	36.50	11	LTAP00000000
B. gaemokensis KCTC 13318T	878	5,616,250	70	35.54	18	LTAQ00000000
B. trypoxylicola KCTC 13244T	9,024	4,347,941	40	35.73	6	LTAO00000000
Bacillus sp. KCTC 13219	1,244	3,853,058	37	38.31	3	LUFJ00000000

Subsamples of 600 Mb were randomly taken from pretreated reads and assembled.

Predicted by antiSMASH 3.0.

KCTC 13219, isolated from Pu-erh tea, was tentatively named “Bacillus nitroreducens” by the submitter but did not lead to a proposal for a novel species (J. S. Lee, personal communication). Very recently, the species Bacillus nitroreducens sp. nov. was proposed for another unrelated bacterium (13). Phylogenetic analysis using the 16S rRNA sequence revealed that KCTC 13219 should be classified into the genus Lysinibacillus (99.93% similarity with “Lysinibacillus fluoroglycofernilyticus” cmg86T). The genome sequencing results will provide insight into the genomic basis of the versatile Bacillus strains and the interspecies interaction between soil bacteria and plants, which can lead to eco-friendly agricultural applications. Additional information is available at http://genoglobe.kr/kribb/four_bacillus_strains_2016.

Nucleotide sequence accession numbers.

These whole-genome shotgun projects have been deposited at DDBJ/EMBL/GenBank, and the accession numbers for all four genome sequences are listed in Table 1.