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Complete genome sequence of Bacillus subtilis BC1-43, a soil bacterium with antagonistic activities against a wide range of plant fungal pathogens
Korean J. Microbiol. 2021;57(4):289-291
Published online December 31, 2021
© 2021 The Microbiological Society of Korea.

Mi-Young Won, Hyejeong Moon, Mee Kyung Sang, Jaekyeong Song, and Hang-Yeon Weon*

Agricultural Microbiology Division, National Institute of Agricultural Sciences, Rural Development Administration (RDA), Wanju 55365, Republic of Korea
Correspondence to: E-mail: why@korea.kr;
Tel.: +82-63-238-3042;Fax: +82-63-238-3834
Received November 17, 2021; Revised December 9, 2021; Accepted December 9, 2021.
Abstract
A Bacillus subtilis strain BC1-43 was isolated from soil, which demonstrated antifungal activity against a wide range of phytopathogenic fungi. The genome of strain BC1-43 was sequenced using the PacBio Sequel system. The draft genome was 4,185,798 bp (chromosome 4,157,766 bp and plasmid 28,032 bp) and the genomic G + C content was 43.6%. In total, the number of coding sequences was 4,170, including 30 rRNAs and 86 tRNAs. We identified gene clusters encoding antimicrobial compounds (bacilysin, subtilosin, bacillibactin, fengycin, bacillaene and surfactin), which are involved in the suppression of phytopathogenic fungi.
Keywords : Bacillus subtilis, genome sequence, biocontrol, phytopathogenic fungi
Body

Athelia rolfsii, a soil-borne plant pathogen, has a wide host range owing to its prolific growth and sclerotia production, and is responsible for substantial financial losses in economically important crops. Chemical control is an arduous task because of the associated technical, economic, and environmental issues (Punja, 1985; Kumari et al., 2021). Strain BC1-43 was isolated from soil in Cheonan, Chungcheongam-do, South Korea, which demonstrated antifungal activity, highlighting its potential as an eco-compatible control agent. Genome-based phylogenetic analysis identified strain BC1-43 as Bacillus subtilis. The strain reduced up to 43.3% of the disease severity caused by A. rolfsii in a hot pepper seedling experiment (unpublished data). In addition, strain BC1-43 showed antifungal activity toward a broad spectrum of pathogenic fungi, including Colletotrichum acutatum, Botrytis cinerea, Fusarium oxysporum, Phytophthora capsici and promoted plant growth in experiments with hot pepper and tomato.

Here, we analyzed the genome sequence of strain BC1-43 to explore the genomic features responsible for its effectiveness as a biocontrol agent. Whole-genome sequencing was performed using the PacBio RSII system by Macrogen. Short-read data sets were used for de novo assembly using a hierarchical genome assembly process (HGAP, v3.0). Bacterial genome annotation was implemented using the NCBI Prokaryotic Genome Annotation Pipeline (Tatusova et al., 2016). The complete genome sequence of strain BC1-43 consisted of one chromosome and one plasmid with 4,157,766 bp and 28,032 bp, respectively. The Rapid Annotation Subsystem (RAST) server was used for more detailed genomic analysis (Aziz et al., 2008). RAST demonstrated that the G + C content of strain BC1-43 was 43.6%. A total of 4,170 genes, including 30 rRNAs, 86 tRNAs, 5 ncRNAs and 74 pseudogenes, were predicted (Table 1).

Genome features of Bacillus subtilis BC1-43

Genome feature Chromosome Plasmid Total
Genome size (bp) 4,157,766 28,032 4,185,798
G + C content (%) 43.6% 39.4% 43.6%
Total genes 4,137 33 4,170
tRNAs 86 0 86
rRNAs (5S, 16S, 23S) 30 (10, 10, 10) 0 30
Pseudogenes 74
GenBank accession No. CP086061 CP086062


Bacillus subtilis is known to possess the ability to produce a variety of antifungal compounds, including bacilysin that is encoded by a gene cluster common to the genus Bacillus (Nannan et al., 2021). Secondary metabolite biosynthesis gene clusters in strain BC1-43 were identified by the antiSMASH web server (https://antismash.secondarymetabolites.org/). According to antiSMASH 6.0 analysis, the bacilysin gene cluster was also predicted to be present in strain BC1-43. Strain BC1-43 genome encodes other antimicrobial compounds including subtilosin, bacillibactin, fengycin, bacillaene, and surfactin (Blin et al., 2021). In all five antimicrobial compounds except surfactin, the regions of strain BC1-43 were 100% consistent with those recorded for the genus Bacillus, whereas the sequence for surfactin demonstrated 82% similarity with that of Bacillus. These compounds are also widely known to inhibit phytopathogenic fungi. According to Chen (2020), bacillomycin, surfactin, and fengycin suppress A. rolfsii when they act as antifungal lipopeptides (Alvarez et al., 2012). In addition, the Comprehensive Antibiotic Resistance Database (https://card.mcmaster.ca/) server predicted that strain BC1-43 harbors an aminoglycoside 6-adenylyltransferase gene that may confer resistance to streptomycin (McArthur et al., 2013). Multidrug efflux gene pump genes blt and ykkCD were also predicted with a matching rate of 100%.

Identifying the function of microorganisms through genetic analysis can improve study efficiency when experimentally verifying microbes’ activity. Therefore, this study provides beneficial information for developing B. subtilis BC1-43 as a potential biocontrol agent by analyzing the whole-genome sequence. The availability of the complete genome sequence will also enable further study and elucidation of the mechanisms underlying the biocontrol of plant diseases.

Nucleotide sequence accession numbers

The accession number of the draft genome sequence of strain BC1-43 is CP086061.1 and CP086062.1 in the GenBank database server. In addition, strain BC1-43 has been deposited in the Korean Agriculture Collection under accession number KACC 81188BP.

적 요

토양에서 분리한 Bacillus subtilis BC1-43 균주는 다양한 식물병원균에 대한 길항능을 가지고 있었다. 이 균주의 유전체는 4,157,766 bp 크기의 염색체와 28,032 bp 크기의 플라스미드로 구성되었다. 유전체 초안으로부터 단백질 유전자 4,1708개, rRNA 유전자 30개, tRNA 유전자 86개를 확인하였다. 유전자 분석 결과 Bacillus BC1-43은 antifungal compounds인 bacilysin, subtilosin, bacillibactin, fengycin, bacillaene 및 surfactin과 같은 다양한 2차 대사 산물을 생산하는 유전자가 확인되었다. Bacillus subtilis BC1-43 균주의 유전체 분석은 식물병원균의 생물적 방제 기작을 이해하는 데 기여할 것이다.

Acknowledgments

This study was carried out with the support of “Research Program for Agricultural Science & Technology Development (Project No. PJ01505102)” from the National Institute of Agricultural Sciences, Rural Development Administration, Republic of Korea.

Conflict to Interest

The researcher claims no conflicts of interest.

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  • National Institute of Agricultural Sciences, Rural Development Administration
     
      PJ01505102