Complete Genome of Bacillus velezensis CMT-6 and Comparative Genome Analysis Reveals Lipopeptide Diversity

  • Qi Deng
  • Rundong Wang
  • Dongfang Sun
  • Lijun SunEmail author
  • Yaling WangEmail author
  • Yuehua Pu
  • Zhijia Fang
  • Defeng Xu
  • Ying Liu
  • Riying Ye
  • Sanjun Yin
  • Sisi Xie
  • Ravi Gooneratne
Original Article


The complete genome sequence of Bacillus velezensis type strain CMT-6 is presented for the first time. A comparative analysis between the genome sequences of CMT-6 with the genome of Bacillus amyloliquefaciens DSM7T, B. velezensis FZB42, and Bacillus subtilis 168 revealed major differences in the lipopeptide synthesis genes. Of the above, only the CMT-6 strain possessed an integrated synthetase gene for synthesizing surfactin, iturin, and fengycin. However, CMT-6 shared 14, 12, and 10 other lipopeptide-producing genes with FZB42, DSM7T, and 168 respectively. The largest numbers of non-synonymous mutations were detected in 205 gene sequences that produced these three lipopeptides in CMT-6 and 168. Comparing CMT-6 with DSM7T, 58 non-synonymous mutations were detected in gene sequences that contributed to produce lipopeptides. In addition, InDels were identified in yczE and glnR genes. CMT-6 and FZB42 had the lowest number of non-synonymous mutations with 8 lipopeptide-related gene sequences. And InDels were identified in only yczE. The numbers of core genes, InDels, and non-synonymous mutations in genes were the main reasons for the differences in yield and variety of lipopeptides. These results will enrich the genomic resources available for B. velezensis and provide fundamental information to construct strains that can produce specific lipopeptides.


Bacillus Genome Sequence Lipopeptide diversity 



The authors gratefully acknowledge the public research and capacity building program of Guangdong Province (Grant Nos. 2014B020204005 and 2014B020205006) and Guangdong Ocean University higher education program for financial support of two major scientific research projects (Grant Nos. GDOU2013050205, 2013050312).

Compliance with Ethical Standards

Conflict of interest

The authors declare they have no conflicts of interest.

Ethical Approval

No procedures in this study, performed by any of the authors, involved human or animal participants.


  1. Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ (1990) Basic local alignment search tool. J Mol Biol 215(3):403–410. Google Scholar
  2. Ashburner M, Ball CA, Blake JA, Botstein D, Butler H, Cherry JM, Davis AP, Dolinski K, Dwight SS, Eppig JT (2000) Gene ontology: tool for the unification of biology. Nat Genet 25(1):25–29Google Scholar
  3. Bais HP, Fall R, Vivanco JM (2004) Biocontrol of Bacillus subtilis against infection of Arabidopsis roots by Pseudomonas syringae is facilitated by biofilm formation and surfactin production. Plant Physiol 134(1):307–319. Google Scholar
  4. Benson G (1999) Tandem repeats finder: a program to analyze DNA sequences. Nucleic Acids Res 27(2):573. Google Scholar
  5. Besemer J, Lomsadze A, Borodovsky M (2001) GeneMarkS: a self-training method for prediction of gene starts in microbial genomes. Implications for finding sequence motifs in regulatory regions. Nucleic Acids Res 29(12):2607–2618. Google Scholar
  6. Blin K, Medema MH, Kazempour D, Fischbach MA, Breitling R, Takano E, Weber T (2013) antiSMASH 2.0—a versatile platform for genome mining of secondary metabolite producers. Nucleic Acids Res 41(W1):W204–W212. Google Scholar
  7. Chen XH, Koumoutsi A, Scholz R, Eisenreich A, Schneider K, Heinemeyer I, Morgenstern B, Voss B, Hess WR, Reva O (2007) Comparative analysis of the complete genome sequence of the plant growth-promoting bacterium Bacillus amyloliquefaciens FZB42. Nat Biotechnol 25(9):1007–1014. Google Scholar
  8. Chiaromonte F, Yap VB, Miller W (2002) Scoring pairwise genomic sequence alignments. Pac Symp Biocomput 7:115–126Google Scholar
  9. Cimino M, Thomas C, Namouchi A, Dubrac S, Gicquel B, Gopaul DN (2012) Identification of DNA binding motifs of the mycobacterium tuberculosis PhoP/PhoR two-component signal transduction system. PLoS ONE 7(8):e42876. Google Scholar
  10. Core L, Perego M (2003) TPR-mediated interaction of RapC with ComA inhibits response regulator-DNA binding for competence development in Bacillus subtilis. Mol Microbiol 49(6):1509–1522. Google Scholar
  11. Darling AE, Mau B, Perna NT (2010) ProgressiveMauve: multiple genome alignment with gene gain, loss and rearrangement. PLoS ONE 5(6):e11147. Google Scholar
  12. Deng Q, Wang WJ, Sun LJ, Wang YL, Liao JM, Xu DF, Liu Y, Ye RY, Gooneratne R (2017) A sensitive method for simultaneous quantitative determination of surfactin and iturin by LC-MS/MS. Anal Bioanal Chem 409(1):179–191. Google Scholar
  13. DePristo MA, Banks E, Poplin R, Garimella KV, Maguire JR, Hartl C, Philippakis AA, del Angel G, Rivas MA, Hanna M (2011) A framework for variation discovery and genotyping using next-generation DNA sequencing data. Nat Genet 43(5):491–498. Google Scholar
  14. Dong WX, Li SZ, Lu XY, Zhang XY, Wang PP, Ma P, Guo QG (2014) Regulation of fengycin biosynthase by regulator PhoP in the Bacillus subtilis strain NCD-2. Acta Phytopathol Sin 44(2):180–187. Google Scholar
  15. Duitman EH, Hamoen LW, Rembold M, Venema G, Seitz H, Saenger W, Bernhard F, Reinhardt R, Schmidt M, Ullrich C, Stein T, Leenders F, Vater J (1999) The mycosubtilin synthetase of Bacillus subtilis ATCC6633: a multifunctional hybrid between a peptide synthetase, an amino transferase, and a fatty acid synthase. Proc Natl Acad Sci USA 96(23):13294–13299. Google Scholar
  16. Eijlander RT, Holsappel S, de Jong A, Ghosh A, Christie G, Kuipers OP (2016) SpoVT: from fine-tuning regulator in Bacillus subtilis to essential sporulation protein in Bacillus cereus. Front Microbiol 7:1–11. Google Scholar
  17. Fisher SH (1999) Regulation of nitrogen metabolism in Bacillus subtilis: vive la difference. Mol Microbiol 32(2):223–232. Google Scholar
  18. Guo QG, Li SZ, Lu XY, Li BQ, Ma P (2010) PhoR/PhoP two component regulatory system affects biocontrol capability of Bacillus subtilis NCD-2. Genet Mol Biol 33(2):333–340. Google Scholar
  19. Gupta M, Rao KK (2014) Phosphorylation of DegU is essential for activation of amyE expression in Bacillus subtilis. J Biosci 39(5):747–752. Google Scholar
  20. Harris RS (2007) Improved pairwise alignment of genomic DNA. Dissertation, The Pennsylvania State UniversityGoogle Scholar
  21. Jin Q, Jiang QY, Zhao L, Su CZ, Li SS, Si FY, Li SS, Zhou CH, Mu YL, Xiao M (2017) Complete genome sequence of Bacillus velezensis S3-1, a potential biological pesticide with plant pathogen inhibiting and plant promoting capabilities. J Biotechnol 259:199–203. Google Scholar
  22. Kanehisa M (1997) A database for post-genome analysis. Trends Genet 13(9):375. Google Scholar
  23. Kanehisa M, Goto S, Kawashima S, Okuno Y, Hattori M (2004) The KEGG resource for deciphering the genome. Nucleic Acids Res 32(suppl 1):D277–D280. Google Scholar
  24. Kanehisa M, Goto S, Hattori M, Aoki-Kinoshita KF, Itoh M, Kawashima S, Katayama T, Araki M, Hirakawa M (2006) From genomics to chemical genomics: new developments in KEGG. Nucleic Acids Res 34(suppl 1):D354–D357. Google Scholar
  25. Karatas AY, Cetinb S, Ozcengiz G (2003) The effects of insertional mutations in comQ, comP, srfA, spo0H, spo0A and abrB genes on bacilysin biosynthesis in Bacillus subtilis. BBA-Gene Struct Ex 1626(1–3):51–56. Google Scholar
  26. Koumoutsi A, Chen XH, Vater J, Borriss R (2007) DegU and YczE positively regulate the synthesis of bacillomycin D by Bacillus amyloliquefaciens strain FZB42. Appl Environ Microbiol 73(21):6953–6964. Google Scholar
  27. Krzywinski M, Schein J, Birol I, Connors J, Gascoyne R, Horsman D, Jones SJ, Marra MA (2009) Circos: an information aesthetic for comparative genomics. Genome Res 19(9):1639–1645. Google Scholar
  28. Kurtz S, Phillippy A, Delcher AL, Smoot M, Shumway M, Antonescu C, Salzberg SL (2004) Versatile and open software for comparing large genomes. Genome Biol 5(2):R12. Google Scholar
  29. Li W, Godzik A (2006) Cd-hit: a fast program for clustering and comparing large sets of protein or nucleotide sequences. Bioinformatics 22(13):1658–1659. Google Scholar
  30. Li WZ, Jaroszewski L, Godzik A (2001) Clustering of highly homologous sequences to reduce the size of large protein databases. Bioinformatics 17(3):282–283. Google Scholar
  31. Li WZ, Jaroszewski L, Godzik A (2002) Tolerating some redundancy significantly speeds up clustering of large protein databases. Bioinformatics 18(1):77–82. Google Scholar
  32. Medema MH, Blin K, Cimermancic P, de Jager V, Zakrzewski P, Fischbach MA, Weber T, Takano E, Breitling R (2011) antiSMASH: rapid identification, annotation and analysis of secondary metabolite biosynthesis gene clusters in bacterial and fungal genome sequences. Nucleic Acids Res 39:W339–W346. Google Scholar
  33. Menkhaus M, Ullrich C, Kluge B, Vater J, Vollenbroich D, Kamp RM (1993) Structural and functional organization of the surfactin synthetase multienzyme system. J Biol Chem 268(11):7678–7684Google Scholar
  34. Miras M, Dubnau D (2016) A DegU-P and DegQ-dependent regulatory pathway for the K-state in Bacillus subtilis. Front Microbiol 7:1–14. Google Scholar
  35. Naknao MM, Marahiel MA, Zuber P (1988) Identification of a genetic locus required for biosynthesis of the lipopeptide antibiotic surfactin in Bacillus subtilis. J Bacteriol 170(12):5662–5668Google Scholar
  36. Nandi T, Ong C, Singh AP, Boddey J, Atkins T, Sarkar-Tyson M, Essex-Lopresti AE, Chua HH, Pearson T, Kreisberg JF (2010) A genomic survey of positive selection in Burkholderia pseudomallei provides insights into the evolution of accidental virulence. PLoS Pathog 6(4):e1000845. Google Scholar
  37. Piazza F, Tortosa P, Dubnau D (1999) Mutational analysis and membrane topology of ComP, a quorum-sensing histidine kinase of Bacillus subtilis controlling competence development. J Bacteriol 181(15):4540–4548Google Scholar
  38. Pu YH, Sun LJ, Wang YL, Deng Q, Chen DM, Liu HM, Xu DF, Deng CJ, Li JR (2013) Modeling inhibitory activity of a novel antimicrobial peptide AMPNT-6 from Bacillus subtilis against Vibrio parahaemolyticus in shrimp under various environmental conditions. Food Control 33(1):249–253. Google Scholar
  39. Ruckert C, Blom J, Chen XH, Reva O, Borriss R (2011) Genome sequence of B. amyloliquefaciens type strain DSM7(T) reveals differences to plant-associated B. amyloliquefaciens FZB42. J Biotechnol 155(1):78–85. Google Scholar
  40. Saha S, Bridges S, Magbanua ZV, Peterson DG (2008) Empirical comparison of ab initio repeat finding programs. Nucleic Acids Res 36(7):2284–2294. Google Scholar
  41. Schneider J, Taraz K, Budzikiewicz H, Deleu M, Thonart P, Jacques P (1999) The structure of two fengycins from Bacillus subtilis S499. Z Naturforsch 54(11):859–865Google Scholar
  42. Stein T (2005) Bacillus subtilis antibiotics: structures, syntheses and specific functions. Mol Microbiol 56(4):845–857. Google Scholar
  43. Su K, Wang K, Sun LJ, Liu HM, Liu YF (2013) Effect of antimicrobial peptide on growth performance and health of weaning piglets. J Henan Agric Sci 42(9):112–115. Google Scholar
  44. Sun LJ, Wang YL, Liu HM, Xu DF, Nie FH, Zhou ZF, Chen J, Li JR (2012) Hemolytic and mice acute oral toxicity evaluation of a new antimicrobial peptide APNT-6. J Fish China 36(6):974–978. Google Scholar
  45. Sun LJ, Wang YL, Liu HM, Xu DF, Zhang YP, Nie FH (2013) Identification of antimicrobial lipopeptides component produced by isolate from Douchi and its antimicrobial properties. China Biotechnol 33(7):50–56. Google Scholar
  46. Tamura K, Stecher G, Peterson D, Filipski A, Kumar S (2013) MEGA6: molecular evolutionary genetics analysis version 6.0. Mol Biol Evol 30(12):2725–2729. Google Scholar
  47. Tan ZY, Zhang Z, Fu MH, Luo D, Zhong J, Zhou JY, Yang J, Xiao L, Tang H (2015) Effect of feeding on regulatory genes of Bacillus subtilis ZK8 synthesizing iturin A in fermentation process. J Agric Sci Technol 17(3):35–41. Google Scholar
  48. Tatusov RL, Koonin EV, Lipman DJ (1997) A genomic perspective on protein families. Science 278(5338):631–637. Google Scholar
  49. Tatusov RL, Fedorova ND, Jackson JD, Jacobs AR, Kiryutin B, Koonin EV, Krylov DM, Mazumder R, Mekhedov SL, Nikolskaya AN (2003) The COG database: an updated version includes eukaryotes. BMC Bioinform 4(1):41. Google Scholar
  50. Tsuge K, Akiyama T, Shoda M (2001) Cloning, sequencing, and characterization of the iturin A operon. J Bacteriol 183(21):6265–6273. Google Scholar
  51. Vater J, Kablitz B, Wilde C, Franke P, Mehta N, Cameotra SS (2002) Matrix-assisted laser desorption ionization-time of flight mass spectrometry of lipopeptide biosurfactants in whole cells and culture filtrates of Bacillus subtilis C-1 isolated from petroleum sludge. Appl Environ Microbiol 68(12):6210–6219. Google Scholar
  52. Vesth T, Lagesen K, Acar O, Ussery D (2013) CMG-biotools, a free workbench for basic comparative microbial genomics. PLoS ONE 8(4):e60120. Google Scholar
  53. Wang DP, Zhang YB, Zhang Z, Zhu J, Yu J (2010) KaKs_Calculator 2.0: a toolkit incorporating gamma-series methods and sliding window strategies. Genom Proteom Bioinform 8(1):77–80. Google Scholar
  54. Weng J, Wang Y, Li J, Shen QR, Zhang RF (2013) Enhanced root colonization and biocontrol activity of Bacillus amyloliquefaciens SQR9 by abrB gene disruption. Appl Microbiol Biotechnol 97(19):8823–8830. Google Scholar
  55. Zhang N, Pu YH, Sun LJ, Wang YL, Deng Q, Xu DF, Liu Y, Hussain M, Gooneratne R (2017a) Modeling the effects of different conditions on the inhibitory activity of antimicrobial lipopeptide (AMPNT-6) against Staphylococcus aureus growth and enterotoxin production in shrimp meat. Aquacult Int 25(1):50–57. Google Scholar
  56. Zhang Z, Ding ZT, Zhong J, Zhou JY, Shu D, Luo D, Yang J, Tan H (2017b) Improvement of iturin A production in Bacillus subtilis ZK0 by overexpression of the comA and sigA genes. Lett Appl Microbiol 64(6):452–458. Google Scholar
  57. Zhi Y, Wu Q, Xu Y (2017) Genome and transcriptome analysis of surfactin biosynthesis in Bacillus amyloliquefaciens MT45. Sci Rep-UK 7:40976. Google Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Qi Deng
    • 1
  • Rundong Wang
    • 1
  • Dongfang Sun
    • 1
  • Lijun Sun
    • 1
    Email author
  • Yaling Wang
    • 1
    Email author
  • Yuehua Pu
    • 2
  • Zhijia Fang
    • 1
  • Defeng Xu
    • 1
  • Ying Liu
    • 1
  • Riying Ye
    • 1
  • Sanjun Yin
    • 3
  • Sisi Xie
    • 3
  • Ravi Gooneratne
    • 4
  1. 1.Guangdong Provincial Key Laboratory of Aquatic Product Processing and Safety, Key Laboratory of Advanced Processing of Aquatic Products of Guangdong Higher Education Institution, College of Food Science and TechnologyGuangdong Ocean UniversityZhanjiangChina
  2. 2.Guangdong Institute of Special Equipment Inspection and Research Zhanjiang BranchZhanjiangChina
  3. 3.Health Time Gene InstituteShenzhenChina
  4. 4.Department of Wine, Food and Molecular BiosciencesLincoln UniversityLincolnNew Zealand

Personalised recommendations