An Aminoglycoside Antibacterial Substance, S-137-R, Produced by Newly Isolated Bacillus velezensis Strain RP137 from the Persian Gulf

  • Roya Pournejati
  • Ronald Gust
  • Hamid Reza Karbalaei-HeidariEmail author


Given antibiotic resistance in pathogens, finding antibiotics from new sources is always a topic of interest to scientists. In the present study, among various isolates from the Persian Gulf coastal area, the strain RP137 was selected as producer of antibacterial compound. Morphological and biochemical studies along with 16S rDNA sequencing showed that strain RP137 belongs to Bacillus genus and was tentatively named Bacillus velezensis strain RP137. The effect of various carbon and nitrogen sources on optimizing the production of antibacterial compound showed that the low-cost rice starch and potassium nitrate supply to the strain RP137 caused producing of 86.0 ± 8.7 µg/mL extract having the antibacterial activity. The fractionation of the primary methanol extract in different solvents followed by reversed-phase HPLC obtained a pure antibacterial-active sample, S-137-R. Structural analysis of the purified S-137-R with the help of FTIR, HR-MS, 1H-NMR, and 13C-NMR showed that the S-137-R compound is classified as aminoglycoside. Minimum inhibition concentration (MIC) of the pure compound for Gram-positive bacteria, Staphylococcus aureus and methicillin resistant Staphylococcus aureus, showed an average antibacterial effect of about 80 µg/mL and 150 µg/mL, respectively and for Pseudomonas aeruginosa (100 µg/mL), while having very little toxic effect on E. coli. Moreover, low cytotoxicity effect of the S-137-R on cancerous and normal cells as well as the low intensity of the hemolysis of red blood cells in higher concentrations of S-137-R make it an ideal candidate for further structure–activity relationship assessments towards its medical applications.

Graphic Abstract



R. Pournejati was supported by a graduate student fellowship from the Ministry of Science, Research and Technology, Government of Iran. The Austrian Research Promotion Agency FFG [West Austrian BioNMR 858017] is also kindly acknowledged. This work was supported in part by a grant for Scientific Research from the Iran National Science foundation (INSF) under Contract Number 93035486.

Compliance with Ethical Standards

Conflict of interest

The authors declare that there are no conflicts of interest.

Ethical Approval

This study was approved by the ethics committee of Department of Biology at Shiraz University, Iran (ECBD-SU9330270). All experiments and data comparisons were carried out in compliance with relevant laws and guidelines and in accordance with the ethical standards of the Declaration of Helsinki.

Supplementary material

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Supplementary material 1 (DOCX 79 kb)


  1. 1.
    Nathan C, Cars O (2014) Antibiotic resistance–problems, progress, and prospects. N Engl J Med 371(19):1761–1763. Google Scholar
  2. 2.
    Demain AL (2006) From natural products discovery to commercialization: a success story. J Ind Microbiol Biotechnol 33(7):486–495Google Scholar
  3. 3.
    Patridge E, Gareiss P, Kinch MS, Hoyer D (2016) An analysis of FDA-approved drugs: natural products and their derivatives. Drug Discov Today 21(2):204–207Google Scholar
  4. 4.
    Guo Z (2017) The modification of natural products for medical use. Acta Pharmaceutica Sinica B 7(2):119–136Google Scholar
  5. 5.
    Wilson RM, Danishefsky SJ (2006) Small molecule natural products in the discovery of therapeutic agents: the synthesis connection. J Org Chem 71(22):8329–8351Google Scholar
  6. 6.
    Newman DJ (2008) Natural products as leads to potential drugs: an old process or the new hope for drug discovery? J Med Chem 51(9):2589–2599Google Scholar
  7. 7.
    Trindade-Silva AE, Lim-Fong GE, Sharp KH, Haygood MG (2010) Bryostatins: biological context and biotechnological prospects. Curr Opin Biotechnol 21(6):834–842Google Scholar
  8. 8.
    Chen X, Koumoutsi A, Scholz R, Schneider K, Vater J, Süssmuth R, Piel J, Borriss R (2009) Genome analysis of Bacillus amyloliquefaciens FZB42 reveals its potential for biocontrol of plant pathogens. J Biotechnol 140(1–2):27–37Google Scholar
  9. 9.
    Sumi CD, Yang BW, Yeo I-C, Hahm YT (2014) Antimicrobial peptides of the genus Bacillus: a new era for antibiotics. Can J Microbiol 61(2):93–103Google Scholar
  10. 10.
    Demain AL (2014) Importance of microbial natural products and the need to revitalize their discovery. J Ind Microbiol Biotechnol 41(2):185–201Google Scholar
  11. 11.
    Vandamme E (2007) Microbial gems: Microorganisms without frontiers. SIM-News 57(3):81–91Google Scholar
  12. 12.
    Piel J (2004) Metabolites from symbiotic bacteria. Nat Prod Rep 21(4):519–538Google Scholar
  13. 13.
    Clardy J, Fischbach MA, Walsh CT (2006) New antibiotics from bacterial natural products. Nat Biotechnol 24(12):1541Google Scholar
  14. 14.
    Sepcic K, Zalar P, Gunde-Cimerman N (2011) Low water activity induces the production of bioactive metabolites in halophilic and halotolerant fungi. Mar Drugs 9(1):43–58Google Scholar
  15. 15.
    Challinor VL, Bode HB (2015) Bioactive natural products from novel microbial sources. Ann N Y Acad Sci 1354(1):82–97Google Scholar
  16. 16.
    Wilson DN (2013) Ribosome-targeting antibiotics and mechanisms of bacterial resistance. Nat Rev Microbiol 12:35. Google Scholar
  17. 17.
    Wainwright M (1991) Streptomycin: discovery and resultant controversy. Hist Philos Life Sci 13(1):97–124Google Scholar
  18. 18.
    Hermann T (2007) Aminoglycoside antibiotics: old drugs and new therapeutic approaches. Cell Mol Life Sci 64(14):1841–1852Google Scholar
  19. 19.
    Kudo F, Eguchi T (2016) Aminoglycoside Antibiotics: new Insights into the Biosynthetic Machinery of Old Drugs. Chemical record (New York, NY) 16(1):4–18. Google Scholar
  20. 20.
    Chaudhary HS, Soni B, Shrivastava AR, Shrivastava S (2013) Diversity and versatility of actinomycetes and its role in antibiotic production. J Appl Pharm Sci 3(8):S83–S94Google Scholar
  21. 21.
    Mondol M, Shin H, Islam M (2013) Diversity of secondary metabolites from marine Bacillus species: chemistry and biological activity. Mar Drugs 11(8):2846–2872Google Scholar
  22. 22.
    Fickers P (2012) Antibiotic compounds from Bacillus: why are they so amazing. Am J Biochem Biotechnol 8:38–43Google Scholar
  23. 23.
    Kunst F, Ogasawara N, Moszer I, Albertini A, Alloni G, Azevedo V, Bertero M, Bessieres P, Bolotin A, Borchert S (1997) The complete genome sequence of the gram-positive bacterium Bacillus subtilis. Nature 390(6657):249Google Scholar
  24. 24.
    Fan L, Bo S, Chen H, Ye W, Kleinschmidt K, Baumann HI, Imhoff JF, Kleine M, Cai D (2011) Genome sequence of Bacillus subtilis subsp. spizizenii gtP20b, isolated from the Indian ocean. J Bacteriol 193(5):1276–1277Google Scholar
  25. 25.
    Scholz R, Vater J, Budiharjo A, Wang Z, He Y, Dietel K, Schwecke T, Herfort S, Lasch P, Borriss R (2014) Amylocyclicin, a novel circular bacteriocin produced by Bacillus amyloliquefaciens FZB42. J Bacteriol. 196:1842–1852Google Scholar
  26. 26.
    An J, Zhu W, Liu Y, Zhang X, Sun L, Hong P, Wang Y, Xu C, Xu D, Liu H (2015) Purification and characterization of a novel bacteriocin CAMT2 produced by Bacillus amyloliquefaciens isolated from marine fish Epinephelus areolatus. Food Control 51:278–282Google Scholar
  27. 27.
    Fu L, Wang C, Ruan X, Li G, Zhao Y, Wang Y (2018) Preservation of large yellow croaker (Pseudosciaena crocea) by Coagulin L1208, a novel bacteriocin produced by Bacillus coagulans L1208. Int J Food Microbiol 266:60–68Google Scholar
  28. 28.
    Cochrane SA, Vederas JC (2016) Lipopeptides from Bacillus and Paenibacillus spp.: a gold mine of antibiotic candidates. Med Res Rev 36(1):4–31Google Scholar
  29. 29.
    Yamamoto S, Shiraishi S, Suzuki S (2015) Are cyclic lipopeptides produced by Bacillus amyloliquefaciens S13‐3 responsible for the plant defence response in strawberry against Colletotrichum gloeosporioides? Lett Appl Microbiol 60(4):379–386Google Scholar
  30. 30.
    Gond SK, Bergen MS, Torres MS, White JF Jr (2015) Endophytic Bacillus spp. produce antifungal lipopeptides and induce host defence gene expression in maize. Microbiol Res 172:79–87Google Scholar
  31. 31.
    Li Y, Li Z, Yamanaka K, Xu Y, Zhang W, Vlamakis H, Kolter R, Moore BS, Qian P-Y (2015) Directed natural product biosynthesis gene cluster capture and expression in the model bacterium Bacillus subtilis. Sci Rep 5:9383Google Scholar
  32. 32.
    Butcher RA, Schroeder FC, Fischbach MA, Straight PD, Kolter R, Walsh CT, Clardy J (2007) The identification of bacillaene, the product of the PksX megacomplex in Bacillus subtilis. Proc Natl Acad Sci USA 104(5):1506–1509Google Scholar
  33. 33.
    Bhate D (1955) Pumilin, a new antibiotic from Bacillus pumilus. Nature 175(4462):816Google Scholar
  34. 34.
    Kontnik R, Bosak T, Butcher RA, Brocks JJ, Losick R, Clardy J, Pearson A (2008) Sporulenes, heptaprenyl metabolites from Bacillus subtilis spores. Org Lett 10(16):3551–3554Google Scholar
  35. 35.
    Sugawara T, Shibazaki M, Nakahara H, Suzuki K (1996) YM-47522, a novel antifungal antibiotic produced by Bacillus sp. J Antibiot 49(4):345–348Google Scholar
  36. 36.
    Pinchuk IV, Bressollier P, Sorokulova IB, Verneuil B, Urdaci MC (2002) Amicoumacin antibiotic production and genetic diversity of Bacillus subtilis strains isolated from different habitats. Res Microbiol 153(5):269–276Google Scholar
  37. 37.
    Liu S, Han X, Jiang Z, Wu G, Hu X, You X, Jiang J, Zhang Y, Sun C (2016) Hetiamacin B–D, new members of amicoumacin group antibiotics isolated from Bacillus subtilis PJS. J Antibiot 69(10):769Google Scholar
  38. 38.
    Kevany BM, Rasko DA, Thomas MG (2009) Characterization of the complete zwittermicin A biosynthesis gene cluster from Bacillus cereus. Appl Environ Microbiol 75(4):1144–1155Google Scholar
  39. 39.
    Milner JL, Silo-Suh L, Lee JC, He H, Clardy J, Handelsman J (1996) Production of kanosamine by Bacillus cereus UW85. Appl Environ Microbiol 62(8):3061–3065Google Scholar
  40. 40.
    Ota Y, Tamegai H, Kudo F, Kuriki H, Koike-Takeshita A, Eouchi K, Kakinuma T (2000) Butirosin-biosynthetic gene cluster from Bacillus circulans. J Antibiot 53(10):1158–1167Google Scholar
  41. 41.
    Nguyen K-P (2014) Structural studies of Butirosin D and FrbG. University of Illinois at Urbana-Champaign,Google Scholar
  42. 42.
    Mincer TJ, Jensen PR, Kauffman CA, Fenical W (2002) Widespread and persistent populations of a major new marine actinomycete taxon in ocean sediments. Appl Environ Microbiol 68(10):5005–5011Google Scholar
  43. 43.
    Kahlmeter G, Brown D, Goldstein F, MacGowan A, Mouton J, Odenholt I, Rodloff A, Soussy CJ, Steinbakk M, Soriano F (2006) European Committee on Antimicrobial Susceptibility Testing (EUCAST) technical notes on antimicrobial susceptibility testing. Clin Microbiol Infect 12(6):501–503Google Scholar
  44. 44.
    Balouiri M, Sadiki M, Ibnsouda SK (2016) Methods for in vitro evaluating antimicrobial activity: a review. J Pharm Anal 6(2):71–79Google Scholar
  45. 45.
    Alborz M, Jarrahpour A, Pournejati R, Karbalaei-Heidari HR, Sinou V, Latour C, Brunel JM, Sharghi H, Aberi M, Turos E, Wojtas L (2018) Synthesis and biological evaluation of some novel diastereoselective benzothiazole beta-lactam conjugates. Eur J Med Chem 143:283–291. Google Scholar
  46. 46.
    Kudo F, Numakura M, Tamegai H, Yamamoto H, Eguchi T, Kakinuma K (2005) Extended sequence and functional analysis of the butirosin biosynthetic gene cluster in Bacillus circulans SANK 72073. J Antibiot 58(6):373Google Scholar
  47. 47.
    Gurung N, Ray S, Bose S, Rai V (2013) A broader view: microbial enzymes and their relevance in industries, medicine, and beyond. BioMed Res Int 2013:329121Google Scholar
  48. 48.
    Jeong H, Jeong D-E, Kim SH, Song GC, Park S-Y, Ryu C-M, Park S-H, Choi S-K (2012) Draft genome sequence of the plant growth-promoting bacterium Bacillus siamensis KCTC 13613T. J Bacteriol 194(15):4148–4149Google Scholar
  49. 49.
    La Rosa PS, Brooks JP, Deych E, Boone EL, Edwards DJ, Wang Q, Sodergren E, Weinstock G, Shannon WD (2012) Hypothesis testing and power calculations for taxonomic-based human microbiome data. PLoS ONE 7(12):e52078Google Scholar
  50. 50.
    Sansinenea E, Ortiz A (2011) Secondary metabolites of soil Bacillus spp. Biotechnol Lett 33(8):1523–1538Google Scholar
  51. 51.
    Urdaci M, Pinchuk I (2004) Antimicrobial activity of Bacillus probiotics. Bacterial spore formers–probiotics and emerging applications. Horizon Bioscience, Norfolk, pp 171–182Google Scholar
  52. 52.
    Sonenshein AL (2000) Control of sporulation initiation in Bacillus subtilis. Curr Opin Microbiol 3(6):561–566Google Scholar
  53. 53.
    Baruzzi F, Quintieri L, Morea M, Caputo L (2011) Antimicrobial compounds produced by Bacillus spp. and applications in food. Sci Against Microb Pathog 2:1102–1111Google Scholar
  54. 54.
    Ruiz B, Chávez A, Forero A, García-Huante Y, Romero A, Sánchez M, Rocha D, Sánchez B, Rodríguez-Sanoja R, Sánchez S (2010) Production of microbial secondary metabolites: regulation by the carbon source. Crit Rev Microbiol 36(2):146–167Google Scholar
  55. 55.
    Sanchez S, Chávez A, Forero A, García-Huante Y, Romero A, Sánchez M, Rocha D, Sánchez B, Ávalos M, Guzmán-Trampe S (2010) Carbon source regulation of antibiotic production. J Antibiot 63(8):442Google Scholar
  56. 56.
    Abouseoud M, Maachi R, Amrane A, Boudergua S, Nabi A (2008) Evaluation of different carbon and nitrogen sources in production of biosurfactant by Pseudomonas fluorescens. Desalination 223(1–3):143–151Google Scholar
  57. 57.
    Aubert-Pivert E, Davies J (1994) Biosynthesis of butirosin in Bacillus circulans NRRL B3312: identification by sequence analysis and insertional mutagenesis of the butB gene involved in antibiotic production. Gene 147(1):1–11Google Scholar
  58. 58.
    Cundliffe E (1992) Self-protection mechanisms in antibiotic producers. Second Metab 11:199–214Google Scholar
  59. 59.
    Wright GD, Berghuis AM, Mobashery S (1998) Aminoglycoside antibiotics. Resolving the antibiotic paradox. Springer, New York, pp 27–69Google Scholar
  60. 60.
    Cundliffe E, Demain AL (2010) Avoidance of suicide in antibiotic-producing microbes. J Ind Microbiol Biotechnol 37(7):643–672Google Scholar
  61. 61.
    Cohen K, Stott K, Manson A, Munsamy V, Earl A, Pym A (2018) Untapped potential for streptomycin: using genomics to optimize aminoglycoside selection in drug-resistant Mycobacterium tuberculosis. A25. Tuberculosis management: new insights. American Thoracic Society, New York, pp A1152–A1152Google Scholar
  62. 62.
    Heifetz C, Chodubski J, Pearson I, Silverman C, Fisher M (1974) Butirosin compared with gentamicin in vitro and in vivo. Antimicrob Agents Chemother 6(2):124–135Google Scholar
  63. 63.
    Vakulenko SB, Mobashery S (2003) Versatility of aminoglycosides and prospects for their future. Clin Microbiol Rev 16(3):430–450Google Scholar
  64. 64.
    Hermann T, Tor Y (2005) RNA as a target for small-molecule therapeutics. Expert Opin Ther Pat 15(1):49–62Google Scholar
  65. 65.
    Becker B, Cooper MA (2012) Aminoglycoside antibiotics in the 21st century. ACS Chem Biol 8(1):105–115Google Scholar
  66. 66.
    Forge A, Schacht J (2000) Aminoglycoside antibiotics. Audiol Neurotol 5(1):3–22Google Scholar
  67. 67.
    Singh SB, Young K, Silver LL (2017) What is an “ideal” antibiotic? Discovery challenges and path forward. Biochem Pharmacol 133:63–73. Google Scholar
  68. 68.
    Selimoglu E (2007) Aminoglycoside-induced ototoxicity. Curr Pharm Des 13(1):119–126Google Scholar
  69. 69.
    Rybak MJ, Abate BJ, Kang SL, Ruffing MJ, Lerner SA, Drusano GL (1999) Prospective evaluation of the effect of an aminoglycoside dosing regimen on rates of observed nephrotoxicity and ototoxicity. Antimicrob Agents Chemother 43(7):1549–1555Google Scholar
  70. 70.
    Prayle A, Watson A, Fortnum H, Smyth A (2010) Side effects of aminoglycosides on the kidney, ear and balance in cystic fibrosis. Thorax 65(7):654–658Google Scholar

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Authors and Affiliations

  1. 1.Molecular Biotechnology Laboratory, Department of Biology, Faculty of ScienceShiraz UniversityShirazIran
  2. 2.Department of Pharmaceutical Chemistry, Institute of Pharmacy, Center for Molecular Biosciences InnsbruckUniversity of Innsbruck, CCB – Centrum for Chemistry and BiomedicineInnsbruckAustria

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