Isolation of Secondary Metabolites with Antimicrobial Activities from Bacillus amyloliquefaciens LWYZ003

Abstract

The strain LWYZ003, which can restrain multiple pathogens, was screened from the sediment of the ocean and identified as Bacillus amyloliquefaciens. Large-scale fermentation and modern chromatographic separation technologies (macroporous resin column chromatography, silica gel column chromatography, thin-layer chromatography and high-performance liquid chromatography) were used to separate antimicrobial products from the fermentation broth of marine-derived Bacillus amyloliquefaciens LWYZ003. Bioactive-guided separation was used in the term of seeking antimicrobial products from the secondary metabolites of Bacillus amyloliquefaciens LWYZ003. As a result, two natural products cycloheximide (1) and trehalose (2) were obtained. Their structures were elucidated by Fourier transform infrared spectroscopy, high-resolution mass spectrometry, ¹H and ¹³C nuclear magnetic resonance analysis. In the cylinder plate method, compound 1 exhibited stronger antimicrobial activities than compound 2 against Micrococcus luteus, and also exhibited wider antimicrobial spectrum than compound 2. In conclusion, isolation of bioactive secondary metabolites from marine Bacillus sp. has enormous potentials in finding suitable antibiotics to inhibit multiple pathogens.

Keywords

Cycloheximide / Trehalose / Bacillus amyloliquefaciens / Bioactivity-guided isolation / Antibiotic

References

[1] Manivasagan P, Venkatesan J, Sivakumar K, et al. Marine actinobacterial metabolites and their pharmaceutical potential. Springer handbook of marine biotechnology, 2015, Heidelberg: Springer.

[2] Hamdache A, Lamarti A, Aleu J, et al. Non-peptide metabolites from the genus bacillus. Nat Prod, 2011, 74(4): 893-899.

[3] Carlin F. Origin of bacterial spores contaminating foods. Food Microbiol, 2011, 28(2): 177-182.

[4] Han Y, Fan J, Zhou Z, et al. Cloning and efficient expression of bacillus sp. BH072 tasA gene in Escherichia coli. Trans Tianjin Univ, 2015, 21(1): 26-31.

[5] Tan X, Han Y, Xiao H, et al. Pediococcus acidilactici, inhibit biofilm formation of food-borne pathogens on abiotic surfaces. Trans Tianjin Univ, 2017, 23(1): 70-77.

[6] Zhao Y, Liang YQ, Liu L, et al. Medium optimization for antifungal active substance production from Streptomyces lydicus using response surface methodology. Trans Tianjin Univ, 2017, 23(1): 78-86.

[7] Lu C, Yin J, Zhang C, et al. Fed-batch fermentation for spinosad production in an improved reactor. Trans Tianjin Univ, 2017, 23(1): 1-8.

[8] Tan X, Han Y, Xiao H, et al. Green fluorescent protein recombinant nisin as a probe for detection of gram-positive bacteria. Trans Tianjin Univ, 2017, 23(4): 1-6.

[9] Feng Y, Fang Z, Wang H, et al. Synthesis and characterization of degraded gelatin grafted poly (ε-caprolactone) copolymers. Trans Tianjin Univ, 2013, 19(3): 182-187.

[10] Shoda M. Bacterial control of plant diseases. J Biosci Bioeng, 2000, 89(6): 515-521.

[11] Coton M, Denis C, Cadot P, et al. Biodiversity and characterization of aerobic spore-forming bacteria in surimi seafood products. Food Microbiol, 2011, 28(2): 252-260.

[12] Brownlee KA, Delves CS, Dorman M, et al. The biological assay of streptomycin by a modified cylinder plate method. J Gen Microbiol, 1948, 2(1): 40-53.

[13] Perry NB, Blunt JW, Munro MHG. Different solution and solid-state conformations of the antibiotic cycloheximide. Magn Reson Chem, 2011, 27(7): 624-627.

[14] Garcia RC, Cruz-Sánchez S, Méndez R, et al. ¹H and ¹³C NMR study of nonsymmetrical α, α -trehalose derivatives. Carbohyd Res, 1992, 237: 283-287.

[15] Huang SX, Yu Z, Robert F, et al. Cycloheximide and congeners as inhibitors of eukaryotic protein synthesis from endophytic actinomycetes Streptomyces sps. YIM56132 and YIM56141. J Antibiot, 2011, 64(1): 163-166.

[16] Choudhary SK, Walker RM, Powell DM, et al. CXCR4 tropic human immunodeficiency virus type 1 induces an apoptotic cascade in immature infected thymocytes that resembles thymocyte negative selection. Virology, 2006, 352(2): 268-284.

[17] Crance JM, Biziagos E, Passagot J, et al. Inhibition of hepatitis A virus replication in vitro by antiviral compounds. J Med Virol, 1990, 31(2): 155-160.

[18] Widholm JM. Trehalose as cryoprotectant for the freeze preservation of carrot and tobacco cells. Plant Physiol, 1985, 78(2): 430-432.

[19] Crowe JH, Crowe LM, Jackson SA. Preservation of structural and functional activity in lyophilized sarcoplasmic reticulum. Arch Biochem Biophys, 1983, 220(2): 477-484.

[20] Kurenda A, Pieczywek PM, Adamiak A, et al. Effect of cytochalasin B, latrunculin B, colchicine, cycloheximide, dimethyl sulfoxide and ion channel inhibitors on biospeckle activity in apple tissue. Food Biophys, 2013, 8(4): 290-296.

[21] Alvarez F, Castro M, Príncipe A, et al. The plant-associated Bacillus amyloliquefaciens strains MEP218 and ARP23 capable of producing the cyclic lipopeptidesiturin or surfactin and fengycin are effective in biocontrol of sclerotinia stem rot disease. J Appl Microbiol, 2012, 112(1): 159-174.

[22] Yin M, Yan Y, Lohman JR, et al. Cycloheximide and actiphenol production in Streptomyces sp. YIM56141 governed by single biosynthetic machinery featuring an acyltransferase-less type I polyketide synthase. Org Lett, 2014, 16(11): 3072-3075.

[23] Liu XM, Huang CY, Chen Q, et al. Study on the metabolic pathway of trehalose in pleurotus pulmonarius during heat stress recovery. Scientia Agricultura Sinica, 2013, 46(24): 5188-5195.