Abstract
A heterotrophoic Bacillus amyloliquefaciens MTCC12713 isolated from an intertidal macroalga Kappaphycus alverezii displayed promising antibacterial activities against multidrug-resistant bacteria. Genome mining of the bacterium predicted biosynthetic gene clusters coding for antibacterial secondary metabolites. Twenty-one membered macrocyclic lactones, identified as difficidin analogues bearing 6-hydroxy-8-propyl carboxylate, 9-methyl-19-propyl dicarboxylate, 6-methyl-9-propyl dicarboxylate-19-propanone, and (20-acetyl)-6-methyl-9-isopentyl dicarboxylate (compounds 1 through 4) functionalities were purified through bioassay-guided fractionation. The difficidin analogues exhibited bactericidal activities against vancomycin-resistant Enterococcus faecalis, methicillin-resistant Staphylococcus aureus, and other drug-resistant strains, such of Klebsiella pneumonia and Pseudomonas aeruginosa with the minimum inhibitory concentration of about 2–9 × 10−3 μM. A plausible enzyme-catalyzed biosynthetic pathway that is generated through addition of acrylyl initiator unit by repetitive decarboxylative Claisen condensation modules with malonate units was recognized, and their structures were corroborated with gene organization of the dif operon, which could comprehend dif A-O (~ 70 kb). Drug-likeness score for 5-ethoxy-28-methyl-(9-methyl-19-propyl dicarboxylate) difficidin (compound 2, 0.35) was greater than those of other difficidin analogues, which corroborated the potential in vitro antibacterial properties of the former. The present study demonstrated the potential of difficidin analogues for pharmaceutical and biotechnological uses against the bottleneck of emergent drug-resistant pathogens.
Key Points
• Difficidins were isolated from marine alga associated Bacillus amyloliquefaciens.
• Whole-genome mining of bacterial genome predicted biosynthetic gene clusters.
• Greater drug-likeness for difficidin 2 confirmed its potent antibacterial activity.
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Acknowledgements
This work was funded by the Indian Council of Agricultural Research-Central Marine Fisheries Research Institute (ICAR-CMFRI), India, under the project titled as “Development of Bioactive Pharmacophores from Marine Organisms” (grant number MBT/HLT/SUB23). The authors thank the Director, ICAR-CMFRI, and Head, Marine Biotechnology Division of ICAR-CMFRI for guidance and support. The authors are thankful to the Dean, Faculty of Marine Sciences, Lakeside Campus, Cochin University of Science and Technology for providing with necessary support.
Data availability statement
All data generated or analyzed during this study are included in this published article and its supplementary information files. The whole-genome sequence of the candidate bacterium was submitted in the GenBank with an accession number of QKQQ00000000 (biosample code: SAMN09389114; link for publicly available data is as follows: https://www.ncbi.nlm.nih.gov/search/all/?term=QKQQ00000000).
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KC conceived and designed research, acquired funds, conducted experiments, and performed structural analysis. KC and VKK contributed new reagents or analytical tools, analyzed data, and drafted the manuscript. MJ performed purification of the compounds and structural analysis. SD carried out statistical analysis of the data. All authors read and approved the manuscript.
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Chakraborty, K., Kizhakkekalam, V.K., Joy, M. et al. Difficidin class of polyketide antibiotics from marine macroalga-associated Bacillus as promising antibacterial agents. Appl Microbiol Biotechnol 105, 6395–6408 (2021). https://doi.org/10.1007/s00253-021-11390-z
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DOI: https://doi.org/10.1007/s00253-021-11390-z