Abstract
Extensive usage of antibiotics has led to the emergence of drug-resistant strains of pathogens and hence, there is an urgent need for alternative antimicrobial agents. Antimicrobial Peptides (AMPs) of bacterial origin have shown the potential to replace some conventional antibiotics. In the present study, an AMP was isolated from Bacillus subtilis subsp. spizizenii strain Ba49 present on the Allium cepa, the common onion and named as peptide-Ba49. The isolated AMP was purified and characterized. The purified peptide-Ba49, having a molecular weight of ~ 3.3 kDa as determined using mass spectroscopy, was stable up to 121 °C and in the pH range of 5–10. Its interaction with protein degrading enzymes confirmed the peptide nature of the molecule. The peptide exhibited low minimum inhibitory concentration (MIC) against Staphylococcus aureus and its (Methicillin-resistant Staphylococcus aureus) MRSA strains (MIC, 2–16 µM/mL). Further, time kill kinetic assay was performed and analysis of the results of membrane depolarization and permeabilization assays (TEM, DiBAC4 (3) and PI) suggested peptide-Ba49 to be acting through the change in membrane potential leading to disruption of S. aureus membrane. Additionally, cytotoxicity studies of peptide-Ba49, carried out using three mammalian cell lines viz. HEK 293T, RAW 264.7, and L929, showed limited cytotoxicity on these cell lines at a concentration much higher than its MIC values. All these studies suggested that the AMP isolated from strain Ba49 (peptide-Ba49) has the potential to be an alternative to antibiotics in terms of eradicating the pathogenic as well as drug-resistant microorganisms.
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Abbreviations
- AMPs:
-
Antimicrobial peptides
- MRSA:
-
Methicillin-resistant Staphylococcus aureus
- MIC:
-
Minimum inhibitory concentration
- SEM:
-
Scanning electron microscopy
- TEM:
-
Transmission electron microscopy
- DiBAC4 (3):
-
Bis-(1,3-dibutylbarbituric acid) trimethine oxonol
- PI:
-
Propidium iodide
- PBS:
-
Phosphate buffer saline
- RP-HPLC:
-
Reverse phase-high-pressure liquid chromatography
- Tricine SDS-PAGE:
-
Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis
- MALDI-TOF:
-
Matrix-assisted laser desorption/ionization-time of flight
- NCBI:
-
National Center for Biotechnology Information
- CFU:
-
Colony forming unit
- MTT:
-
3-(4,5-Dimethylthoazole-2-yl)-2,5-diphenyltetrazolium bromide
- DMF:
-
Dimethylformamide
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Acknowledgements
We are thankful to Director, CSIR- IMTECH, for providing all the institutional facilities. A special thanks to Mr. Sanpreet Singh (C/o Dr. Javed Agrewala lab) for helping in flow cytometry and mammalian cytotoxicity experiments and Mr. Randeep Sharma, CSIR-IMTECH, Chandigarh for helping to carry out TEM experiments. Financial support to RT as a recipient of DST INSPIRE Fellowship from Department of Science and Technology, Government of India is highly acknowledged.
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DKS, RT conceived the project. RT and MJ isolated, purified, and characterized the compound. RT performed the in vitro experiments with the assistance of AD. MJ performed the MIC experiments in MRSA strains. KB and PBP analysed the whole genome data of the microorganism. RT, MJ, DKS analysed the data. RT and DKS wrote and edited the manuscript, with input from all of the authors.
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Taggar, R., Jangra, M., Dwivedi, A. et al. Bacteriocin isolated from the natural inhabitant of Allium cepa against Staphylococcus aureus. World J Microbiol Biotechnol 37, 20 (2021). https://doi.org/10.1007/s11274-020-02989-x
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DOI: https://doi.org/10.1007/s11274-020-02989-x