Antibacterial Activity, Cytotoxicity, and the Mechanism of Action of Bacteriocin from <b><i>Bacillus subtilis</i></b> GAS101

Abstract

Objective: The aim of this study was to purify and characterize bacteriocin from the soil isolate Bacillus subtilis GAS101, and to determine its antimicrobial as well as antibiofilm potential. The purified bacteriocin was further analyzed and evaluated for mammalian cell cytotoxicity and the possible mode of action. Material and Methods: Bacteriocin from B. subtilis GAS101 (an animal husbandry soil isolate) was partially purified and checked for antimicrobial and antibiofilm activity against gram-positive and gram-negative bacteria. The molecular weight of bacteriocin was determined using tricine SDS-PAGE gel. The stability of bacteriocin was investigated at various temperatures and pH levels, and its sensitivity towards 8 enzymes and 6 chemicals was determined. Cytotoxicity analysis was performed on a Vero cell line by a tetrazolium dye-based assay. Scanning electron microscopy (SEM) of bacteriocin-treated bacteria was carried out to determine the possible mode of action. Results: Bacteriocin from B. subtilis GAS101 was a potential inhibitor of both the indicator organisms (Staphylococcus epidermidis and Escherichia coli), and had a molecular weight of approximately 6.5 kDa. An in situ gel assay showed a zone of inhibition corresponding to the estimated protein band size. Bacteriocin was stable and showed antibacterial activity in broad ranges of temperature (30–121°C) and pH (2–12). It was sensitive to 4 proteolytic enzymes, which indicated its proteinaceous nature. Bacteriocin showed > 70% cell viability on the mammalian Vero cell line. SEM depicted that the bacteriocin was able to disrupt the bacterial cell membrane as its probable mode of action. Conclusion: Thermostable and pH-tolerant bacteriocin from B. subtilis GAS101, of about 6.5 kDa, showed broad-spectrum antimicrobial and antibiofilm activity.

Identification, Recombinant Expression, and Characterization of LGH2, a Novel Antimicrobial Peptide of Lactobacillus casei HZ1

“…In this paper, we found that LGH2 has a special antimicrobial activity against S. aureus . Different from antibiotics, most AMPs inhibit pathogenic bacteria by interfering with the integrity of the cell membrane [ 34 , 35 ]. In order to understand the mode of LGH2 against S. aureus , we used the dye leakage assays CLSM, SEM, and FCM to analyze the mode of LGH2 against the Gram-positive bacterium S. aureus .…”

CHARACTERIZATION OF ANTIMICROBIAL SUBSTANCE WITH ANTIBIOFILM ACTIVITY FROM Pediococcus acidilactici

“…Cell free supernatant (CFS) was collected by centrifuging overnight grown culture of bacterial isolates at 10,000g/ 15 min/ 4°C. 10µl of CFS was poured in agar well and incubated at 37°C for 16 h and zone of inhibition was measured according to the CLSI guidelines (Sharma, 2018). Each experiment was repeated thrice in triplicates.…”

Bacteriocins and Bacteriophages: Therapeutic Weapons for Gastrointestinal Diseases?

“…They have antimicrobial activity against gram-negative bacteria that infect foods, and this property could be empowered by combining bacteriocins with other compounds (e.g., organic acids, phenolic compounds). Other antimicrobial bacteriocins involved in food protection belong to the class I and II bacteriocins of Bacillus subtilis GAS101 and act against some gram-positive bacterial species such as Staphylococcus Epidermidis [62]. Bacteriocins are overall able to reduce the costs of food treatments and at the same time increase the product shelf-life.…”