Evaluation of Antibacterial and Antiadenoviral Activity of Bacillus Clausii Supernatant and Its Effect on the Expression of Apoptotic Regulatory Genes Bax, Bcl-2 and MiR-145 in HeLa Cancer Cells
Evaluation of Antibacterial and Antiadenoviral Activity of Bacillus Clausii Supernatant and Its Effect on the Expression of Apoptotic Regulatory Genes Bax, Bcl-2 and MiR-145 in HeLa Cancer Cells
Zahra Fateminasab,1,*Mohammad Shayestehpour,2Mohammad Reza Zolfaghari,3
1. Department of Microbiology, Qom Branch, Islamic Azad University, Qom, Iran 2. Autoimmune Diseases Research Center, Kashan University of Medical Sciences, Kashan, Iran 3. Department of Microbiology, Qom Branch, Islamic Azad University, Qom, Iran
Introduction: Bacillus clausii is being studied as a probiotic candidate. There is insufficient information on the antimicrobial and anticancer effects of B. clausii cell-free supernatant. The current study aimed to evaluate the antibacterial and antiadenoviral activity of B. clausii supernatant and its effect on the expression of apoptotic regulatory genes bax, Bcl-2 and miR-145 in HeLa cancer cells.
Methods: First, the cell-free supernatant (CFS) was prepared from a 24-hour bacterial culture, then its antibacterial effect on several gram-positive and gram-negative bacteria was evaluated using the microdilution method to calculate the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). Adenovirus 5 (Ad5) espouse to the CFS under pre-treatment (incubation of cells with the CFS followed by Ad5 inoculation), pre-incubation (a mixture of co-incubated CSF/Ad5 added to cells), competition (adding Ad5 and CFS into cells at the same time) and post-treatment (cells inoculated with Ad5 and subsequently incubated with CFS) assays. Titer of virus and E1A level calculated by tissue culture infectious dose 50 (TCID50) and real-time polymerase chain reaction (real-time PCR) methods, respectively. After exposure to the CFS, expression levels of Bax, Bcl-2 and MiR-145 in HeLa cancer cells were evaluated using a real-time PCR method.
Results: B. clausii supernatant showed antibacterial activity against E. faecalis, S. aureus, MRSA, E. coli, P. aeruginosa, and A. baumannii. Ad5 titers in pre-treatment, pre-incubation, competition, and post-treatment assays with CFS (dilution: ) decreased by about 4.61, 4, 3.9, and 3.1 Log10 TCID50/ml in comparison to control, respectively. Comparison of E1A expression levels between experimental assays and control showed similar findings of the viral titration. B. clausii supernatant during 48 h exposure to HeLa cells increased the expression level of Bax, Bcl-2 and miR-145 genes to 9.1, 2.3 and 55-fold, respectively, in comparison to the untreated condition
Conclusion: The supernatant of B. clausii had an antibacterial effect against some gram-positive and gram-negative bacteria. It inhibits replication of Ad5 and increases the bax/Bcl-2 ratio in cervical cancer cell line. The bacterial supernatant upregulates the tumor suppressor miR-145. Therefore, B. clausii can be considered as a potent antimicrobial and anticancer agent. However, further studies are needed to determine its spectrum of antibacterial, antiviral, and anticancer effects.