Dispersal of pathogen-associated multispecies biofilm by novel probiotic Bacillus subtilis in a contact-dependent manner.

Abstract

Biofilms are involved in pathogenesis of various bacterial infections. Treatment of biofilm-related bacterial infection remains a major challenge due to the reduced efficacy of antibiotics and associated antibiotic resistance. Given the high prevalence of Enterotoxigenic Escherichia coli (ETEC), Salmonella Typhimurium (S. Typhimurium) and methicillin-resistant Staphylococcus aureus (MRSA)-related infections and associated drug resistance, it is imperative to develop alternative strategies for treatment and prevention. The current study investigated antibiofilm activity of a recently isolated Bacillus subtilis (B. subtilis-9) against these pathogens. Crystal violet staining showed that treatment with B. subtilis-9 significantly reduced biofilm biomass of ETEC (60%-80%), S. Typhimurium (68%-73%) and MRSA (66%-82%). In addition, B. subtilis-9 significantly reduced pre-formed biofilm biomass of ETEC (59%), S. Typhimurium (62%), MRSA (65%) and multispecies (58%). Fluorescence microscopy revealed that B. subtilis-9 treatment significantly reduced the thickness of biofilm and viability of the embedded bacteria. Additionally, B. subtilis-9 significantly reduced planktonic cell growth of ETEC (92%), S. Typhimurium (94%) and MRSA (93%). Interestingly, transwell assay showed that B. subtilis-9 exhibited antibiofilm properties in a cell-to-cell contact-dependent manner and significantly reduced mRNA expression of biofilm-related genes, bssS, luxS and ihfB in ETEC. Novel B. subtilis-9 exhibits a strong inhibitory activity against ETEC, S. Typhimurium and MRSA biofilm formation and adhesion to abiotic surfaces. With further investigations, our study could bring forward a novel Bacillus-based probiotic intervention strategy to combat pathogenic biofilms, in clinical and agricultural settings. Probiotic bacteria propose a potential alternative in combating biofilm-related infections, however, data on the efficacy and strain selection are limited. Data from this study are critical in further developing Bacillus-based novel probiotic applications that may reduce the use of antibiotics in biofilm-related infections in humans and animals.