Evaluating the Impact of Bacillus subtilis on Salmonella Biofilm Formation: A Co-Culture Study

Abstract

Background: Biofilms are structured bacterial communities that enhance microbial survival and significantly contribute to persistent, antibiotic-resistant infections in clinical settings. Aims: This study aimed at investigating the interaction between Salmonella spp. and Bacillus subtilis (ATCC 6633) during biofilm formation and assessing the potential antagonistic effects of B. subtilis in co-culture systems. Study Design: Quantitative and qualitative analysis of biofilms formed by Salmonella isolates in mono- and dual-species cultures with B. subtilis In vitro. Place and Duration of Study: The study was conducted at the Livestock Laboratory, Department of Livestock Production, Faculty of Agricultural Sciences, Sabaragamuwa University of Sri Lanka. Methodology: Ten Salmonella isolates obtained from the laboratory repository maintained at -800 C were cultured as mono and dual-species biofilms with B. subtilis in 96 wells micro titer plates. Quantification of the biofilm formation was done by using a microtiter plate assay according to the standard protocol. As a qualitative assay, the fluorescence microscopy was used to visualize cellulose components in the biofilm matrices following the Calcofluor white staining procedure. Results: Eight out of ten (8/10) Salmonella isolates exhibited a significant reduction (P < 0.05) in biofilm formation when co-cultured with B. subtilis, with reductions ranging from 21.6% to 64.9%. Fluorescence microscopy confirmed disrupted biofilm architecture and reduced cellulose content in dual-species biofilms compared to the mono species biofilm. Two isolates showed no significant reduction in biofilm formation when co cultured.