Effects of Mn2+ and Ca2+ on biofilm formation and signal molecule AI-2 activity in Lactobacillus fermentum 733 during fermentation: A proteomic analysis

Biofilm formation in bacteria is influenced by several environmental factors, including the presence of divalent cations. This study examined the impact of Mn 2+ and Ca 2+ on biofilm formation and quorum sensing in Lactobacillus fermentum 733. The findings revealed that 0.15 mg/mL MnCl 2 and 0.1 mg/mL CaCl 2 can significantly enhance biofilm production, AI-2 signal molecule levels, extracellular polysaccharide content, cell hydrophobicity, and self-aggregation. Both Mn 2+ and Ca 2+ , added individually or in combination with each other, upregulated the expression of LuxS , DapB , LamC , and FtsH while downregulating PfsF . Specifically, Ca 2+ primarily modulated LuxS (involved in AI-2 production) and LamC (linked to biofilm formation), whereas Mn 2+ predominantly influenced biofilm-related genes such as DapB and FtsH . Proteomic analysis revealed that Mn 2+ , both alone and in combination with Ca 2+ , significantly affected bacterial growth, metabolism, and catalytic activity. Meanwhile, Ca 2+ alone had minimal impact on protein expression. These results underscore the distinct regulatory roles of Mn 2+ and Ca 2+ in quorum sensing and biofilm formation in lactic acid bacteria, offering valuable insights into the mechanisms underlying these processes. • Divalent Cations Influence Quorum Sensing and Biofilm Formation : Mn 2+ and Ca 2+ enhance biofilm formation and AI-2 production in L. fermentum 733. • Gene Expression Modulation : Upregulate genes LuxS , DapB , LamC , FtsH , and downregulate PfsF. • Distinct Roles of Mn 2+ and Ca 2+ : Ca 2+ mainly affects LuxS and LamC ; Mn 2+ targets DapB and FtsH. • Proteomic Insights : Mn 2+ impacts growth, metabolism; Ca 2+ has minimal effect on proteins. • Synergistic Effects : Combined Mn 2+ and Ca 2+ boost growth and metabolic activity synergistically.