Temperature Adaptive Biofilm Formation in Yersinia enterocolitica in Response to pYV Plasmid and Calcium

<b>Background/Objectives</b>: <i>Yersinia enterocolitica</i> is a pathogenic bacterium that forms biofilms, enhancing its persistence and resistance to antimicrobial agents. Biofilm formation in <i>Y. enterocolitica</i> is influenced by environmental factors such as temperature, calcium, and the presence of the virulence plasmid pYV. This study aims to explore how temperature, calcium, and pYV modulate biofilm formation in <i>Y. enterocolitica</i>, with a focus on motility and extracellular polymeric substance (EPS) production as key factors. <b>Methods</b>: <i>Y. enterocolitica</i> strains with and without the pYV plasmid were cultured at two different temperatures (26 °C and 37 °C). The effect of calcium (5 mM) on biofilm formation was tested at both temperatures. Biofilm formation was measured using crystal violet staining, motility was assessed using soft agar plates, and EPS production was quantified to determine its role in biofilm stabilization. <b>Results</b>: At 26 °C, biofilm formation increased in pYV-negative strains, driven primarily by motility and flagellar expression. In contrast, at 37 °C, pYV-positive strains showed strong biofilm formation despite reduced growth, with EPS production as the key stabilizing factor. Calcium modulated biofilm formation in a temperature-dependent manner: at 26 °C, 5 mM calcium modestly reduced biofilm formation in pYV-negative strains, while at 37 °C, it significantly suppressed both EPS production and biofilm formation by approximately 50% in pYV-positive strains. <b>Conclusions</b>: This study reveals a novel regulatory switch where temperature, calcium, and pYV modulate biofilm formation in <i>Y. enterocolitica</i>. These findings suggest that <i>Y. enterocolitica</i> can adapt between motility- and EPS-dominated biofilm strategies depending on environmental conditions. Understanding these mechanisms offers potential targets for controlling biofilm-related persistence in clinical and food safety contexts.