Serotype-Specific and Temperature-Dependent Biofilm Formation in Salmonella: Limited Impact of Antimicrobial Resistance or Source

Salmonella is a major zoonotic and foodborne pathogen whose ability to form biofilms enhances antimicrobial tolerance and environmental persistence. This study aimed to investigate how temperature-dependent biofilm formation is influenced by serotype, multidrug resistance (MDR) status, and source of isolation. A total of 114 Salmonella isolates from food and humans (including clinical cases and asymptomatic carriers) were assessed for biofilm-forming capacity. Biofilms were stained with crystal violet and quantified by measuring the OD 570 after incubation at 15 °C, 25 °C, and 37 °C for 3 days. Isolates were categorized into weak, moderate, and strong biofilm formers. Associations with serotype, MDR status, and isolation source were analyzed using ANOVA and multiple regression. PCR screening was performed to examine the distribution of 28 biofilm-related genes. Biofilm formation peaked at 25°C across all sources, indicating a strong temperature effect. Distinct serotype-specific patterns of biofilm capacity were observed, while antimicrobial resistance and source showed no significant associations. Statistical analysis identified temperature (F = 36.46, p = 6.40 × 10 -15 ) and serotype (F = 3.31, p = 6.91 × 10 -8 ) as primary determinants of biofilm variability. Among the 28 genes screened, 24 were universally present, whereas the distribution of rcK , fimH , steB , and pefA was serotype-dependent but independent of extent of biofilm formation. Biofilm formation in Salmonella was primarily driven by temperature and serotype, while no significant association was observed with antimicrobial resistance or isolation source in this study. These findings provide new insights into the ecological adaptation of Salmonella biofilms and have implications for controlling persistence and transmission under fluctuating environmental conditions. • Biofilm formation in Salmonella is enhanced at 25°C across all sources • Serotype and temperature mainly determine biofilm-forming capacity • Antimicrobial resistance and source show little effect impact on biofilm formation • Most biofilm genes (23/28) are universally present in all Salmonella isolates • Genes like rck and fimH are serotype-linked, not related to biofilm strength