The T6SS effector Tldi1 enhances Salmonella enterica serovar Typhimurium virulence by regulating motility and stress adaptation

The type VI secretion system (T6SS) enhances stress tolerance, antimicrobial resistance in Gram-negative bacteria, contributing to chronic infections and complicating clinical management. Understanding its regulatory mechanisms governing bacterial persistence and pathogenicity has become a critical research priority. Tldi1 is an immune protein encoded by Salmonella enterica serovar Typhimurium, serving as a “protector” of T6SS function. Its existence is a prerequisite for the safe deployment of effector proteins and efficient competition in bacteria, but little is known about its functional significance. In this study, we generated Tldi1-knockout and complemented strains to elucidate its role and compared their phenotypic traits with those of wild-type bacteria. In vitro assays demonstrated that Tldi1 contributes to bacterial fitness by modulating motility, biofilm formation, and stress responses (including resistance to acid/alkali, osmotic pressure, ethanol, and bile salts). In a murine infection model, Tldi1-deficient Salmonella exhibited attenuated virulence, resulting in prolonged host survival, reduced bacterial loads (liver, spleen, cecum), and less severe histopathological damage. Furthermore, Tldi1 deletion suppressed the production of proinflammatory cytokines (IL-12, IL-15, IL-18) in infected tissues, suggesting its involvement in immune modulation. Tldi1 also influences the diversity and structure of the mouse gut microbiota and plays a crucial role in pathogenicity. Collectively, our findings demonstrate that Tldi1 enhances Salmonella pathogenicity by promoting environmental adaptability and host inflammatory responses. These results provide mechanistic insights into T6SS-mediated virulence and highlight Tldi1 as a potential therapeutic target for anti- Salmonella interventions.