Factors Required for Adhesion of Salmonella enterica Serovar Typhimurium to Corn Salad (Valerianella locusta)

Abstract

Salmonella enterica is a foodborne pathogen often leading to gastroenteritis and is commonly acquired by consumption of contaminated food of animal origin. However, frequency of outbreaks linked to the consumption of fresh or minimally processed food of nonanimal origin is increasing. New infection routes of S. enterica by vegetables, fruits, nuts, and herbs have to be considered. This leads to special interest in S. enterica interactions with leafy products, e.g., salads, that are mainly consumed in a minimally processed form. The attachment of S. enterica to salad is a crucial step in contamination, but little is known about the bacterial factors required and mechanisms of adhesion. S. enterica possesses a complex set of adhesive structures whose functions are only partly understood. Potentially, S. enterica may deploy multiple adhesive strategies for adhering to various salad species and other vegetables. In this study, we systematically analyzed the contributions of the complete adhesiome, of lipopolysaccharide (LPS), and of flagellum-mediated motility of S. enterica serovar Typhimurium (STM) in adhesion to Valerianella locusta (corn salad). We deployed a reductionist, synthetic approach to identify factors involved in the surface binding of STM to leaves of corn salad, with particular regard to the expression of all known adhesive structures, using the Tet-on system. This work reveals the contribution of Saf fimbriae, type 1 secretion system-secreted BapA, an intact LPS, and flagellum-mediated motility of STM in adhesion to corn salad leaves. IMPORTANCE Transmission of gastrointestinal pathogens by contaminated fresh produce is of increasing relevance to human health. However, the mechanisms of contamination of, persistence on, and transmission by fresh produce are poorly understood. We investigated the contributions of the various adhesive structures of STM to the initial event in transmission, i.e., binding to the plant surface. A reductionist system was used that allowed experimentally controlled surface expression of individual adhesive structures and analyses of the contribution to binding to leave surfaces of corn salad under laboratory conditions. The model system allowed the determination of the relative contributions of fimbrial and nonfimbrial adhesins, the type 3 secretion systems, the O antigen of lipopolysaccharide, the flagella, and chemotaxis of STM to binding to corn salad leaves. Based on these data, future work could reveal the mechanism of binding and the relevance of interaction under agricultural conditions.

Factors required for adhesion ofSalmonella entericaserovar Typhimurium to lettuce (Lactuca sativa)

“…Alleles of bapA with deletions of BIg1, BIg28, or BIg1-28 were expressed by AHT induction. Synthesis and secretion of mutant forms of BapA were confirmed by flow cytometry in prior work (28), and revealed that deletion of BIg1-28 ablated surface expression of BapA. The deletion of BIg1 showed a similar significantly increased adhesion as observed before for full-length BapA, indicating no contribution of BIg1 in adhesion to lettuce.…”

“…In a previous study (28) we deployed a reductionist, synthetic approach to identify factors that contribute to the surface binding of STM to leaves of corn salad. We now investigate factors that contribute to surface binding of STM to leaves of lettuce.…”

“…Enhanced surface expression of SiiE was achieved by AHT-induced expression of hilD , the central transcriptional activator of SPI1/SPI4 genes (31), as generation of a vector for Tet-on expression of the sii operon failed. The frequency of SiiE-expressing STM increased from approximately 12% under native conditions, to over 80% after AHT-induced hilD expression (28). The increased surface expression of SiiE in SR11 Δ12 correlated with significantly decreased adhesion to lettuce (Figure 3A), while a SPI4-deletion strain showed significantly increased adhesion.…”

“…AHT-induced expression of bapABCD led to significantly increased adhesion to lettuce, whereas deletion of bapABCD did not affect adhesion (Figure 3B). To achieve a better understanding of BapA binding to lettuce leaves, we used various truncated forms of BapA (28). Alleles of bapA with deletions of BIg1, BIg28, or BIg1-28 were expressed by AHT induction.…”

“…Therefore, we used a synthetic system based on the tetA promoter for controlled expression of various STM adhesins (23). We tested the adhesive structures for involvement in binding to lettuce leave surfaces by a reductionist approach described before (28). Our data will support the classification of adhesive structures commonly involved in binding to salad species, in order to subsequently develop strategies for prevention or reduction of adhesion of S. enterica to salad leaves.…”