Human Bile-Mediated Regulation of Salmonella Curli Fimbriae

Abstract

Typhoid fever is caused primarily by Salmonella enterica serovar Typhi. Approximately 3% to 5% of individuals infected with S Typhi become chronic carriers with the gallbladder (GB) as the site of persistence, as gallstones within the GB are a platform on which the bacteria form a biofilm. S Typhi is a human-restricted pathogen; therefore, asymptomatic carriers represent a critical reservoir for further spread of disease. To examine the dynamics of the Salmonella biofilm during chronic carriage, the human gallstone (GS) environment was simulated by growing biofilms on cholesterol-coated surfaces in the presence of bile, and the transcriptional profile was determined. Some of the most highly activated genes corresponded to the curli fimbria operon, with the major structural component csgA upregulated >80-fold. The curli protein polymer is a major component of the extracellular matrix (ECM) in Salmonella biofilms. The upregulation of curli fimbriae by human bile was validated through reverse transcription-quantitative PCR (qRT-PCR), microscopy, and Western blotting. Interestingly, this activation appears human specific, as qRT-PCR showed repression of csgA in biofilms grown in mouse or ox bile. Comparative transcriptional studies of the two divergent csg operons suggest an early activation of both operons in minimal medium complemented with glucose that quickly diminishes as the biofilm matures. However, in the presence of human bile, there is a modest activation of both operons that steadily increases as the biofilm matures. Understanding the effect of the GB environment on key biofilm-associated factors can help target antibiofilm therapeutics or other preventative strategies to eradicate chronic carriage.IMPORTANCE Typhoid fever is caused by Salmonella enterica serovar Typhi, and 3% to 5% of patients become chronic gallbladder (GB) carriers (also known as "Typhoid Marys"). We have previously demonstrated a role for Salmonella biofilm formation on gallstones as a primary mechanism of carriage. In this study, we found that the important biofilm extracellular matrix component curli fimbria is induced in an in vitro human GB model system. This induction is specific to human bile and increases as the biofilm matures. We also found that the biofilm and curli regulator CsgD play a key role in this observed induction. This work further enhances our understanding biofilm-mediated chronic carriage and provides a potential target for eliminating persistent GB infection by S Typhi.

Keywords: Salmonella; bile; biofilm; chronic carriage; curli fimbriae.