These results illuminate the overlooked clinical manifestation of persistent salmonellosis that can serve as a human reservoir for NTS infections. Additionally, we demonstrate mechanisms of in-host microevolution and exhibit their potential to shape Salmonella pathogenicity, antimicrobial resistance and host-pathogen interactions.

c Salmonella enterica serovar Typhimurium is a facultative intracellular human and animal bacterial pathogen posing a major threat to public health worldwide. Salmonella pathogenicity requires complex coordination of multiple physiological and virulence pathways. DksA is a conserved Gram-negative regulator that belongs to a distinct group of transcription factors that bind directly to the RNA polymerase secondary channel, potentiating the effect of the signaling molecule ppGpp during a stringent response. Here, we established that in S. Typhimurium, dksA is induced during the logarithmic phase and DksA is essential for growth in minimal defined medium and plays an important role in motility and biofilm formation. Furthermore, we determined that DksA positively regulates the Salmonella pathogenicity island 1 and motility-chemotaxis genes and is necessary for S. Typhimurium invasion of human epithelial cells and uptake by macrophages. In contrast, DksA was found to be dispensable for S. Typhimurium host cell adhesion. Finally, using the colitis mouse model, we found that dksA is spatially induced at the midcecum during the early stage of the infection and required for gastrointestinal colonization and systemic infection in vivo. Taken together, these data indicate that the ancestral stringent response regulator DksA coordinates various physiological and virulence S. Typhimurium programs and therefore is a key virulence regulator of Salmonella. Salmonella enterica is a facultative intracellular human and animal bacterial pathogen responsible for global pandemics of food-borne infections that pose a major threat to public health. This highly versatile pathogen can infect a broad range of hosts and causes different clinical outcomes, ranging from asymptomatic carriage to systemic life-threatening disease (1). The single species S. enterica includes more than 2,600 serovars that share high sequence similarity and are taxonomically classified into six subspecies (2). Estimations suggest that Salmonella causes 93.8 million human infections and 155,000 deaths annually worldwide (3). The majority of nontyphoidal Salmonella (NTS) infections in humans present as gastroenteritis; however, about 5% may be invasive and manifest as bacteremia or other extraintestinal infections (4). Many of the NTS serovars are capable of colonizing the intestines of livestock with potential risk of contaminating the food chain, and therefore, salmonellosis is often associated with animal products and produce (5). One of the most common serovars worldwide is S. enterica serovar Typhimurium, ranking first in prevalence in North America and Oceania (6).Salmonella intestinal colonization is a complex phenotype essential for establishing disease. Salmonella colonization requires synchronized function of both conserved and host-specific colonization factors such as fimbrial adhesins, invasion factors (e.g., SiiE, MisL, and ShdA) and genes encoded within Salmonella pathogenicity island 1 (SPI-1) and SPI-2 (reviewed in references 7 and 8). Both SPIs e...

Salmonella enterica serovars Typhi and Paratyphi A are human-restricted pathogens and the leading causative agents of enteric fever. The Typhi colonization factor (Tcf) is a chaperone-usher fimbria, thought to play a role in the host-specificity of typhoidal serovars. Here we show that the tcf cluster (tcfABCD tinR tioA) is present in at least 25 non-typhoidal Salmonella (NTS) serovars and demonstrate its native expression in clinically-important serovars including Schwarzengrund, 9,12:l,v:-, Choleraesuis, Bredeney, Heidelberg, Montevideo, Virchow and Infantis. Although the genetic organization of the tcf cluster is well conserved, the N-terminal half of the fimbrial adhesin, TcfD is highly diverse, suggesting different binding properties of distinct tcfD variants. Comparison of tcfA expression in typhoidal and NTS serovars demonstrated unexpected differences in its expression profiles, with the highest transcription levels in S. Typhi, S. Choleraesuis and S. Infantis. In the latter, tcf is induced in rich broth and under microaerobic conditions, characterizing the intestines of warm blooded animals. Furthermore, Tcf is negatively regulated by the ancestral leucine-responsive transcriptional regulator (Lrp). Using the colitis mouse model, we demonstrate that during mice infection tcfA is expressed at higher levels by S. Infantis than S. Schwarzengrund or S. Heidelberg. Moreover, while Tcf is dispensable for S. Schwarzengrund and S. Heidelberg mouse colonization, Tcf is involved in cecum and colon colonization by S. Infantis. Taken together, our results establish that Tcf is broadly encoded by multiple NTS serovars, but presents variable expression profiles and contributes differently to their virulence.