LsrR-Mediated Quorum Sensing Controls Invasiveness of Salmonella typhimurium by Regulating SPI-1 and Flagella Genes

Bacterial cell-to-cell communication, termed quorum sensing (QS), controls bacterial behavior by using various signal molecules. Despite the fact that the LuxS/autoinducer-2 (AI-2) QS system is necessary for normal expression of Salmonella pathogenicity island-1 (SPI-1), the mechanism remains unknown. Here, we report that the LsrR protein, a transcriptional regulator known to be involved in LuxS/AI-2-mediated QS, is also associated with the regulation of SPI-1-mediated Salmonella virulence. We determined that LsrR negatively controls SPI-1 and flagella gene expressions. As phosphorylated AI-2 binds to and inactivates LsrR, LsrR remains active and decreases expression of SPI-1 and flagella genes in the luxS mutant. The reduced expression of those genes resulted in impaired invasion of Salmonella into epithelial cells. Expression of SPI-1 and flagella genes was also reduced by overexpression of the LsrR regulator from a plasmid, but was relieved by exogenous AI-2, which binds to and inactivates LsrR. These results imply that LsrR plays an important role in selecting infectious niche of Salmonella in QS dependent mode

Crosstalk between Virulence Loci: Regulation of Salmonella enterica Pathogenicity Island 1 (SPI-1) by Products of the std Fimbrial Operon

Invasion of intestinal epithelial cells is a critical step in Salmonella infection and requires the expression of genes located in Salmonella pathogenicity island 1 (SPI-1). A key factor for SPI-1 expression is DNA adenine (Dam) methylation, which activates synthesis of the SPI-1 transcriptional activator HilD. Dam-dependent regulation of hilD is postranscriptional (and therefore indirect), indicating the involvement of unknown cell functions under Dam methylation control. A genetic screen has identified the std fimbrial operon as the missing link between Dam methylation and SPI-1. We show that all genes in the std operon are part of a single transcriptional unit, and describe three previously uncharacterized ORFs (renamed stdD, stdE, and stdF). We present evidence that two such loci (stdE and stdF) are involved in Dam-dependent control of Salmonella SPI-1: in a Dam− background, deletion of stdE or stdF suppresses SPI-1 repression; in a Dam+ background, constitutive expression of StdE and/or StdF represses SPI-1. Repression of SPI-1 by products of std operon explains the invasion defect of Salmonella Dam− mutants, which constitutively express the std operon. Dam-dependent repression of std in the ileum may be required to permit invasion, as indicated by two observations: constitutive expression of StdE and StdF reduces invasion of epithelial cells in vitro (1,000 fold) and attenuates Salmonella virulence in the mouse model (>60 fold). In turn, crosstalk between std and SPI-1 may play a role in intestinal infections by preventing expression of SPI-1 in the caecum, an intestinal compartment in which the std operon is known to be expressed

The Cost of Virulence: Retarded Growth of Salmonella Typhimurium Cells Expressing Type III Secretion System 1

Virulence factors generally enhance a pathogen's fitness and thereby foster transmission. However, most studies of pathogen fitness have been performed by averaging the phenotypes over large populations. Here, we have analyzed the fitness costs of virulence factor expression by Salmonella enterica subspecies I serovar Typhimurium in simple culture experiments. The type III secretion system ttss-1, a cardinal virulence factor for eliciting Salmonella diarrhea, is expressed by just a fraction of the S. Typhimurium population, yielding a mixture of cells that either express ttss-1 (TTSS-1+ phenotype) or not (TTSS-1− phenotype). Here, we studied in vitro the TTSS-1+ phenotype at the single cell level using fluorescent protein reporters. The regulator hilA controlled the fraction of TTSS-1+ individuals and their ttss-1 expression level. Strikingly, cells of the TTSS-1+ phenotype grew slower than cells of the TTSS-1− phenotype. The growth retardation was at least partially attributable to the expression of TTSS-1 effector and/or translocon proteins. In spite of this growth penalty, the TTSS-1+ subpopulation increased from <10% to approx. 60% during the late logarithmic growth phase of an LB batch culture. This was attributable to an increasing initiation rate of ttss-1 expression, in response to environmental cues accumulating during this growth phase, as shown by experimental data and mathematical modeling. Finally, hilA and hilD mutants, which form only fast-growing TTSS-1− cells, outcompeted wild type S. Typhimurium in mixed cultures. Our data demonstrated that virulence factor expression imposes a growth penalty in a non-host environment. This raises important questions about compensating mechanisms during host infection which ensure successful propagation of the genotype

Detection of hilA gene sequences in serovars of Salmonella enterica sufigbspecies enterica

hilA gene promoter, component of the Salmonella Pathogenicity Island 1, has been found in Salmonella serovar Typhimurium, being important for the regulation of type III secretion apparatus genes. We detected hilA gene sequences in Salmonella serovars Typhi, Enteritidis, Choleraesuis, Paratyphi A and B, and Pullorum, by polymerase chain reaction (PCR) and hybridization techniques. The primers to carry out PCR were designed according to hilA sequence. A low stringency hybridization with the probe pVV441 (hilA open-reading-frame plasmid) was carried out. To find hilA gene sequences in other Salmonella sp. suggest that these serovars could have similar sequences of this kind of virulence genes