Typhimurium Strain SL1344 Research Articles

Salmonella enterica serovar Typhimurium Induces Rapid Migration of Dendritic Cells into the Gut Lumen

Dendritic cells (DC)s are key elements for the immune responses and can operate as antigen sampling cells. We report now that they can also migrate into the gut lumen upon bacterial challenge. Isolated intestinal loops in BALB/c mice, C57/BL6 and C57/BL6 MyD88 knock out (KO) mice were infected with non invasive GFP‐labelled S. Typhimurium strain SL1344 (?SPI1, rpsM‐gfp), a derivative of the same strain lacking SPI1 and both fliC and fljB flagellin encoding genes, E. coli DH5a (/pZEP16, rpsM‐gfp), or PBS. After three hours, luminal content was harvested, washed and DCs were separated. FACS was carried out to characterize and enumerate the recovered cells. Confocal microscopy (CM) and TEM were also performed.Approximately 7 × 104 cells were present in loops treated with flagellated S. typhimurium SL1344, but not in loops belonging to the C57/BL6 MyD88 KO mice. Phenotypic analysis showed the presence of CD11c+, CX3CR1+, MHCII+, CD8a− and CD11b− cells associated with Salmonella. CM confirmed the increase of CD11c+ DCs in the gut lamina propria. TEM and CM showed bacteria associated with luminal DCs. These data show that Salmonella induces flagellin‐ and Toll‐like receptor‐dependent DC migration into the gut lumen.

During infection of epithelial cells Salmonella enterica serovar Typhimurium undergoes a time-dependent transcriptional adaptation that results in simultaneous expression of three type 3 secretion systems

The biogenesis of the Salmonella-containing vacuole within mammalian cells has been intensively studied over recent years. However, the ability of Salmonella to sense and adapt to the intracellular environment of different types of host cells has received much less attention. To address this issue, we report the transcriptome of Salmonella enterica serovar Typhimurium SL1344 within epithelial cells and show comparisons with Salmonella gene expression inside macrophages. We report that S. Typhimurium expresses a characteristic intracellular transcriptomic signature in response to the environments it encounters within different cell types. The signature involves the upregulation of the mgtBC, pstACS and iro genes for magnesium, phosphate and iron uptake, and Salmonella pathogenicity island 2 (SPI2). Surprisingly, in addition to SPI2, the invasion-associated SPI1 pathogenicity island and the genes involved in flagellar biosynthesis were expressed inside epithelial cells at later stages of the infection, while they were constantly downregulated in macrophage-like cells. To our knowledge, this is the first report of the simultaneous transcription of all three Type Three Secretion Systems (T3SS) within an intracellular Salmonella population. We discovered that S. Typhimurium strain SL1344 was strongly cytotoxic to epithelial cells after 6 h of infection and hypothesize that the time-dependent changes in Salmonella gene expression within epithelial cells reflects the bacterial response to host cells that have been injured by the infection process.

The Lactobacillus plantarum strain ACA-DC287 isolated from a Greek cheese demonstrates antagonistic activity in vitro and in vivo against Salmonella enterica serovar Typhimurium

The purpose of this study was to investigate the antibacterial activity of the Xynotyri cheese isolate Lactobacillus plantarum ACA-DC287 using a set of in vitro and in vivo assays. The co-culture of L. plantarum strain ACA-DC287 and Salmonella enterica serovar Typhimurium strain SL1344 results in the killing of the pathogen. The killing activity was produced mainly by non-lactic acid molecule(s) that were present in the cell-free culture supernatant of the L. plantarum strain ACA-DC287. The culture of the L. plantarum strain ACA-DC287 inhibited the penetration of S. typhimurium SL1344 into cultured human enterocyte-like Caco-2/TC7 cells. In conventional mice infected with S. typhimurium SL1344, the intake of L. plantarum strain ACA-DC287 results in a decrease in the levels of Salmonella associated with intestinal tissues or those present in the intestinal contents. In germ-free mice, the L. plantarum strain ACA-DC287 colonized the gastrointestinal tract. The L. plantarum strain ACA-DC287 strain exerts anti-Salmonella activity similar that of the established probiotic strains Lactobacillus rhamnosus GG, Lactobacillus casei Shirota YIT9029 and Lactobacillus johnsonii La1. The observation that a selected cheese Lactobacillus strain exerted antibacterial activity that was similar to those of probiotic Lactobacillus strains, is of interest for the use of this strain as an adjunct strain for the production of health-giving cheeses.

Microarray-Based Detection of Salmonella enterica Serovar Typhimurium Transposon Mutants That Cannot Survive in Macrophages and Mice

DNA microarrays provide an opportunity to combine the principles of signature-tagged mutagenesis (STM) with microarray technology to identify potentially important bacterial virulence genes. The scope of DNA microarrays allows for less laborious screening on a much larger scale than possible by STM alone. We have adapted a microarray-based transposon tracking strategy for use with a Salmonella enterica serovar Typhimurium cDNA microarray in order to identify genes important for survival and replication in RAW 264.7 mouse macrophage-like cells or in the spleens of BALB/cJ mice. A 50,000-CFU transposon library of S. enterica serovar Typhimurium strain SL1344 was serially passaged in cultured macrophages or intraperitoneally inoculated into BALB/cJ mice. The bacterial genomic DNA was isolated and processed for analysis on the microarray. The novel application of this approach to identify mutants unable to survive in cultured cells resulted in the identification of components of Salmonella pathogenicity island 2 (SPI2), which is known to be critical for intracellular survival and replication. In addition, array results indicated that a number of SPI1-associated genes, currently not associated with intracellular survival, are negatively selected. However, of the SPI1-associated mutants individually tested for intracellular survival, only a sirA mutant exhibited reduced numbers relative to those of wild-type bacteria. Of the mutants unable to survive in mice, significant proportions are either components of the SPI2 pathogenicity island or involved in lipopolysaccharide synthesis. This observation is in agreement with results obtained in the original S. enterica serovar Typhimurium STM screen, illustrating the utility of this approach for the high-throughput identification of virulence factors important for survival in the host.

Effect of Inactivation of degS on Salmonella enterica Serovar Typhimurium In Vitro and In Vivo

The alternative sigma factor (RpoE sigma(E)) enables Salmonella enterica serovar Typhimurium to adapt to stressful conditions, such as oxidative stress, nutrient deprivation, and growth in mammalian tissues. Infection of mice by Salmonella serovar Typhimurium also requires sigma(E). In Escherichia coli, activation of the sigma(E) pathway is dependent on proteolysis of the anti-sigma factor RseA and is initiated by DegS. DegS is also important in order for E. coli to cause extraintestinal infection in mice. We constructed a degS mutant of the serovar Typhimurium strain SL1344 and compared its behavior in vitro and in vivo with those of its wild-type (WT) parent and an isogenic rpoE mutant. Unlike E. coli degS strains, the Salmonella serovar Typhimurium degS strain grew as well as the WT strain at 42 degrees C. The degS mutant survived very poorly in murine macrophages in vitro and was highly attenuated compared with the WT strain for both the oral and parenteral routes of infection in mice. However, the degS mutant was not as attenuated as the serovar Typhimurium rpoE mutant: 100- to 1,000-fold more degS bacteria than rpoE bacteria were present in the livers and spleens of mice 24 h after intraperitoneal challenge. In most assays, the rpoE mutant was more severely affected than the degS mutant and a sigma(E)-dependent reporter gene was more active in the degS mutant than the rpoE strain. These findings indicate that degS is important for activation of the sigma(E) pathway in serovar Typhimurium but that alternative pathways for sigma(E) activation probably exist.

Characterization of Salmonella pathogenicity island 1 type III secretion-dependent hemolytic activity in Salmonella enterica serovar Typhimurium

A number of bacteria that are pathogenic for animals and plants possess a type III secretion system (TTSS) to translocate virulence-associated proteins into host cells. In several bacteria, it has been reported that the TTSS is correlated with an ability to cause contact-dependent hemolysis in vitro. Here, we showed that the Salmonella enterica serovar Typhimurium strain SL1344 exhibited Salmonella pathogenicity island 1 type III secretion-dependent, contact-mediated, hemolytic activity. Mutations with a single deletion in genes encoding putative pore-forming proteins, SipB and SipC, secreted by the TTSS abolished hemolytic activity. In addition, the osmoprotection studies revealed that molecules larger than PEG2000 conferred significant protection against lysis induced by Salmonella. These results indicate that the hemolysis generated by Salmonella is due to the formation of pores within the erythrocyte membrane.

Oral immunization with an rfaH mutant elicits protection against salmonellosis in mice.

Loss of the transcriptional antiterminator RfaH results in virulence attenuation (>10(4)-fold increase in 50% lethal dose) of the archetypal Salmonella enterica serovar Typhimurium strain SL1344 by both orogastric and intraperitoneal routes of infection in BALB/c mice. Oral immunization with the mutant efficiently protects mice against a subsequent oral infection with the wild-type strain. Interestingly, in vitro immunoreactivity is not confined to strain SL1344; rather, it is directed also towards other serovars of S. enterica and even Salmonella bongori strains.