TolA mediates the differential detergent resistance pattern between the Salmonella enterica subsp. enterica serovars Typhi and Typhimurium.

The tol-pal genes are essential for maintaining the outer membrane integrity and detergent resistance in various Gram-negative bacteria, including Salmonella. The role of TolA has been well established for the bile resistance of Salmonella enterica subsp. enterica serovar Typhimurium. We compared the bile resistance pattern between the S. enterica serovars Typhi and Typhimurium and observed that Typhi is more resistant to bile-mediated damage. A closer look revealed a significant difference in the TolA sequence between the two serovars which contributes to the differential detergent resistance. The tolA knockout of both the serovars behaves completely differently in terms of membrane organization and morphology. The role of the Pal proteins and difference in LPS organization between the two serovars were verified and were found to have no direct connection with the altered bile resistance. In normal Luria broth (LB), S. Typhi ΔtolA is filamentous while S. Typhimurium ΔtolA grows as single cells, similar to the wild-type. In low osmolarity LB, however, S. Typhimurium ΔtolA started chaining and S. Typhi ΔtolA showed no growth. Further investigation revealed that the chaining phenomenon observed was the result of failure of the outer membrane to separate in the dividing cells. Taken together, the results substantiate the evolution of a shorter TolA in S. Typhi to counteract high bile concentrations, at the cost of lower osmotic tolerance.

A safe vaccine (DV-STM-07) against Salmonella infection prevents abortion and confers protective immunity to the pregnant and new born mice.

Pregnancy is a transient immuno-compromised condition which has evolved to avoid the immune rejection of the fetus by the maternal immune system. The altered immune response of the pregnant female leads to increased susceptibility to invading pathogens, resulting in abortion and congenital defects of the fetus and a subnormal response to vaccination. Active vaccination during pregnancy may lead to abortion induced by heightened cell mediated immune response. In this study, we have administered the highly attenuated vaccine strain DeltapmrG-HM-D (DV-STM-07) in female mice before the onset of pregnancy and followed the immune reaction against challenge with virulent S. Typhimurium in pregnant mice. Here we demonstrate that DV-STM-07 vaccine gives protection against Salmonella in pregnant mice and also prevents Salmonella induced abortion. This protection is conferred by directing the immune response towards Th2 activation and Th1 suppression. The low Th1 response prevents abortion. The use of live attenuated vaccine just before pregnancy carries the risk of transmission to the fetus. We have shown that this vaccine is safe as the vaccine strain is quickly eliminated from the mother and is not transmitted to the fetus. This vaccine also confers immunity to the new born mice of vaccinated mothers. Since there is no evidence of the vaccine candidate reaching the new born mice, we hypothesize that it may be due to trans-colostral transfer of protective anti-Salmonella antibodies. These results suggest that our vaccine DV-STM-07 can be very useful in preventing abortion in the pregnant individuals and confer immunity to the new born. Since there are no such vaccine candidates which can be given to the new born and to the pregnant women, this vaccine holds a very bright future to combat Salmonella induced pregnancy loss.

lac repressor is an antivirulence factor of Salmonella enterica: its role in the evolution of virulence in Salmonella.

The genus Salmonella includes many pathogens of great medical and veterinary importance. Bacteria belonging to this genus are very closely related to those belonging to the genus Escherichia. lacZYA operon and lacI are present in Escherichia coli, but not in Salmonella enterica. It has been proposed that Salmonella has lost lacZYA operon and lacI during evolution. In this study, we have investigated the physiological and evolutionary significance of the absence of lacI in Salmonella enterica. Using murine model of typhoid fever, we show that the expression of LacI causes a remarkable reduction in the virulence of Salmonella enterica. LacI also suppresses the ability of Salmonella enterica to proliferate inside murine macrophages. Microarray analysis revealed that LacI interferes with the expression of virulence genes of Salmonella pathogenicity island 2. This effect was confirmed by RT-PCR and Western blot analysis. Interestingly, we found that SBG0326 of Salmonella bongori is homologous to lacI of Escherichia coli. Salmonella bongori is the only other species of the genus Salmonella and it lacks the virulence genes of Salmonella pathogenicity island 2. Overall, our results demonstrate that LacI is an antivirulence factor of Salmonella enterica and suggest that absence of lacI has facilitated the acquisition of virulence genes of Salmonella pathogenicity island 2 in Salmonella enterica making it a successful systemic pathogen.

SopB of Salmonella enterica serovar Typhimurium is a potential DNA vaccine candidate in conjugation with live attenuated bacteria.

The immune response against Salmonella is multi-faceted involving both the innate and the adaptive immune system. The characterization of specific Salmonella antigens inducing immune response could critically contribute to the development of epitope based vaccines for Salmonella. We have tried to identify a protective T cell epitope(s) of Salmonella, as cell mediated immunity conferred by CD8+ T cells is the most crucial subset conferring protective immunity against Salmonella. It being a proven fact that secreted proteins are better in inducing cell mediated immunity than cell surface and cytosolic antigens, we have analyzed all the genbank annotated Salmonella pathogenicity island 1 and 2 secreted proteins of Salmonella enterica serovar Typhimurium (S. typhimurium) and S. enterica serovar Typhi (S. typhi). They were subjected to BIMAS and SYFPEITHI analysis to map MHC-I and MHC-II binding epitopes. The huge profile of possible T cell epitopes obtained from the two classes of secreted proteins were tabulated and using a scoring system that considers the binding affinity and promiscuity of binding to more than one allele, SopB and SifB were chosen for experimental confirmation in murine immunization model. The entire SopB and SifB genes were cloned into DNA vaccine vectors and were administered along with live attenuated Salmonella and it was found that SopB vaccination reduced the bacterial burden of organs by about 5-fold on day 4 and day 8 after challenge with virulent Salmonella and proved to be a more efficient vaccination strategy than live attenuated bacteria alone.

Differentially evolved genes of Salmonella pathogenicity islands: insights into the mechanism of host specificity in Salmonella.

BACKGROUND: The species Salmonella enterica (S. enterica) includes many serovars that cause disease in avian and mammalian hosts. These serovars differ greatly in their host range and their degree of host adaptation. The host specificity of S. enterica serovars appears to be a complex phenomenon governed by multiple factors acting at different stages of the infection process, which makes identification of the cause/s of host specificity solely by experimental methods difficult. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we have employed a molecular evolution and phylogenetics based approach to identify genes that might play important roles in conferring host specificity to different serovars of S. enterica. These genes are 'differentially evolved' in different S. enterica serovars. This list of 'differentially evolved' genes includes genes that encode translocon proteins (SipD, SseC and SseD) of both Salmonella pathogenicity islands 1 and 2 encoded type three secretion systems, sptP, which encodes an effector protein that inhibits the mitogen-activated protein kinase pathway of the host cell, and genes which encode effector proteins (SseF and SifA) that are important in placing the Salmonella-containing vacuole in a juxtanuclear position. CONCLUSIONS/SIGNIFICANCE: Analysis of known functions of these 'differentially evolved genes' indicates that the products of these genes directly interact with the host cell and manipulate its functions and thereby confer host specificity, at least in part, to different serovars of S. enterica that are considered in this study.