Typhi selleck chemicals llc STH2370 was the most cytotoxic strain among all bacteria tested. This result suggests that the SseJ effector protein decreased S. Typhi cytoxicity when bacteria interact with human cell lines, resulting in increased cell permeability. Figure 5 Analyses of cytotoxicity HT-29 infected with complemented and wild type S . Typhi strains. HT-29 cells were grown in transwells for

12-15 days. Polarised HT-29 cells were apically infected with the S. Typhi wild type or the respective complemented strains. Released LDH was measured 3 h post-infection and reported as percentage relative to the S. Typhi wild type. The values correspond to the means CP673451 manufacturer ± SD of three independent experiments, each performed in duplicate. The percentages of each S. Typhimurium 14028s, S. Typhi STH2370/pNT005 and S. Typhi STH2370/pNT006, have significantly differences respect S. Typhi

STH2370 wild type. LDH release from infected cells with S. Typhi carrying empty plasmid (pSU19 or pCC1) showed no differences with respect to the wild type strain (data not shown). The presence of sseJ STM in S. Typhi increased bacterial intracellular retention/proliferation within HEp-2 cells It has been reported that sseJ contributes to the intracellular proliferation of S. Typhimurium [31, 38]. Moreover, the decreased cell death produced by the presence of sseJ STM in S. Typhi strains (Figure 5) may lead to an increased proliferation of intracellular bacteria because of a decreased cytotoxicity. A less cytotoxic pathogen should be retained inside eukaryotic cells over time, allowing an increased bacterial proliferation. If this hypothesis is correct, S. Typhi carrying selleck inhibitor sseJ STM should exhibit increased CFUs in the gentamicin protection assay (see Materials Amisulpride and Methods). As expected, Figure 6 shows

that the presence of sseJ STM yielded a significantly increase in the CFUs recovered from the infected cells compared to the wild type. Figure 6 Gentamicin protection assay of complemented and wild type strains of S. Typhi. HEp-2 cells were grown and infected with the S. Typhimurium 14028s, S. Typhi STH2370 or the respective S. Typhi complemented strains. The recovered CFUs were counted 3 h post-infection. The values correspond to the means ± SD of three different experiments, each performed in triplicate. The CFUs recovered from infected cells with S. Typhi with each empty plasmid (pSU19 or pCC1) showed no differences with respect to the wild type strain (data not shown). Discussion In the process of adaptation to humans, genes no longer compatible with the lifestyle of S. Typhi within the host were selectively inactivated. These inactivated genes are called “”antivirulence genes”" and their loss of function results in the adaptation to a given host [39]. S. Typhi is a facultative bacterial pathogen that has accumulated a high number of pseudogenes (approximately 5% of the genome) and over 75% of them have completely lost their functions [7, 16].