Project description:The Salmonella enterica serovar Typhimurium (ST) mutant lacking the msbB gene (ΔmsbB) has been widely studied as a candidate for attenuated bacterial vectors in therapeutic applications. Deletion of msbB results in LPS with under-acylated lipid A, which lowers endotoxicity while maintaining structural integrity. This attenuation has traditionally been attributed to reduced TLR4 activation due to weaker interaction between the modified lipid A and TLR4. In our study, we confirmed that ΔmsbB ST was less lethal than wild-type (WT) ST in a mouse sepsis model. However, this difference persisted even in TLR4- and caspase-11-deficient mice, suggesting that LPS signaling is not the primary determinant of virulence. In vitro, bone marrow–derived macrophages (BMDMs) from TLR4- or caspase-11-deficient mice showed only modest reductions in ST-induced cell death and cytokine production. Importantly, ΔmsbB ST behaved similarly to WT ST in these assays, further indicating that LPS-mediated signaling is not central to the observed attenuation. Additionally, the mutant exhibited increased outer membrane permeability, likely contributing to its heightened antibiotic sensitivity—and reduced motility due to lower flagellin protein levels.

2025-12-09 | GSE304712 | GEO

qRT-PCR of ex vivo Salmonella enterica in TLR4 minus/plus macrophage background

Project description:A major virulence factor in S. enterica is the pathogenicity island Spi2. The goal of the experiment was to determine the impact of the toll-like receptor 4 (TLR-4) on expression of Spi2 in S. enterica wild type (WT) and triple mutant strain (TMS) with deletions in SPI-1 TTSS, SPI-2 TTSS, flagella. The stain 14028 was used for the experiment. Cells were grown in a TLR-4 wildtype (plus) or deleted (minus) macrophage background. Total RNA was extracted and processed by qRT-PCR. Two strains (WT and TMS) grown under 2 conditions (TLR-4 wildtype (plus) or TLR-4 deleted (minus)) - total 4 samples were analyzed in the study. All PCRs were done in quadruplicates, located on the same PCR plate. For each quadruplicates the median value was calculated and used as a data point for further normalization. A set of controls was selected for all data sets using the following criteria: the data points for the controls should have Cp <40 and be present in all the samples.

2011-06-13 | E-GEOD-26804 | biostudies-arrayexpress

qRT-PCR of ex vivo Salmonella enterica in NRAMP-1 minus/plus macrophage background

Project description:A major virulence factor in S. enterica is the pathogenicity island Spi2. The goal of the experiment was to determine the impact of the NRAMP-1 (natural resistance-associated macrophage protein-1) on expression of Spi2 in S. enterica wild type (WT) and triple mutant strain (TMS) with deletions in SPI-1 TTSS, SPI-2 TTSS, flagella. The stain 14028 was used for the experiment. Cells were grown in a NRAMP-1 wildtype (plus) or deleted (minus) macrophage background. Total RNA was extracted and processed by qRT-PCR. Two strains ( WT and TMS) grown under 2 conditions (NRAMP-1 wildtype (plus) or NRAMP-1 deleted (minus)) - total 4 samples were analyzed in the study. All PCRs were done in duplicates, located on the same PCR plate. For each duplicate the mean value was calculated and used as a data point for further normalization. We averaged the Cp values for all the data points in a sample and then subtracted this sample average Cp from all the data points of the sample. Only 192 primer pairs from 288 on the platform GPL11111 were used.

2011-06-13 | E-GEOD-26806 | biostudies-arrayexpress