Regulation of phenotypic heterogeneity permits Salmonella evasion of the host caspase-1 inflammatory response.

Sensing and adapting to the environment is one strategy by which bacteria attempt to maximize fitness in an unpredictable world; another is the stochastic generation of phenotypically distinct subgroups within a genetically clonal population. In culture, Salmonella Typhimurium populations are bistable for the expression of flagellin. We report that YdiV controls this expression pattern by preventing transcription of the sigma factor that recruits RNA polymerase to the flagellin promoter. Bistability ensues when the sigma factor is repressed in a subpopulation of cells, resulting in two phenotypes: flagellin expressors and flagellin nonexpressors. Although the ability to swim is presumably a critical survival trait, flagellin activates eukaryotic defense pathways, and Salmonella restrict the production of flagellin during systemic infection. Salmonella mutants lacking YdiV are unable to fully repress flagellin at systemic sites, rendering them vulnerable to caspase-1 mediated colonization restriction. Thus, a regulatory mechanism producing bistability also impacts Salmonella virulence.

The lipid A phosphate position determines differential host Toll-like receptor 4 responses to phylogenetically related symbiotic and pathogenic bacteria.

The human symbiont Bacteroides thetaiotaomicron promotes intestinal function and health, whereas the phylogenetically related pathogen Porphyromonas gingivalis is associated with the chronic oral inflammatory disease periodontitis. Although both B. thetaiotaomicron and P. gingivalis synthesize lipopolysaccharides (LPS) consisting of penta-acylated, monophosphorylated lipid A in addition to immunologically silent, nonphosphorylated lipid A, they elicit strikingly distinct Toll-like receptor 4 (TLR4) responses. We show that the phosphate position of penta-acylated, monophosphorylated lipid A is a key feature for determining the differential TLR4 responses elicited by these evolutionarily related bacteria. B. thetaiotaomicron produces TLR4-stimulatory lipid A bearing a 1-phosphate, in contrast to P. gingivalis, which produces TLR4-evasive lipid A bearing a 4'-phosphate. Confirming these observations, recombinant Escherichia coli LPS containing penta-acylated, 1-phosphorylated lipid A is more TLR4 stimulatory than LPS containing 4'-phosphorylated lipid A. The specific capacity of a Gram-negative bacterium to alert or evade the host innate immune defense system through TLR4-dependent signaling is currently recognized as a critical aspect defining the relationship between the host and the bacterium. We propose that the distinct lipid A phosphate positions observed for the B. thetaiotaomicron and P. gingivalis LPS contributes to the manifestation of these bacteria as commensal or pathogen within the human host.

The structurally similar, penta-acylated lipopolysaccharides of Porphyromonas gingivalis and Bacteroides elicit strikingly different innate immune responses.

Lipid A structural modifications can substantially impact the host's inflammatory response to bacterial LPS. Bacteroides fragilis, an opportunistic pathogen associated with life-threatening sepsis and intra-abdominal abscess formation, and Bacteroides thetaiotaomicron, a symbiont pivotal for proper host intestinal tissue development, both produce an immunostimulatory LPS comprised of penta-acylated lipid A. Under defined conditions, Porphyromonas gingivalis, an oral pathogen associated with periodontitis, also produces an LPS bearing a penta-acylated lipid A. However, this LPS preparation is 100-1000 times less potent than Bacteroides LPS in stimulating endothelial cells. We analyzed Bacteroides and P. gingivalis lipid A structures using MALDI-TOF MS and gas chromatography to determine the structural basis for this phenomenon. Even though both Bacteroides and P. gingivalis lipid A molecules are penta-acylated and mono-phosphorylated, subtle differences in mass and fatty acid content could account for the observed difference in LPS potency. This fatty acid heterogeneity is also responsible for the peak "clusters" observed in the mass spectra and obfuscates the correlation between LPS structure and immunostimulatory ability. Further, we show the difference in potency between Bacteroides and P. gingivalis LPS is TLR4-dependent. Altogether, the data suggest subtle changes in lipid A structure may profoundly impact the host's innate immune response.