Abstract
BackgroundPathogens tolerate stress conditions that include low pH, oxidative stress, high salt and high temperature in order to survive inside and outside their hosts. Lipopolysaccharide (LPS), which forms the outer-leaflet of the outer membrane in Gram-negative bacteria, acts as a permeability barrier. The lipid A moiety of LPS anchors it to the outer membrane bilayer. The MsbB enzyme myristoylates the lipid A precursor and loss of this enzyme, in Salmonella, is correlated with reduced virulence and severe growth defects that can both be compensated with extragenic suppressor mutations.ResultsWe report here that msbB (or msbB somA) Salmonella are highly sensitive to physiological CO2 (5%), resulting in a 3-log reduction in plating efficiency. Under these conditions, msbB Salmonella form long filaments, bulge and lyse. These bacteria are also sensitive to acidic pH and high osmolarity. Although CO2 acidifies LB broth media, buffering LB to pH 7.5 did not restore growth of msbB mutants in CO2, indicating that the CO2-induced growth defects are not due to the effect of CO2 on the pH of the media. A transposon insertion in the glucose metabolism gene zwf compensates for the CO2 sensitivity of msbB Salmonella. The msbB zwf mutants grow on agar, or in broth, in the presence of 5% CO2. In addition, msbB zwf strains show improved growth in low pH or high osmolarity media compared to the single msbB mutant.ConclusionThese results demonstrate that msbB confers acute sensitivity to CO2, acidic pH, and high osmolarity. Disruption of zwf in msbB mutants restores growth in 5% CO2 and results in improved growth in acidic media or in media with high osmolarity. These results add to a growing list of phenotypes caused by msbB and mutations that suppress specific growth defects.
Highlights
Pathogens tolerate stress conditions that include low pH, oxidative stress, high salt and high temperature in order to survive inside and outside their hosts
Bacteria were rapidly cleared from the peripheral blood of humans and targeting to human tumors was only observed in few patients at the highest dose levels of 3 × 108 colony forming units (CFUs)/m2 and 1 × 109/m2 [6]
By plating more cells, the presence of a few resistant colonies could be detected, as we obtained 3.3 × 108 CFU/ml on plates incubated in air and 1.7 × 105 CFU/ml on plates incubated in the presence of 5% CO2, a greater than 3 log reduction
Summary
Pathogens tolerate stress conditions that include low pH, oxidative stress, high salt and high temperature in order to survive inside and outside their hosts. Lipopolysaccharide (LPS), which forms the outer-leaflet of the outer membrane in Gram-negative bacteria, acts as a permeability barrier. The MsbB enzyme myristoylates the lipid A precursor and loss of this enzyme, in Salmonella, is correlated with reduced virulence and severe growth defects that can both be compensated with extragenic suppressor mutations. Lipopolysaccharide (LPS), the most abundant molecule on the surface of Gram-negative bacteria, acts as a permeability barrier and renders the outer-leaflet of the outer membrane (OM) relatively impermeable to hydrophobic antibiotics, detergents [1], and host complement [2]. It has been shown that msbB Salmonella serovar Typhimurium exhibits severe growth defects in LB and sensitivity to bile salts (MacConkey) and EGTA-containing media. Toso et al [6] noted that YS1646 (suppressed msbB strain, see below) grew best in air without added CO2
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