LPS consists of three major components: lipid A, core polysaccharides and O-linked polysaccharides. Lipid A, with its fatty acid anchors [lauric, myristic and sometimes palmitic acid], is an endotoxin primarily responsible for TNFα-mediated septic shock. The addition of myristic

acid to the lipid A precursor AMN-107 in vivo is catalyzed by the enzyme MsbB [3]. 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. However, compensatory suppressor mutants can be isolated that grow under these conditions. One of these suppressor phenotypes results from a mutation in somA, a gene of unknown function [4]. msbB Salmonella Typhimurium Cell Cycle inhibitor strains have recently been developed as potential anti-cancer agents that possess impressive anti-tumor activity in mice [5]. In a phase I clinical study msbB Salmonella were shown to be safe in humans when administered i.v. However, 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 CFU/m2 and 1 × 109/m2 [6]. Toso et al. [6] noted that YS1646 (suppressed msbB strain, see below) grew

best in air without added CO2. The potential to grow in acidic and CO2-rich environments is a hallmark of pathogenic bacteria, enhancing persistence within phagocytes and survival inside the host. Sensitivity to CO2 and low pH of msbB Salmonella strains might explain poor colonization of tumors, which often contain

high levels of CO2 and lactic acid [7, 8] due to the Warburg effect, also known as aerobic glycolysis, whereby glucose uptake is elevated while oxidative phosphorylation is reduced, even in the presence of click here oxygen. Our previous work on suppressors of msbB Salmonella raised the possibility that secondary mutations could suppress sensitivity to 5% CO2 and acidic conditions. Here we report that the growth of msbB Salmonella is highly inhibited (greater than 3-log reduction in plating efficiency) in a 5% CO2 atmosphere in LB media as well as under low pH conditions Teicoplanin when compared to wild-type Salmonella. Furthermore, several CO2 resistant clones were selected from an msbB Salmonella transposon library (Tn5). Three mutations were mapped and all were shown to contain the Tn5 marker in the zwf gene, which encodes the enzyme glucose-6-phosphate-dehydrogenase and is tightly linked to the msbB gene. Results CO2 sensitivity of msbB Salmonella CO2 sensitivity was first observed when YS1646, an msbB purI Suwwan deletion strain of Salmonella Typhimurium, was plated on blood or LB plates and incubated in a 5% CO2 incubator (Caroline Clairmont, personal communication; Toso et al., 2002).