High levels of σS impair the growth of E. coli on poor carbon sources or under nutrient limitation [28]. Stress resistance is not constant amongst all E. coli strains [28–30] also indicating possible variation in gene expression relating to RpoS and/or ppGpp. We demonstrate here that strain variation in ppGpp is one of several factors that contribute

to the difference in the level of σS across the species E. coli and discuss the polymorphisms at the core of bacterial regulation. Results The goal of this study is three-fold: to provide evidence that rpoS polymorphism and variation in σS levels are widespread in the species E. coli; to show that the genes that control ppGpp synthesis and degradation are also subject to variation and finally to demonstrate that the different levels of RpoS are at least partially dependent on variability of endogenous ppGpp. Strain BMN 673 in vivo variation in RpoS levels in the species E. coli To test the extent of variation in RpoS levels, we analysed 31 strains from the ECOR collection of E. coli isolates from various locations and environments [31]. The 72 ECOR strains are divided into five phylogenetic groups (A, B1, B2,

D and E). Nine of the strains tested here belonged to group this website A, 7 to group B1, 10 to group B2 and 5 to group D. The K-12 strain MG1655 was used as a control reference. As shown in Figure 1, the cellular content of RpoS was highly variable in standardised overnight cultures. Nine isolates had no detectable RpoS, another five had

RpoS level 3 to 7-fold above that of the laboratory K-12 strain MG1655. The remainder of strains had levels within a 2-fold range around MG1655. The absence of RpoS from the nine strains was confirmed by screening for σS-related phenotypes (glycogen accumulation [32] and catalase activity [33]; results not shown). Figure 1 Quantitation of RpoS. Overnight bacterial cultures grown in LB were harvested, lysed and their total protein content resolved by SDS-PAGE. Proteins were immunoblotted with anti-RpoS monoclonal antibodies. The bands were scanned and quantified. Densitometric measurements were normalised against ECOR 56 PAK6 to which was assigned 100 units. Relative values represent the mean ± S.E. of at least three independent experiments. rpoS sequences in ECOR strains Variation in the rpoS locus was already indicated by the observation that PCR amplification of the rpoS region resulted in fragments of three different sizes, as shown in Table 1. These differences were consistent with the genomic variation in the rpoS-mutS region in the species E. coli [34]. The size of fragments and sequence matches correspond to previously described rpoS regions, with the 1.3 Kb fragment like that in E. coli K-12, and the 4.2 Kb and 3.4 Kb products similar to those found in [35] and [36] respectively.