4 kDa). The ferric aerobactin transport system is a well-known virulence factor in E. coli strains causing extraintestinal infections (reviewed in [22]), such as urinary tract infections [23]. Although its role as a virulence determinant in

intestinal E. coli is not well understood, it has been proposed that it contributes to the strong colonizing capacity of those strains carrying the aerobactin genes [24]. For this reason, we evaluated the contribution of this iron transport system in the colonization capabilities of E. coli O104:H4. Figure 2 Detection of differentially www.selleckchem.com/products/netarsudil-ar-13324.html expressed surface proteins in E. coli O104:H4 strains 15% SDS-PAGE of heat-extracted proteins from E. coli O104:H4 strain 2050 (lanes 1), 2071 (lanes 2), and C3493 (lanes 3) grown on LB or MacConkey agar. The arrows indicate the location of the aerobactin MNK inhibitor transport receptor (Arrow A) and the chain A, dipeptide-binding protein (Arrow B). Low iron concentration

in MacConkey induces aerobactin receptor BI 10773 in vitro expression MacConkey agar is considered a low iron-containing medium which has been used to identify high-affinity iron and zinc uptake systems [25]. Therefore, expression of the aerobactin receptor in the E. coli O104:H4 wild type and the iutA mutant was investigated by using heat-extracted preparations of bacteria grown on agar plates with and without the addition of the iron chelator 2,2’-dipyridyl (DP). Expression was monitored on MacConkey as well as LB agar supplemented with DP, because the addition of Buspirone HCl the iron chelator is known to induce expression of iron transport systems in E. coli[17]. No production of IutA (the 80.9 kDa aerobactin receptor) was observed on Coomassie-stained 12.5% SDS-PAGE gels containing LB agar-recovered bacterial extracts, while abundant IutA was evident in samples from MacConkey plates (Figure 3, panel

A). In contrast, the iutA mutant lacked detectable expression of IutA on either media tested. To confirm that aerobactin receptor expression responded to iron depletion, the media was supplemented with 200 μM of DP. As shown in Figure 3, panel A, iron chelation resulted in the expression of IutA in bacteria grown on LB + DP as well as MacConkey + DP. As expected, the aerobactin receptor was absent in heat extracts obtained from the CSS001 strain (iutA::cat) grown on either of the iron-depleted media. However, for reasons that remain unclear, the expression of the IutA receptor does not appear to be further induced on MacConkey agar supplemented with DP. Figure 3 IutA protein induction and qRT-PCR analysis of iutA expression. A. Heat-extracted proteins of E. coli O104:H4 strains C3493 (German isolate) and CCSS001 (iutA::cat) grown on MacConkey (MC) or LB agar in the absence (MC or LB) or presence (MC + DP or LB + BS) of 2,2’-dipyridil (DP) were separated in 12.5% SDS-PAGE gels and stained with Coomassie brilliant blue. Molecular mass markers are indicated on the left and the heat-extracted IutA protein is depicted by an arrow on the right. B. E.