Phage insertion in mlrA and variations in rpoS limit curli expression and biofilm formation in Escherichia coli serotype O157 : H7

Abstract

Biofilm formation in Escherichia coli is a tightly controlled process requiring the expression of adhesive curli fibres and certain polysaccharides such as cellulose. The transcriptional regulator CsgD is central to biofilm formation, controlling the expression of the curli structural and export proteins and the diguanylate cyclase adrA, which indirectly activates cellulose production. CsgD itself is highly regulated by two sigma factors (RpoS and RpoD), multiple DNA-binding proteins, small regulatory RNAs and several GGDEF/EAL proteins acting through c-di-GMP. One such transcription factor MlrA binds the csgD promoter to enhance the RpoS-dependent transcription of csgD. Bacteriophage, often carrying the stx 1 gene, utilize an insertion site in the proximal mlrA coding region of E. coli serotype O157 : H7 strains, and the loss of mlrA function would be expected to be the major factor contributing to poor curli and biofilm expression in that serotype. Using a bank of 55 strains of serotype O157 : H7, we investigated the consequences of bacteriophage insertion. Although curli/biofilm expression was restored in many of the prophagebearing strains by a wild-type copy of mlrA on a multi-copy plasmid, more than half of the strains showed only partial or no complementation. Moreover, the two strains carrying an intact mlrA were found to be deficient in biofilm formation. However, RpoS mutations that attenuated or inactivated RpoS-dependent functions such as biofilm formation were found in .70 % of the strains, including the two strains with an intact mlrA. We conclude that bacteriophage interruption of mlrA and RpoS mutations provide major obstacles limiting curli expression and biofilm formation in most serotype O157 : H7 strains.

The rpoS Gene Is Predominantly Inactivated during Laboratory Storage and Undergoes Source-Sink Evolution in Escherichia coli Species

“…In collections of natural E. coli and Salmonella isolates originating from the environment or various hosts, either as commensal bacteria or as pathogens, the frequency of inactivating alleles of rpoS is very variable and ranges from less than 1% to more than 70% of strains (25)(26)(27)(28)(29)(30). However, these data are difficult to interpret, because inactivation can depend on laboratory storage conditions, as discussed above.…”

“…Interestingly, the rpoS gene is located on the lagging strand, where the rate of point mutations is higher than on the leading strand, probably due to conflicts between replication and transcription (3). rpoS is also located close to the MR gene mutS, in a region that has high genomic variability as a result of horizontal gene transfer (23,24).In collections of natural E. coli and Salmonella isolates originating from the environment or various hosts, either as commensal bacteria or as pathogens, the frequency of inactivating alleles of rpoS is very variable and ranges from less than 1% to more than 70% of strains (25)(26)(27)(28)(29)(30). However, these data are difficult to interpret, because inactivation can depend on laboratory storage conditions, as discussed above.…”

Elucidating the Function of the RpoS Regulon

“…Since Crl effects are most pronounced at low RpoS concentrations and in the presence of RpoD [35], it is likely relevant in the exponential phase and may help explain why many genes are controlled by RpoS at this stage of growth [37]. Crl-mediated control of exponential phase genes may be relevant for pathogenesis, as RpoS levels are much higher in most O157:H7 strains in all phases of growth [38], although rpoS mutations appear common in many strains of this pathogenic serotype [39].…”

The Polymorphic Aggregative Phenotype of Shiga Toxin-Producing Escherichia coli O111 Depends on RpoS and Curli

“…Uhlich et al reported that in many strains of STEC O157:H7, the inability to produce curli correlated with mutations in rpoS (37). Thus, it appears that regulation of curli expression in STEC O111 is similar to that in K-12 and STEC O157:H7, with the exception of certain strains, such as those implicated in the spinach-linked outbreak in 2006, for which the csg operons are expressed in an RpoS Ϫ background (29).…”