No abstract

SummaryWe report the functional characterization of BipA, a GTPase that undergoes tyrosine phosphorylation in an enteropathogenic Escherichia coli (EPEC) strain. BipA ¹ mutants adhere to cultured epithelial cells but fail to trigger the characteristic cytoskeletal rearrangements found in cells infected with wild-type EPEC. In contrast, increased expression of BipA enhances actin remodelling and results in the hyperformation of pseudopods. BipA appears to be the first example of a new class of virulence regulator, as it also controls flagellamediated cell motility and resistance to the antibacterial effects of a human host defence protein. Its striking sequence similarity to ribosome-binding elongation factors suggests that it uses a novel mechanism to modulate gene expression.

SummaryBipA is a novel member of the ribosome binding GTPase superfamily and is widely distributed in bacteria and plants. We report here that it regulates multiple cell surface-and virulence-associated components in the enteropathogenic Escherichia coli (EPEC) strain E2348/69. The regulated components include bacterial flagella, the espC pathogenicity island and a type III secretion system specified by the locus of enterocyte effacement (LEE). BipA positively regulated the espC and LEE gene clusters through transcriptional control of the LEE-encoded regulator, Ler. Additionally, it affected the pattern of proteolysis of intimin, a key LEE-encoded adhesin specified by the LEE. BipA control of the LEE operated independently of the previously characterized regulators Per, integration host factor and H-NS. In contrast, it negatively regulated the flagella-mediated motility of EPEC and in a Ler-independent manner. Our results indicate that the BipA GTPase functions high up in diverse regulatory cascades to co-ordinate the expression of key pathogenicity islands and other virulenceassociated factors in E. coli .

Salmonella adaptation to low pH is a critical survival response and essential for virulence. Here, we show that another key virulence-associated process, flagella-mediated cell motility, is co-regulated by low pH via the PhoPQ signal transduction system. Using a proteomic approach, we found that phase 1 and phase 2 flagellin were specifically down-regulated when acid-adapted (pH 5.0) Salmonella SL1344 cells were exposed to pH 3.0. Decreased flagellin expression and cell motility was dependent on activation of the PhoPQ pathway, which directly or indirectly negatively regulated transcription of the flagellin gene fliC. In contrast, the general stress sigma factor RpoS (sigma s) positively regulated flagellar gene expression. Low external pH had no effect on the level of H-NS protein, a further regulator of flagellar gene expression. We suggest that flagellar repression at low pH conserves ATP for survival processes and helps to limit the influx of protons into the cytosol. These results highlight the power of proteomics to reveal unanticipated links between relatively well-characterised regulatory systems in bacteria.

Aims:  The aims of this study were to develop media to cultivate actinomycetes, screen the resulting isolates with Actinobacteria‐specific primers, and examine the efficacy of detection of the actinobacterial isolates with universal primers. Methods and Results:  Soil‐extract medium was developed for a terrestrial bluff environment. Recovered isolates were subjected to polymerase chain reaction (PCR) with taxon‐specific primers to identify Actinobacteria. Universal bacterial primers 24f and 1492r (modified and original versions) were used to amplify the 16S rRNA gene from the putative Actinobacteria. While both reverse primers failed to provide amplification products from 20% to 50% of the isolates, the 1492r primer detected Actinobacteria more effectively than 1492r‐mod. The region of the gene containing the annealing site for the 1492r primers from 15 isolates that failed to amplify showed no differences in nucleotide sequence to the original 1492r primer. Conclusions:  Universal 16S rRNA gene primers are not capable of amplifying this gene from all bacteria within an environmental sample. Some Actinobacteria may share 100% sequence similarity to universal primers but remain undetected. Significance and Impact of the Study:  These findings are important for studies of particular taxa in environmental samples where reactions utilizing universal primers may not reveal the extent of their presence and diversity.