The type 1 pili regulator gene fimX and pathogenicity island PAI-X as molecular markers of uropathogenic Escherichia coli

Abstract

Uropathogenic Escherichia coli (UPEC) fall within a larger group of isolates producing extraintestinal disease. UPEC express type 1 pili as a critical virulence determinant mediating adherence to and invasion into urinary tract tissues. Type 1 pili expression is under regulation by a family of site-specific recombinases, including FimX, which is encoded from a genomic island called PAI-X for pathogenicity island of FimX. Using a new multiplex PCR, fimX and the additional PAI-X genes were found to be highly associated with UPEC (144/173 = 83.2 %), and more prevalent in UPEC of lower urinary tract origin (105/120 = 87.5 %) than upper urinary tract origin (39/53 = 74 %; P<0.05) or commensal isolates (28/78 = 36 %; P≤0.0001). The Fim-like recombinase gene fimX is the only family member that has a significant association with UPEC compared to commensal isolates. Our results indicate PAI-X genes, including the type 1 pili regulator gene fimX, are highly prevalent among UPEC isolates and have a strong positive correlation with genomic virulence factors, suggesting a potential role for PAI-X in the extraintestinal pathogenic E. coli lifestyle.

Fig. 1.

Overview of the genomic location, features, and organization of PAI-X among sequenced E. coli isolates. (a) Genomic organization and conservation of PAI-X among sequenced isolates from EHEC (EDL933) and UPEC (UTI89) pathotypes. MG1655 is used as a K-12, commensal reference. (b) Percentage G+C trace of PAI-X_UTI89 and surrounding flanking regions. Percentage G+C is plotted as area under the curve for the entire region (betT through ykgE) using a 100 bp window. The average G+C across the entire UTI89 genome is plotted as a grey dotted line. The average % G+C content across PAI-X and flanking sequences is plotted as a black stepped line. (c) Grey arrows represent ORFs detected by multiplex PCR. Directionality of the ORFs, representing forward or reverse strand orientation, is shown. Amplicons are indicated as a black line (_). The genomic sequence for ipuA and ipuB is shown for CFT073 while fimB, fimE and PAI-X sequences are shown for the prototypic cystitis isolate UTI89. (d) Representative results of the multiplex PCR for the Fim-like recombinases and associated genomic islands using multiplex PCR. UTI89 and CFT073 are shown as prototypic UPEC strains to show visualization of all bands. (e) All E. coli isolates (n = 259), regardless of clinical syndrome, were scored based on the number of selective VFs present by PCR analysis and evaluated for the correlation between E. coli total VFs and the presence of PAI-X. Aggregate VFs refers to the number of total VFs present (out of the five assayed). Sample sizes varied between groups: 0 VFs (n = 30), 1 VF (n = 44), 2 VFs (n = 68), ≥3 VFs (n = 117).

Fig. 2.

Clonal group variation and Fim-like recombinase prevalence by isolate source and syndrome. (a) Phylogenetic associations among commensal and pathogenic isolates. The percentage of isolates in each of the four clonal groups is shown. Clonal groups were assigned based on a previously published multiplex PCR for chuA, yjaA, and genomic fragment TSPE4.C2 (Clermont et al., 2000; Gordon et al., 2008). ECOR strain clonal groups were determined previously (Johnson et al., 2001) and are in accordance with our experimental results. CY, cystitis; PY, pyelonephritis; UTI-BL, urosepsis. (b) The percentage of isolates that encode one of four recombinase profiles observed, grouped as follows: fimB-E only (similar to MG1655); fimB-E-X only (similar to UTI89); fimB-E ipuA-B (rarely observed); and all five recombinases present, fimB-E-X ipuA-B (similar to CFT073). Two human commensal isolate and one ASB isolate were excluded from analysis as they did not carry the fimB gene by PCR analysis.