Pathogenic potential, genetic diversity, and population structure of Escherichia coli strains isolated from a forest-dominated watershed (Comox Lake) in British Columbia, Canada

Abstract

Escherichia coli isolates (n = 658) obtained from drinking water intakes of Comox Lake (2011 to 2013) were screened for the following virulence genes (VGs): stx1 and stx2 (Shiga toxin-producing E. coli [STEC]), eae and the adherence factor (EAF) gene (enteropathogenic E. coli [EPEC]), heat-stable (ST) enterotoxin (variants STh and STp) and heat-labile enterotoxin (LT) genes (enterotoxigenic E. coli [ETEC]), and ipaH (enteroinvasive E. coli [EIEC]). The only genes detected were eae and stx2, which were carried by 37.69% (n = 248) of the isolates. Only eae was harbored by 26.74% (n = 176) of the isolates, representing potential atypical EPEC strains, while only stx2 was detected in 10.33% (n = 68) of the isolates, indicating potential STEC strains. Moreover, four isolates were positive for both the stx2 and eae genes, representing potential EHEC strains. The prevalence of VGs (eae or stx2) was significantly (P < 0.0001) higher in the fall season, and multiple genes (eae plus stx2) were detected only in fall. Repetitive element palindromic PCR (rep-PCR) fingerprint analysis of 658 E. coli isolates identified 335 unique fingerprints, with an overall Shannon diversity (H') index of 3.653. Diversity varied among seasons over the years, with relatively higher diversity during fall. Multivariate analysis of variance (MANOVA) revealed that the majority of the fingerprints showed a tendency to cluster according to year, season, and month. Taken together, the results indicated that the diversity and population structure of E. coli fluctuate on a temporal scale, reflecting the presence of diverse host sources and their behavior over time in the watershed. Furthermore, the occurrence of potentially pathogenic E. coli strains in the drinking water intakes highlights the risk to human health associated with direct and indirect consumption of untreated surface water.

FIG 1

Map showing Comox Lake and sampling locations. (Reprinted from Google Maps.)

FIG 2

Dendrogram showing the relatedness of E. coli strains isolated from Comox Lake during different seasons, as determined by rep-PCR fingerprint analysis using the BOX A1R primer. Because of the large population size (2011, n = 134 [A]; 2012, n = 143 [B]; 2013, n = 58 [C]), a condensed dendrogram (using a 60% cutoff value) is presented. DNA fingerprint similarities were calculated by using the curve-based Pearson coefficient, and dendrograms were generated by UPGMA. Data are shown as season (number of isolates); the total number of isolates in each cluster and percent similarities are given next to the clusters. F, fall; S, summer; W, winter; Sp, spring. Note that the cluster numbers mentioned in the text are presented on the clusters.

FIG 3

MANOVA plot of rep-PCR DNA fingerprint patterns of E. coli strains isolated from Comox Lake in 2011 (n = 215), 2012 (n = 230), and 2013 (n = 213) by year (A) and season (B to D). A binary band-matching character table was analyzed by MANOVA, accounting for variance. Only the first two discriminants are presented in this graph, as shown by the distances along the x and y axes.

FIG 4

MANOVA plot of rep-PCR DNA fingerprint patterns of E. coli strains isolated from Comox Lake during different months of the fall season in 2011 (n = 121) (A), 2012 (n = 143) (B), and 2013 (n = 137) (C). A binary band-matching character table was analyzed by MANOVA, accounting for variance. Only the first two discriminants are presented in this graph, as shown by the distances along the x and y axes.