Virulence behavior of uropathogenic Escherichia coli strains in the host model Caenorhabditis elegans

Abstract

Urinary tract infections (UTIs) are among the most common bacterial infections in humans. Although a number of bacteria can cause UTIs, most cases are due to infection by uropathogenic Escherichia coli (UPEC). UPEC are a genetically heterogeneous group that exhibit several virulence factors associated with colonization and persistence of bacteria in the urinary tract. Caenorhabditis elegans is a tiny, free-living nematode found worldwide. Because many biological pathways are conserved in C. elegans and humans, the nematode has been increasingly used as a model organism to study virulence mechanisms of microbial infections and innate immunity. The virulence of UPEC strains, characterized for antimicrobial resistance, pathogenicity-related genes associated with virulence and phylogenetic group belonging was evaluated by measuring the survival of C. elegans exposed to pure cultures of these strains. Our results showed that urinary strains can kill the nematode and that the clinical isolate ECP110 was able to efficiently colonize the gut and to inhibit the host oxidative response to infection. Our data support that C. elegans, a free-living nematode found worldwide, could serve as an in vivo model to distinguish, among uropathogenic E. coli, different virulence behavior.

Keywords: Caenorhabditis elegans; Escherichia coli; oxidative stress; urinary tract infections; uropathogenic strains.

Figure 1

HEp‐2 cell monolayers were infected by adding logarithmically grown of Escherichia coli strains at a multiplicity of infection of approximately 10 bacteria per cell. Bacterial invasion was measured using a gentamicin protection assay before lysis and plating as described in Materials and methods (*p < 0.05)

Figure 2

Minimum spanning tree based on multilocus concatenated alleles showing the phylogenetic relationship between ECP45 and ECP110 strains and uropathogenic Escherichia coli (UPEC), commensal and environmental origin E. coli strains. Genetic distances, expressed as amount of single nucleotide variants, are showed as number in small circles

Figure 3

(a) Kaplan–Meier survival plots of worms infected with the indicated Escherichia coli strains are shown. Infections were performed at 25°C, and worm mortality was monitored every day. E. coli CFT073‐fed worms were taken as control. (b) Survival of Caenorhabditis elegans fed heat‐killed E. coli strains. n = 60. Statistical analysis was evaluated by Log‐rank (Mantel‐Cox) test; asterisks indicate significant differences (**p < 0.01; ***p < 0.001; ns: not significant)

Figure 4

(a) Colonization of uropathogenic Escherichia coli strains within the nematode gut. coli strains within the nematode gut. Asterisks indicate significant differences (^* p < 0.05; ^** p < 0.01; ^*** p < 0.001). (b) Fluorescence photomicrographs of 10 representative nematodes infected with the GFP‐expressing CFT073, ECP45 and ECP110 strains for 2 days are reported (scale bar, 100 µm)

Figure 5

(a) Fluorescence microscopy of SOD3::GFP worm strain after 48 hr of Escherichia coli strains infection. Scale bar = 100 μm. (b) MFI represents mean fluorescence intensity of Caenorhabditis elegans SOD3::GFP transgenic strain fed different E. coli strains (*p < 0.05; ***p < 0.001; ns: not significant)

Figure 6

ROS production in H₂DCFDA‐stained N2 worms after exposure to pathogenic Escherichia coli strains for 2 days. Statistical analysis was performed with respect to animals fed with CFT073 strain (*p < 0.05; **p < 0.01; ns: not significant)