Non-Melibiose Fermentation and Tellurite Resistance by Shigatoxigenic and Enteropathogenic Escherichia coli O80:H2 from Diseased Calves: Comparison with Human Shigatoxigenic E. coli O80:H2

Abstract

Despite their prevalence in Europe, the source of contamination of humans by Attaching-Effacing Shigatoxigenic Escherichia coli (AE-STEC) O80:H2 remains unidentified. This study aimed to assess a procedure based on non-melibiose fermentation and resistance to tellurite to isolate AE-STEC and enteropathogenic (EPEC) O80:H2 from healthy cattle. The genome sequences of 40 calf and human AE-STEC and EPEC O80:H2 were analyzed: (i) none harbored the mel operon, but the 70mel DNA sequence instead; (ii) the ter-type 1 operon was detected in 16 EPEC and stx1a or stx2a AE-STEC, while no ter-type 1 operon was detected in the remaining 24 EPEC and stx2d AE-STEC. The 21 calf AE-STEC and EPEC O80:H2 were tested phenotypically: (i) none fermented melibiose on melibiose-MacConkey agar plates; (ii) ten of the 11 ter-type 1-positive strains had Minimal Inhibitory Concentrations (MIC) ≥ 128 µg/mL to potassium tellurite; (iii) conversely, the ten ter-negative strains had MIC of two µg/mL. Accordingly, enrichment broths containing two µg/mL of potassium tellurite and inoculated with one high MIC (≥256 µg/mL) stx1a AE-STEC O80:H2 tested positive with the O80 PCR after overnight growth, but not the enrichment broths inoculated with one low MIC (two µg/mL) EPEC. Nevertheless, neither AE-STEC nor EPEC O80:H2 were recovered from 96 rectal fecal samples collected from healthy cattle at one slaughterhouse after overnight growth under the same conditions. In conclusion, this procedure may help to isolate stx1a and stx2a AE-STEC and EPEC O80:H2, but not stx2d AE-STEC that are tellurite sensitive, and new surveys using different procedures are necessary to identify their animal source, if any.

Keywords: Shigatoxigenic E. coli; enteropathogenic E. coli; healthy cattle; melibiose fermentation; serotype O80:H2; tellurite resistance; zoonosis.

Figure 1

Minimal Inhibitory Concentration (MIC) determination to potassium tellurite in presence of bromocresol purple as a pH indicator. The change of color from yellow-orange to dark purple marks bacterial growth and by deduction the MIC (one strain per row). ¹ C+: positive growth control (column 1); C-: negative growth control (column 2). ² Potassium tellurite concentrations (μg/mL) in columns 3 to 12.