Study of an ELISA method for the detection of E. coli O157 in food

Abstract

A new ELISA assay, so-called Transia Plate E. coli O157, was developed for monitoring E. coli O157 contamination in food samples. The performances of this new method were compared to the gold standard method (immunomagnetic separation combined with agar plate) based on an enrichment step in buffer peptone water, followed by an immunoseparation with magnetic beads, then streaking onto Cefixime Tellurite Sorbitol Mac Conkey agar (CT-SMAC). The ELISA was composed of 3 steps, with incubation times of 20, 10 and 5 minutes respectively. This immunoassay was demonstrated specific for E. coli O157 and showed a limit of detection of around 5×10 4 CFU/ml with several strains. Among the different other strains evaluated (47 non-E. coli O157 strains), only two Salmonella , belonging to the serogroup N, were crossreacting with the ELISA. The best enrichment protocol was combining either mTSBn or mECn, with an incubation time between 16 to 20 hours at a temperature of 41.5°C. This temperature allowed an easier confirmation of the positive ELISA samples once isolated onto CT-SMAC plate. The limit of detection with artificially contaminated samples was assessed in the range of 1 to 3 CFU/25g of food for both methods (ELISA and gold standard). Finally, naturally contaminated samples (n=47, raw meat and dairy products) were analysed with the Transia Plate E. coli O157 method. Two positive samples were detected in the raw meat samples, and were confirmed as E. coli O157 without any verotoxin production. The Transia Plate E. coli O157 method was representing a good alternative to the IMS method as it was highly specific for E. coli O157 and proposed an objective analysis of food samples, without any cumbersome step such as immunocapture. This method improved also the safety as the ELISA was done with heat treated samples. It allowed an effective screening of food samples within 45 minutes, and avoided streaking and reading of agar plates for all the samples.