P-072. Optimization of Raw Milk and Raw Milk Products Enrichment Step before E. coli O26 Detection

D. Thevenot, M. Bouvier, A. Gleizal, C. Vernozy-Rozand;
Ecole Natl. Vét. de Lyon, Marcy l'Etoile, FRANCE.

Background: EHEC O26 strains constitute the most important common non-O157 EHEC group associated with diarrhea and haemolytic uremic syndrome in Europe. E. coli O26 was involved in several outbreaks and sporadic cases world-wide. An outbreak was reported in France in 2005 implicating raw milk cheese. Consequently, it appears essential to develop methods to detect E. coli O26 from such complex foods. The aim of this study was to optimize the enrichment step for the E. coli O26 detection from raw milk and raw milk products. Methods: a total of 12 different raw milk and ruminant (goat, caw) raw milk products (soft and hard cheeses) were experimentally inoculated with 1 to 5 CFU/25g of a single E. coli O26 strain. Three transformed E. coli O26 strains (with a plasmid vector carrying the green fluorescent protein) were used with a view to numerate E. coli O26 strains among the abundant background microflora. Several enrichment protocols (one and two steps) and enrichment conditions were tested. Results: the study pointed out that an 8 hours enrichment in buffered peptone water (BPW) supplemented with acryflavine (5mg/ml) allowed the growth of E. coli O26 (104 to 105 CFU/ml) for most of the raw milk cheese except for the “Camembert” type cheeses. For them, 24 hours of incubation at 41.5°C and the addition of cefixime and tellurite in the BPW medium were required. This enrichment protocol was proved to be suitable for all the different type of raw milk cheese manufactured in France. Conclusion: the optimised enrichment step described for the recovery of very low number of E. coli O26 in dairy complex products could be used before an immunological or a genetic detection of the target.