Duplex PCR for specific detection of Escherichia coli and its differentiation from other Enterobacteriaceae

Escherichia coli is a normal inhabitant of gastrointestinal tract of humans and animals and one of the most important causes of bovine mastitis. Definitive identification of E. coli from other members of Enterobacteriaceae remains ambiguous. The present study describes a duplex PCR, targeting 2 housekeeping genes, the lacy (lactose permease) and phoA (alkaline phosphatase) for the reliable detection of E. coli that enables its differentiation from biochemically and phylogenetically related bacteria. The assay was evaluated employing 19 ATCC (American type culture collection) reference strains of Enterobacteriaceae family. Validation of the assay with E. coli (154) isolated from milk and faeces rendered the assay to be specific. The results suggest that the technique can be used for accurate detection of E. coli and thus can be adapted for testing bacteriological safety of milk, for field applications, and in laboratories handling clinical samples. This PCR (polymerase chain reaction) can successfully distinguish E. coli including E. coli O157 from Shigella spp and other related enterobacteria, emphasizing its relevance and utility in studies related to E. coli infection.