Identification of shiga toxin producing Escherichia coli O157:H7 in raw cow milk samples from dairy farms in Mashhad using multiplex PCR assay

In this study 130 bulk tank milk samples which were delivered to the Pegah Pasturisation Factory in
Mashhad were collected randomly during the summer months. Samples were firstly enriched in modified
trypticase soy broth containing novobiocin, followed by plating onto sorbitol MacConkey agar supplemented
with cefixime and potassium tellurite for isolation of Escherichia coli O157:H7. Consequently the suspected
non-sorbitol fermenting (NSF) colonies were confirmed by biochemical tests as Escherichia coli and then
were used for multiplex-PCR assay, using primers specific for O157 and H7 antigens genes and then primers
specific for stx1 and stx2 genes. NSF Escherichia coli colonies were recovered from 8 samples, and in multiplex-PCR assay one sample (0.77%) was confirmed as Escherichia coli O157:H7. The second multiplex
PCR assay showed that the isolate was harboring the stx2 gene. The PCR assay used in this study may be a
possible alternative to immunological assays which detect somatic and flagellar antigens. Besides, this
procedure determines the potential of isolates for toxin production.