Isolation of Shiga toxin-producing Escherichia coli O157 and non-O157 from retail imported frozen beef marketed in Saudi Arabia using immunomagnetic separation and multiplex PCR.

Abstract

Introduction: Shiga toxin-producing Escherichia coli (STEC), particularly E. coli O157:H7, is a major contributor to foodborne outbreaks globally. Both E. coli O157 and non-O157 strains can lead to severe health issues, including hemolytic colitis and hemolytic uremic syndrome, which can result in kidney failure.

Methods: Two hundred and one frozen beef samples were purchased from various supermarkets located in the Eastern Province of Saudi Arabia and subsequently enriched in tryptic soy broth (TSB). From the enriched samples in TSB, 1 mL portion was mixed with immunomagnetic beads (IMB) coated with specific antibodies targeting the E. coli O157 O antigen. The beads, which contained the captured bacteria, were then streaked onto CHROMagar O157 and Sorbitol MacConkey (SMAC) agar. The DNA extracted from these samples was examined using multiplex PCR to identify potential virulence gene markers, specifically stx-1, stx-2, and eae.

Results: Of the 201 examined samples, 88 (43.8%) and 106 (52.7%) were positive for E. coli and produced colorless and mauve colonies on SMAC agar and CHROMagar O157, respectively. Out of 298 isolates in total, 174 isolates of E. coli were isolated with IMB enrichment. The highest detection rate of virulence gene markers was found among isolates that had been isolated using IMB enrichment, where 25 (8.4%), 2 (0.7%) and 12 (4%) isolates tested positive for stx1, stx2, and eae genes respectively. Among 42 isolates harboring potential virulence gene markers, 11 isolates were identified as E. coli O157 (stx ₁ ⁺/eae ⁺ or stx ₂ ⁺/eae ⁺). ERIC-PCR genotyping was able to determine the genetic relatedness among 42 isolates of E. coli O157 and E. coli non-O157 into 10 types with four identical related clusters and a genetic similarity rate above 90% homology from the identified isolates.

Conclusions: The present study gives a clear perspective on STEC contamination in imported frozen beef marketed in Saudi Arabia. Because of the many possibilities of STEC contamination in imported frozen beef, further studies on the spread of STEC at various levels of imported frozen meat are needed on a long-term basis.