Molecular detection of Shiga toxin-producing Escherichia coli (STEC) O157 in sheep, goats, cows and buffaloes

Abstract

Background: Shiga toxin-producing E. coli (STEC) are important foodborne pathogens that causing serious public health consequences worldwide. The present study aimed to estimate the prevalence ratio and to identify the zoonotic potential of E. coli O157 isolates in slaughtered adult sheep, goats, cows and buffaloes.

Materials and methods: A total of 400 Recto-anal samples were collected from two targeted sites Rawalpindi and Islamabad. Among them, 200 samples were collected from the slaughterhouse of Rawalpindi included sheep (n = 75) and goats (n = 125). While, 200 samples were collected from the slaughterhouse of Islamabad included cows (n = 120) and buffalos (n = 80). All samples were initially processed in buffered peptone water and then amplified by conventional PCR. Samples positive for E. coli O157 were then streaked onto SMAC media plates. From each positive sample, six different Sorbitol fermented pink-colored colonies were isolated and analyzed again via conventional PCR to confirm the presence of rfbE O157 gene. Isolates positive for rfbE O157 gene were then further analyzed by multiplex PCR for the presence of STEC other virulent genes (sxt1, stx2, eae and ehlyA) simultaneously.

Results: Of 400 RAJ samples only 2 (0.5%) showed positive results for E. coli O157 gene, included sheep 1/75 (1.33%) and buffalo 1/80 (1.25%). However, goats (n = 125) and cows (n = 120) found negative for E. coli O157. Only 2 isolates from each positive sample of sheep (1/6) and buffalo (1/6) harbored rfbE O157 genes, while five isolates could not. The rfbE O157 isolate (01) of sheep sample did not carry any of STEC genes, while the rfbE O157 isolate (01) of buffalo sample carried sxt1, stx2, eae and ehlyA genes simultaneously.

Conclusion: It was concluded that healthy adult sheep and buffalo are possibly essential carriers of STEC O157. However, rfbE O157 isolate of buffalo RAJ sample carried 4 STEC virulent genes, hence considered an important source of STEC infection to humans and environment which should need to devise proper control systems.

Keywords: Conventional PCR; Escherichia coli (STEC) O157; Multiplex PCR; RAJ.