Molecular Detection of Heat Labile, Heat Stable and Shiga-Toxin E. Coli Genes in Sheep and Goats Research Article

Article Sidebar

PDF
Published: 2025-09-30
DOI: https://doi.org/10.31580/pjmls.v8i3.3242
Keywords:
Escherichia coli, Enterotoxigenic E. coli, Goats, PCR, Sheep, Shiga-toxin producing E. coli

Main Article Content

Sadaf Ijaz
Center for Advance Studies Vaccinology and Biotechnology (CASVAB), University of Balochistan, Quetta, Pakistan
Irfan Shahzad Sheikh
Center for Advance Studies Vaccinology and Biotechnology (CASVAB), University of Balochistan, Quetta, Pakistan, 87300
Muhammad Tahir
Department of Pediatrics (Ward-II), Bolan Medical Complex Hospital, Quetta, Pakistan
Fahmida Naheed
Department of Gynecology, Bolan Medical Complex Hospital, Quetta, Pakistan
Muhammad Nadeem
Faculty of Veterinary and Animals Sciences, PMAS Arid Agriculture University, Rawalpindi, sub-campus Khushab, Pakistan
Tariq Hussain
Department of Basic Sciences, College of Veterinary and Animal Science, Jhang, Pakistan
Daud Khan
Livestock and Dairy Development Department, Balochistan, Pakistan, 87300.
Hina Ali Ahmed
Department of Zoology, Faculty of Life Sciences, Sardar Bahadur Khan Women’s University, Quetta, Pakistan
Komal Abid
Center for Advance Studies Vaccinology and Biotechnology (CASVAB), University of Balochistan, Quetta, Pakistan, 87300.
Saima Asif
Institute of Molecular Biology and Biotechnology, Bahauddin Zakariya University, Multan, Pakistan
Euzah Shahzadi
Faculty of Veterinary and Animal Sciences, The Islamia University of Bahawalpur, Bahawalpur, Pakistan
Hafiz Muhammad Ali
Faculty of Veterinary and Animal Sciences, The Islamia University of Bahawalpur, Bahawalpur, Pakistan
Abdul Samad
Center for Advance Studies Vaccinology and Biotechnology (CASVAB), University of Balochistan, Quetta, Pakistan, 87300.
Ishtiaq Ahmad
Internal Medicine Resident, Geisinger Wyoming Valley Medical Center Wilkes-Barre, PA, United States

Abstract

E. coli cause diarrhea in sheep and goats through its virulent strains, enterotoxigenic E. coli (ETEC) and shiga-toxin producing E. coli (STEC). The present study was conducted for molecular detection of E. coli and its pathogenic strains, including heat labile (elt), heat stable (est) and shiga-toxin (stx1, stx2) producing genes from diarrheal fecal samples of small ruminants. About 155 fecal diarrheal fecal samples were randomly collected from sheep (n=88) and goats (n=67) and checked for the presence of E. coli by microbiological techniques. The inoculation of samples onto EMB agar revealed green sheen colonies that appeared pink, rod-shaped bacilli on Gram staining. DNA extraction was performed through boiling and the phenol-chloroform method, and a PCR assay was done for the detection of the uidA gene of E. coli, along with the elt, est, stx1 and stx2 genes of ETEC and STEC strains. PCR assay confirmed the presence of E. coli in 96% isolates by the uidA gene in sheep and goats. The results revealed the presence of est (84.5%), stx1 (81.8%) and stx2 (61.1%) virulence genes, but none of the isolates detected elt gene. The findings indicate that sheep and goat are a reservoir of pathogenic E. coli possessing est, stx1 and stx2 coding genes and thus, the presence of these virulence factors is a potential harm for human health.

Article Details

Issue

Vol. 8 No. 3 (2025): July-September 2025

Section

Research Article
Creative Commons License

This work is licensed under a Creative Commons Attribution 4.0 International License.

How to Cite

Molecular Detection of Heat Labile, Heat Stable and Shiga-Toxin E. Coli Genes in Sheep and Goats: Research Article. (2025). Pak-Euro Journal of Medical and Life Sciences, 8(3), 723-730. https://doi.org/10.31580/pjmls.v8i3.3242

References

1. Lee W, Kim MH, Sung S, Kim E, An ES, Kim SH, Kim HY. Genome-based characterization of hybrid Shiga toxin-producing and enterotoxigenic Escherichia coli (STEC/ETEC) strains isolated in South Korea, 2016–2020. Microorganisms, 2023;11(5):1285.

2. Malabadi RB, Sadiya MR, Kolkar KP, Chalannavar RK. Pathogenic Escherichia coli (E. coli) food borne outbreak: Detection methods and controlling measures. Magna Scientia Advanced Research and Reviews, 2024;10(1):052-085.

3. Croxen M, Finlay B. Molecular mechanisms of Escherichia coli pathogenicity. Nature Reviews Microbiology. 2010;8: 26–38.

4. Pokharel P, Dhakal S, Dozois CM. The Diversity of Escherichia coli Pathotypes and Vaccination Strategies against This Versatile Bacterial Pathogen. Microorganisms. 2023;11(2):344.

5. Aslam J, Ali HM, Hussain S, Ahmad MZ, Siddique AB, Shahid M, Shahzad MI, Fatima H, Tariq S, Sadiq F, Aslam M, Farooq U, Zia S, Aljaluod RS, Alarjani KM. Effectiveness of cephalosporins in hydrolysis and inhibition of Staphylococcus aureus and Escherichia coli biofilms. Journal of Veterinary Science. 2024;25(3):e47.

6. Goulart DB, Mellata M. (2022) Escherichia coli Mastitis in Dairy Cattle: Etiology, Diagnosis, and Treatment Challenges. Frontiers in Microbiology. 13:928346.

7. Wang F, Chen L, Li Y, Gao G, Wang Y, Shi R, Zhang H, Chen Y, Cao R, Shi Q, Wu T. Lacticaseibacillus paracasei subsp. paracasei Q- 1 Exhibits Good Safety and Effectively Prevents Escherichia coli K99-Induced Diarrhea in Mice. Probiotics and antimicrobial proteins, 2025;10.1007/s12602-025-10552-z.

8. Navidinia M, Karimi A, Rahbar M, Fallah F, Ahsani RR, Malekan MA, Jahromi MH, Gholinejad Z. Study Prevalence of Verotoxigenic E.coli Isolated from Urinary Tract Infections (UTIs) in an Iranian Children Hospital. Open Microbiology Journal. 2012;6:1-4.

9. Li Y, Xia S, Jiang X, Feng C, Gong S, Ma J, Yin Y. Gut microbiota and diarrhea: an updated review. Frontiers in Cellular and Infection Microbiology. 2021;11:625210.

10. Ramatla T, Tutubala M, Motlhaping T, de Wet L, Mokgokong P, Thekisoe O, Lekota K. Molecular detection of Shiga toxin and extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli isolates from sheep and goats. Molecular Biology Reports. 2024;51(1):57.

11. Gurjar T, Singathia R, Sharma DK, Gaurav A, Solanki S, Kumari M, Rathore K. Escherichia coli in diarrhoeic lambs: Prevalence, virulence and antibiotic resistance. Polish Journal of Veterinary Sciences. 2024;451-457.

12. Butt S, Saleh M, Gagnon J. Impact of the Escherichia coli heat-stable enterotoxin b (STb) on gut health and function. Toxins. 2020;12(12):760.

13. Wang, H., Zhong, Z., Luo, Y., Cox, E., & Devriendt, B. Heat-stable enterotoxins of enterotoxigenic Escherichia coli and their impact on host immunity. Toxins. 2019;11(1):24.

14. Do KH, Seo K. Animal models of Shiga toxin producing Escherichia coli infection, Journal of Applied Animal Research. 2024;52(1):2300625.

15. Gambushe SM, Zishiri OT, El Zowalaty ME. Review of Escherichia coli O157:H7 Prevalence, Pathogenicity, Heavy Metal and Antimicrobial Resistance, African Perspective. Infection and Drug Resistance. 2022;15:4645-4673.

16. Berry ED, Wells JE. 2010. Chapter 4 - Escherichia coli O157:H7: Recent Advances in Research on Occurrence, Transmission, and Control in Cattle and the Production Environment. Editor(s): Steve L. Taylor. Advances in Food and Nutrition Research. Academic Press 60:67-117.

17. Paray AA, Singh M, Mir MA, Kaur A. Gram staining: a brief review. International Journal of Research and Review. 2023;10(9):336-341.

18. Ali HM, Urbinati G, Chapuis H, Desmaële D, Bertrand JR, Couvreur P, Massaad-Massade L. Effects of siRNA on RET/PTC3 junction oncogene in papillary thyroid carcinoma: from molecular and cellular studies to preclinical investigations. PLoS ONE 2014;9(4):e95964.

19. Urbinati G, de Waziers I, Slamiç M, Foussignière T, Ali HM, Desmaële D, Couvreur P, Massaad-Massade L. Knocking down TMPRSS2-ERG fusion oncogene by siRNA could be an alternative treatment to flutamide. Molecular Therapy - Nucleic Acids. 2016:5:e301.

20. He F. E. coli genomic DNA extraction. Bio-protocol 2011;1(14): e97.

21. Cui S, Wei Y, Li C, Zhang J, Zhao Y, Peng X, Sun F. Visual loop mediated isothermal amplification (LAMP) assay for rapid on-site detection of Escherichia coli O157: H7 in milk Products. Foods. 2024;13(13):2143.

22. Al-Sabawi AH, Jwher DM. Isolation and characterization of stx1 and stx2 toxin producing Escherichia coli in neonatal lambs with diarrhea in Nineveh governorate, Iraq. Journal of Applied Veterinary Sciences, 2022;7(4):23-27.

23. Wang L, Wakushima M, Kamata Y, Nishikawa Y. Exhaustive isolation of diarrhoeagenic Escherichia coli by a colony hybridization method using hydrophobic grid‐membrane filters in combination with multiplex real‐time PCR. Letters in Applied Microbiology. 2011;53(3):264-270.

24. Osman KM, Mustafa AM, Elhariri M, Abdelhamed GS. The distribution of Escherichia coli serovars, virulence genes, gene association and combinations and virulence genes encoding serotypes in pathogenic E. coli recovered from diarrhoeic calves, sheep and goat. Transboundary and Emerging Diseases. 2012;60(1):69-78.

25. Mokhbatly AA, Elsheikh N, Ghazy EW, Elgamal AM, Hegazy YM, Assar DH. Prevalence of Shiga toxin-producing Escherichia coli and Salmonellae and some associated hematologic and biochemical profile alterations in lambs. Veterinary Research Forum. 2022;13(2):155-162.

26. Hafez AA. Virulence and antimicrobial resistance genes of E. coli isolated from diarrheic sheep in the North-West Coast of Egypt. Systematic Reviews in Pharmacy. 2020;11(11):609 617.

27. Deng YF, Wang K, Zhang LJ, Yan K, He SJ. Detection of virulence genes, antibiotic resistance genes and antibiotic resistance of Escherichia coli isolated from diarrheic lambs in Anhui Province, China. Pesquisa Veterinária Brasileira. 2024;44:e07346.

28. Türkyılmaz S, Eskiizmirliler S, Tunaligil S, Bozdogan B. Identification, characterization and molecular epidemiology of Escherichia coli isolated from lamb and goat kids with diarrhoea. Acta Veterinaria Brno. 2014;82(4):357-362.

29. Ferreira MR, Silva TDS, Stella AE, Conceição FR, Reis EFD, Moreira CN. Detection of virulence factors and antimicrobial resistance patterns in shiga toxin-producing Escherichia coli isolates from sheep. Pesquisa Veterinária Brasileira. 2015;35(9):775-780.

30. El-Dabae WH, Syame SM, Elgabry EA, Hassan MM, Farahat E. Detection of Escherichia coli and Associated Shiga Toxin Genes in Feces of Cattle and Sheep Reared in Egyptian Research Farm. Egyptian Journal of Veterinary Sciences. 2025;56(8):1673-1679.

31. van Hoek AHAM, Lee S, van den Berg RR, Rapallini M, van Overbeeke L, Opsteegh M, Bergval I, Wit B, van der Weijden C, van der Giessen J, van der Voort M. Virulence and antimicrobial resistance of Shiga toxin-producing Escherichia coli from dairy goat and sheep farms in The Netherlands. Journal of Applied Microbiology. 2023;134(6):lxad119.

32. Manzoor R, Shah MI, Wani SA, Pandit F, Dar PA, Mir MI. Prevalence, serodiversity and antibiogram of enterotoxigenic Escherichia coli (ETEC) in diarrhoeic calves and lambs of Kashmir valley (J&K), India. Journal of Applied and Natural Science. 2015;7(1):477-481.

33. Njoroge S, Muigai AWT, Njiruh PN, Kariuki S. Molecular characterisation and antimicrobial resistance patterns of Escherichia coli isolates from goats slaughtered in parts of Kenya. East African Medical Journal. 2013:90(3):72-83.