Development of a multiplex PCR protocol for the detection of several bacteria causing acute diarrhea in pig

Authors

  • Nguyen Tuan Anh Khoa Thuong Biotechnology Company
  • Nguyen Thi Minh Tam Khoa Thuong Biotechnology Company
  • Trinh Thị Thanh Huyen Center of Biotechnology Applications, Dong Nai
  • Ngo Quang Huong Center of Biotechnology Applications, Dong Nai
  • Phi Thi Thu Hien Center of Biotechnology Applications, Dong Nai
  • Nguyen Thi Hoang Center of Biotechnology Applications, Dong Nai

DOI:

https://doi.org/10.15625/0866-7160/v36n1se.4334

Keywords:

Clostridium perfringens, Escherichia coli, Salmonella, diarrhea, multiplex PCR.

Abstract

We established a multiplex PCR-based protocol for the detection of three bacteria Salmonella spp., Clostridium perfringens and E. coli ETEC causing acute diarrhea in pig from field samples, feces and intestinal biopsies. The PCR protocol was built up with technical parameters, such as primer concentrations that were 350 nM (Salmonella spp.), 200 nM (C. perfringens), 200 nM (E. coli ETEC), 50 nM internal control primer (porcine β-actin) and the annealing temperature was 61oC for all of primers used. The sensitivity of the protocol was found to be 5´103 copies/reaction. The protocol was also screened for the presence of the bacteria in parallelly comparing with commercial kits on field samples and indicated that it could reliably detect three bacterial species with the Kappa values correlatively to be 1.0, 0.45 and 0.56. Finally, the protocol was set up successfully and could become a convenient, rapid, specific and accurate diagnostic kit to detect simultaneously three kinds of bacteria in field samples. Moreover, it could be used as a tool to study bacterial infection epidemiology in pig population.

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Published

04-08-2014

How to Cite

Anh, N. T., Minh Tam, N. T., Thanh Huyen, T. T., Huong, N. Q., Thu Hien, P. T., & Hoang, N. T. (2014). Development of a multiplex PCR protocol for the detection of several bacteria causing acute diarrhea in pig. Academia Journal of Biology, 36(1se), 8–14. https://doi.org/10.15625/0866-7160/v36n1se.4334

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