Rapid Detection of Enterotoxigenic Staphylococcus aureus by Polymerase Chain Reaction

중합효소 연쇄반응에 의한 식중독성 황색포도상구균의 신속한 검출

  • Kim, Eun-Seon (Department of Biology, Chonnam National University) ;
  • Jhon, Deok-Young (Department of Food and Nutrition, Chonnam National University)
  • 김은선 (전남대학교 생물학과) ;
  • 전덕영 (전남대학교 식품영양학과)
  • Published : 1996.12.30

Abstract

Staphylococcal food poisoning is the major cause of bacterial food poisoning occurring in this country. Therefore government regulates commercial foods through Official Dictionary of Food that there should be free of enterotoxigenic Staphylococcus aureus in Korean rice cakes, bread, and a box lunch. Since at least 5 days are required to identify the S. aureus by the official method in the Dictionary it is difficult to prevent the food poisoning and the investigation of the outbreaks. In this report an improved determination method of the S. aureus has been developed using polymerase chain reaction (PCR) technique. Sense and antisense primers for specific amplification of genes encoding staphylococcal enterotoxins were designed and synthesized for the PCR. Rapid chromosomal DNA isolation method was also developed from S. aureus using lysostaphin. The PCR condition was developed as follows. Reaction solution $(50\;{\mu}l)$ consisted of target DNA $2\;{\mu}l$ (about 20ng), 10X buffer $5\;{\mu}l$, primer 100pmole, dNTP (10 mM) $4\;{\mu}l$ and Taq DNA polymerase 2.5 unit in a thin-wall tube. Operation condition of the PCR was 5 min pre-denaturation at $94^{\circ}C$, 15 sec denaturation at $94^{\circ}C$, 15 sec annealing at $50^{\circ}C$, 20 sec extension at $72^{\circ}C$, and 5 min post-extension at $72^{\circ}C$, and 30 cycles of denaturation-annealing- extension. Using the PCR with Perkin Elmer GeneAmp PCR system 2400, types of enterotoxigenic S. aureus could be identified from Ddok or bread in a day.

Keywords

Staphylococcus aureus;enterotoxin detection;polymerase chain reaction (PCR);Korean rice cake