세균성 설사 원인균의 신속검출을 위한 다중 중합효소연쇄반응 검사의 대변검체에 대한 직접 적용
Direct Application of Multiplex PCR on Stool Specimens for Detection of Enteropathogenic Bacteria
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배경: 감염성 설사 원인균의 진단은 주로 배양에 이해 이루어졌으나 이러한 방법은 민감도가 상대적으로 낮고 시간이오래 걸리는 단점을 가진다. 본 연구에서는 다양한 설사 원인균의 신속 동정을 위해 다중 중합효소연쇄반응을 환자의대변 검체에 직접 적용하여 평가하였다. 방법: 2009년 6월부터 9월까지 대변배양이 의뢰된 173건의 설사 검체에 SeeplexⓇ Diarrhea ACE Detection kit (Seegene,Seoul, Korea)를 이용하여 다중 중합효소연쇄반응을 실시하였다. 모든 검체에 대하여 다중 중합효소연쇄반응 결과와Salmonella, Shigella, Vibrio, Yersinia를 검출하기 위한 통상적인 배양법 결과를 비교하였다. 후반부의 50 검체에 대하여는Campylobacter 배양이 이루어졌다. 다중 중합효소연쇄반응에서 verotoxin 생산 Escherichia coli (VTEC)가 양성인 검체는장출혈성 E. coli O157:H7 진단을 위해 계대배양을 실시하였다. 또한 환자의 의무기록을 후향적으로 조사하여 임상 증상과 검사결과를 확인하였다. 결과: 총 173건의 검체 중 다중 중합효소연쇄반응과 배양방법에 의해 각각 36검체(20.8%)와 8검체(4.6%)에서 설사원인균이 동정되었다. 다중 중합효소연쇄반응에서는 Salmonella 5건, Campylobacter 15건, Vibrio 1건, Clostridium difficile toxin B 4건, Clostridium perfringens 5건, Yersinia enterocolitica 1건, Aeromonas 5건, VTEC 2건으로 검출되었고, 배양검사법으로는 Samonella 5건, Vibrio 1건, Y. enterocolitica 1건, Aeromonas 1건, E. coli O157:H7 2건이 진단되었다. 결론: 다중 중합효소연쇄반응 방법은 Campylobacter, VTEC and C. perfringens 등과 같은 배양이 어려운 장병원성 균의검출에 매우 유용하였을 뿐만 아니라, 일반적인 장병원성균인 Salmonella, Shigella, Vibrio, Y. enterocolitica에 대해서는통상배양법과 유사한 민감도를 나타냈다. 통상 배양검사와 더불어 다중 중합효소연쇄반응을 대변 검체에 직접 적용하는것은 세균성 장염 원인균 진단의 민감도를 크게 향상시킬 수 있을 것이다. [대한임상미생물학회지 2010;13:162-168]
Abstract
Background: Causative bacterial agents of infectious diarrheal disease were traditionally diagnosed by stool cultures. Stool culture, however, has a problem because of relatively low sensitivity and long turnaround time. In this study, we evaluated multiplex PCR applied on stool specimens directly to diagnose enteropathogenic bacteria. Methods: From June to September 2009, 173 diarrheal stools submitted for stool cultures were tested by SeeplexⓇ Diarrhea ACE Detection kit (Seegene,Korea) to detect 10 enteropathogenic bacteria. Specimens were cultured for Salmonella, Shigella, Vibrio,and Yersinia. Late 50 specimens were also cultured for Campylobacter. The specimens positive for verotoxin-producing Escherichia coli (VTEC) were further subcultured for detecting enterohaemorrhagic Escherichia coli O157:H7. Electronic medical records were reviewed for clinical and laboratory findings. Results: Of 173 specimens, multiplex PCR and cultures identified enteropathogens in 36 (20.8%) and 8 specimens (4.6%), respectively. While multiplex PCR detected 5 Salmonella, 15 Campylobacter, 1Vibrio, 4 Clostridium difficiles toxin B, 5 Clostridium perfringens, 1 Yersinia enterocolitica, 5 Aeromonas,and 2 VTEC, cultures detected 5 Salmonella, 1Vibrio, 1 Y. enterocolitica, 1 Aeromonas, and 2 E. coli O157:H7. Conclusion: Multiplex PCR would be useful to detect Campylobacter, VTEC and C. perfringens, as well as have equivalent sensitivity to conventional culture for ordinary enteropathogens such as Salmonella,Shigella, Vibrio, Y. enterocolitica. Direct application of multiplex PCR combined with conventional cultures on stool warrants remarkable improvement of sensitivity to diagnose enteropathogenic bacteria. (Korean J Clin Microbiol 2010;13:162-168)
참고문헌 (28)
- Baldi F, "Focus on acute diarrhoeal disease", World J Gastroenterol, 15, pp.3341~3348 (2009)
- Vernacchio L, "Diarrhea in American infants and young children in the community setting: incidence, clinical presentation and microbiology", Pediatr Infect Dis J, 25, pp.2~7 (2006)
- Farthing MJ, "Diarrhoea: a significant worldwide problem", Int J Antimicrob Agents, 14, pp.65~69 (2000)
- Tarr PI, "Shiga-toxin-producing Escherichia coli and haemolytic uraemic syndrome", Lancet, 365, pp.1073~1086 (2005)
- Thielman NM, "Clinical practice. Acute infectious diarrhea", N Engl J Med, 350, pp.38~47 (2004)
- Guerrant RL, "Thielman NM, Slutsker L, Tauxe RV, et al. Practice guidelines for the management of infectious diarrhea", Clin Infect Dis, 32, pp.331~351 (2001)
- Cheng AC, "Infectious diarrhea in developed and developing countries", J Clin Gastroenterol, 39, pp.757~773 (2005)
- Thapar N, "Diarrhoea in children: an interface between developing and developed countries", Lancet, 363, pp.641~653 (2004)
- Gadewar S, "Current concepts in the evaluation, diagnosis and management of acute infectious diarrhea", Curr Opin Pharmacol, 5, pp.559~565 (2005)
- Shin HB, "Isolation trend of enteropathogenic bacteria in 1969-1998", Korean J Clin Microbiol, 4, pp.87~95 (2001)
- "The prevalence and characteristics of bacteria causing acute diarrhea in Korea"[URL]
- Iijima Y, "Improvement in the detection rate of diarrhoeagenic bacteria in human stool specimens by a rapid real-time PCR assay", J Med Microbiol, 53, pp.617~622 (2004)
- Reller ME, "Yield of stool culture with isolate toxin testing versus a two-step algorithm including stool toxin testing for detection of toxigenic Clostridium difficile", J Clin Microbiol, 45, pp.3601~3605 (2007)
- Chapela MJ, "Comparison between a TaqMan polymerase chain reaction assay and a culture method for ctx-positive Vibrio cholerae detection", J Agric Food Chem, 58, pp.4051~4055 (2010)
- Nhung PH, "Rapid and specific identification of 5 human pathogenic vibrio species by multiplex polymerase chain reaction targeted to dnaJ gene", Diagn Microbiol Infect Dis, 59, pp.271~275 (2007)
- Daube G, "Hybridization of 2,659 Clostridium perfringens isolates with gene probes for seven toxins (alpha, beta, epsilon, iota, theta, mu, and enterotoxin) and for sialidase", Am J Vet Res, 57, pp.496~501 (1996)
- 장윤하, "Toxigenic Clostridium difficile 배양에서 종동정과 독소확인 다중 PCR 검사의 적용", 대한임상미생물학회지, 12(1), pp.11~16 (2009), 대한임상미생물학회
- Paton AW, "Multiplex PCR for direct detection of Shiga toxigenic Escherichia coli strains producing the novel subtilase cytotoxin", J Clin Microbiol, 43, pp.2944~2947 (2005)
- Kim JS, "A novel multiplex PCR assay for rapid and simultaneous detection of five pathogenic bacteria: Escherichia coli O157:H7, Salmonella, Staphylococcus aureus, Listeria monocytogenes, and Vibrio parahaemolyticus", J Food Prot, 70, pp.1656~1662 (2007)
- O'Leary J, "Comparison of the EntericBio multiplex PCR system with routine culture for detection of bacterial enteric pathogens", J Clin Microbiol, 47, pp.3449~3453 (2009)
- Butzler JP, "Campylobacter, from obscurity to celebrity", Clin Microbiol Infect, 10, pp.868~876 (2004)
- Thorney JP, "Virulence properties of clinically significant Aeromonas species: evidence for pathogenicity", Rev Med Microbiol, 8, pp.61~72 (1997)
- Choi JP, "Clinical significance of spontaneous Aeromonas bacterial peritonitis in cirrhotic patients: a matched case-control study", Clin Infect Dis, 47, pp.66~72 (2008)
- 강재명, "Aeromonas 균혈증의 임상적 특징과 예후와 관련된 위험요인", Infection and Chemotherapy, 37(3), pp.161~166 (2005), 대한감염학회
- Labbe R, "Clostridium perfringens in Foodborne Bacterial Pathogens", pp.191~234 (1989), Marcel Dekker, Inc.
- Kokai-Kun JF, "Comparison of western immunoblots and gene detection assays for identification of potentially enterotoxigenic isolates of Clostridium perfringens", J Clin Microbiol, 32, pp.2533~2539 (1994)
- Lee SW, "A study on epidemiological characteristics and control methods of EHEC infection in Korea", Korean J Epidemiol, 27, pp.37~52 (2005)
- Oh YH, "Use of polymerase chain reaction and serum antibodies for diagnosis of enterohemorrhagic Escherichia coli", J Korean Soc Microbiol, 33, pp.99~110 (1998)