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

다국어 초록 (Multilingual 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 (Reference)

1 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" 45 : 3601-3605, 2007

2 Thorney JP, "Virulence properties of clinically significant Aeromonas species: evidence for pathogenicity" 8 : 61-72, 1997

3 Oh YH, "Use of polymerase chain reaction and serum antibodies for diagnosis of enterohemorrhagic Escherichia coli" 33 : 99-110, 1998

4 장윤하, "Toxigenic Clostridium difficile 배양에서 종동정과 독소확인 다중 PCR 검사의 적용" 대한임상미생물학회 12 (12): 11-16, 2009

5 Guerrant RL, "Thielman NM, Slutsker L, Tauxe RV, et al. Practice guidelines for the management of infectious diarrhea" 32 : 331-351, 2001

6 Korea Center for Disease Control & Prevention, "The prevalence and characteristics of bacteria causing acute diarrhea in Korea"

7 Tarr PI, "Shiga-toxin-producing Escherichia coli and haemolytic uraemic syndrome" 365 : 1073-1086, 2005

8 Nhung PH, "Rapid and specific identification of 5 human pathogenic vibrio species by multiplex polymerase chain reaction targeted to dnaJ gene" 59 : 271-275, 2007

9 Paton AW, "Multiplex PCR for direct detection of Shiga toxigenic Escherichia coli strains producing the novel subtilase cytotoxin" 43 : 2944-2947, 2005

10 Shin HB, "Isolation trend of enteropathogenic bacteria in 1969-1998" 4 : 87-95, 2001

11 Cheng AC, "Infectious diarrhea in developed and developing countries" 39 : 757-773, 2005

12 Iijima Y, "Improvement in the detection rate of diarrhoeagenic bacteria in human stool specimens by a rapid real-time PCR assay" 53 : 617-622, 2004

13 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" 57 : 496-501, 1996

14 Baldi F, "Focus on acute diarrhoeal disease" 15 : 3341-3348, 2009

15 Farthing MJ, "Diarrhoea: a significant worldwide problem" 14 : 65-69, 2000

16 Thapar N, "Diarrhoea in children: an interface between developing and developed countries" 363 : 641-653, 2004

17 Vernacchio L, "Diarrhea in American infants and young children in the community setting: incidence, clinical presentation and microbiology" 25 : 2-7, 2006

18 Gadewar S, "Current concepts in the evaluation, diagnosis and management of acute infectious diarrhea" 5 : 559-565, 2005

19 Kokai-Kun JF, "Comparison of western immunoblots and gene detection assays for identification of potentially enterotoxigenic isolates of Clostridium perfringens" 32 : 2533-2539, 1994

20 O'Leary J, "Comparison of the EntericBio multiplex PCR system with routine culture for detection of bacterial enteric pathogens" 47 : 3449-3453, 2009

21 Chapela MJ, "Comparison between a TaqMan polymerase chain reaction assay and a culture method for ctx-positive Vibrio cholerae detection" 58 : 4051-4055, 2010

22 Labbe R, "Clostridium perfringens in Foodborne Bacterial Pathogens" Marcel Dekker, Inc. 191-234, 1989

23 Choi JP, "Clinical significance of spontaneous Aeromonas bacterial peritonitis in cirrhotic patients: a matched case-control study" 47 : 66-72, 2008

24 Thielman NM, "Clinical practice. Acute infectious diarrhea" 350 : 38-47, 2004

25 Butzler JP, "Campylobacter, from obscurity to celebrity" 10 : 868-876, 2004

26 강재명, "Aeromonas 균혈증의 임상적 특징과 예후와 관련된 위험요인" 대한감염학회 37 (37): 161-166, 2005

27 Lee SW, "A study on epidemiological characteristics and control methods of EHEC infection in Korea" 27 : 37-52, 2005

28 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" 70 : 1656-1662, 2007