JCM Accepts, published online ahead of print on 9 July 2008

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J. Clin. Microbiol. doi:10.1128/JCM.00027-08
Copyright (c) 2008, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Comparison of Conventional, Nested, and Real-time PCR for Rapid and Accurate Diagnosis of Patients with Vibrio vulnificus

Hyong Sun Kim, Dong-Min Kim^*, Ganesh Prasad Neupane, Yu-mi Lee, Nam-Woong Yang, Sook Jin Jang, Sook-In Jung, Kyung-Hwa Park, Hae-Ryoung Park, Chang Seop Lee, and Sun Hee Lee

Division of Infectious Disease, Department of Internal Medicine, and Department of Microbiology, and Department of Laboratory Medicine, Chosun University College of Medicine, Gwang-ju; Department of Internal Medicine, Chonnam National University Medical School, Gwang-ju; Dental Science Research Institute and BK21 project for dental School, Chonnam National university, Gwangju; Department of Internal Medicine, Chonbuk National University Medical School, Jeonju; Department of Internal Medicine, College of Medicine, Pusan National University, Pusan, Republic of Korea

^* To whom correspondence should be addressed. Email: drongkim{at}chosun.ac.kr.

We conducted a prospective study to target toxR in the blood of patients with skin and soft tissue infections who were admitted to four tertiary hospitals, to assess the clinical usefulness of Real-time quantitative PCR as a diagnostic technique. We performed Conventional PCR (C-PCR), Nested PCR (N-PCR) and Real-time quantitative PCR (Q-PCR) and compared the results to the "gold standard" microbiologic culture. The lower detection limit of Q-PCR was 5x10⁰ copies/µl. Using the blood of patients with skin and soft tissue infections, the sensitivities of C-PCR and N-PCR against the target toxR gene of V. vulnificus as diagnostic tools were 45% and 86%, respectively. The C-PCR and N-PCR assays had specificities of 100% and 73%, respectively. When we adopted a crossing point (cp) cut off value < 38 cp as a positive result, the Q-PCR assay had 100% sensitivity and specificity. Q-PCR to detect V. vulnificus- specific genes is not only the most sensitive and specific techniques, but is also the most rapid diagnostic method. Therefore the appropriate application of the Q-PCR assay using blood is useful for rapid diagnosis and subsequent treatment of V. vulnificus sepsis.