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KMID : 0829320090120010011
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Korean Journal of Clinical Microbiology
2009 Volume.12 No. 1 p.11 ~ p.16
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Implementation of Multiplex PCR for Species Identification and Toxin Typing in Toxigenic Clostridium difficile Culture
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Jang Yun-Ha
Kim Mi-Na
Chung Jae-Woo
Sung Heung-Sup
Park Sook-Ja
Baek Seung-Mi
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Abstract
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Background: We evaluated multiplex PCR for species identification and toxin typing to improve the sensitivity and turnaround time of toxigenic Clostridium difficile culture (TCDC).
Methods: We performed multiplex PCR using primers targeting the species-specific gene, tpi, and the toxin genes, tcdA and tcdB. From January to March 2008, 528 stool specimens were tested with direct toxin assay (DT) using C. difficile Tox A/B II (Techlab, Blacksburg, USA) and TCDC. For 288 specimens from early study period, toxin production by C. difficile isolates of TCDC was measured by enzyme immunoassay with culture supernatants using VIDAS C. difficile Toxin A&B (CDAB; bioMerieux, Marcy-l’Etoile, France) and multiplex PCR with isolated colonies.For 240 specimens from late period, only multiplex PCR was used to test toxin production by the isolates.
Results: During the early period, 29 C. difficile were isolated and their toxin-positive rates were 65.5% by PCR and 44.8% by CDAB (P＜0.05). Among 528 stool specimens, the results of DT+/TCDC+, DT+/TCDC-, and DT-/TCDC+ were 32 (6.1%), 33 (6.3%), and 10 (1.9%), respectively, when tested with PCR. 13.3% of total 75 positive specimens was detected only by TCDC. Of the 42 toxigenic C. difficile isolates, all were positive for tpi, 30 (71.4%) were tcdA+/tcdB+, and 12 (28.6%) were tcdA-/tcdB+.
Conclusion: TCDC using multiplex PCR for species identification and toxin typing is sensitive and rapid to be used as a routine diagnostic test
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KEYWORD
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Clostridium difficile, Toxin, Polymerase, chain reaction
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