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Title: Evaluation of multiplex PCR with enhanced spore germination for detection of clostridium difficile from stool samples of the hospitalized patients.
Authors: Surang Chankhamhaengdecha
Piyapong Hadpanus
Amornrat Aroonnual
Puriya Ngamwongsatit
Darunee Chotiprasitsakul
Piriyaporn Chongtrakool
Tavan Janvilisri
Mahidol University. Faculty of Science. Department of Biology
Mahidol University. Faculty of Tropical Medicine. Department of Tropical Nutrition and Food Science.
Mahidol University. Faculty of Veterinary Science. Department of Clinical Sciences and Public Health.
Mahidol University. Faculty of Medicine Ramathibodi Hospital. Department of Medicine.
Mahidol University. Faculty of Medicine Ramathibodi Hospital. Department of Pathology.
Mahidol University. Faculty of Science. Department of Biochemistry.
Keywords: multiplex PCR;spore;germination;Clostridium;Clostridium;Patients;hospitalized patients;Open Access article
Issue Date: 2013
Citation: BioMed research international. Vol.2013 (2013), 1-6
Abstract: Clostridium difficile poses as the most common etiologic agent of nosocomial diarrhea. Although there are many diagnostic methods to detect C. difficile directly from stool samples, the nucleic acid-based approach has been largely performed in several laboratories due to its high sensitivity and specificity as well as rapid turnaround time. In this study, a multiplex PCR was newly designed with recent accumulated nucleotide sequences. The PCR testing with various C. difficile ribotypes, other Clostridium spp., and non-Clostridium strains revealed 100% specificity with the ability to detect as low as ~22 genomic copy number per PCR reaction. Different combinations of sample processing were evaluated prior to multiplex PCR for the detection of C. difficile in fecal samples from hospitalized patients. The most optimal condition was the non-selective enrichment at 37 °C for 1 h in brain heart infusion broth supplemented with taurocholate, followed by the multiplex PCR. The detection limit after sample processing was shown as being 5 spores per gram of fecal sample. Two hundred and thirty-eight fecal samples collected from the University affiliated hospital were analyzed by the enrichment multiplex PCR procedure. The results suggested that the combination of sample processing with the high-performance detection method would be applicable for routine diagnostic use in clinical setting.
ISSN: 2314-6133
Appears in Collections:SC-Article

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