| Research Title |
RAPID DETECTION AND DIFFERENTIATION OF CLONORCHIS SINENSIS AND OPISTHORCHIS VIVERRINI EGGS IN HUMAN FECAL SAMPLES USING A DUPLEX REAL-TIME FLUORESCENCE RESONANCE ENERGY TRANSFER PCR AND MELTING CURVE ANALYSIS |
| Date of Distribution |
29 March 2013 |
| Conference |
| Title of the Conference |
The Second Symposium of Specific Health Problem in Greater Mekong Sub-region (SHeP) |
| Organiser |
Health Cluster, The National Research Project, Khon Kaen University |
| Conference Place |
Auditorium 1, Faculty of Medicine , Khon Kaen University |
| Province/State |
Khon Kaen University, Thailand |
| Conference Date |
29 March 2013 |
| To |
29 March 2013 |
| Proceeding Paper |
| Volume |
2 |
| Issue |
1 |
| Page |
57 |
| Editors/edition/publisher |
Health Cluster, The National Research Project, Khon Kaen University |
| Abstract |
Introduction: Clonorchis sinensis and Opisthorchis viverrini are both important pathogens of food-borne parasitic zoonoses causing serious public health issue in several endemic countries in Asia. The endemic areas of both parasites are closely adjacent to each other with the discovery of co-endemic areas in Thailand.
Methods: To develop a single step duplex real-time fluorescence resonance energy transfer (FRET) PCR merged with melting curve analysis for the fast detection and differentiation of C. sinensis and O. viverrini eggs in human fecal samples. Two species of mitochondrial NADH dehydrogenase subunit 2 (nad2) DNA elements, the 165-bp nad2 product of C. sinensis and the 209-bp nad2 product of O. viverrini were amplified by species-specific primers and the fluorescence melting curve analyses were generated from hybrid of amplicons and two pairs of species-specific fluorophore-labeled probes.
Results: By their different fluorescence channels and melting temperatures, both C. sinensis and O. viverrini eggs in infected human fecal samples were detected and differentiated with high (100%) sensitivity and specificity. Detection limit was as little as a single C. sinensis egg and two O. viverrini eggs in 100 mg of fecal sample. The assay could distinguish the DNA of both parasites from the DNA of negative fecal samples and fecal samples with other parasitosis, as well as from the well-defined genomic DNA of human leukocytes and other parasites.
Conclusions: It can reduce labor time of microscopic examination and is not prone to carry over contamination of agarose electrophoresis. Our duplex real-time FRET PCR method would be useful to determine the accurate range of endemic areas and/or to discover the co-endemic areas of two liver flukes, C. sinensis and O. viverrini, in Asia. This method also would be helpful for the differential diagnosis of the suspected cases of liver fluke infections among travelers who had visited the endemic countries of those parasites. |
| Author |
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| Peer Review Status |
มีผู้ประเมินอิสระ |
| Level of Conference |
ชาติ |
| Type of Proceeding |
Abstract |
| Type of Presentation |
Poster |
| Part of thesis |
true |
| Presentation awarding |
false |
| Attach file |
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| Citation |
0
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Publication
Journal Publication
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