Clonorchis sinensis and Opisthorchis viverrini: Development of a mitochondrial-based multiplex PCR for their identification and discrimination
| Autor: | Nguyen Van De, Thanh Hoa Le, Donald P. McManus, David Blair, Paiboon Sithithaworn |
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| Rok vydání: | 2006 |
| Předmět: |
Immunology
DNA Mitochondrial Opisthorchiasis Polymerase Chain Reaction Sensitivity and Specificity law.invention Fish Diseases Species Specificity law Cricetinae Opisthorchis Multiplex polymerase chain reaction medicine Animals Humans Opisthorchis viverrini Polymerase chain reaction Life Cycle Stages Clonorchis sinensis Mesocricetus biology Fishes General Medicine DNA Helminth medicine.disease biology.organism_classification Molecular biology genomic DNA Infectious Diseases Clonorchiasis Parasitology |
| Zdroj: | Experimental Parasitology. 112:109-114 |
| ISSN: | 0014-4894 |
| Popis: | We report a single, one-step PCR approach for detection and discrimination of Clonorchis sinensis and Opisthorchis viverrini in different life-stage forms (adults, metacercariae, and eggs) from fish intermediate hosts and from infected patients. Primers designed for species-specific PCR, amplifying portions of the mitochondrial (mt) genome, were also suitable for a multiplex PCR. The latter was a single, one-step reaction under high stringency conditions, using simultaneously 2 pairs of primers (1 pair for C. sinensis--product size 612 bp, and 1 pair for O. viverrini--product size 1357 bp). Assays using serially diluted templates demonstrated that as little as 0.78 ng of genomic DNA of either species could yield amplicons. Genomic DNA extracted from different life-stage forms including adult worms (of both species), eggs (of O. viverrini), eggs possibly of several trematode species (collected from patients infected with C. sinensis in Vietnam) and mixed metacercariae of common trematodes (collected from fishes in the C. sinensis endemic areas), yielded specific bands of the correct size and their identity was confirmed by sequence analysis. The multiplex PCR approach described here proved to be a species-specific, sensitive and fast tool for accurate diagnosis of clonorchiasis and/or opisthorchiasis, permitting the detection of their metacercariae in infected fishes or adult/eggs from patients in endemic areas. |
| Databáze: | OpenAIRE |
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