Development of a real-time PCR assay with fluorophore-labelled hybridization probes for detection of Schistosoma mekongi in infected snails and rat feces.
Schistosoma mekongi, a blood-dwelling fluke, is a water-borne parasite that is found in communities along the lower Mekong River basin, i.e. Cambodia and Lao People's Democratic Republic. This study developed a real-time PCR assay combined with melting-curve analysis to detect S. mekongi in laboratory setting conditions, in experimentally infected snails, and in fecal samples of infected rats. The procedure is based on melting-curve analysis of a hybrid between an amplicon from S. mekongi mitochondrion sequence, the 260 bp sequence specific to S. mekongi, and specific fluorophore-labelled probes. This method could detect as little as a single cercaria artificially introduced into a pool of 10 non-infected snails, a single cercaria in filtered paper, and 2 eggs inoculated in 100 mg of non-infected rat feces. All S. mekongi-infected snails and fecal samples from infected rats were positive. Non-infected snails, non-infected rat feces, and genomic DNA of other parasites were negative. The method gave high sensitivity and specificity, and could be applied as a fast and reliable tool for cercarial location in water environments in endemic areas and for epidemiological studies and eradication programmes for intermediate hosts.
Research and Diagnostic Center for Emerging Infectious Diseases, Khon Kaen University, Khon Kaen 40002, Thailand.
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Sanpool
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PM
Intapan
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Thanchomnang
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P
Sri-Aroon
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Lulitanond
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Sadaow
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W
Maleewong
D000818:Animals; D015342:DNA Probes / Q000737:chemistry; D005243:Feces / Q000469:parasitology; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D051381:Rats; D060888:Real-Time Polymerase Chain Reaction; D012376:Rodent Diseases / Q000175:diagnosis*; D012547:Schistosoma / Q000235:genetics / Q000502:physiology*; D012552:Schistosomiasis / Q000175:diagnosis*; D012680:Sensitivity and Specificity; D012908:Snails / Q000469:parasitology*
D015342:DNA Probes