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Korean J Parasitol. 2013 Dec;51(6):645-50. doi: 10.3347/kjp.2013.51.6.645. Epub 2013 Dec 31.

Rapid detection and identification of Wuchereria bancrofti, Brugia malayi, B. pahangi, and Dirofilaria immitis in mosquito vectors and blood samples by high resolution melting real-time PCR.

Author information

1
Research and Diagnostic Center for Emerging Infectious Diseases, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand. ; Faculty of Medicine, Mahasarakham University, Mahasarakham 44000, Thailand.
2
Research and Diagnostic Center for Emerging Infectious Diseases, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand. ; Department of Parasitology, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand.
3
Research and Diagnostic Center for Emerging Infectious Diseases, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand. ; Division of Cell Biology, Department of Preclinical Sciences, Faculty of Medicine, Thammasat University, Rangsit Campus, Pathum Thani 12121, Thailand.
4
Research and Diagnostic Center for Emerging Infectious Diseases, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand. ; Department of Microbiology, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand.
5
Parasitology Unit, Department of Pathology, Faculty of Veterinary Science, Chulalongkorn University, Bangkok 10330, Thailand.
6
Research and Diagnostic Center for Emerging Infectious Diseases, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand. ; Department of Biochemistry, Faculty of Medical Science, Naresuan University, Phitsanulok 65000, Thailand.
7
Department of Parasitology, Faculty of Medicine, Chiang Mai University, Chiang Mai 50200, Thailand.

Abstract

A simple, rapid, and high-throughput method for detection and identification of Wuchereria bancrofti, Brugia malayi, Brugia pahangi, and Dirofilaria immitis in mosquito vectors and blood samples was developed using a real-time PCR combined with high-resolution melting (HRM) analysis. Amplicons of the 4 filarial species were generated from 5S rRNA and spliced leader sequences by the real-time PCR and their melting temperatures were determined by the HRM method. Melting of amplicons from W. bancrofti, B. malayi, D. immitis, and B. pahangi peaked at 81.5±0.2℃, 79.0±0.3℃, 76.8±0.1℃, and 79.9±0.1℃, respectively. This assay is relatively cheap since it does not require synthesis of hybridization probes. Its sensitivity and specificity were 100%. It is a rapid and technically simple approach, and an important tool for population surveys as well as molecular xenomonitoring of parasites in vectors.

KEYWORDS:

Brugia malayi; Brugia pahangi; Dirofilaria immitis; Wuchereria bancrofti; dog; high resolution melting real-time PCR; mosquito

PMID:
24516268
PMCID:
PMC3916452
DOI:
10.3347/kjp.2013.51.6.645
[Indexed for MEDLINE]
Free PMC Article

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