A fluorescence-based polymerase chain reaction-linked single-strand conformation polymorphism (F-PCR-SSCP) assay for the identification of Fasciola spp.

Citation:
Alasaad, S., R. C. Soriguer, M. Abu-Madi, A. M. ElBehairy, P. D. Baños, A. Píriz, J. Fickel, and Xing-Qu, "A fluorescence-based polymerase chain reaction-linked single-strand conformation polymorphism (F-PCR-SSCP) assay for the identification of Fasciola spp.", Parasitology Research, vol. 108, issue 6, pp. 1513 -1517, 2011.

Abstract:

The present study aimed to establish a fluorescencebased polymerase chain reaction-linked single-strand
conformation polymorphism (F-PCR-SSCP) assay for the identification of Fasciola spp. Based on the sequences
of the second internal transcribed spacer (ITS-2) of the nuclear ribosomal DNA, we designed a set of genusspecific primers for the amplification of Fasciola ITS-2, with an estimated size of 140 bp. These primers were labelled by fluorescence dyes, and the PCR products were analyzed by capillary electrophoresis under nondenaturing conditions (F-PCR-SSCP). Capillary electrophoresis analysis of the fluorescence-labelled DNA fragments displayed three different peak profiles that allowed the accurate identification of Fasciola species: one single peak specific for either Fasciola hepatica or Fasciola gigantica and a doublet peak corresponding to the
“intermediate” Fasciola. Validation of our novel method was performed using Fasciola specimens from different
host animals from China, Spain, Nigeria, and Egypt. This F-PCR-SSCP assay provides a rapid, simple, and robust tool for the identification and differentiation between Fasciola spp.