title: A fluorescence-based polymerase chain reaction-linked single-strand conformation polymorphism (F-PCR-SSCP) assay for the identification of Fasciola spp.
abstract: The present study aimed to establish a fluorescence-based 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 genus-specific 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 non-denaturing 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.
authors.affiliation: Estación Biológica de Doñana, Consejo Superior de Investigaciones Científicas (CSIC), Avda. Américo Vespucio s/n, 41092, Seville, Spain. samer@ebd.csic.es
authors.forename: Samer
authors.initials: S
authors.lastname: Alasaad
authors.forename: Ramón C
authors.initials: RC
authors.lastname: Soriguer
authors.forename: Marawan
authors.initials: M
authors.lastname: Abu-Madi
authors.forename: Ahmed
authors.initials: A
authors.lastname: El Behairy
authors.forename: Pablo Díez
authors.initials: PD
authors.lastname: Baños
authors.forename: Ana
authors.initials: A
authors.lastname: Píriz
authors.forename: Joerns
authors.initials: J
authors.lastname: Fickel
authors.forename: Xing-Quan
authors.initials: XQ
authors.lastname: Zhu
mesh_terms: D000818:Animals; D017931:DNA Primers / Q000235:genetics; D018509:DNA, Helminth / Q000032:analysis / Q000235:genetics; D021903:DNA, Ribosomal Spacer; D019075:Electrophoresis, Capillary / Q000379:methods; D005209:Fasciola / Q000235:genetics*; D005456:Fluorescent Dyes; D016133:Polymerase Chain Reaction / Q000379:methods*; D018807:Polymorphism, Single-Stranded Conformational
chemical_list: D017931:DNA Primers; D018509:DNA, Helminth; D021903:DNA, Ribosomal Spacer; D005456:Fluorescent Dyes