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. 2012 May;19(2):177-82.
doi: 10.1051/parasite/2012192177.

Molecular identification of Fasciola spp. (Digenea: Fasciolidae) in Egypt

Y Dar  1 , S Amer, A Mercier, B Courtioux, G Dreyfuss
Affiliations

Molecular identification of Fasciola spp. (Digenea: Fasciolidae) in Egypt

Y Dar et al. Parasite. 2012 May.

Abstract
in English, French

A total of 134 Egyptian liver flukes were collected from different definitive hosts (cattle, sheep, and buffaloes) to identify them via the use of PCR-RFLP and sequence analysis of the first nuclear ribosomal internal transcribed spacer (ITS1). Specimens of F. hepatica from France, as well as F. gigantica from Cameroon were included in the study for comparison. PCR products of ITS1 were subjected for digestion by RsaI restriction enzyme and visualized on agarose gel. According to RFLP pattern, Egyptian flukes were allocated into two categories. The first was identical to that of French hepatica flukes to have a pattern of 360, 100, and 60 (bp) band size, whereas the second resembled to that of Cameroonian gigantica worms to have a profile of 360, 170, and 60 bp in size. Results of RFLP analysis were confirmed by sequence analysis of representative ITS1 amplicons. No hybrid forms were detected in the present study. Taken together, this study concluded that both species of Fasciola are present in Egypt, whereas the hybrid form may be not very common.

Des douves égyptiennes (134 spécimens) provenant de différents hôtes définitifs (bovins, buffles et moutons) ont été analysées par PCR-RFLP et l’étude de la séquence ITS-1 pour identifier les espèces locales de Fasciola (F. hepatica, F. gigantica, ou les formes hybrides entre ces deux espèces). Des douves provenant de France (F. hepatica) et du Cameroun (F. gigantica) ont été utilisées comme références. Deux types de bandes ont été trouvés dans les fragments de la séquence ITS-1 : le premier type est identique à celui des F. hepatica de France (trois bandes de 60, 100 et 360 bp) tandis que le second ressemble à celui des F. gigantica du Cameroun (trois bandes de 60, 170 et 360 bp). De plus, le séquençage des amplicons ITS-1 confirme cette différence en montrant la présence de six sites nucléotidiques variables, ce qui permet de discriminer F. hepatica de F. gigantica. Aucune forme intermédiaire entre les deux Fasciola n’a été trouvée dans les spécimens analysés. Pris ensemble, cette étude permet de conclure que les deux espèces de Fasciola sont présentes en Égypte, alors que la forme hybride pourrait être pas très commune.

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Figures

Fig. 1.
Fig. 1.
Agarose gel electrophoresis of amplified ITS1 ribosomal region. Lanes 1-10 denote to different fluke samples amplified as a single band of 680 bp; lane 11, negative control; lane M, 100 bp ladder molecular weight marker.
Fig. 2.
Fig. 2.
RsaI restriction enzyme digestion products of ITS1 fragment. Lanes 1-5 denote to those of Fasciola gigantica isolated from cattle at Yaoundé, Cameron (1) and Cairo, Egypt (2, 3); as well as from sheep at Cairo, Egypt (4, 5), respectively. Lanes 6-10 denote to those of Fasciola hepatica isolated from sheep and cattle (6, 7) at Cairo, Egypt, and sheep and cattle (8, 9) at Tanta, Egypt, respectively, as well as from cattle at Limoges, France (10).
Fig. 3.
Fig. 3.
Nucleotide sequence alignment of ITS1 fragments of Fasciola species from Egypt (Egy) and four other countries: Cameroon (Cam), France (Fra), Ireland (Irl), and Zambia (Zam). The restriction sites of RsaI endonuclease are underlined and shaded in gray. The nucleotide differences between the two Fasciola species are denoted in bold. Dots indicate that the nucleotides are identical to those in the upper line. ‘Y’ represents a nucleotide indicating the presence of a double peak, C and T. (Irl), GenBank accession number: AB514850 for Fasciola hepatica from Ireland. (Zam), GenBank accession number: AB514855 for Fasciola gigantica from Zambia. Underline denote to primer position.
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References

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