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Identification and differentiation of Fasciola hepatica and Fasciola gigantica using a simple PCR-restriction enzyme method

Rokni, M.B. and Mirhendi, H. and Mizani, A. and Mohebali, M. and Sharbatkhori, M. and Kia, E.B. and Abdoli, H. and Izadi, S. (2010) Identification and differentiation of Fasciola hepatica and Fasciola gigantica using a simple PCR-restriction enzyme method. Experimental Parasitology, 124 (2). pp. 209-213. ISSN 00144894 (ISSN)

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Abstract

Accurate morphological differentiation between the liver fluke species Fasciola hepatica and Fasciola gigantica is difficult. We evaluated PCR-restriction enzyme profiles of internal transcribed spacer 1 (ITS1) that could aid in their identification. Fifty F. hepatica and 30 F. gigantica specimens were collected from different hosts in three provinces of Iran. For DNA extraction, we crushed fragments of the worms between two glass slides as a new method to break down the cells. DNA from the crushed materials was then extracted with a conventional phenol-chloroform method and with the newly developed technique, commercial FTA cards. A primer pair was selected to amplify a 463-bp region of the ITS1 sequence. After sequencing 14 samples and in silico analysis, cutting sites of all known enzymes were predicted and TasI was selected as the enzyme that yielded the most informative profile. Crushing produced enough DNA for PCR amplification with both the phenol-chloroform and commercial FTA card method. The DNA extracted from all samples was successfully amplified and yielded a single sharp band of the expected size. Digestion of PCR products with TasI allowed us to distinguish the two species. In all samples, molecular identification was consistent with morphological identification. Our PCR-restriction enzyme profile is a simple, rapid and reliable method for differentiating F. hepatica and F. gigantica, and can be used for diagnostic and epidemiological purposes. © 2009 Elsevier Inc. All rights reserved.

Item Type: Article
Additional Information: Unmapped bibliographic data: LA - English [Field not mapped to EPrints] J2 - Exp. Parasitol. [Field not mapped to EPrints] C2 - 19769969 [Field not mapped to EPrints] AD - Department of Medical Parasitology and Mycology, School of Public Health, Institute of Public Health Research, Tehran, Iran [Field not mapped to EPrints] AD - Department of Medical Parasitology, School of Medicine, Golestan University of Medical Sciences, Gorgan, Iran [Field not mapped to EPrints] AD - Molecular Biology Laboratory, Institute of Public Health Research, Isfahan Station, Isfahan, Iran [Field not mapped to EPrints] DB - Scopus [Field not mapped to EPrints]
Uncontrolled Keywords: Fasciola gigantica, Fasciola hepatica, Identification, PCR-restriction enzyme, TasI, chloroform, internal transcribed spacer 1, phenol, restriction endonuclease, ribosome DNA, accuracy, article, computer analysis, controlled study, DNA extraction, Fasciola gigantica, Fasciola hepatica, Iran, microbial morphology, nonhuman, nucleotide sequence, parasite identification, polymerase chain reaction, priority journal, restriction fragment length polymorphism, restriction mapping, sequence analysis, species differentiation, species distribution, species identification, Abattoirs, Animals, Base Sequence, Buffaloes, Cattle, Diagnosis, Differential, DNA Primers, DNA Restriction Enzymes, DNA, Helminth, Electrophoresis, Agar Gel, Fasciola, Fasciola hepatica, Fascioliasis, Goats, Iran, Liver, Molecular Sequence Data, Polymerase Chain Reaction, Polymorphism, Restriction Fragment Length, Restriction Mapping, Sequence Alignment, Sheep, Fasciola gigantica, Fasciola hepatica
Subjects: مقالات نمایه شده محققین دانشگاه در سایت ,Web of Science ,Scopus
موارد کلی
Divisions: معاونت تحقیقات و فناوری
Depositing User: GOUMS
Date Deposited: 18 Apr 2015 04:12
Last Modified: 05 Apr 2017 10:06
URI: http://eprints.goums.ac.ir/id/eprint/2225

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