Molecular Identification and Phylogenetic Analysis of Fasciola Hepatica Isolates From Cattle and Sheep in Golestan Province, Northeast of Iran.

Abstract Background: Fascioliasis in livestock and humans, caused by Fasciola hepatica and Fasciola gigantica, has socioeconomic importance worldwide. Iran is one of the important endemic foci for fascioliasis. Molecular approaches can be precise and reliable for the identification and characterization of Fasciola spp. The aims of this study were molecular identification of Fasciola hepatica isolates in Golestan province, north of Iran, and then comparative analysis of results using GenBank sequences.Material and Method: Fasciola flukes were isolated from the livers of infected livestock. DNA was extracted from samples using the phenol-chloroform technique. RFLP-PCR using TasI restriction enzyme was performed along with morphometric evaluations for the detection of F. hepatica species. PCR was performed to amplify partial fragments of the internal transcribed spacer 1 (ITS1), cytochrome c oxidase sub.1 (CO1), and NADH dehydrogenase sub.1 (ND1) genes for samples that were confirmed as F. hepatica. Twenty-eight PCR products were sequenced. The phylogenetic tree with MEGA 7 software was drawn for all three genes. Results: Out of 271 flukes collected from sheep and cattle, 126 confirmed as F. hepatica by the PCR-RFLP method. Results based on PCR-RFLP analysis were confirmed by sequence analysis. Only one haplotype for the CO1 gene and four different haplotypes for the ND1 gene were identified. Seven sequences of each gene registered on GenBank and accession numbers were received.Conclusions: This study showed that F. hepatica is widely distributed among livestock in Golestan province. It was also found that genetic diversity among the ND1 region in isolates of our study was considerably higher than the CO1 sequence region.