Molecular Identification And Phylogenetic Analysis of Fasciola Species Isolated from Sheep And Cattle Using The Its2 Rdna Marker in Duhok Province, Kurdistan Region, Iraq

Abstract

The aims of the current study were molecular identification and phylogenetic analysis of Fasciola spp. isolated from infected cattle and sheep using the ITS2 region of rDNA as a molecular marker in Duhok province, Kurdistan region, Iraq. DNA was extracted from 40 adult flukes isolated from the livers of infected sheep (23) and cattle (17) slaughtered at the abattoirs of Duhok Province. The amplification of the rDNA ITS2 region of these 40 specimens was done by PCR using primers (FITS2-F and FITS2-R) designed for this study by the authors. ITS2 fragments (approximately 314 bp) of the rDNA were successfully amplified, and produced monoclear bands, that were confirmed by gel electrophoresis. From these isolates, 5 amplicons of ITS2 (2 sheep and 3 cattle) were selected for sequencing, and then the obtained sequences were compared to those in NCBI-GenBank for genotyping and phylogenetic analysis. The blast alignment of NCBI-GenBank revealed four sequences belonging to Fasciola hepatica and one belonging to Fasciola gigantica, both with 100% similarity. The genetic distance between F. hepatica and F. gigantica was 0.0127. Phylogenetic analysis revealed that the sequences in this study were clustered into two main clades (F. hepatica and F. gigantica), with other sequences available in GenBank. The study concluded that ITS2 rDNA sequences were appropriate to identify both Fasciola species isolated from the infected sheep and cattle in Duhok province, which were identical to those of neighboring countries (Iran and Turkey). The primers of ITS2 rDNA region that designed for the current study can be useful for the diagnosis of animals and humans fasciolosis