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. 2016 Jan-Mar;11(1):52-64.

Genetic Characterization of Fasciola Isolates from West Azerbaijan Province Iran Based on ITS1 and ITS2 Sequence of Ribosomal DNA

Hossein Galavani  1 , Saber Gholizadeh  2 , Khosrow Hazrati Tappeh  3
Affiliations

Genetic Characterization of Fasciola Isolates from West Azerbaijan Province Iran Based on ITS1 and ITS2 Sequence of Ribosomal DNA

Hossein Galavani et al. Iran J Parasitol. 2016 Jan-Mar.

Abstract

Background: Fascioliasis, caused by Fasciola hepatica and F. gigantica, has medical and economic importance in the world. Molecular approaches comparing traditional methods using for identification and characterization of Fasciola spp. are precise and reliable. The aims of current study were molecular characterization of Fasciola spp. in West Azerbaijan Province, Iran and then comparative analysis of them using GenBank sequences.

Methods: A total number of 580 isolates were collected from different hosts in five cities of West Azerbaijan Province, in 2014 from 90 slaughtered cattle (n=50) and sheep (n=40). After morphological identification and DNA extraction, designing specific primer were used to amplification of ITS1, 5.8s and ITS2 regions, 50 samples were conducted to sequence, randomly.

Result: Using morphometric characters 99.14% and 0.86% of isolates identified as F. hepatica and F. gigantica, respectively. PCR amplification of 1081 bp fragment and sequencing result showed 100% similarity with F. hepatica in ITS1 (428 bp), 5.8s (158 bp), and ITS2 (366 bp) regions. Sequence comparison among current study sequences and GenBank data showed 98% identity with 11 nucleotide mismatches. However, in phylogenetic tree F. hepatica sequences of West Azerbaijan Province, Iran, were in a close relationship with Iranian, Asian, and African isolates.

Conclusions: Only F. hepatica species is distributed among sheep and cattle in West Azerbaijan Province Iran. However, 5 and 6 bp variation in ITS1 and ITS2 regions, respectively, is not enough to separate of Fasciola spp. Therefore, more studies are essential for designing new molecular markers to correct species identification.

Keywords: Fasciola hepatica; ITS1; ITS2; Iran; West Azerbaijan.

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Figures

Fig. 1:
Fig. 1:
Location of the sites where Fasciola isolates were collected in West Azerbaijan Province, Iran
Fig. 2:
Fig. 2:
Multiple sequence alignments of the rDNA ITS1, 5.8s, and ITS2 regions of F. hepatica (JF708027) and F. gigantica (JF432073) as well as representative F. hepatica sequences of the current study [KF531639 (ITS1), KF531689 (5.8s), KF531739 (ITS2)]. Bold sequences belong to 5.8s region flanking to ITS1 and ITS2
Fig. 3:
Fig. 3:
The phylogenetic relationship of the Fasciola spp. isolates collected from cattle and sheep in West Azerbaijan Province of Iran and other Fasciola spp. isolates in different locations based on ITS1 sequence estimated by neighbor-joining algorithms
Fig. 4:
Fig. 4:
The phylogenetic relationship of the Fasciola spp. isolates collected from cattle and sheep in West Azerbaijan Province of Iran and other Fasciola spp. isolates in different locations based on ITS2 sequence estimated by neighbor-joining algorithms
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