Morphological and molecular characterization of Paramphistomum epiclitum of small ruminants.

Morphological and molecular identification can pave the way to design the most effective control measures against the Paramphistomum epiclitum in small ruminants. Morphology of the flukes had described the features of Paramphistomum genus. Body was conical with concave ventral and convex dorsal surface, tegumental spines all around the body in the immature stage, terminal funnel shape oral sucker, sub-terminal acetabulum, blind caeca with a serpentine course touching the anterior level of the acetabulum. Vitelline glands were at the lateral margins of the body extended from the pharynx to the posterior sucker. Testes were lobed and tandem, wavy post-testicular uterus and genital pore behind intestinal bifurcation. Sequence analyses of internal transcribed spacer (ITS)-2+ (PCR products of approximately 500 bp) of 10 flukes yielded 2 genotypes, Navsari isolate 1 and 2. In BLAST analysis, ITS-2+ genotypes were 97.3-99% similar with published sequences (KF564870, JF834888, KF642983 and JX678254) of P. epiclitum of Paramphistomatidae. Two genotypes depicted 4 single nucleotide polymorphisms (NPs) in the form of transitions (C-T at 10 and 18; G-A at 255; A-G at 367 locus), 1 triple NPs (CGT-GAA between 21-23 loci) and missing A base at codon 40 in the genotype 1. Average AT and GC content was 49.61% and 50.38%, respectively. Trees topology inferred by Neighbor Joining and Maximum Likelihood methods of ITS2+ of trematodes were similar, with small difference of bootstrap values. Navsari genotypes formed a tight cluster with the P. epiclitum, originated from different location with high bootstrap value and 0.004-0.011 estimated evolutionary divergence.