Helminths infection of Schizothorax niger in Kashmir, India: morphological and molecular characterization

The identification of helminth parasites in Schizothorax spp. from Kashmir, including Schyzocotyle acheilognathi, Pomphorhynchus kashmirensis, and Adenoscolex oreini, is hindered by morphological limitations and high intraspecific variation. While previous studies have relied on morphological diagnosis, a comprehensive molecular characterization is lacking. To address these limitations and achieve more accurate species identification, this study employs molecular techniques such as DNA sequencing and phylogenetic analysis.

Genomic DNA was extracted using standardized protocols. Partial amplification of 18S, ITS, and 28S rRNA was carried out using specific primer sets, followed by Sanger sequencing. Genetic characteristics were analyzed using bioinformatic approaches, focusing on Nucleotide divergence and Phylogenetic relationships.

Phylogenetic analysis confirmed the monophyly of S. acheilognathi, revealing close relationships with S. nayarensis. Interspecific variation ranged from 2.0 - 4.1% for 18S and 0.8 - 2.2% for 28S rDNA, while P. kashmirensis showed high genetic similarity to P. tereticollis and P. laevis with variations ranging from 0.1 - 1.5% for 18S and 2.3-6.0% for ITS rDNA. A. oreini forms a monophyletic lineage separate from Caryophyllaeides and Caryophyllaeus species.

Our study resolved persistent taxonomic ambiguities within the order Bothriocephalidea, resulting in the reassignment of Schyzocotyle acheilognathi (previously Bothriocephalus acheilognathi) to the reinstated genus Schyzocotyle. This study underscores the utility of ribosomal gene sequencing in characterizing fish parasites and highlights the need for taxonomic re-evaluation of A. oreini within the Capingentidae family.