Molecular identification and characterization of Trichinella spiralis from a leopard in India

Present study describes species level identification of Trichinella spiralis of leopard origin from India using multiplex PCR and molecular characterization of the parasite based on sequencing of multiple genetic markers viz. 5S ribosomal DNA intergenic spacer region (5S ISR), partial mitochondrial large ribosomal subunit (Mt-lsr) and partial mitochondrial cytochrome c oxidase I (COI) genes. A single amplicons of 173 bp, indicative of T. spiralis was obtained in multiplex PCR. Further, specific PCR amplifications viz. 750 bp (5S ISR), 445 bp (Mt-lsr) and 850 bp (COI) were obtained for selected genetic markers. Homology search analysis of 5S-ISR, Mt-lsr gene and COI gene showed highest 99.6% identity with sequences originating from China (KT894074, T. spiralis), 98.6% similarity with T. spiralis China isolates (GU339127, GU339147) and 99.8% sequence homology with T. spiralis sequences originating from Belarus MH119334), respectively. In the phylogenetic analysis, sequences of each selected genetic marker clustered together with published T. spiralis isolates only, which further confirmed species level identification of detected larvae as T. spiralis, although very few differences were noted with reference to relative positions. This is the first study from India, which provide molecular evidence on circulation of T. spiralis in wild animals.