Molecular Characterization of β-Tubulin Isotype-1 Gene of Trichostrongylus colubriformis

Benzimidazole resistance (BZR) is known to be associated with single nucleotide polymorphisms (SNPs) on beta-tubulin isotype 1 gene at any of F200Y, F167Y and E198A positions. Characterization of beta-tubulin isotype 1 gene is prerequisite for designing molecular diagnostic method for ascertaining BZR at the early stage of development. Present study was designed to characterize beta tubulin isotype-1 gene of Trichostrongylus colubriformis of small ruminants. Total RNA was extracted from male T. colubriformis. The β-tubulin gene complementary DNA (cDNA) was reverse transcribed from total RNA. The truncated β-tubulin gene of 1202 bp was PCR amplified from complementary DNA (cDNA), cloned in pGEM®-T easy TA cloning vector, custom sequenced and analysed. The analysis of deduced amino acid sequence revealed that the Uttarakhand isolate is susceptible to benzimidazole resistance as the amino acid on 200th position is phenylalanine (F200Y). The amino acid sequences showed 98.3% to 99% identity with the published T. colubriformis sequence as well as other trichstrongyle species. The phylogenetic analysis revealed that Uttarakhand isolate clustered with other trichostrongyles species. The information of beta tubulin isotype-1 gene of T. colubriformis provided an idea for development of molecular tools to diagnosis of benzimidazole resistance at the early stage.