Molecular detection of benzimidazole resistance in Haemonchus contortus of sheep in Andhra Pradesh, India

A study was carried out to detect the mutation in β-tubulin isotype 1 gene of Haemonchus contortus worms collected from sheep slaughtered in different abattoirs of Rayalaseema region (Chittoor, YSR Kadapa, Anantapur and Kurnool) and adjoining SPS Nellore district of Andhra Pradesh, India. The isolated genomic DNA from all the 150 male worms were separately used to amplify β-tubulin isotype 1 gene that yielded PCR product of 305 bp. Digestion of PCR product with restriction endonuclease revealed three different RFLP patterns. Individual worms with 257 and 48 bp bands were genotyped as ‘rr’ (homozygous resistant), those of 305, 257 and 48 bp bands as ‘rS’ (heterozygous) and those without restriction site as ‘SS’ (homozygous susceptible). The genotypic frequencies varied significantly (P<0.01) in all regions under study except in Kurnool region. The frequency of ‘SS’ genotypes were high in all the regions except in SPS Nellore and Anantapur. The frequency of ‘rr’ genotypes were more in SPS Nellore (46.67%) whereas the frequency of ‘rS’ was more in Anantapur (100%). The overall prevalence of benzimidazole susceptible allele was found significantly (P<0.01) higher (60%) as compared to resistant allele. Results revealed the presence of variation in β-tubulin gene at codon 200 in H. contortus in sheep.