Molecular Detection of Benzimidazole Resistance in Haemonchus contortus Larvae of Goats in Chhattisgarh, India

摘要

Background: Benzimidazole resistance is one of the key problem in small ruminant production. A rapid, truthful and responsive system is required for detection benzimidazole resistance so that proper regulatory measure can be applied. Allele specific PCR is one of the tools to understand the mechanism and origin of benzimidazole resistance. Methods: A total 198 larvae of Haemonchus contortus were isolated from goats of Chhattisgarh region, central India were genotyped by allele specific polymerase chain reaction (AS-PCR). Faecal samples of goats were collected from three Government farms and adjoining field goats and were subjected for faecal culture, separately. DNA of third stage larva was used for nested PCR for amplification of beta-tubulin gene. Restriction fragment length polymorphism (RFLP) was applied on nested PCR product for species identification with RsaI enzyme. AS-PCR was applied on the nested-PCR product to know the genotypic and allelic frequency. Result: The nested PCR amplified product showed approximately 820 bp in all cases and PCR-RFLP revealed 462 bp, 211 bp and 147 bp fragments, which confirmed the species as H. Contortus. Frequency of resistant allele ('r') was 49.7 and 50.3 for susceptible allele ('S'). Frequency of homozygous resistant (rr), heterozygous susceptible (rS) and homozygous susceptible (SS) genotype were 33.83 per cent, 31.81 per cent and 34.34 per cent, respectively. The frequency of homozygous resistant (rr) genotype was low (19.61) in field compare to farm (48.96) indicating refugia in field region.