The aim of the present study was to investigate the occurrence of Haemonchus contortus and Haemonchus placei in beef cattle and the frequency of single nucleotide polymorphisms associated with benzimidazole (BZ)-resistance in Haemonchus spp. in Brazil. For such, fecal samples were collected from 61 beef cattle ranches in 11 Brazilian states. Third-stage larvae (L3) were produced for morphological species identification and DNA extraction. PCR was performed for the analysis of the isotype 1 β-tubulin gene and the products were sequenced to confirm the presence of H. placei and H. contortus. For each field population, pyrosequencing assays were performed to quantify the frequency of the F167Y, E198A and F200Y polymorphisms in the isotype-1 β-tubulin gene. The results of the morphometric analysis of 2345 larvae showed that H. placei was present on all ranches. The analysis of the isotype 1 β-tubulin gene confirmed 100% prevalence for H. placei and 23.7% for H. contortus. Pyrosequencing assays demonstrated single-nucleotide polymorphisms (SNPs) associated with BZ-resistance in all three codons (F167Y, E198A and F200Y) of the isotype 1 β-tubulin gene in H. placei field populations. Frequencies of resistance-associated alleles above background (≥ 15%) were found for at least one codon in 11.4% of the field isolates and maximum frequencies of 30, 21 and 29% were found for codons 167, 198 and 200, respectively, on individual ranches. This study confirms the presence of H. contortus in beef cattle in the major livestock farming states in Brazil and demonstrates that genotypes associated with BZ resistance are present in field populations of Haemonchus spp..
Keywords: Haemonchus contortus; Haemonchus placei; Larvae; Pyrosequencing; Ruminants; β-Tubulin.
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