Standardization and application of the tetraprimer ARMS-PCR technique for screening of the E198A SNP in the β-tubulin gene of hookworm populations in Brazil.
The tetraprimer ARMS-PCR technique is efficient for SNP detection and can be used to search for polymorphisms associated with drug resistance. However, the establishment of this methodology is not always straightforward because of the constraints on primer design due to the restrictions of the polymorphic regions. Here, we describe the standardization of the tetraprimer ARMS-PCR methodology for the detection of a SNP at codon 198 of the Ancylostoma caninum β-tubulin gene. This SNP is associated with resistance to albendazole in various nematodes. The methodology was used to screen 327 individuals from 6 different locations. No mutation was found in any of the samples. This methodology will be useful for screening for the E198A SNP in the β-tubulin gene of canine hookworms in a broader population to determine whether this SNP is associated with benzimidazole resistance in this species. The method could also be adapted for the analysis of other SNPs in other nematode species.
Universidade Federal de Minas Gerais, Instituto de Ciências Biológicas, Departamento de Parasitologia, L4 237, Laboratório de Parasitologia Molecular, Pampulha, CEP 31270-901, Belo Horizonte, Minas Gerais, Brazil.
Luis Fernando Viana
LFV
Furtado
Universidade Federal de Minas Gerais, Instituto de Ciências Biológicas, Departamento de Parasitologia, L4 237, Laboratório de Parasitologia Molecular, Pampulha, CEP 31270-901, Belo Horizonte, Minas Gerais, Brazil.
William Pereira
WP
Alves
Universidade Federal de Minas Gerais, Instituto de Ciências Biológicas, Departamento de Parasitologia, L4 237, Laboratório de Parasitologia Molecular, Pampulha, CEP 31270-901, Belo Horizonte, Minas Gerais, Brazil.
Thayse Batista
TB
Moreira
Universidade Federal do Maranhão, Centro de Ciências da Saúde, Departamento de Patologia, Av. dos Portugueses, 1966, 65500000, São Luís, Maranhão, Brazil.
Livio Martins
LM
Costa Junior
Universidade Estadual Vale do Acaraú, Centro de Ciências Agrárias e Biológicas, Av. da Universidade, 850, Betânia, 62040-370, Sobral, Ceará, Brazil.
Rodrigo Rodrigues Cambraia
RRC
Miranda
Universidade Federal de Minas Gerais, Instituto de Ciências Biológicas, Departamento de Parasitologia, L4 237, Laboratório de Parasitologia Molecular, Pampulha, CEP 31270-901, Belo Horizonte, Minas Gerais, Brazil. Electronic address: elidam.rabelo@gmail.com.
Élida Mara Leite
ÉML
Rabelo
D015766:Albendazole / Q000494:pharmacology; D000722:Ancylostoma / Q000235:genetics; D000723:Ancylostomatoidea / Q000187:drug effects / Q000235:genetics; D000818:Animals; D000871:Anthelmintics / Q000494:pharmacology; D001938:Brazil; D017931:DNA Primers / Q000235:genetics / Q000592:standards; D004283:Dog Diseases / Q000469:parasitology*; D004285:Dogs; D004351:Drug Resistance / Q000235:genetics*; D006725:Hookworm Infections / Q000469:parasitology / Q000662:veterinary*; D010274:Parasitology / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000592:standards / Q000662:veterinary*; D020641:Polymorphism, Single Nucleotide / Q000235:genetics; D014404:Tubulin / Q000235:genetics*
D000871:Anthelmintics; D017931:DNA Primers; D014404:Tubulin; D015766:Albendazole
Drug resistance; Hookworms; Tetraprimer ARMS-PCR; β-tubulin