Costa-Junior Livio M, Chaudhry Umer N, Skuce Philip J, Stack Seamus, Sargison Neil D
Pathology Department, Federal University of Maranhão, São Luís, Maranhão Brazil.
Royal (Dick) School of Veterinary Studies, University of Edinburgh, Roslin, Midlothian, EH25 9RG UK.
J Parasit Dis. 2022 Mar;46(1):47-55. doi: 10.1007/s12639-021-01414-w. Epub 2021 Jul 5.
Development of sustainable gastrointestinal nematode (GIN) control strategies depends on the ability to identify the frequencies of drug-susceptible and resistant genotypes in GIN populations arising from management practices undertaken on individual farms. Resistance to BZ drugs in GINs has been shown to be conferred by the presence of defined SNPs in the isotype 1 β-tubulin locus. Loop-mediated isothermal amplification (LAMP) assays are amenable to use on a range of DNA templates and are potentially adaptable to use in practical, cost-effective, pen-side diagnostic platforms that are needed to detect anthelmintic resistance in the field. In this study, we designed primers and examined LAMP assays to detect each of the three major isotype 1 β-tubulin SNPs conferring genetic susceptibility to BZ drugs. We used artificial pools of synthetic DNA, containing different proportions of susceptible and resistant SNPs to determine reproducibility of the assays. We demonstrated the detection of each of the isotype 1 β-tubulin SNPs conferring susceptibility to BZ drugs using the optimal LAMP assay. Isotype 1 β-tubulin SNP typing was effective in detecting BZ susceptibility, but the accuracy was reduced in samples with less than 60 % susceptible DNA. Our results show the potential for LAMP SNP typing to detect genetic susceptibility or resistance to anthelmintic drugs in livestock GINs, and some of the limitations in our approach that will need to be overcome in order to evaluate this assay using field samples.
The online version contains supplementary material available at 10.1007/s12639-021-01414-w.
可持续胃肠道线虫（GIN）控制策略的发展取决于能否识别因各个农场的管理措施而产生的GIN群体中药物敏感和耐药基因型的频率。已证明GIN对苯并咪唑类（BZ）药物的耐药性是由同型1β-微管蛋白基因座中特定单核苷酸多态性（SNP）的存在所导致。环介导等温扩增（LAMP）检测适用于一系列DNA模板，并且有可能适用于实际的、具有成本效益的现场诊断平台，这些平台是检测野外驱虫抗性所必需的。在本研究中，我们设计了引物并检测了LAMP检测方法，以检测赋予对BZ药物遗传易感性的三种主要同型1β-微管蛋白SNP中的每一种。我们使用了含有不同比例敏感和耐药SNP的合成DNA人工池来确定检测方法的可重复性。我们使用最佳LAMP检测方法证明了对赋予对BZ药物易感性的每种同型1β-微管蛋白SNP的检测。同型1β-微管蛋白SNP分型在检测BZ易感性方面是有效的，但在敏感DNA含量低于60%的样本中准确性会降低。我们的结果表明LAMP SNP分型在检测家畜GIN中对驱虫药物的遗传易感性或耐药性方面具有潜力，以及我们的方法中一些需要克服的局限性，以便使用现场样本评估该检测方法。
在线版本包含可在10.1007/s12639-021-01414-w获取的补充材料。
