Alvarez-Sánchez M A, Pérez-García J, Cruz-Rojo M A, Rojo-Vázquez F A
Dpt. Patología Animal (Sanidad Animal), Facultad de Veterinaria, Universidad de León, C/Profesor Pedro Cármenes S/N, 24071 León, Spain.
Vet Parasitol. 2005 May 15;129(3-4):291-8. doi: 10.1016/j.vetpar.2005.02.004.
This report describes a new molecular method for the diagnosis of benzimidazole susceptibility or resistance in three main species of trichostrongylids of sheep (Haemonchus contortus, Teladorsagia circumcincta and Trichostrongylus vitrinus). This assay is based on the use of real time polymerase chain reaction (PCR) to detect mutations of residue 200 on isotype 1 of beta-tubulin. The technique allows calculation of the proportion of each allelic variant as it combines kinetic (real time quantitative) PCR with allele-specific amplification and requires no post-PCR processing. The level of resistance in the population is determined by the proportion of the beta-tubulin codon 200 TAC allele. Hence, it was observed that the proportion of the resistant allele in susceptible strains ranged between 24% and 32.3%; in resistant strains this value increased to between 71.3% and 86.3%. Furthermore, there was a good correlation between real time PCR, faecal egg count reduction test, egg hatch assay and conventional allele-specific PCR, in both resistant and susceptible strains. A sensitive, rapid, highly reproducible and inexpensive technique for detecting resistance to benzimidazoles in a worm population has been developed.
本报告描述了一种用于诊断绵羊三种主要毛圆科线虫（捻转血矛线虫、环形泰勒虫和透明毛圆线虫）对苯并咪唑敏感性或抗性的新分子方法。该检测方法基于使用实时聚合酶链反应（PCR）来检测β-微管蛋白1型异构体第200位残基的突变。该技术将动力学（实时定量）PCR与等位基因特异性扩增相结合，可计算每个等位基因变体的比例，且无需PCR后处理。群体中的抗性水平由β-微管蛋白密码子200的TAC等位基因比例决定。因此，观察到敏感品系中抗性等位基因的比例在24%至32.3%之间；在抗性品系中，该值增加到71.3%至86.3%之间。此外，在抗性和敏感品系中，实时PCR、粪便虫卵计数减少试验、虫卵孵化试验与传统等位基因特异性PCR之间均具有良好的相关性。已开发出一种用于检测蠕虫群体对苯并咪唑抗性的灵敏、快速、高度可重复且廉价的技术。
