绵羊细颈线虫耐伊维菌素虫株三期幼虫P-gps基因表达分析

Analysis of P-gps gene expression on the third stage larvae of ivermectin-resistant Nematodirus in sheep

关键词:  绵羊  细颈线虫  伊维菌素  耐药性  P-gps基因  荧光定量PCR

Key words:  sheep  Nematodirus oiratianus  ivermectin  resistance   P-gps gene  RT-qPCR

摘要:

为分析奥丽春细颈线虫(Nematodirus oiratianus)三期幼虫耐伊维菌素虫株外输泵P-gps基因表达水平，分别在低浓度(3.90 μg/mL)和高浓度(19.98 μg/mL)伊维菌素药物作用下，通过RT-qPCR方法检测药物作用0、3、6、12和24 h 后，外输泵P-gps基因mRNA的表达水平。结果显示:奥丽春细颈线虫三期幼虫在不同时间点分别作用于伊维菌素2 种浓度，其P-gp-1基因均不表达；随着药物作用时间的延长，P-gp-2、P-gp-3、P-gp-11基因表达量均显著增加(P<0.05)，且均于12 h时达峰值。在低浓度药物作用时，P-gp-9.1基因表达量在3 h时显著增加了4.71 倍(P<0.05)，P-gp-12基因表达量却一直很低；但在高浓度药物作用时，随时间的增加，P-gp-12表达量逐渐上调。综上，伊维菌素药物作用耐伊维菌素细颈线虫三期幼虫后，其P-gp-2、P-gp-3、P-gp-9.1、P-gp-11和P-gp-12基因在某些时间点出现基因过表达现象，为研究奥丽春细颈线虫耐药机理以及分子检测耐药性方法的建立提供基础数据。

Abstract:

In order to study the P-gps gene expression level of the third stage larvae of Nematodirus oiratianus resistant to ivermectin, RT-qPCR was used to detected the expression level of P-gps gene mRNA at low and high concentrations of ivermectin after 0, 3, 6, 12 and 24 h treatment, respectively. The results showed that the P-gp-1 gene was not expressed at different time points under two concentrations of ivermectin; The expressions of P-gp-2, P-gp-3 and P-gp-11 genes were increased significantly with the prolongation of drug treatment time, and reached the peak at 12 h. The expression of P-gp-9. 1 gene was increased significantly by 4. 71 times at 3 h under the treatment of low concentration drug and the expression level of P-gp-12 gene was very low all the time. However, the P-gp-12 gene expression was gradually increased at a high concentration with the increase of time. In conclusion, P-gp-2, P-gp-3, P-gp-9. 1, P-gp-11, and P-gp-12 gene were over-expressed at some time points after the treatment of the third stage larvae with ivermectin resistant drugs. This study provided basic data for the study of resistance mechanism and the establishment of molecular detection method of resistance of N. oiratianus.