Department of Biochemical Sciences, Faculty of Pharmacy, Charles University, Heyrovského 1203, 500 05, Hradec Králové, Czech Republic.
Vet Res. 2021 Sep 30;52(1):124. doi: 10.1186/s13567-021-00980-4.
The parasitic gastrointestinal nematode Haemonchus contortus causes serious economic losses to agriculture due to infection and disease in small ruminant livestock. The development of new therapies requires appropriate viability testing, with methods nowadays relying on larval motility or development using procedures that involve microscopy. None of the existing biochemical methods, however, are performed in adults, the target stage of the anthelmintic compounds. Here we present a new test for the viability of H. contortus adults and exsheathed third-stage larvae which is based on a bioluminescent assay of ATP content normalized to total protein concentration measured using bicinchoninic acid. All the procedure steps were optimized to achieve maximal sensitivity and robustness. This novel method can be used as a complementary assay for the phenotypic screening of new compounds with potential antinematode activity in exsheathed third-stage larvae and in adult males. Additionally, it might be used for the detection of drug-resistant isolates.
寄生性胃肠道线虫捻转血矛线虫会导致小型反刍动物家畜感染和患病，从而造成严重的农业经济损失。新疗法的开发需要进行适当的生存能力测试，目前的方法依赖于幼虫的运动或使用涉及显微镜的程序进行发育。然而，现有的生化方法都没有在成虫中进行，而成虫是驱虫化合物的靶标阶段。在这里，我们提出了一种新的捻转血矛线虫成虫和刚蜕皮的第三期幼虫生存能力检测方法，该方法基于生物发光法测定三磷酸腺苷（ATP）含量，并使用二喹啉甲酸（bicinchoninic acid）法测定总蛋白浓度进行归一化。优化了所有步骤以实现最大的灵敏度和稳健性。这种新方法可作为一种补充方法，用于筛选具有潜在抗线虫活性的新化合物，包括刚蜕皮的第三期幼虫和雄性成虫。此外，它还可用于检测耐药分离株。
