Target screening using RNA interference in the sheep abomasal nematode parasite Haemonchus contortus

IF 1.4 4区医学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Molecular and biochemical parasitology Pub Date : 2024-07-14 DOI:10.1016/j.molbiopara.2024.111648

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Abstract

RNA interference (RNAi) on parasitic nematodes has been described as a valuable tool for screening putative targets that could be used as novel drug and/or vaccine candidates. This study aimed to set up a pipeline to identify potential targets using RNAi for vaccine/anti-parasite therapy development against Haemonchus contortus, a blood-feeding abomasal nematode parasite. The available H. contortus sequence data was mined for targets, which were tested for essentiality using RNAi electroporation assays. A total of 56 genes were identified and tested for knockdown using electroporation of first-stage larvae (L1) H. contortus with the target double-stranded RNA. Electroporation of L1 proved to be effective overall; 17 targets had a strong phenotype and significant reduction in alive H. contortus, and another 24 had a moderate phenotype with a significant reduction in larvae development. A total of 28 targets showed a significant reduction in the development of H. contortus larvae to the infective stage (L3) following the RNAi assay. Down-regulation of target transcript levels was evaluated in some targets by semi-quantitative PCR. Four out of five genes tested showed complete knockdown of mRNA levels via semi-quantitative PCR, whereas the knockdown was partial for one. In conclusion, the results indicate that the RNAi pathway is confirmed in H. contortus and that several target genes have the potential to be investigated further as possible vaccine candidates.

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利用 RNA 干扰羊腹腔线虫寄生虫 Haemonchus contortus 的目标筛选。

据介绍，对寄生线虫进行 RNA 干扰（RNAi）是筛选可用作新型药物和/或候选疫苗的潜在靶点的重要工具。本研究旨在建立一个管道，利用 RNAi 确定潜在靶点，以开发针对血食性腹腔线虫（Haemonchus contortus）的疫苗/抗寄生虫疗法。我们从现有的柯氏血吸虫序列数据中挖掘了目标基因，并利用 RNAi 电穿孔实验对这些目标基因的本质进行了检测。共确定了 56 个基因，并利用目标双链 RNA 对第一阶段幼虫（L1）进行电穿孔，以测试基因敲除情况。事实证明，电穿孔 L1 总体上是有效的；17 个目标基因具有强表型，能显著减少活着的 H. contortus，另外 24 个目标基因具有中度表型，能显著减少幼虫的发育。共有 28 个靶标在进行 RNAi 试验后显示，幼虫发育到感染期（L3）时，线虫数量明显减少。通过半定量 PCR 评估了一些靶标转录本水平的下调情况。在测试的五个基因中，有四个基因的 mRNA 水平通过半定量 PCR 被完全敲除，而有一个基因的 mRNA 水平被部分敲除。总之，研究结果表明，RNAi 途径已在霍乱弧菌中得到证实，一些靶基因有可能作为候选疫苗得到进一步研究。

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来源期刊

Molecular and biochemical parasitology 医学-寄生虫学

CiteScore

2.90

自引率

0.00%

发文量

审稿时长

63 days

期刊介绍： The journal provides a medium for rapid publication of investigations of the molecular biology and biochemistry of parasitic protozoa and helminths and their interactions with both the definitive and intermediate host. The main subject areas covered are: • the structure, biosynthesis, degradation, properties and function of DNA, RNA, proteins, lipids, carbohydrates and small molecular-weight substances • intermediary metabolism and bioenergetics • drug target characterization and the mode of action of antiparasitic drugs • molecular and biochemical aspects of membrane structure and function • host-parasite relationships that focus on the parasite, particularly as related to specific parasite molecules. • analysis of genes and genome structure, function and expression • analysis of variation in parasite populations relevant to genetic exchange, pathogenesis, drug and vaccine target characterization, and drug resistance. • parasite protein trafficking, organelle biogenesis, and cellular structure especially with reference to the roles of specific molecules • parasite programmed cell death, development, and cell division at the molecular level.