microRNAs are small non-coding RNAs that are important regulators of gene expression in a range of animals, including nematodes. We have analysed a cluster of four miRNAs from the pathogenic nematode species Haemonchus contortus that are closely linked in the genome. We find that the cluster is conserved only in clade V parasitic nematodes and in some ascarids, but not in other clade III species nor in clade V free-living nematodes. Members of the cluster are present in parasite excretory-secretory products and can be detected in the abomasum and draining lymph nodes of infected sheep, indicating their release in vitro and in vivo. As observed for other parasitic nematodes, H. contortus adult worms release extracellular vesicles (EV). Small RNA libraries were prepared from vesicle-enriched and vesicle-depleted supernatants from both adult worms and L4 stage larvae. Comparison of the miRNA species in the different fractions indicated that specific miRNAs are packaged within vesicles, while others are more abundant in vesicle-depleted supernatant. Hierarchical clustering analysis indicated that the gut is the likely source of vesicle-associated miRNAs in the L4 stage, but not in the adult worm. These findings add to the growing body of work demonstrating that miRNAs released from parasitic helminths may play an important role in host-parasite interactions.

microRNA是小的非编码RNA，它们是许多动物（包括线虫）中基因表达的重要调节剂。我们已经分析了来自基因组中紧密联系的致病线虫物种Haemonchus contortus的四个miRNA的簇。我们发现该簇仅在进化枝V寄生线虫和某些scar虫中保守，而在其他进化枝III物种和进化枝V自由活动线虫中均不保守。在集群的成员存在于寄生虫排泄-分泌产物，可以在受感染的羊的皱胃和引流淋巴结被检测到，表明其释放的体外和体内。如针对其他寄生线虫所观察到的，H。扭曲成年蠕虫释放细胞外囊泡（EV）。小RNA文库是从成虫和L4期幼虫的富含囊泡和耗尽囊泡的上清液中制备的。比较不同部分中的miRNA种类表明，特定的miRNA被包装在囊泡中，而其他的miRNA在囊泡耗尽的上清液中更为丰富。分层聚类分析表明，肠道是L4期囊泡相关miRNA的可能来源，而在成虫中则不是。这些发现增加了工作量，表明寄生虫蠕虫释放的miRNA可能在宿主-寄生虫相互作用中起重要作用。