Microbe and Pathogen Biology, Institute for Global Food Security, School of Biological Sciences, Queen's University Belfast, Belfast, UK.
Department of Animal, Plant and Soil Sciences and Centre for AgriBioscience, La Trobe University, Bundoora, Victoria, Australia.
Vet Parasitol. 2020 Oct;286:109244. doi: 10.1016/j.vetpar.2020.109244. Epub 2020 Sep 12.
The surface tegument of Fasciola hepatica is a crucial tissue due to its key role at the host-parasite interface. We characterised three novel proteins, termed Fhteg1, Fhteg5 and Fhteg8, that are found in the tegument membrane fraction of adult F. hepatica. Bioinformatic analysis of proteomic datasets identified Fhteg5 and Fhteg8 as tegumental glycoproteins and revealed that Fhteg1, Fhteg5 and Fhteg8 are associated with exosomes of adult F. hepatica. Fhteg1, Fhteg5 and Fhteg8 appear to be related to uncharacterised sequences in F. gigantica, Fasciolopsis buski, Echinostoma caproni, Clonorchis sinensis, Opisthorchis viverrini, Schistosoma japonicum and S. mansoni, although F. hepatica appears to have expanded this family. Fhteg1 and Fhteg5 were characterised in detail. The Fhteg1 and Fhteg5 gene transcripts each demonstrate significant upregulation in juvenile fluke 2-4 days post-excystment, with transcript levels maintained during development over 3 weeks in vitro. RNAseq data showed that both Fhtegs are expressed in the adult life stage, although the transcript levels were about 8 fold lower than those in juveniles (3 week post infection). Using immunocytochemistry, Fhteg1 and Fhteg5 were each shown to be expressed in cells adjacent to the muscle layer as well as on the surface of 1 week old juveniles, whilst Fhteg5 was also present in cells at the base of the pharynx. RNAi mediated knockdown of Fhteg1 and Fhteg5 transcripts in 4-10 day old juveniles had no effect on parasite survival, movement or growth in vitro. Although no IgG responses were observed for Fhteg1 or Fhteg5 during infection in sheep and cattle, both proteins elicited a low IgG response in a proportion of infected rats. Rats vaccinated with Fhteg1 and Fhteg5 showed good IgG responses to both proteins and a mean 48.2 % reduction in worm burden following parasite challenge. Although vaccination of cattle with both proteins induced a range of IgG responses, no protection was observed against parasite challenge. This is the first study to provide insights into the molecular properties of two novel, developmentally regulated surface tegument proteins in F. hepatica.
肝片形吸虫的表皮层是一种至关重要的组织，因为它在宿主-寄生虫界面中起着关键作用。我们鉴定了三种新型蛋白质，分别称为 Fhteg1、Fhteg5 和 Fhteg8，它们存在于成虫肝片形吸虫的表皮膜部分。蛋白质组数据集的生物信息学分析表明，Fhteg5 和 Fhteg8 是表皮糖蛋白，并揭示了 Fhteg1、Fhteg5 和 Fhteg8 与成虫肝片形吸虫的外泌体有关。Fhteg1、Fhteg5 和 Fhteg8 似乎与 F. gigantica、Fasciolopsis buski、Echinostoma caproni、Clonorchis sinensis、Opisthorchis viverrini、Schistosoma japonicum 和 S. mansoni 中的未描述序列有关，尽管肝片形吸虫似乎扩展了这个家族。我们详细研究了 Fhteg1 和 Fhteg5。Fhteg1 和 Fhteg5 的基因转录本在囊蚴孵化后 2-4 天的幼体中均显著上调，并且在体外 3 周的发育过程中维持转录水平。RNAseq 数据显示，这两种 Fhtegs 在成虫生活阶段都有表达，尽管转录水平比幼体低约 8 倍（感染后 3 周）。通过免疫细胞化学，Fhteg1 和 Fhteg5 均在靠近肌肉层的细胞以及 1 周龄幼体的表面表达，而 Fhteg5 也存在于咽底的细胞中。在 4-10 日龄的幼体中，用 RNAi 介导的 Fhteg1 和 Fhteg5 转录本敲低对寄生虫的存活、运动或体外生长没有影响。尽管在绵羊和牛感染期间没有观察到针对 Fhteg1 或 Fhteg5 的 IgG 反应，但这两种蛋白质在一部分感染大鼠中引起了低水平的 IgG 反应。用 Fhteg1 和 Fhteg5 接种的大鼠对这两种蛋白质均表现出良好的 IgG 反应，在寄生虫挑战后，蠕虫负荷平均降低了 48.2%。尽管用这两种蛋白质接种牛可诱导多种 IgG 反应，但在寄生虫挑战中没有观察到保护作用。这是首次研究提供了肝片形吸虫两种新型、发育调节的表皮层蛋白的分子特性的见解。
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