RNA sequencing of gilthead sea bream with a mild Sparicotyle chrysophrii infection

Background: Monogenean flatworms are the main fish ectoparasites inflicting serious economic losses in aquaculture. The polyopisthocotylean Sparicotyle chrysophrii parasitizes the gills of gilthead sea bream (GSB, Sparus aurata) causing systemic anaemia, lamellae fusion and sloughing of epithelial cells, with the consequent hypoxia, emaciation, lethargy and mortality. Currently no preventive or curative measures against this disease exist and therefore information on the host-parasite interaction is crucial to find mitigation solutions for sparicotylosis. The knowledge about gene regulation in monogenean-host models mostly comes from freshwater monopysthocotylean monogeneans and almost nothing is known about polyopisthocotyleans. The current study aims to decipher the host response at local (gills) and systemic (spleen, liver) levels in farmed GSB with a mild natural S. chrysophrii infection by transcriptomic analysis.Results: Using Illumina RNA sequencing and transcriptomic analysis, a total of 2,581 differentially expressed transcripts were identified in infected fish when compared to uninfected controls. Gill tissues in contact with the parasite (P gills) displayed regulation of fewer genes (700) than gill portions not in contact with the parasite (NP gills) (1,235), most likely due to a local silencing effect of the parasite. The systemic reaction in the spleen was much higher than that at the parasite attachment site (local) (1,240), and higher than in liver (334). NP gills displayed a strong enrichment of genes mainly related to immune response and apoptosis. Processes such as apoptosis, inflammation and cell proliferation dominated gills, whereas inhibition of apoptosis, autophagy, platelet activation, signalling and aggregation, and inflammasome were observed in spleen. Proteasome markers were increased in all tissues, whereas hypoxia related genes were down-regulated in gills and spleen.Conclusions: Contrasting forces seem to be acting at local and systemic levels. The splenic down-regulation could be part of a hypometabolic response, to counteract the hypoxia induced by the parasite damage to the gills and to concentrate the energy on defence and repair responses. Alternatively, it can be also interpreted as the often observed action of helminths to modify host immunity in its own interest. These results provide the first toolkit for future studies towards understanding and management of this parasitosis.

背景：单殖吸虫扁形动物（Monogenean flatworms）是鱼类主要的体外寄生虫（ectoparasites），给水产养殖业（aquaculture）造成了严重的经济损失。多后盘目（Polyopisthocotylea）的金鲷单殖吸虫（Sparicotyle chrysophrii）寄生于金头鲷（gilthead sea bream，简称GSB，学名Sparus aurata）的鳃部，引发全身性贫血、鳃丝融合以及上皮细胞脱落，进而导致缺氧、消瘦、嗜睡甚至死亡。目前尚无针对该疾病的预防或治疗手段，因此解析宿主-寄生虫（host-parasite interaction）互作机制，对于探寻金鲷单殖吸虫病（sparicotylosis）的防控方案至关重要。目前关于单殖吸虫-宿主模型中基因调控的认知，大多来源于淡水单后盘目（Monopysthocotylea）单殖吸虫，而对多后盘目单殖吸虫的相关研究几乎空白。本研究旨在通过转录组学（transcriptomic）分析，解析轻度自然感染S. chrysophrii的养殖金头鲷，其局部（鳃部）与系统性（脾脏、肝脏）层面的宿主免疫应答反应。
结果：通过Illumina RNA测序与转录组学分析，相较于未感染的对照组，感染组金头鲷共鉴定出2581个差异表达转录本（differentially expressed transcripts）。与寄生虫直接接触的鳃组织（P鳃）所调控的基因数量（700个），少于未接触寄生虫的鳃组织（NP鳃）的1235个，这一现象大概率是由寄生虫的局部免疫沉默效应所致。脾脏的系统性免疫应答反应强度远高于寄生虫附着的局部鳃组织（1240个差异基因），也高于肝脏（334个差异基因）。未接触寄生虫的鳃组织中，大量富集的基因主要与免疫应答及细胞凋亡（apoptosis）相关。鳃组织中主导的生物学过程为细胞凋亡、炎症反应与细胞增殖；而脾脏中则观察到细胞凋亡、自噬（autophagy）、血小板活化、信号传导与聚集以及炎症小体（inflammasome）的表达受到抑制。所有组织中的蛋白酶体（proteasome）相关标志物表达均上调，而鳃与脾脏中缺氧相关基因的表达则出现下调。
结论：局部与系统性层面似乎存在截然不同的调控机制。脾脏中的基因表达下调可能属于低代谢应答的一部分，用以抵消寄生虫损伤鳃部所引发的缺氧，并将能量集中于防御与修复反应。此外，这一现象也可被解释为蠕虫为自身利益而调控宿主免疫的常见策略。本研究结果为未来解析该寄生虫病的致病机制与防控策略提供了首个研究工具包。