Department of Laboratory Medicine & Molecular Diagnostics, Division of Microbiology, Sunnybrook Health Sciences Centre, Toronto, Ontario, Canada.
Department of Pathology and Microbiology, Atlantic Veterinary College, University of Prince Edward Island, Charlottetown, Prince Edward Island, Canada.
PLoS Negl Trop Dis. 2020 Apr 6;14(4):e0008105. doi: 10.1371/journal.pntd.0008105. eCollection 2020 Apr.
Crimean-Congo hemorrhagic fever virus (CCHFV) is a tick-borne virus that can cause a hemorrhagic fever in humans, with a case fatality rate of up to 40%. Cases of CCHFV have been reported in Africa, Asia, and southern Europe; and recently, due to the expanding range of its vector, autochthonous cases have been reported in Spain. Although it was discovered over 70 years ago, our understanding of the pathogenesis of this virus remains limited. We used RNA-Seq in two human liver cell lines (HepG2 and Huh7) infected with CCHFV (strain IbAr10200), to examine kinetic changes in host expression and viral replication simultaneously at 1 and 3 days post infection. Through this, numerous host pathways were identified that were modulated by the virus including: antiviral response and endothelial cell leakage. Notably, the genes encoding DDX60, a cytosolic component of the RIG-I signalling pathway and OAS2 were both shown to be dysregulated. Interestingly, PTPRR was induced in Huh7 cells but not HepG2 cells. This has been associated with the TLR9 signalling cascade, and polymorphisms in TLR9 have been associated with poor outcomes in patients. Additionally, we performed whole-genome sequencing on CCHFV to assess viral diversity over time, and its relationship to the host response. As a result, we have demonstrated that through next-generation mRNA deep-sequencing it is possible to not only examine mRNA gene expression, but also to examine viral quasispecies and typing of the infecting strain. This demonstrates a proof-of-principle that CCHFV specimens can be analyzed to identify both the virus and host biomarkers that may have implications for prognosis.
克里米亚-刚果出血热病毒（CCHFV）是一种蜱媒病毒，可引起人类出血热，病死率高达 40%。该病毒已在非洲、亚洲和南欧报告病例；最近，由于其传播媒介范围的扩大，在西班牙也报告了本地病例。尽管该病毒被发现已有 70 多年，但我们对其发病机制的理解仍然有限。我们使用 RNA-Seq 在两种感染 CCHFV（IbAr10200 株）的人源肝细胞系（HepG2 和 Huh7）中进行检测，同时在感染后 1 天和 3 天检查宿主表达和病毒复制的动力学变化。通过这种方法，鉴定了许多被病毒调节的宿主途径，包括抗病毒反应和内皮细胞渗漏。值得注意的是，病毒编码 DDX60（RIG-I 信号通路的细胞质成分）和 OAS2 的基因都被证明失调。有趣的是，PTPRR 在 Huh7 细胞中被诱导，但在 HepG2 细胞中没有。这与 TLR9 信号级联有关，TLR9 基因的多态性与患者预后不良有关。此外，我们对 CCHFV 进行了全基因组测序，以评估随时间推移的病毒多样性及其与宿主反应的关系。结果表明，通过下一代 mRNA 深度测序，不仅可以检测 mRNA 基因表达，还可以检测病毒准种和感染株的分型。这证明了一个原理，即可以对 CCHFV 标本进行分析，以鉴定可能对预后有影响的病毒和宿主生物标志物。
