Molecular characterisation of influenza B virus from the 2017/18 season in primary models of the human lung reveals improved adaptation to the lower respiratory tract

The 2017/18 influenza season was characterized by unusual high numbers of severe infections and hospitalizations. Instead of influenza A viruses, this season was dominated by infections with influenza B viruses of the Yamagata lineage. While this IBV/Yam dominance was associated with a vaccine mismatch, a contribution of virus intrinsic features to the clinical severity of the infections was speculated. Here, we performed a molecular and phenotypic characterization of three IBV isolates from patients with severe flu symptoms in 2018 and compared it to an IBV/Yam isolate from 2016 using experimental models of increasing complexity, including human lung explants, lung organoids, and alveolar macrophages. Viral genome sequencing revealed the presence of clade but also isolate specific mutations in all viral genes, except NP, M1, and NEP. Comparative replication kinetics in different cell lines provided further evidence for improved replication fitness, tolerance towards higher temperatures, and the development of immune evasion mechanisms by the 2018 IBV isolates. Most importantly, immunohistochemistry of infected human lung explants revealed an impressively altered cell tropism, extending from AT2 to AT1 cells and macrophages. Finally, transcriptomics of infected human lung explants demonstrated significantly reduced amounts of type I and type III IFNs by the 2018 IBV isolate, supporting the existence of additional immune evasion mechanisms. Our results show that the severeness of the 2017/18 Flu season was not only the result of a vaccine mismatch but was also facilitated by improved adaptation of the circulating IBV strains to the environment of the human lower respiratory tract.

2017/2018年流感季以异常高发的重症感染与住院病例为显著特征。本季的主流感染病原体并非甲型流感病毒，而是山形系（Yamagata lineage）乙型流感病毒（influenza B viruses, IBV）。尽管这种IBV/山形系毒株的优势流行与疫苗株匹配不佳有关，但学界曾推测病毒本身的固有特性也会加剧感染的临床严重程度。本研究针对2018年3名出现重症流感症状患者分离得到的3株IBV分离株开展了分子与表型特征分析，并以2016年的1株IBV/山形系分离株作为对照，所用实验模型的复杂度逐步提升，涵盖人肺外植体、肺类器官与肺泡巨噬细胞。病毒基因组测序结果显示，除NP、M1与NEP基因外，所有病毒基因均存在进化分支特异性突变以及分离株特异性突变。不同细胞系中的复制动力学对比实验进一步证实，2018年的IBV分离株具备更优异的复制适配能力、对更高温度的耐受性，且演化出了免疫逃逸相关机制。最为关键的是，感染后人肺外植体的免疫组化分析结果显示，病毒的细胞嗜性发生了显著改变，感染范围从AT2细胞扩展至AT1细胞与巨噬细胞。最后，感染后人肺外植体的转录组学分析表明，2018年IBV分离株可显著下调I型与III型干扰素（interferons, IFNs）的表达水平，这进一步支持了该毒株存在额外免疫逃逸机制的结论。本研究结果显示，2017/2018年流感季的重症化态势不仅源于疫苗株与流行毒株匹配不佳，还与当时流行的IBV毒株对人类下呼吸道微环境的适应性增强密切相关。