Wellcome-Wolfson Institute for Experimental Medicine, Queen's University Belfast, Belfast BT9 7BL, UK.
Medical Research Council-University of Glasgow Centre for Virus Research, University of Glasgow, Glasgow G61 1QH, UK.
Viruses. 2022 Feb 5;14(2):325. doi: 10.3390/v14020325.
SARS-CoV-2 can efficiently infect both children and adults, albeit with morbidity and mortality positively associated with increasing host age and presence of co-morbidities. SARS-CoV-2 continues to adapt to the human population, resulting in several variants of concern (VOC) with novel properties, such as Alpha and Delta. However, factors driving SARS-CoV-2 fitness and evolution in paediatric cohorts remain poorly explored. Here, we provide evidence that both viral and host factors co-operate to shape SARS-CoV-2 genotypic and phenotypic change in primary airway cell cultures derived from children. Through viral whole-genome sequencing, we explored changes in genetic diversity over time of two pre-VOC clinical isolates of SARS-CoV-2 during passage in paediatric well-differentiated primary nasal epithelial cell (WD-PNEC) cultures and in parallel, in unmodified Vero-derived cell lines. We identified a consistent, rich genetic diversity arising in vitro, variants of which could rapidly rise to near fixation within two passages. Within isolates, SARS-CoV-2 evolution was dependent on host cells, with paediatric WD-PNECs showing a reduced diversity compared to Vero (E6) cells. However, mutations were not shared between strains. Furthermore, comparison of both Vero-grown isolates on WD-PNECs disclosed marked growth attenuation mapping to the loss of the polybasic cleavage site (PBCS) in Spike, while the strain with mutations in Nsp12 (T293I), Spike (P812R) and a truncation of Orf7a remained viable in WD-PNECs. Altogether, our work demonstrates that pre-VOC SARS-CoV-2 efficiently infects paediatric respiratory epithelial cells, and its evolution is restrained compared to Vero (E6) cells, similar to the case of adult cells. We highlight the significant genetic plasticity of SARS-CoV-2 while uncovering an influential role for collaboration between viral and host cell factors in shaping viral evolution and ultimately fitness in human respiratory epithelium.
SARS-CoV-2 能够有效地感染儿童和成人，尽管发病率和死亡率与宿主年龄的增加和合并症的存在呈正相关。SARS-CoV-2 继续适应人类种群，导致出现了具有新型特性的几种关注变种（VOC），例如 Alpha 和 Delta。然而，驱动 SARS-CoV-2 在儿科人群中适应性和进化的因素仍未得到充分探索。在这里，我们提供的证据表明，病毒和宿主因素共同作用，塑造了从儿童中分离出的原代气道细胞培养物中 SARS-CoV-2 的基因型和表型变化。通过对病毒全基因组进行测序，我们在儿科分化良好的鼻上皮细胞（WD-PNEC）培养物中研究了两个预 VOC 临床分离株的遗传多样性随时间的变化，同时在未修饰的 Vero 衍生细胞系中进行了平行研究。我们发现了一种一致的、丰富的遗传多样性在体外产生，其中的变体可以在两次传代内迅速接近固定。在分离株内，SARS-CoV-2 的进化取决于宿主细胞，与 Vero（E6）细胞相比，儿科 WD-PNEC 显示出较低的多样性。然而，突变在株间没有共享。此外，对在 WD-PNEC 上生长的两种 Vero 分离株的比较揭示了明显的生长衰减，其映射到 Spike 中的多碱性切割位点（PBCS）缺失，而在 WD-PNEC 中仍具有生存能力的分离株具有 Nsp12（T293I）、 Spike（P812R）和 Orf7a 截断的突变。总的来说，我们的工作表明，预 VOC SARS-CoV-2 能够有效地感染儿科呼吸道上皮细胞，其进化受到限制，与 Vero（E6）细胞相比，与成人细胞的情况相似。我们强调了 SARS-CoV-2 的显著遗传可塑性，同时揭示了病毒和宿主细胞因素之间的合作在塑造病毒进化并最终在人类呼吸道上皮中适应度方面的重要作用。
