Contemporary seasonal human coronaviruses display differences in cellular tropism compared to laboratory-adapted reference strains.

IF 3.8 2区 医学 Q2 VIROLOGY
Journal of Virology Pub Date : 2025-09-23 Epub Date: 2025-08-27 DOI:10.1128/jvi.00684-25
Matthew J Gartner, Monique L Smith, Clyde Dapat, Yi Wen Liaw, Thomas Tran, Randy Suryadinata, Joseph Chen, Guizhi Sun, Rory A Shepherd, George Taiaroa, Michael Roche, Wen Shi Lee, Philip Robinson, Jose M Polo, Kanta Subbarao, Jessica A Neil
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引用次数: 0

Abstract

Seasonal human coronaviruses (sHCoVs) cause 15%-30% of common colds. The reference strains used for research were isolated decades ago and have been passaged extensively, but contemporary sHCoVs have been challenging to study as they are notoriously difficult to grow in standard immortalized cell lines. Here, we addressed these issues by utilizing primary human nasal epithelial cells (HNECs) and immortalized human bronchial epithelial cells (BCi) differentiated at an air-liquid interface, as well as human embryonic stem cell-derived alveolar type II (AT2) cells to recover contemporary sHCoVs from human nasopharyngeal specimens. From 21 specimens, we recovered four HCoV-229e, three HCoV-NL63, and eight HCoV-OC43 viruses. All contemporary sHCoVs showed sequence differences from lab-adapted CoVs, particularly within the spike gene. Evidence of nucleotide changes in the receptor binding domains within HCoV-229e and detection of recombination for both HCoV-229e and HCoV-OC43 isolates was also observed. Importantly, we developed methods for the amplification of high-titer stocks of HCoV-NL63 and HCoV-229e that maintained sequence identity, and we established methods for the titration of contemporary sHCoV isolates. Comparison of lab-adapted and contemporary strains in immortalized cell lines and airway epithelial cells revealed differences in cell tropism, growth kinetics, and cytokine production between lab-adapted and contemporary sHCoV strains. These data confirm that contemporary sHCoVs differ from lab-adapted reference strains and, using the methods established here, should be used for the study of CoV biology and evaluation of medical countermeasures.IMPORTANCEZoonotic coronaviruses have caused significant public health emergencies. The occurrence of a similar spillover event in the future is likely, and efforts to further understand coronavirus biology should be a high priority. Several seasonal coronaviruses circulate within the human population. Efforts to study these viruses have been limited to reference strains isolated decades ago due to the difficulty in isolating clinical isolates. Here, we use human airway and alveolar epithelial cultures to recover contemporary isolates of human coronaviruses HCoV-NL63, HCoV-229e, and HCoV-OC43. We establish methods to make high-titer stocks and titrate HCoV-229e and HCoV-NL63 isolates. We show that contemporary isolates of HCoV-NL63 and HCoV-OC43 have a different tropism within the respiratory epithelium compared to lab-adapted strains. Although HCoV-229e clinical and lab-adapted strains similarly infect the respiratory epithelium, differences in host response and replication kinetics are observed. Using the methods developed here, future research should include contemporary isolates when studying coronavirus biology.

与实验室适应的参考菌株相比,当代季节性人类冠状病毒在细胞趋向性方面表现出差异。
季节性人类冠状病毒(sHCoVs)导致15%-30%的普通感冒。用于研究的参考菌株是在几十年前分离出来的,并且已经广泛传代,但当代shcov的研究一直具有挑战性,因为它们在标准的永活细胞系中生长非常困难。在这里,我们通过利用原代人鼻上皮细胞(HNECs)和在气液界面分化的永生人支气管上皮细胞(BCi)以及人胚胎干细胞衍生的肺泡II型(AT2)细胞从人鼻咽标本中恢复当代shcov来解决这些问题。从21份标本中分离出4种HCoV-229e、3种HCoV-NL63和8种HCoV-OC43病毒。所有当代shcov都显示出与实验室适应的cov的序列差异,特别是在刺突基因内。在HCoV-229e和HCoV-OC43分离株中,还观察到HCoV-229e受体结合域核苷酸变化和重组检测的证据。重要的是,我们开发了保持序列一致性的HCoV-NL63和HCoV-229e高滴度库存扩增方法,并建立了当代sHCoV分离株的滴定方法。在永生化细胞系和气道上皮细胞中比较实验室适应株和当代株,发现实验室适应株和当代株在细胞趋向性、生长动力学和细胞因子产生方面存在差异。这些数据证实,当代shcov不同于实验室适应的参考菌株,应使用本文建立的方法进行冠状病毒生物学研究和医学对策评估。人畜共患冠状病毒已造成重大公共卫生突发事件。未来可能会发生类似的溢出事件,进一步了解冠状病毒生物学应该是重中之重。几种季节性冠状病毒在人群中传播。由于临床分离的困难,研究这些病毒的努力仅限于几十年前分离的参考菌株。在这里,我们使用人类气道和肺泡上皮培养物来恢复人类冠状病毒HCoV-NL63、HCoV-229e和HCoV-OC43的当代分离株。建立了HCoV-229e和HCoV-NL63分离株的高滴度测定方法。我们发现,与实验室适应菌株相比,当代分离的HCoV-NL63和HCoV-OC43在呼吸道上皮内具有不同的倾向。虽然HCoV-229e临床和实验室适应菌株相似地感染呼吸道上皮,但观察到宿主反应和复制动力学的差异。使用这里开发的方法,在研究冠状病毒生物学时,未来的研究应该包括当代分离株。
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来源期刊
Journal of Virology
Journal of Virology 医学-病毒学
CiteScore
10.10
自引率
7.40%
发文量
906
审稿时长
1 months
期刊介绍: Journal of Virology (JVI) explores the nature of the viruses of animals, archaea, bacteria, fungi, plants, and protozoa. We welcome papers on virion structure and assembly, viral genome replication and regulation of gene expression, genetic diversity and evolution, virus-cell interactions, cellular responses to infection, transformation and oncogenesis, gene delivery, viral pathogenesis and immunity, and vaccines and antiviral agents.
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