Isolation and Characterization of an HKU4-related coronavirus circulating in Malayan pangolin

As the mostly trafficked mammal, pangolins may play important role in coronaviruses zoonotic transmission cycle. Here, we report the circulation of a Tylonycteris bat coronavirus HKU4-related coronavirus from smuggled Malayan pangolins ( Manis javanica ), including 4 viral nucleotide positive and another 7 viral seropositives from 86 animals. Four almost identical (99%) genomes were identified and a MjHKU4r-CoV- 1 virus was isolated, which uses human dipeptidyl peptidase-4 (hDPP4) as receptor and human proteases for cell infection, and this infection is enhanced by a furin cleavage site that is absent in the bat HKU4r-CoVs. In addition, the MjHKU4r-CoV-1 spike showed higher binding affinity to the hDPP4 or wider host range than the bat HKU4-CoV. Finally, MjHKU4r-CoV-1 is productively infectious and pathogenic in human airway, intestinal organs or in hDPP4 transgenic mice. Our work highlighted the importance of pangolins as susceptible hosts of bat-related coronaviruses that poised for human emergence.

作为全球非法贸易规模最大的哺乳类野生动物，穿山甲或在冠状病毒的人畜共患传播循环中扮演关键角色。本研究针对86只走私马来穿山甲（Manis javanica）开展检测，从样本中检出4份病毒核酸阳性与7份病毒血清学阳性，报道了一株与伏翼蝠属（Tylonycteris）蝙蝠冠状病毒HKU4相关的冠状病毒在穿山甲中的传播情况。研究团队获得4条序列相似度高达99%的近乎完全一致的病毒基因组，并分离得到MjHKU4r-CoV-1毒株；该病毒以人二肽基肽酶-4（human dipeptidyl peptidase-4, hDPP4）作为细胞受体，依赖人类蛋白酶完成细胞侵染，其基因组中存在一处蝙蝠HKU4相关冠状病毒所缺失的弗林蛋白酶（furin）切割位点，该位点可增强病毒的细胞侵染效率。此外，相较于蝙蝠HKU4冠状病毒，MjHKU4r-CoV-1的刺突蛋白与人二肽基肽酶-4的结合亲和力更强，宿主范围也更广。最后，实验证实MjHKU4r-CoV-1可在人类气道、肠道器官以及表达人二肽基肽酶-4的转基因小鼠中实现高效增殖侵染并致病。本研究凸显了穿山甲作为蝙蝠相关冠状病毒易感宿主的重要性，这类病毒具备向人类跨物种传播的潜在风险。