Phylogenetic and experimental reconstruction of zoonotic transmission of human coronaviruses

自从2002/2003年中国会议上出现SARS冠状病毒以来，冠状病毒引起了人们的极大关注。SARS冠状病毒在人中的感染表明，冠状病毒可以跨越物种屏障，从动物宿主传播到人类，并在人类中引起严重和致命的疾病。2012年MERS冠状病毒的出现表明，人畜共患病传播冠状病毒并不罕见。这两种病毒在蝙蝠中都有近亲，此外，还提出了中间宿主，如浣熊、果子狸冠状病毒(SARS-CoV)和单峰骆驼(MERS-CoV)，以促进人畜共患病的传播。单峰骆驼携带MERS冠状病毒，并可能作为蝙蝠和人类之间的桥梁水库的发现，为病毒出现的生态学提供了一个重要的范例。有趣的是，人类冠状病毒229E(HCoV-229E；在人类中引起普通感冒)似乎有类似的人畜共患病传播史，因为在蝙蝠和骆驼物种(羊驼和单峰骆驼)中发现了近亲。虽然MERS-CoV的人畜传播历史很近，从人和骆驼分离的病毒之间的差异很小，但骆驼-229E-like-CoV和HCoV-229E已经有了很大的分歧。本项目的目的是以HCoV-229E系统为模型，识别和表征CoV人畜共患病传播的关键物种屏障。我们将分析病毒进入步骤(刺激物-受体相互作用)和细胞内复制过程中的物种障碍。重要的是，我们的研究涉及蝙蝠、骆驼和人类229E病毒及其正宗宿主的原代呼吸道上皮细胞培养，使我们能够模拟上呼吸道的跨物种感染。具体而言，我们将侧重于：(A)跨物种传播的宿主决定因素，即先天性免疫反应(I型干扰素系统)，我们将评估非编码RNA(长的非编码RNA和微RNA)是否可能构成关键的物种屏障，以及(B)通过跟随病毒适应新宿主而导致跨物种传播的病毒决定因素，以及确定在适应过程中最不稳定的基因组区域/病毒结构域。相关性：蝙蝠已被证明携带各种各样的冠状病毒物种，包括人类冠状病毒的近亲(HCoV-229E、SARS-CoV、MERS-CoV)。因此，重要的是要确定影响跨物种传播的关键病毒-宿主相互作用。几个全长BAT-229E-Like-CoV基因组序列的获得，骆驼-229E-Like-CoV的发现，以及来自真实宿主的原代上皮靶细胞的可获得性，为研究CoV在人畜共患传播中的关键物种障碍提供了前所未有的机会。这些信息对于评估未来可能发生的人畜共患传播的风险以及为有效的抗病毒干预提供一个概念性的框架是非常有价值的。

Coronaviruses have received considerable attention since the emergence of SARS-CoV in China 2002/2003. SARS-CoV infection in people exemplified that coronaviruses can cross the species barrier from an animal reservoir to humans and cause severe and lethal disease in humans. Theemergence of MERS-CoV in 2012 showed that zoonotic transmission of coronaviruses is not a rare event. Both viruses have close relatives in bats, and moreover, intermediate hosts, such asracoon dogs, civet cats (SARS-CoV) and dromedary camels (MERS-CoV) have been proposed to facilitate zoonotic transmission. The discovery that dromedary camels harbour MERS-CoV and may act as a bridge reservoir between bats and humans provides an important paradigm for the ecology of viral emergence. Interestingly, the human coronavirus 229E (HCoV-229E; causing common cold in humans), appears to have a similar history of zoonotic transmission, since close relatives were detected in bats as well as in camelid species (alpacas and dromedary camels). While MERS-CoV has a very recent history of zoonotic transmission and differences between viruses isolated from humans and camels are rather small, camel-229E-like-CoV and HCoV-229E have already diverged considerably.The aim of the proposed project is to identify and characterize critical species barriers of CoV zoonotic transmission by using the HCoV-229E system as a model. We will analyse species barriers at the virus entry step (spike-receptor interaction) and during intracellular replication. Importantly, our studies involve bat- camel- and human 229E viruses and primary airway epithelial cultures of their authentic hosts, allowing us to mimic cross-species infection of the upper airways. Specifically, we will focus on:(a) host determinants of cross species transmission, namely innate immune responses (type I interferon system), and we will assess if non-coding RNAs (long non-coding RNA and micro-RNAs) may constitute a critical species barrier, and(b) virus determinants of cross species transmission by following virus adaptation to a new host, and identification of genomic regions/viral domains that are most volatile during adaptation. Relevance: Bats have been shown to harbour a wide variety of diverse CoV species, including close relatives of human coronaviruses (HCoV-229E, SARS-CoV, MERS-CoV). Therefore it is important to define critical virus-host interactions that impact on cross-species transmission. The availability of several full-length bat-229E-like-CoV genome sequences, the discovery of a camel-229E-like-CoV, and the availability of primary epithelial target cells from the authentic hosts offers an unprecedented opportunity to study critical species barriers for the zoonotic transmission of CoVs. This information is extremely valuable to assess the risk of possible future zoonotic transmissions and provide a conceptional framework for efficacious antiviral intervention.