Table_1_Evidence of SARS-CoV-2 Related Coronaviruses Circulating in Sunda pangolins (Manis javanica) Confiscated From the Illegal Wildlife Trade in Viet Nam.DOCX

Despite the discovery of several closely related viruses in bats, the direct evolutionary progenitor of SARS-CoV-2 has not yet been identified. In this study, we investigated potential animal sources of SARS-related coronaviruses using archived specimens from Sunda pangolins (Manis javanica) and Chinese pangolins (Manis pentadactyla) confiscated from the illegal wildlife trade, and from common palm civets (Paradoxurus hermaphroditus) raised on wildlife farms in Viet Nam. A total of 696 pangolin and civet specimens were screened for the presence of viral RNA from five zoonotic viral families and from Sarbecoviruses using primers specifically designed for pangolin coronaviruses. We also performed a curated data collection of media reports of wildlife confiscation events involving pangolins in Viet Nam between January 2016 and December 2020, to illustrate the global pangolin supply chain in the context of Viet Nam where the trade confiscated pangolins were sampled for this study. All specimens from pangolins and civets sampled along the wildlife supply chains between February 2017 and July 2018, in Viet Nam and tested with conventional PCR assays designed to detect flavivirus, paramyxovirus, filovirus, coronavirus, and orthomyxovirus RNA were negative. Civet samples were also negative for Sarbecoviruses, but 12 specimens from seven live pangolins confiscated in Hung Yen province, northern Viet Nam, in 2018 were positive for Sarbecoviruses. Our phylogenetic trees based on two fragments of the RdRp gene revealed that the Sarbecoviruses identified in these pangolins were closely related to pangolin coronaviruses detected in pangolins confiscated from the illegal wildlife trade in Yunnan and Guangxi provinces, China. Our curated data collection of media reports of wildlife confiscation events involving pangolins in Viet Nam between January 2016 and December 2020, reflected what is known about pangolin trafficking globally. Pangolins confiscated in Viet Nam were largely in transit, moving toward downstream consumers in China. Confiscations included pangolin scales sourced originally from Africa (and African species of pangolins), or pangolin carcasses and live pangolins native to Southeast Asia (predominately the Sunda pangolin) sourced from neighboring range countries and moving through Viet Nam toward provinces bordering China.

尽管在蝙蝠中发现了多种密切相关病毒，但SARS-CoV-2的直接进化始祖尚未确定。本研究通过分析从非法野生动物贸易中查获的苏门答腊穿山甲（Manis javanica）和中华穿山甲（Manis pentadactyla）的存档样本，以及越南野生动物养殖场饲养的普通椰子猫（Paradoxurus hermaphroditus）的样本，调查了与SARS相关的冠状病毒的潜在动物来源。共对696个穿山甲和椰子猫样本进行了筛查，以检测五种动物源性病毒家族以及Sarbecoviruses的病毒RNA，筛查过程中使用了针对穿山甲冠状病毒特异性设计的引物。此外，我们还对2016年1月至2020年12月期间越南发生的涉及穿山甲的野生动物查获事件的媒体报道进行了精心整理的数据收集，以描绘越南在全球穿山甲供应链中的地位，其中被查获的穿山甲样本用于本研究。在2017年2月至2018年7月期间，越南野生动物供应链中的穿山甲和椰子猫样本，以及使用旨在检测黄病毒、副粘病毒、丝状病毒、冠状病毒和正粘病毒RNA的常规PCR检测方法进行检测，结果均为阴性。椰子猫样本对Sarbecoviruses也为阴性，但2018年在越南北部河静省查获的七只活穿山甲中的12个样本对Sarbecoviruses呈阳性。基于RdRp基因两个片段构建的进化树显示，这些穿山甲中发现的Sarbecoviruses与在中国云南和广西省非法野生动物贸易中查获的穿山甲中检测到的穿山甲冠状病毒密切相关。我们对2016年1月至2020年12月期间越南涉及穿山甲的野生动物查获事件的媒体报道的精心整理数据收集反映了全球穿山甲走私的情况。在越南查获的穿山甲主要处于运输途中，正运往中国的下游消费者。查获的穿山甲鳞片最初源自非洲（以及非洲穿山甲物种），或源自东南亚（主要是苏门答腊穿山甲）的穿山甲尸体和活穿山甲，这些穿山甲来自邻近的分布国，通过越南运往与中国接壤的省份。