Fatal infection of a novel canine/human reassortant H3N2 influenza A virus in the zoo-housed golden monkeys

In June 2022, seven golden monkeys (Rhinopithecus roxellana) developed flu-like symptoms in succession at a zoo in Jiangsu Province, China, and two died of respiratory distress. Histopathological and immunostaining results supported a diagnosis of pulmonary infection with influenza virus. An H3N2 canine/human reassortment virus, designated A/golden monkey/Jiangsu/1/2022 (Gm-1), was isolated from the lungs of deceased animals. Sequencing analysis revealed that except for PB1 gene that was 99% similar to that from human H1N1 virus (A/WSN/1933, WSN), the other 7 segments of Gm-1 were entirely derived from previously circulating canine H3N2 virus (A/canine/Jiangsu/06/2010, JS06). To determine whether the human-origin PB1 segment conferred a virulence advantage to Gm-1, we reconstructed this reassortant event using reverse genetics and generated two reassortment viruses, rGm-1 and rJS06. Experimental infection of mice revealed that the rGm-1 virus induced more severe pathological damage along with higher levels of proinflammatory cytokines in the lungs, suggesting that replacement of the PB1 segment of JS06 by PB1 of WSN enhanced viral virulence. Furthermore, we identified a unique cytotoxic motif (68I, 69L and 70V) in PB1-F2 protein that was present in Gm-1 but absent in JS06, potentially enhancing viral virulence. This is the first report confirming IAV infection in golden monkeys. Our finding highlight the the importance of enhanced biosecurity surveillance for this endangered species.

2022年6月，中国江苏省某动物园内7只川金丝猴（Rhinopithecus roxellana）相继出现流感样症状，其中2只因呼吸窘迫死亡。组织病理学与免疫染色结果支持其为流感病毒肺部感染的诊断结论。研究人员从死亡个体的肺部组织中分离得到一株H3N2型犬源/人源重配病毒，命名为A/golden monkey/Jiangsu/1/2022（简称Gm-1）。测序分析显示，除PB1基因与人类H1N1病毒（A/WSN/1933，简称WSN）的同源性达99%外，Gm-1其余7个基因片段均来源于此前流行的犬源H3N2病毒（A/canine/Jiangsu/06/2010，简称JS06）。为明确人源PB1基因片段是否赋予Gm-1毒力优势，本研究通过反向遗传学（reverse genetics）技术重构了该重配事件，获得两株重配病毒rGm-1与rJS06。小鼠实验感染结果表明，rGm-1可诱导更严重的病理损伤，并在肺部引发更高水平的促炎细胞因子，提示将JS06的PB1基因替换为WSN来源的PB1可增强病毒毒力。此外，本研究在Gm-1的PB1-F2蛋白中发现了一处独特的细胞毒性基序（68位异亮氨酸、69位亮氨酸与70位缬氨酸），该基序在JS06中不存在，或可增强病毒毒力。本研究首次证实金丝猴可感染甲型流感病毒（Influenza A Virus, IAV）。本研究结果凸显了加强针对这一濒危物种的生物安全监测的重要性。