Wang Yanwen, Zhong Qiu, Meng Fei, Cheng Zhang, Zhang Yijie, Song Zuchen, Zhang Yali, Feng Zijian, Zhai Yujia, Chen Yan, Qiao Chuanling, Yang Huanliang
State Key Laboratory for Animal Disease Control and Prevention, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150069, China.
Viruses. 2025 Jul 20;17(7):1018. doi: 10.3390/v17071018.
The H1N1 swine influenza viruses CQ91 and CQ445, isolated from pigs in China, exhibited distinct virulence in mice despite sharing similar genomic constellations. CQ91 demonstrated higher pathogenicity (MLD: 5.4 log EID) and replication efficiency in mice compared to CQ445 (MLD: 6.6 log EID). Through reverse genetics, we found that the attenuation of CQ445 was due to a single substitution of glutamic acid (E) with lysine (K) at position 343 in the PB2 protein. Introducing the CQ445-PB2 (343K) into CQ91 significantly reduced viral replication and pathogenicity in mice, while replacing CQ445-PB2 with CQ91-PB2 (343E) restored virulence. In vitro studies showed that the K343E mutation impaired viral replication in MDCK and A549 cells and reduced polymerase activity in minigenome assays. Mechanistically, the amino acid at position 343 in the PB2 affects the transcription stage of the viral replication process. Structural modeling indicated that the charge reversal caused by E343K altered local electrostatic interactions without major conformational changes. Phylogenetic analysis revealed that PB2-343E is highly conserved (>99.9%) in human and swine H1/H3 influenza viruses, suggesting that PB2-343E confers an adaptive advantage. This study identifies PB2-343E as a critical determinant of influenza virus pathogenicity in mammals, highlighting its role in host adaptation.
从中国猪群中分离出的H1N1猪流感病毒CQ91和CQ445，尽管基因组构成相似，但在小鼠中表现出不同的毒力。与CQ445（半数致死剂量：6.6 log EID）相比，CQ91在小鼠中表现出更高的致病性（半数致死剂量：5.4 log EID）和复制效率。通过反向遗传学，我们发现CQ445的减毒是由于PB2蛋白第343位的谷氨酸（E）被赖氨酸（K）单一位点替换所致。将CQ445-PB2（343K）导入CQ91可显著降低病毒在小鼠中的复制和致病性，而用CQ91-PB2（343E）替换CQ445-PB2则恢复了毒力。体外研究表明，K343E突变损害了病毒在MDCK和A549细胞中的复制，并降低了微型基因组试验中的聚合酶活性。从机制上讲，PB2蛋白第343位的氨基酸影响病毒复制过程的转录阶段。结构建模表明，E343K引起的电荷反转改变了局部静电相互作用，而没有重大构象变化。系统发育分析显示，PB2-343E在人和猪的H1/H3流感病毒中高度保守（>99.9%），表明PB2-343E赋予了适应性优势。本研究确定PB2-343E是流感病毒在哺乳动物中致病性的关键决定因素，突出了其在宿主适应性中的作用。
