一株猪流感病毒A/swine/Shanghai/01/2019(H1N1)的分离鉴定及遗传进化分析

Identification and evolution analysis of a swine influenza virus A/swine/Shanghai/01/2019(H1N1)

为鉴定上海地区猪流感病毒的流行株及其分子生物学特征,本研究从上海地区养殖场采集疑似流感症状的猪咽拭子样品40份,采用套式PCR结合鸡胚分离鉴定方法,从中分离到1株H1N1亚型猪流感病毒(SIV),命名为A/swine/Shanghai/01/2019(H1N1)。经全基因组测定和遗传进化分析结果显示该分离株的8个基因节段与欧亚类禽H1N1 SIV同源性最高,其PB2、PB1、PA基因节段可能来源于A/swine/Jiangsu/49/2012(H1N1)病毒,NA、NP和M基因节段可能来源于A/swine/Shanghai/3/2014(H1N1)病毒。该分离病毒的HA蛋白裂解位点为PSIQSR↓GLFGAI,为低致病性流感病毒的分子特征。该分离株NA蛋白未出现与神经氨酸酶抑制剂相关的氨基酸突变,M2蛋白S31N突变表明该病毒可能对金刚烷胺类药物具有一定的耐药性。PA蛋白224S和PB2蛋白701N突变提示该分离株对哺乳动物的适应性增强,可能导致其致病性增强,需要加强对此类SIV的监测。结合近年来的监测表明,上海地区猪群中H1N1亚型SIV持续存在,需要持续跟踪监测SIV变异趋势。本研究为防控猪流感提供实验依据。

A strain of H1N1 subtype swine influenza virus(SIV) was isolated and identified from 40 throat swab samples of pigs with cough and running nose by nested PCR and SPF chicken embryos inoculation, named as A/swine/Shanghai/01/2019(H1N1). Whole genome sequencing and genetic analysis showed that eight segments of this strain had the highest homology with Avian-like H1N1 SIV. PB2, PB1 and PA genes were most likely derived from A/swine/Jiangsu/49/2012(H1N1) strain, while NA, NP and M genes were most likely derived from A/swine/Shanghai/3/2014(H1N1) strain. The cleavage site sequence of its HA protein was PSIQSR↓GLFGAI, indicating the typical molecular characteristics of low pathogenic influenza virus. Amino acid substitutions associated with resistance to neuraminidase inhibitor were not found in NA protein, while 31N mutation of M2 protein showed a resistance to amantadine. The 224S of PA protein and 701 N of PB2 protein associated with mammalian adaption were identified and may increase its pathogenicity in mammals. Combining with the surveillance data on SIV in Shanghai in recent years, H1N1 SIV persisted in Shanghai for these years. Our study will benefit our better understanding of mutation and control of SIV.