Pathology of Equine Influenza virus (H3N8) in Murine Model

Equine influenza viruses (EIV)—H3N8 continue to circulate in equine population throughout the world. They evolve by the process of antigenic drift that leads to substantial change in the antigenicity of the virus, thereby necessitating substitution of virus strain in the vaccines. This requires frequent testing of the new vaccines in the in vivo system; however, lack of an appropriate laboratory animal challenge model for testing protective efficacy of equine influenza vaccine candidates hinders the screening of new vaccines and other therapeutic approaches. In the present investigation, BALB/c mouse were explored for suitability for conducting pathogenecity studies for EIV. The BALB/c mice were inoculated intranasally @ 2×106.24 EID50 with EIV (H3N8) belonging to Clade 2 of Florida sublineage and monitored for setting up of infection and associated parameters. All mice inoculated with EIV exhibited clinical signs viz. loss in body weights, lethargy, dyspnea, etc, between 3 and 5 days which commensurate with lesions observed in the respiratory tract including rhinitis, tracheitis, bronchitis, bronchiolitis, alveolitis and diffuse interstitial pneumonia. Transmission electron microscopy, immunohistochemistry, virus quantification through titration and qRT-PCR demonstrated active viral infection in the upper and lower respiratory tract. Serology revealed rise in serum lactate dehydrogenase levels along with sero-conversion. The pattern of disease progression, pathological lesions and virus recovery from nasal washings and lungs in the present investigations in mice were comparable to natural and experimental EIV infection in equines. The findings establish BALB/c mice as small animal model for studying EIV (H3N8) infection and will have immense potential for dissecting viral pathogenesis, vaccine efficacy studies, preliminary screening of vaccine candidates and antiviral therapeutics against EIV.

马流感病毒（Equine influenza viruses, EIV）H3N8亚型目前仍在全球马群中持续传播。该病毒通过抗原漂移机制不断进化，导致病毒抗原性发生显著改变，因此需要更新疫苗所使用的病毒毒株。这就需要频繁在体内系统中对新型疫苗进行测试；然而，目前缺乏用于评估马流感疫苗候选株保护效力的合适实验动物攻击模型，这阻碍了新型疫苗及其他治疗手段的筛选工作。

本研究探索了BALB/c小鼠用于开展马流感病毒致病性研究的适用性。研究人员以属于佛罗里达亚系分支2的H3N8亚型马流感病毒，以2×10^6.24 EID50的剂量经鼻内接种BALB/c小鼠，并监测感染建立情况及相关检测指标。所有接种马流感病毒的小鼠均在接种后3至5天内出现临床症状，包括体重下降、精神萎靡、呼吸困难等，这些症状与呼吸道出现的病变相符，病变类型涵盖鼻炎、气管炎、支气管炎、细支气管炎、肺泡炎及弥漫性间质性肺炎。

通过透射电子显微镜、免疫组织化学、病毒滴定定量及实时定量反转录聚合酶链反应（qRT-PCR）检测，证实小鼠上下呼吸道均存在活跃的病毒感染。血清学检测结果显示，小鼠血清乳酸脱氢酶水平升高，同时出现血清转换现象。本研究中小鼠的疾病进展模式、病理损伤特征以及从鼻洗液和肺组织中分离到病毒的情况，与马自然感染和实验感染马流感病毒的表现相当。

本研究结果证实BALB/c小鼠可作为研究H3N8亚型马流感病毒感染的小型动物模型，在解析病毒致病机制、开展疫苗效力研究、初步筛选疫苗候选株以及研发抗马流感病毒治疗手段等方面具有巨大应用潜力。