Construct novel influenza A (H1N1) virus chimeric HA genes for development of bro

描述（由申请人提供）：甲型流感病毒在多种鸟类和哺乳动物宿主（包括人类和猪）中引起高度传染性呼吸道疾病。最近出现的新型猪源性甲型H1N1流感病毒2009 A （H1N1）的出现和大流行分类对公众健康构成重大威胁。人们担心该病毒可能与季节性人类流感病毒重新组合，从而增加传播性或致病性。特别是，这些病毒似乎保留了传播回猪的潜力，因此继续与猪病毒重组可能会产生更强毒的病毒。需要确定和实施有效的战略，以预防当前和未来的疫情。控制流感病毒流行的主要手段是疫苗接种，针对表面糖蛋白HA的中和抗体是目前大多数许可的流感病毒疫苗的靶标。然而，疫苗接种对流感病毒的效力受到血凝素基因内快速抗原变异（抗原漂移）的严重限制。针对目前流行的季节性人类A （H1N1）病毒培养的雪貂感染后抗血清与2009年A （H1N1）猪源染色株没有反应。从历史上看，每个流感亚型包含一个以上的分离物受到抗原限制，包括刺激免疫所需的透明质酸量、总透明质酸含量耐受性和一种疫苗株对另一种疫苗株的免疫优势潜力。为了开发一种可以广泛有效地对抗各种病毒株的疫苗，并限制新型猪源H1N1流感病毒的传播，这项拟议的研究将测试一种分子育种（DNA洗牌）方法的可行性，以创建H1N1流感病毒的新型HA基因，用于开发广泛保护性疫苗。这种方法将通过结合猪源性和人源性H1N1流感病毒的HA基因来创造新的HA基因，这些HA基因代表了五个不同的系统发育集群。重组嵌合HA抗原诱导的保护效果将通过猪攻击模型进行评估。具体目的是：1)对甲型H1N1流感病毒5个主要系统发育簇的HA进行分子育种（DNA重组），构建HA嵌合基因；2)评价重组嵌合HA抗原在猪攻毒模型中作为亚单位疫苗的潜力。这项研究将产生新的血凝素抗原，并测试它们诱导针对猪和人类来源的甲型H1N1流感病毒的广泛免疫反应的能力。以这种方式开发疫苗将证明，对人类和动物病毒分离物诱导免疫反应的疫苗将提供广泛的免疫，并防止未来从动物来源出现新的HAs进入人类群体。研究团队将招募积极性高的本科生参与项目，在研究方法、疾病机制、疾病预防等方面提供宝贵的培训（见附件：本科生培训）。

DESCRIPTION (provided by applicant): Influenza A virus causes highly contagious respiratory diseases in a variety of avian and mammalian hosts, including humans and pigs. The recent emergence and pandemic classification of novel swine-origin influenza A H1N1 virus, 2009 A (H1N1) poses a significant threat to public health. There are concerns that the virus may reassort with seasonal human influenza viruses giving rise to either increased transmissibility or pathogenicity. In particular, these viruses appear to retain the potential to transmit back to swine and thus continued reassortment with swine viruses may generate more virulent viruses. Identification and implementation of effective strategies to prevent current and future outbreaks are needed. The primary means for controlling influenza virus epidemics is vaccination, with neutralizing antibody against the surface glycoprotein HA being the target of most of the currently licensed influenza virus vaccines. However, the efficacy of vaccination towards influenza virus is severely limited by rapid antigenic variations within the HA gene (antigenic drift). Ferret postinfection antisera raised against the currently circulating seasonal human A (H1N1) viruses did not react with the 2009 A (H1N1) swine-origin stains. Historically, inclusion of more than one isolate per subtype of influenza has been limited by antigenic constraints, including the amount of HA required to stimulate immunity, the total HA content tolerance, and the potential for immunodominance by one vaccine strain over the other. In order to develop a vaccine that can be broadly effective against various strains of virus, and limit the spread of novel swine-origin H1N1 influenza viruses, this proposed study will test the feasibility of a molecular breeding (DNA shuffling) approach to create novel HA genes of influenza H1N1 viruses for development of broadly protective vaccines. This approach will create the novel HA genes by combination of HA genes from swine and human- origin influenza H1N1 viruses representing five distinct phylogenetic clusters. The efficacy of the shuffled chimeric HA antigen-induced protection will be assessed using a pig challenge model. Specific aims are: 1) To create chimeric HA genes by molecular breeding (DNA shuffling) of influenza HAs from five major phylogenic clusters of influenza A H1N1 viruses; 2) To evaluate the potential of using the shuffled chimeric HA antigen as a subunit vaccine in a pig challenge model. This study will generate novel HA antigens and test their ability to induce broad immune responses against swine and human-origin influenza A H1N1 viruses. Development of a vaccine in this manner will serve as a proof of principle that vaccines inducing immune response toward both human and animal viral isolates will provide broad immunity, and prevent future emergence of novel HAs from animal sources into the human population. The research team will recruit highly motivated undergraduate students to participate in the project, which provides valuable training in research methods, disease mechanisms and disease prevention (refer to attached document: Undergraduate student training). PUBLIC HEALTH RELEVANCE: Seasonal influenza virus infections are associated with 3-5 million hospitalizations and 250-500,000 deaths on an annual basis in the industrialized world. The recent emergence of the swine-origin influenza A (H1N1) virus that is genetically divergent from human seasonal influenza vaccine strain exemplifies the need for development of an effective vaccine in a short period of time. This proposed study will use molecular breeding approach by DNA shuffling and screening to generate chimeric influenza HA antigens that have the capacity to induce a broadly protective immune response against swine and human-origin influenza A H1N1 viruses. The technology established in this study can be applied to chimeric HA genes within (or between) other subtypes of influenza virus. The long term goal of our study is to develop universal vaccines that can elicit broad immunity to prevent future pandemics.