预先存在的免疫力对学生人群甲型H1N1流感发病的影响和变异抗原表位的识别

After experiencing influenza A (H1N1)pdm09 pandemic of 2009-2010, the general population, especially students，obtained a high level of the antibody against the virus. During the post-pandemic period, influenza A (H1N1)pdm09 continue to circulate as seasonal influenza, and influenza A (H1N1)pdm09 outbreaks in school have occurred occasionally. In early phase of the next influenza A (H1N1)pdm09 wave, we will enroll 15-30 patients aged 10-17 years who was confirmed with influenza A (H1N1)pdm09 infection during the pandemic (i.e. the former cases). After interview on vaccination information, we will collect them 10ml anticoagulant blood. In early phase of the next influenza A (H1N1)pdm09 wave, we will enroll 50-70 new confirmed patients aged 10-17 years from the new influenza A (H1N1)pdm09 outbreaks in school. These new cases will be asked some information on the onset of illness and vaccination, and be collected nasopharyngeal swabs and 10 ml of anticoagulant blood(acute phase) within 3 days of the onset. They will be collected 10 ml anticoagulant blood in 4-6 weeks after the onset(convalescent phase). The blood from the acute phase of the new cases and from the former cases respectively will be isolated to serum and peripheral blood cells. Serum will be used to test neutralizing antibodies. We stimulate peripheral blood cells using ELISPOT and tetramer staining (H1N1) to measure memory T cell responses by conservative epitope peptide, non-conservative epitope peptide, specific epitope peptide derived from influenza A (H1N1)pdm09. And then we compare these indicators to test their statistical difference by the two cohorts. We will explore the correlation of pre-existing humoral and cellular immunity with influenza A (H1N1)pdm09. The influenza A (H1N1)pdm09 strain isolated from nasopharyngeal swabs will be conduct sequencing and antigen analysis, and identify variation epitopes by immune epitope mapping and tetrimer-guided epitope mapping, the peripheral blood cells from the convalescent phase of new cases in vitro experiment will be stimulated to test the T cell response to the variant immunodominant epitope peptide derived from the new influenza A (H1N1)pdm09 strain.

2009-10年甲型H1N1流感大流行后，人群尤其学生获得高水平免疫抗体，甲型H1N1呈季节流行，学校暴发时有发生。在下次甲型H1N1流行初期，征集10-17岁、大流行期间确诊的甲型H1N1病例（即往病例）15-30例，收集疫苗接种等信息并采集10ml抗凝血；流行期间，征集学校暴发同年龄段、发病3日内的新甲型H1N1病例50-70例，收集相关信息采集鼻咽拭子和10ml抗凝血（急性期），采集病后4-6周10ml抗凝血（恢复期）。新病例急性期和即往病例抗凝血分离出血清和外周血细胞，检测血清的中和抗体，外周血细胞采用ELISPOT和四聚体染色法分别测定甲型H1N1保守、非保守、特异的表位肽记忆性T细胞反应，并比较两组各指标，探讨预先存在的体液和细胞免疫对甲型H1N1发病的影响。分离的病毒进行序列、抗原分析，识别变异的抗原表位，新病例恢复期外周血细胞体外实验确认变异的、免疫显性表位肽的T细胞反应。