Immune Responses Against SARS CoV Induced By Virus-like Particles And Studies On The Baculoviruse As The Live Virus Vaccine Vector

This thesis contains two parts, in which we studied the immune responses againstSARS CoV induced by virus-like particles and the feasibility of the Baculoviruse as thelive virus vaccine vector.First Part: Baculoviruses' characteristics of biosafety, large capacity, ease to bemanipulated and low cost make them be engineered as expression vector and bedeveloped and improved constantly. In this thesis, we got the SARS CoV virus-likeparticles in insect cell using Bac-to-Bac baculovirus express system and studied theimmunogenic by immunizing mice with purified VLPs, expecting to provide a new safetyvaccine strategy for SARS CoV and a tool for study the virus assembly.Chapter one gave an overview outlines the character of SARS CoV, major on themoleculebiology research, especially on the progress in SARS CoV vaccine.In chapter two we constructed two recombinant baculoviruses: vAcEM, containingSARS CoVE and M gene; vAcS, containing SARS CoV S gene.In chapter three insect cells were co-infected with vAcME and vAcS, and SARS CoVVLPs can be observed in the infected cells by electron microscope. Immunogold labelingresults showed S protein was assembled into the VLPs. The VLPs were purified bycentrifugation.In chapter four we immunized mice with purified VLPs, identified humoral andcell-mediated immune response against SARS CoV and neutralizing antibodies againstSARS CoV were analyzed using SARS CoV S protein-pseudotyped murine leukemiavirus (MLV) vector particles. Our findings demonstrate that SARS CoV VLPs are immunogenic and can elicit strong SARS CoV-specific humoral and cellular immuneresponses in mice.Second Part: Baculovirus AcMNPV has shown the ability of high efficientlytransducing into a variety of mammalian cell types and achived a high-level expression offoreign gene under the mammalian promotor. So it may express a foreign gene in vivo andinduced immune response. In this part, we constructed two recombinant baculoviruses,expressing HA and NA gene of AIV H5N1 under the mammalian promotor, injected therecombinant AcMNPV into mice and tested the titer change of antibody against AIVH5N1.Chapter one is a brief review recent advance on AIV molecular characteristics andvaccine. Another part is about the Baculovirus can transduce into some mammaliankidney cells and express foreign gene under heterogonous promoter.In chapter two we constructed two pFastBac DUAL plasmid, containing HA and NAgene ofAIV H5N 1 under the mammalian promoter.In chapter three we got tWO recombinant baculoviruses, expressing HA and NA geneof AIV H5N1. After tested the efficiency of AcMNPV by transducing into BHK cells, theAcMNPV were injected into mice. The titer change of antibody was tested by ELISA.Our findings demonstrate that recombinant baculoviruses can elicit strong AIV-specifichumoral immune responses in mice.