The Study Of Recombinant Newcastle Disease Viruses Expressing Porcine Circovirus Type2Cap Protein

Porcine circovirus type2(PCV2), a member of the family Circoviridae, is recognized asthe major causative agent of the postweaning multisystemic wasting syndrome (PMWS),various other clinical manifestations including reproductive failure and porcine dermatitisand nephropathy syndrome (PDNS). All these clinical conditions were namedPCV-associated diseases (PCVAD). After first reported in1991in Canada, by now thePWMS has spread all over the world and caused major economic loss in the global pigindustries.PCV2is non-enveloped single-stranded circular DNA virus with a1.76kb ambiencegenome. The capsid protein (Cap) is the major immunogenic protein and associated withthe production of PCV2-specific neutralizing antibodies. The Cap is also an ideal targetantigen in the design of a new generation of vaccines against PCV2infection. To date, thespread of commercially available vaccines now are restricted by its tedious process andexpensive price.The application of reverse-genetic techniques based on Newcastle disease virus (NDV)opens the way for the NDV as a vaccine vector for the control of mammal diseases. Thisresearch aims to evaluate the expression level of PCV2capsid protein by recombinantNDV and develop a novel potential vaccine vector against PCV2infection.By the reverse genetics techniques system based on NDV La Sota vaccine strain, therecombinant NDV rLa-PCV2/Cap expressing the Cap protein was successfully constructed.To improve the expression of Cap, two recombinant NDVs rLa-PCV2/CapΔ41andrLa-PCV2/CapΔ18expressing truncated Caps with the nuclear localization signal (NLS)or part of NLS removed were constructed respectively. Furthermore, in order to improvethe immunogenicity of recombinant NDV in pig the rLa-PCV2/CapΔ18IL2SP which could express the CapΔ18-IL2fusion protein was constructed. The expression of Capprotein and all kinds of modified Cap proteins were confirmed by indirectimmunofluorescence assay. To study the biological properties of recombinant NDVs thegrowth property in chicken eggs, the MDT, ICPI and IVPI were evaluated.These results indicated that all of PCV2Cap and modified Caps were successfullyexpressed in BHK-21cells. After truncation CapΔ41and CapΔ18were both successfullyreleased from inside the nucleus. All those recombinant viruses retained the La Sotastrain’s high-titer growth propertyin chicken eggs and low pathogenicity. Theimmunogenicity of those recombinant viruses need to be further detected.