Orthopoxvirus vector has a broad prospect in recombinant vaccine research, but the rarely severe side-effect impedes its development. Vaccinia virus and Cowpox virus of Orthopoxvirus have broad host range, and they have typical host range genes as K1L, CP77 and C7L. These three genes affect host range of Vaccinia virus, disturb the cell signaling pathways, suppress immune response and are related to virulence.
Cell Line ; Cowpox virus ; genetics ; immunology ; pathogenicity ; physiology ; Genetic Vectors ; Host Specificity ; genetics ; Orthopoxvirus ; genetics ; immunology ; pathogenicity ; physiology ; Signal Transduction ; Vaccines, Synthetic ; immunology ; Vaccinia virus ; genetics ; immunology ; pathogenicity ; physiology ; Viral Proteins ; genetics ; metabolism ; Viral Vaccines ; immunology ; Virulence

BACKGROUNDTo investigate expression of IL-6 in non?replicating vaccinia virus and its immune effects on recombinant virus.

METHODSThe recombinant non replicating vaccinia virus RVJ123 delta CK11 beta 75IL6 was constructed with non?replicating vaccinia virus vector pNEOCK11beta75IL6 and replicating vaccinia virus RVJ123. In animal model, immunization with the recombinant virus was carried out and its immune response was studied.

RESULTSThe recombinant virus could express IL-nd HBsAg simultaneously. Southern blot analysis demonstrated that the genes between vaccinia virus Hind? C and K fragments were deleted and IL-6 gene was integrated stably. Given intranasal inocula of the virus to immunize BALB/c mouse and New Zealand Rabbit, the elevated anti-HBsAg IgA and IgG antibody secreting cells in mouse lung lymphoid to vectors expressing IL-6 was at about two?fold higher level than those elicited by control virus at day 14 after immunization. Authors also could detect elevated anti-HBsAg IgA and IgG antibody conversion in mouse serum and lung fluid, rabbits serum, lung fluid, saliva, vagina and nasal washing samples.

CONCLUSIONSIL-6 expressed by non-replicating recombinant vaccinia virus could enhance the induced?immune effects, it could serve as the effective adjuvant for recombinant vector vaccine.

Adjuvants, Immunologic ; Animals ; Cells, Cultured ; Chick Embryo ; Female ; Genetic Vectors ; Humans ; Immunoglobulin A ; analysis ; Immunoglobulin G ; analysis ; Interleukin-6 ; biosynthesis ; genetics ; immunology ; Lung ; immunology ; Mice ; Mice, Inbred BALB C ; Rabbits ; Recombination, Genetic ; Vaccinia virus ; immunology ; metabolism ; physiology ; Virus Replication

OBJECTIVETo generate a human papillomavirus (HPV16) prophylactic and therapeutic vaccine candidate for cervical cancer.

METHODSHPV16 major capsid protein L1 gene/minor capsid protein L2 gene and HPV16 early E6/E7 genes were inserted into a vaccinia virus expression vector. A strain of non-recombinant vaccinia virus containing the sequences was obtained through a homologous recombination and identified.

RESULTSDNA hybridization confirmed that the HPV16L1/L2/E6/E7 genes were integrated into vaccinia virus DNA. Western Blot result showed that full-length L1/L2/E6/E7 proteins were co-expressed in CEF cells infected with the recombinant virus.

CONCLUSIONNTVJE6E7CKL1L2 could be taken as a candidate of prophylactic and therapeutic vaccine for HPV-associated tumors and their precancerous transformations.

Animals ; Blotting, Western ; Capsid Proteins ; genetics ; metabolism ; Cells, Cultured ; Chick Embryo ; Cloning, Molecular ; Female ; Gene Expression ; Genetic Vectors ; genetics ; Humans ; Oncogene Proteins, Viral ; genetics ; metabolism ; Papillomaviridae ; genetics ; immunology ; Papillomavirus E7 Proteins ; Papillomavirus Infections ; immunology ; prevention & control ; virology ; Papillomavirus Vaccines ; genetics ; immunology ; therapeutic use ; Recombinant Fusion Proteins ; genetics ; immunology ; metabolism ; Repressor Proteins ; genetics ; metabolism ; Transfection ; Tumor Virus Infections ; immunology ; prevention & control ; virology ; Uterine Cervical Neoplasms ; immunology ; prevention & control ; virology ; Vaccinia virus ; genetics ; Virus Replication

Vaccinia virus complement control protein (VCP) is one of the proteins encoded by vaccinia virus to modulate the host inflammatory response. VCP modulates the inflammatory response and protects viral habitat by inhibiting the classical and the alternative pathways of complement activation. The extended structure of VCP, mobility between its sequential domains, charge distribution and type of residues at the binding regions are factors that have been identified to influence its ability to bind to complement proteins. We report that a Lister strain of vaccinia virus encodes a VCP homolog (Lis VCP) that is functional, glycosylated, has two amino acids less than the well-characterized VCP from vaccinia virus WR strain (WR VCP), and the human smallpox inhibitor of complement enzymes (SPICE) from variola virus. The glycosylated VCP of Lister is immunogenic in contrast to the weak immunogenicity of the nonglycosylated VCP. Lis VCP is the only orthopoxviral VCP homolog found to be glycosylated, and we speculate that glycosylation influences its pattern of complement inhibition. We also correlate dimerization of VCP observed only in mammalian and baculovirus expression systems to higher levels of activity than monomers, observed in the yeast expression system.
Amino Acid Sequence ; Animals ; Cell Line ; Cercopithecus aethiops ; Complement Activation ; drug effects ; immunology ; Complement System Proteins ; metabolism ; Dimerization ; Gene Expression ; Glycosylation ; Humans ; Molecular Sequence Data ; Protein Binding ; Protein Structure, Tertiary ; Recombinant Proteins ; genetics ; metabolism ; Smallpox ; immunology ; metabolism ; Structure-Activity Relationship ; Vaccinia virus ; chemistry ; immunology ; metabolism ; Variola virus ; chemistry ; immunology ; metabolism ; Viral Proteins ; genetics ; metabolism ; pharmacology

A recombinant modified vaccinia Ankara (MVA) virus expressing mature viral protein 2 (VP2) of the infectious bursal disease virus (IBDV) was constructed to develop MVA-based vaccines for poultry. We demonstrated that this recombinant virus was able to induce a specific immune response by observing the production of anti-IBDV-seroneutralizing antibodies in specific pathogen-free chickens. Besides, as the epitopes of VP2 responsible to induce IBDV-neutralizing antibodies are discontinuous, our results suggest that VP2 protein expressed from MVA-VP2 maintained the correct conformational structure. To our knowledge, this is the first report on the usefulness of MVA-based vectors for developing recombinant vaccines for poultry.
Animals ; Antibodies, Viral ; Birnaviridae Infections/prevention & control/*veterinary ; Cells, Cultured ; Chick Embryo ; Chickens ; Fibroblasts/metabolism ; Infectious bursal disease virus/*immunology ; Poultry Diseases/*prevention & control/virology ; Specific Pathogen-Free Organisms ; Vaccinia virus/*genetics/immunology/metabolism ; Viral Structural Proteins/genetics/*immunology/metabolism ; Viral Vaccines/*immunology

Modified ORF5 (MORF5) and ORF6 gene of PRRSV were cloned into two multiple cloning sites of MVA transfer vector pLR-gpt to construct the recombinant plasmid pLR-MORF5/ORF6. Homologous recombination between pLR-MORF5/ORF6 and the wtMVA on BHK-21 cell line was mediated with liposome by infecting the cell with 0.01 MOI wtMVA two hours before transfecting the recombinant plasmid into the cell. When the cytopathic effect (CPE) was obvious, virus was collected from the cell plate and the recombinant virus was selected with drug selecting medium (2% MXHAT). After 12 cycles of selection, rMVA with a selection marker Eco gpt was obtained and named as rMVAgpt-MGP5/M. By infecting BHK-Cre expressing Cre recombinant enzyme, the Eco gpt marker in rMVAgpt-MGP5/M was deleted and this rMVA was named as rMVA-MGP5/M. The insertion of MORF5 and ORF6 into the MVA genome was confirmed with PCR analysis and the expression of MGP5 and M protein was identified with Western blot and IFA. Through biological study on the recombinant MVA, no obvious difference was observed between rMVA-MGP5/M and the wtMVA regarding to the CPE and growth curve. The recombinant MVA constructed in this study could coexpress the modified GP5 and M protein and the expressed product had good immunocompetence. Furthermore, the insertion of the MORF5 and ORF6 into MVA genome had no obvious effect on the replication and biological characteristics of this virus.
Animals ; Cell Line ; Genetic Vectors ; genetics ; Porcine Reproductive and Respiratory Syndrome ; prevention & control ; Porcine respiratory and reproductive syndrome virus ; genetics ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; immunology ; Recombination, Genetic ; Swine ; Transfection ; Vaccines, DNA ; genetics ; immunology ; Vaccinia virus ; classification ; genetics ; metabolism ; Viral Envelope Proteins ; biosynthesis ; genetics ; Viral Matrix Proteins ; biosynthesis ; genetics