ABSTRACT
Therapies that selectively target cancer cells for death have been the center of intense research recently. One potential therapy may involve apoptin proteins, which are able to induce apoptosis in cancer cells leaving normal cells unharmed. Apoptin was originally discovered in the Chicken anemia virus (CAV); however, human gyroviruses (HGyV) have recently been found that also harbor apoptin-like proteins. Although the cancer cell specific activity of these apoptins appears to be well conserved, the precise functions and mechanisms of action are yet to be fully elucidated. Strategies for both delivering apoptin to treat tumors and disseminating the protein inside the tumor body are now being developed, and have shown promise in preclinical animal studies.
Subject(s)
Antineoplastic Agents/pharmacology , Capsid Proteins/pharmacology , Drug Delivery Systems/methods , Animals , Antineoplastic Agents/administration & dosage , Apoptosis/drug effects , Capsid Proteins/physiology , Cell Death/drug effects , Chicken anemia virus/chemistry , Gyrovirus/chemistry , Humans , Viral Proteins/isolation & purification , Viral Proteins/pharmacologyABSTRACT
The presence of infectious chicken anemia virus (CAV) was detected in a previous study by nested-PCR as a contaminant in seven commercial vaccines, produced in the 1990s by three different manufacturers, prepared against the most relevant virus etiologies. In order to phylogenetically characterize the genome and compare it to CAV isolates from Brazil and other parts of the world, sequences of approximately 675 bp of the gene encoding the hypervariable region of VP1 protein of three CAV vaccine contaminant strains were studied. The CAV genome in contaminated vaccines showed high similarity (> 98.9%) with the Brazilian BR91/99 and Argentinian ArgA001028 (> 99%) strains. However, the comparison with the Cuxhaven-1 vaccine strain showed a lower identity of between 96.8% and 97.7%, and comparing it with the CAV26P4 vaccine strain showed an identity between 97.2% and 98.2%; both are available in Brazil. Such differences might be relevant for the highly conserved CAV genome. CAV contaminants were positioned in the same genetic group (clusters) with the Brazilian strain BR91/99 and Argentinian strain ArgA001028. Results indicated that the contamination of live vaccines by CAV may have influenced CAV epidemiology in the Brazilian and Argentinian poultry industry.
Subject(s)
Chicken anemia virus/genetics , Chicken anemia virus/immunology , Chickens , Viral Vaccines/genetics , Viral Vaccines/immunology , Animals , Capsid Proteins/chemistry , Capsid Proteins/genetics , Capsid Proteins/metabolism , Chicken anemia virus/chemistry , Molecular Sequence Data , Phylogeny , Polymerase Chain Reaction , Polymorphism, Genetic , Sequence Analysis, DNA , Sequence Analysis, Protein , Sequence Homology , Vaccines, Attenuated/geneticsABSTRACT
Purification of chicken anemia virus (CAV) VP3 protein, expressed in a prokaryotic expression system as histidine-tagged fusion protein is demonstrated in the present study. CAV particle was obtained from infected liver of chicken and DNA was extracted. The VP3 protein gene was amplified from the extracted DNA by polymerase chain reaction (PCR) and cloned. The recombinant expression construct (pTrc-VP3) was identified by PCR and sequencing analysis. Expression of VP3 protein with a molecular mass of approximately 21kDa was confirmed by Western blotting analysis with CAV-specific antibodies. The in vitro expressed VP3 protein was purified to near homogeneity by elution from the gel, as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. The purified VP3 protein was recognized by CAV antibodies in a Western blotting assay. This finding indicates that recombinant VP3 expressed in the pTrcHis2 vector system can be used as antigen to detect anti-CAV antibodies.
Subject(s)
Capsid Proteins/genetics , Chicken anemia virus/genetics , Cloning, Molecular , Gene Expression , Animals , Antibodies, Viral/blood , Antibodies, Viral/immunology , Blotting, Western , Capsid Proteins/immunology , Capsid Proteins/isolation & purification , Chicken anemia virus/chemistry , Chickens , Circoviridae Infections/diagnosis , Circoviridae Infections/veterinary , DNA, Viral/genetics , Escherichia coli/genetics , Genetic Vectors , Poultry Diseases/diagnosis , Recombinant Proteins/genetics , Recombinant Proteins/isolation & purificationABSTRACT
Chicken anaemia virus (CAV) was detected by a Nested-PCR assay in field samples from different regions of Brazil. The 539 bp amplified fragments of vp1 gene from 44 field samples were sequenced and 10 new nucleotide sequences of CAV were observed. These sequences were phylogenetically analysed by Mega2 using neighbour joining distance methods with 1000 bootstrap replications. Phylogenetic analysis did not show correlation between CAV pathology pattern and genetic groups. The 10 nucleotide sequences of the Brazilian samples were also analysed together with 30 sequences of CAV strains previously described from other countries. The genetic variability observed was not related to the geographical distribution. Amino acid substitutions were detected at 9 positions of the Brazilian sequences and two of them had not been observed before, (65)R replacing the Q residue and (98)F replacing Y residue.