Classical swine fever virus E2 glycoprotein antigen produced in adenovirally transduced PK-15 cells confers complete protection in pigs upon viral challenge.
E2 is the major envelope glycoprotein present on the outer surface of the classical swine fever virus (CSFV). It is exposed as a homodimer originated by disulfide linkage and represents an important target for the induction of neutralizing immune responses against the viral infection. The E2his glycoprotein nucleotide sequence used in this work contains the CSFV E2 extracellular domain preceded by the tissue plasminogen signal peptide and a hexa-histidine tag in the 3' terminus. The recombinant antigen was produced at a range of 120-150 microg/mL in the culture media of epithelial kidney pig cells, transduced with a replication defective adenoviral vector (Ad-E2his) generated by means of cloning the E2his sequence in the vector genome. The glycoprotein was obtained from clarified culture media as a homodimer of 110 kDa with purity over 95% after a single affinity chromatography step in Ni-NTA Agarose column. The E2his characterization by lectin-specific binding assay showed the presence of N-linked oligosaccharides of both hybrid and complex types. The protective capacity of E2his was demonstrated in two immunization and challenge experiments in pigs using doses of 15 or 30 microg of the glycoprotein, emulsified in Freund's adjuvant. The intramuscular immunization followed by a unique boost three weeks later, elicited high titers of neutralizing antibodies between the second and the fourth week after the primary vaccination. The immunized animals were fully protected from the viral infection after challenge with 10(5) PLD(50) of homologous CSFV "Margarita" strain administered by intramuscular injection. Consequently, no clinical signs of the disease or viral isolation from lymphocytes were detected in the vaccinated pigs. These results suggest that the E2his antigen produced in mammalian cells may be a feasible vaccine candidate for CSF prevention.
Sánchez
Oliberto
O
Animal Biotechnology Department, Centre for Genetic Engineering and Biotechnology (CIGB), Havana, Cuba.
Barrera
Maritza
M
Rodríguez
María P
MP
Frías
María T
MT
Figueroa
Nancy E
NE
Naranjo
Paula
P
Montesino
Raquel
R
Farnos
Omar
O
Castell
Sara
S
Venereo
Alina
A
Ganges
Llilianne
L
Borroto
Carlos
C
Toledo
Jorge R
JR
D000256:Adenoviridae / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D000914:Antibodies, Viral / Q000097:blood; D002478:Cells, Cultured; D006691:Classical Swine Fever / Q000401:mortality / Q000517:prevention & control* / Q000821:virology; D006692:Classical Swine Fever Virus / Q000276:immunology* / Q000472:pathogenicity; D007668:Kidney / Q000166:cytology / Q000378:metabolism* / Q000821:virology; D013552:Swine; D014161:Transduction, Genetic; D014611:Vaccination / Q000662:veterinary*; D014759:Viral Envelope Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D014765:Viral Vaccines / Q000008:administration & dosage / Q000235:genetics / Q000276:immunology
D000914:Antibodies, Viral; D014759:Viral Envelope Proteins; D014765:Viral Vaccines; C087086:glycoprotein E2, classical swine fever virus