Liposomes Containing Recombinant Gp85Protein Vaccine Against ALV-J In Chickens

Subgroup J avian leukosis virus (ALV-J), which was first isolated from the meat-typechickens in1990s, mainly caused myeloid leukosis (ML). In1999, the virus was isolatedfrom the white feather broiler chickens in China. Up to date, it has been reported that the virushas spread to broilers, commercial layers, as well as local Chinese chicken breeds in China,inducing serious losses to the poultry industry.To study the potential of liposome vaccines in the clinical prevention of ALV-J, the effectof recombinant gp85protein of ALV-J entrapped by liposomes in chickens against ALV-Jinfection was investigated in this study.The method of prokaryotic expression was adopted to prepare the recombinant gp85protein. Gene of gp85was transformed into the PET-28a vector and then expressed inRosetta(DE3) cells using0.5mM IPTG at37℃. The cells were harvested at6h postinductionand then purfied. And the protein concentration was determined as2mg/mL.The method of reverse-phase evaporation was used to prepare the liposome vaccine, ofwhich the encapsulation efficicency was evaluated by ways of protamine aggregation. Simply,the encapsulation efficicency was determined as40%under the following condition: the ratioof phospholipid-to-cholesterol(M)1:1, the concentration of recombinant protein2mg/mL,Tween-803mL, the ratio of solvent-to-aqueous phase3:1.Sixty Hy-Line Brown chickens were randomly divided into five groups: the controlgroup (containing challenge group and no challenge group), the gp85+L (inoculation once)group, the gp85+L(inoculation twice) group, the gp85+F (inoculation once) group, and thegp85+F (inoculation twice) group. The chickens were inoculated intramuscularly either onceor twice with the liposomes or with Freund’s adjuvant emulsion containing recombinant gp85protein at1week old and3weeks old. Moreover, blood samples were collected from eachchicken at weekly intervals for serum antibody analyses. Results showed that positive serumantibodies (S/P value>0.6) could be induced in all groups regardless of the frequency ofinoculation but improved significantly in the twice-inoculated groups. As well, high levels of antibodies emerged earlier in the Freund’s adjuvant groups but persisted longer in theliposome groups.All of the chickens, except those in the control group(no challenge group), werechallenged intraperitoneally with a102.450%tissue culture infective dose (TCID50) of ALV-JNX0101strain at5weeks old. For ALV viremia detection, blood samples were asepticallycollected at weekly intervals for3weeks. Results indicated that the liposomes provide betterprotection against ALV-J than Freund’s adjuvant emulsion and that this protection is directlyinfluenced by serum antibody levels.In a word, this study revealed the potential of liposome vaccines containing recombinantgp85protein in the clinical prevention of ALV-J, which will play an important role in thecontrol and eradication of ALV-J in China.