Genetic Variation Of S2 Gene Of The Donkey Leukocyte-attenuated Vaccine Strain Of Equine Infectious Anemia Virus In Experimentally Infected Ponies

Equine infectious anemia virus (EIAV) is a member of the lentivirus subfamily of retroviruses and causes a persistent infection and chronic disease in horses worldwide. In the early 1970s, Chinese scientists successfully developed a donkey-leukocyte-attenuated EIAV vaccine, EIAVDLV121, which is the first lentiviral vaccine with a successful application. EIAVDLV121 can induce immune protection and can be safely used for vaccination. Thus, an understanding of the mechanismthat underlies the attenuation of virulence and the induction of immune protection imparted by this vaccine will elucidate the immune protective mechanism that is responsible for lentivirus infection and facilitate the development of lentivirus vaccines.The genetic organization of EIAV is relatively simple compared with that of other lentiviruses in that the EIAV genome contains only three accessory genes, initially termed S1, S2, and S3, in addition to the standard gag, pol, and env genes common to all retroviruses. The EIAV S2 gene is in fact an important determinant of viral replication and pathogenic properties in vivo. During EIAVDLV121 attenuated in vitro, S2 gene is one of large variable regions.Thus,the variation of S2 gene may caused the depression of virulence. Dynamic Evolution of gp90 Gene of EIAVDLV121 has been analyzed in experimentally Infected ponies,but the variation of S2 gene has not been investigated in vivo. The in vivo variation of S2 gene of the equine infectious anemia virus (EIAV) attenuated vaccine adapted to cultivated donkey leukocytes was investigated by analyzing experimentally infected horses. Four horses (#1,#2,#5 and #8) were inoculated with the EIAV vaccine strain EIAVDLV121 and another two horses were injected with saline as health controls. Body temperature, platelet and viral copies of these horses were monitored for 5 months. The viral copies were evaluated by real time RT PCR and the S2 gene fragment of EIAVDLV121 was amplified by RT-PCR from plasma of horses at indicated time points and were cloned and sequenced.The Results showed that all the horses didn't appear typical signs of equine infectious anemia. After inoculation. The number of viral copies were below 1×10⁶ copies/ml, simultaneously body temperature and platelets were within normal limits.S2 gene had a lot of mutations in experimentally infected horses, which caused substitutions of translated amino acid residues in EIAV replicated in the hosts. substitutions of translated amino acid residues were in the 17th, 37th, 39th, 41st, 51st and 55th about #1 horse, in the 17th, 22nd, 37th, 39th, 41st, 51st and 55th about #2 horse, in the 6th, 18th, 22nd, 37th, 39th, 41st, 51st and 55th about #5 horse and 37th, 39th, 41st, 51st and 55th about #8 horseIn addition, the S2 gene sequences of samples at the 3rd, 7th and 8th from horse #1 and samples at the 1st from horse #8 appeared shortened divergence to the consensus S2 sequence of EIAV pathogenic strains, EIAVLN40. The average differences from EIAV_LN40 were respectively 0.4 %,0.5 %, 0.3 %, 0.5 % about nucleotide and 0.9 %, 1.3 %, 0.8 %, 1.3 % about amino acid. The results above indicated that Chinese equine infectious anemia virus attenuated vaccine have a favorable safety. The variation of S2 gene in vivo couldn'tcause increase of viral virulence. Our data on the EIAV S2 evolution in vivo will facilitate the studies on procedures of infection of pathogenic EIAV and vaccine strains.