| Equine infectious anemia virus (EIAV), a member of lentivirus genus of retrovirusfamily, is the etiological agent of equine infectious anemia (EIA). Lentivirus brings greatchallenges to the health of human and animals, so researchers worldwide do their greateffort focusing on the preventive immunization against tentivirus. Chinese donkeyleukocyte attenuated strain of equine infectious anemia virus (DLA-EIAV) was the onlysuccessful lentivirus vaccine, which has been applied extensively in China against EIA fortwenty years. DLA-EIAV can be served as the animal model for studying on humanimmunodeficiency virus (HIV). The vaccine was developed by in vivo passaging EIAVstrain Liaoning (EIAV L) in donkeys (a donkey-adapted equine infectious anemia virus,DA-EIAV was obtained) and then in vitro passaging it in donkey leukocytes for many times.The DLA-EIAV inoculated animals could resist challenge from homogeneous orheterogeneous virulent FLAy. To elucidate the molecular basis of the attenuation ofDLA-EIAV in virulence thus provide theory foundation for designing HIV vaccine, thewhole genomes of DLA-EIAV and EJAV L provirus were cloned and sequenced. Aninfectious molecular clone derived from DLA-ELAV was constructed and characterized.Three DLA-EIAV/ L chimeric viruses were constructed. Promoting efficacy of the longterminal repeat (LTR) of the DLA-EIAV was charactenzed.In this study, proviral DNA was extracted from peripheral blood lymphocytes of ahorse infected with EIAV L, and donkey leukocvte culture infected by DLA-EIAV. Afterproviral DNA fragments were amplified by PCR, these PCR products were cloned andsequenced. The complete nucleotide sequences of DLA-EIAV and EIAV L proviral DNAwere determined. The proviral genomes of EIAV L and DLA-EIAV were 8235 bp and 8266in length, respectively. The DNasis analyzing results showed that the sequence otDLA-EIAV was 97.0% and 97.5% in homology with that of DLA-EIAV and DA-EJAV,respectively. The significant sequence change was found in LTR (including U3 13.2%, R7.5% and U5 5.1%), env (2.7%) and ORE S2 (3.9%) between DLA-EIAV and E]AV L,while the predicted amino acid sequence difference of Env and S2 protein from the twostrains were 4.4% and 8.8%, respectively. There was a high difference in nucleotidesequence between EIAV L and other alien ELAV strains, the homology between them wasonly about 79%.The amino acid homology of RTase and Gag protein between EIAV L andother alien strains were 86% and 79% ,respectively. These results shown that there was adistant relation between EIAV L and other alien strains.Through comparison of predicted gp9O amino acid sequence among EIAV LDA-EIAV. DLA-EIAV and other alien ELAV strains, we found that the virulence of EIIAV iscomcident with increased N-like glycosylation sites in gp9O coding region.There is a bHLH transcription factor binding consensus sequence, E-box in U3 ofDLA-EIAV but not in U3 of EIAV L. Through comparison of TAR among DLA-EIAV,EIAV L and other strains, we found there was variation in the origination site of TAR stemin DLA-EIAV. Virus gene expression is regulated by cellular transcriptional factors/cis-activating motifs and virus encoded trans-activating protein (Tat). Based on the importantroles of the transcriptional factors/cis-activating motifs in virus gene expression, theincorporation of E-box in U3 and variation of structure of TAR might result in attenuationof the DLA-EIAV.A recombinant plasmid (designated as EIAV-p8.2) containing entire DLA-EIAVprovira] genome was obtained by ligating cloned PCR products from DLA-EIAV. Thecomplete nucleotide sequence of EIAV-p8.2 was determined. The EIAV-p8.2 wastransfected into donkey leukocyte. Following an incubation period, reverse transcriptaseactivity was detected in cell culture supematants...
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