{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,10]],"date-time":"2026-10-10T00:25:33Z","timestamp":1791591933571,"version":"4.3.4"},"reference-count":33,"publisher":"MDPI AG","issue":"4","license":[{"start":{"date-parts":[[2019,4,24]],"date-time":"2019-04-24T00:00:00Z","timestamp":1556064000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["31672533"],"award-info":[{"award-number":["31672533"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["31302066"],"award-info":[{"award-number":["31302066"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["31672578"],"award-info":[{"award-number":["31672578"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Viruses"],"abstract":"<jats:p>As the only widely used live lentiviral vaccine, the equine infectious anima virus (EIAV) attenuated vaccine was developed by in vitro passaging of a virulent strain for 121 generations. In our previous study, we observed that the attenuated vaccine was gradually selected under increased environmental pressure at the population level (termed a quasispecies). To further elucidate the potential correlation between viral quasispecies evolution and pathogenesis, a systematic study was performed by sequencing env using several methods. Some key mutations were identified within Env, and we observed that increased percentages of these mutations were accompanied by an increased passage number and attenuated virulence. Phylogenetic analysis revealed that env mutations related to the loss of virulence might have occurred evolutionarily. Among these mutations, deletion of amino acid 236 in the V4 region of Env resulted in the loss of one N-glycosylation site that was crucial for virulence. Notably, the 236-deleted sequence represented a \u201cvaccine-specific\u201d mutation that was also found in wild EIAVLN40 strains based on single genome amplification (SGA) analysis. Therefore, our results suggest that the EIAV attenuated vaccine may originate from a branch of quasispecies of EIAVLN40. Generally, the presented results may increase our understanding of the attenuation mechanism of the EIAV vaccine and provide more information about the evolution of other lentiviruses.<\/jats:p>","DOI":"10.3390\/v11040380","type":"journal-article","created":{"date-parts":[[2019,4,25]],"date-time":"2019-04-25T03:02:59Z","timestamp":1556161379000},"page":"380","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":9,"title":["Characterization of EIAV env Quasispecies during Long-Term Passage In Vitro: Gradual Loss of Pathogenicity"],"prefix":"10.3390","volume":"11","author":[{"given":"Cong","family":"Liu","sequence":"first","affiliation":[{"name":"State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150069, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xue-Feng","family":"Wang","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150069, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yan","family":"Wang","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150069, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jie","family":"Chen","sequence":"additional","affiliation":[{"name":"Key Laboratory of Special Animal Epidemic Disease of Ministry of Agriculture, Institute of Special Animal and Plant Sciences, Chinese Academy of Agricultural Sciences, Changchun 130112, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0778-5053","authenticated-orcid":false,"given":"Zhaohua","family":"Zhong","sequence":"additional","affiliation":[{"name":"Department of Microbiology, Harbin Medical University, Harbin 150069, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yuezhi","family":"Lin","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150069, China"},{"name":"Department of Microbiology, Harbin Medical University, Harbin 150069, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4521-4099","authenticated-orcid":false,"given":"Xiaojun","family":"Wang","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150069, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2019,4,24]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"241","DOI":"10.1007\/s12250-012-3253-x","article-title":"The comparison of genetic variation in the envelope protein between various immunodeficiency viruses and equine infectious anemia virus","volume":"27","author":"Yuan","year":"2012","journal-title":"Virol. Sin."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"181","DOI":"10.1016\/j.vetmic.2013.09.031","article-title":"Equine infectious anemia and equine infectious anemia virus in 2013: A review","volume":"167","author":"Cook","year":"2013","journal-title":"Vet. Microbiol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"96","DOI":"10.1016\/j.virol.2010.10.032","article-title":"A proviral derivative from a reference attenuated eiav vaccine strain failed to elicit protective immunity","volume":"410","author":"Ma","year":"2011","journal-title":"Virology"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"353","DOI":"10.1007\/s00705-010-0877-8","article-title":"Genomic comparison between attenuated chinese equine infectious anemia virus vaccine strains and their parental virulent strains","volume":"156","author":"Wang","year":"2011","journal-title":"Arch. Virol."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"9","DOI":"10.1186\/s12977-016-0240-6","article-title":"Genetic evolution during the development of an attenuated EIAV vaccine","volume":"13","author":"Wang","year":"2016","journal-title":"Retrovirology"},{"key":"ref_6","doi-asserted-by":"crossref","unstructured":"Bingham, R.J., Dykeman, E.C., and Twarock, R. (2017). RNA virus evolution via a quasispecies-based model reveals a drug target with a high barrier to resistance. Viruses, 9.","DOI":"10.3390\/v9110347"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1990","DOI":"10.1093\/gbe\/evu150","article-title":"Evolution of functional diversification within quasispecies","volume":"6","author":"Colizzi","year":"2014","journal-title":"Genome Biol. Evol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"012714","DOI":"10.1103\/PhysRevE.91.012714","article-title":"Quasispecies theory for evolution of modularity","volume":"91","author":"Park","year":"2015","journal-title":"Phys. Rev. E Stat. Nonlin. Soft Matter Phys."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"159","DOI":"10.1128\/MMBR.05023-11","article-title":"Viral quasispecies evolution","volume":"76","author":"Domingo","year":"2012","journal-title":"Microbiol. Mol. Biol. Rev."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"46","DOI":"10.1016\/j.virol.2015.03.022","article-title":"Viral quasispecies","volume":"479\u2013480","author":"Andino","year":"2015","journal-title":"Virology"},{"key":"ref_11","first-page":"51","article-title":"Viruses as quasispecies: Biological implications","volume":"299","author":"Domingo","year":"2006","journal-title":"Curr. Top. Microbiol."},{"key":"ref_12","doi-asserted-by":"crossref","unstructured":"Craigo, J.K., Ezzelarab, C., Cook, S.J., Chong, L., Horohov, D., Issel, C.J., and Montelaro, R.C. (2013). Envelope determinants of equine lentiviral vaccine protection. PLoS ONE, 8.","DOI":"10.1371\/journal.pone.0066093"},{"key":"ref_13","doi-asserted-by":"crossref","unstructured":"Du, J.S., Wang, X.F., Ma, J., Wang, J.X., Qin, Y.Y., Zhu, C.H., Liu, F., Shao, Y.M., Zhou, J.H., and Qiao, W.T. (2014). Structural and biochemical insights into the v\/i505t mutation found in the eiav gp45 vaccine strain. Retrovirology, 11.","DOI":"10.1186\/1742-4690-11-26"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"64","DOI":"10.1016\/j.virol.2016.01.010","article-title":"Structural and functional characterization of EIAV gp45 fusion peptide proximal region and asparagine-rich layer","volume":"491","author":"Duan","year":"2016","journal-title":"Virology"},{"key":"ref_15","doi-asserted-by":"crossref","unstructured":"Wang, X.F., Liu, Q., Wang, Y.H., Wang, S., Chen, J., Lin, Y.Z., Ma, J., Zhou, J.H., and Wang, X. (2018). Characterization of equine infectious anemia virus long terminal repeat quasispecies in vitro and in vivo. J. Virol., 92.","DOI":"10.1128\/JVI.02150-17"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"292","DOI":"10.1016\/j.antiviral.2011.08.016","article-title":"An attenuated eiav vaccine strain induces significantly different immune responses from its pathogenic parental strain although with similar in vivo replication pattern","volume":"92","author":"Lin","year":"2011","journal-title":"Antiviral Res."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"235","DOI":"10.1016\/j.virusres.2011.04.007","article-title":"C-terminal truncation of the transmembrane protein of an attenuated lentiviral vaccine alters its in vitro but not in vivo replication and weakens its potential pathogenicity","volume":"158","author":"Jiang","year":"2011","journal-title":"Virus Res."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"207","DOI":"10.3928\/01484834-20140325-04","article-title":"Understanding poisson regression","volume":"53","author":"Hayat","year":"2014","journal-title":"J. Nurs. Educ."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"1870","DOI":"10.1093\/molbev\/msw054","article-title":"Mega7: Molecular evolutionary genetics analysis version 7.0 for bigger datasets","volume":"33","author":"Kumar","year":"2016","journal-title":"Mol. Biol. Evol."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"81","DOI":"10.2174\/157016210790416398","article-title":"Eiav envelope diversity: Shaping viral persistence and encumbering vaccine efficacy","volume":"8","author":"Craigo","year":"2010","journal-title":"Curr. HIV Res."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"46","DOI":"10.1016\/j.jviromet.2015.01.006","article-title":"Single genome amplification and standard bulk pcr yield hiv-1 envelope products with similar genotypic and phenotypic characteristics","volume":"214","author":"Etemad","year":"2015","journal-title":"J. Virol. Methods"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"4265","DOI":"10.3390\/v6114265","article-title":"A unique evolution of the s2 gene of equine infectious anemia virus in hosts correlated with particular infection statuses","volume":"6","author":"Wang","year":"2014","journal-title":"Viruses"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"2544","DOI":"10.1128\/JCM.00045-09","article-title":"Quantitative detection of the m204v hepatitis B virus minor variants by amplification refractory mutation system real-time PCR combined with molecular beacon technology","volume":"47","author":"Ntziora","year":"2009","journal-title":"J. Clin. Microbiol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"9604","DOI":"10.3748\/wjg.v22.i43.9604","article-title":"New real-time-pcr method to identify single point mutations in hepatitis C virus","volume":"22","author":"Chen","year":"2016","journal-title":"World J. Gastroenterol."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"3952","DOI":"10.1128\/JVI.02660-07","article-title":"Deciphering human immunodeficiency virus type 1 transmission and early envelope diversification by single-genome amplification and sequencing","volume":"82","author":"Bailes","year":"2008","journal-title":"J. Virol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"316","DOI":"10.1007\/s12250-010-3122-4","article-title":"Indinavir resistance evolution in one human immunodeficiency virus type 1 infected patient revealed by single-genome amplification","volume":"25","author":"Geng","year":"2010","journal-title":"Virol. Sin."},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Guinoiseau, T., Moreau, A., Hohnadel, G., Ngo-Giang-Huong, N., Brulard, C., Vourc\u2019h, P., Goudeau, A., and Gaudy-Graffin, C. (2017). Deep sequencing is an appropriate tool for the selection of unique hepatitis C virus (HCV) variants after single genomic amplification. PLoS ONE, 12.","DOI":"10.1371\/journal.pone.0174852"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"3226","DOI":"10.3390\/v7062768","article-title":"Rna viruses and rnai: Quasispecies implications for viral escape","volume":"7","author":"Presloid","year":"2015","journal-title":"Viruses"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"7","DOI":"10.1016\/j.cimid.2016.05.007","article-title":"Evolutionary characteristics of morbilliviruses during serial passages in vitro: Gradual attenuation of virus virulence","volume":"47","author":"Liu","year":"2016","journal-title":"Comp. Immunol. Microbiol. Infect. Dis."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"344","DOI":"10.1038\/nature04388","article-title":"Quasispecies diversity determines pathogenesis through cooperative interactions in a viral population","volume":"439","author":"Vignuzzi","year":"2006","journal-title":"Nature"},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"1222","DOI":"10.1016\/j.vaccine.2010.12.001","article-title":"Lights and shades on an historical vaccine canine distemper virus, the rockborn strain","volume":"29","author":"Martella","year":"2011","journal-title":"Vaccine"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"814","DOI":"10.1007\/s11262-016-1382-2","article-title":"Amino acid mutations in the env gp90 protein that modify n-linked glycosylation of the chinese eiav vaccine strain enhance resistance to neutralizing antibodies","volume":"52","author":"Han","year":"2016","journal-title":"Virus Genes"},{"key":"ref_33","doi-asserted-by":"crossref","unstructured":"Glebova, O., Knyazev, S., Melnyk, A., Artyomenko, A., Khudyakov, Y., Zelikovsky, A., and Skums, P. (2017). Inference of genetic relatedness between viral quasispecies from sequencing data. BMC Genomics, 18.","DOI":"10.1186\/s12864-017-4274-5"}],"container-title":["Viruses"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1999-4915\/11\/4\/380\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T12:46:51Z","timestamp":1760186811000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1999-4915\/11\/4\/380"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2019,4,24]]},"references-count":33,"journal-issue":{"issue":"4","published-online":{"date-parts":[[2019,4]]}},"alternative-id":["v11040380"],"URL":"https:\/\/doi.org\/10.3390\/v11040380","relation":{},"ISSN":["1999-4915"],"issn-type":[{"value":"1999-4915","type":"electronic"}],"subject":[],"published":{"date-parts":[[2019,4,24]]}}}