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      Construction and Characterization of Highly Infectious Full-Length Molecular Clones of a HIV-1 CRF07_BC Isolate from Xinjiang, China

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          Abstract

          Among the various subtypes of the M group of human immunodeficiency virus type 1 (HIV-1), clade CRF07_BC is the most prevalent in China. To date, no strong replicable CRF07_BC infectious clone has been constructed. Here we report on the construction and characterization of highly replicable infectious molecular clones from the isolate XJDC6291 of this HIV-1 subtype. Four full-length clones pXJDC2-7, pXJDC3-7, pXJDC2-6 and pXJDC3-6 were successfully produced, but only pXJDC2-7 presented detectable infectivity and replication capability. To improve the replication capability of pXJDC2-7, a 4.8 kb region spanning from the pol Integrase to nef gene of the clone was replaced by PCR products of the corresponding fragments from the original isolate XJDC6291, which produced two clones pXJDC13 and pXJDC17 that exhibited strong replication capability. The viral stocks obtained by pXJDC-13 and pXJDC-17 transfection into 293T cells replicated efficiently in human PBMCs, human primary CD4 + T cells and displayed CCR5 tropism. Sequence alignment between pXJDC13, pXJDC17 and pXJDC2-7 suggested that polymorphisms in the V1V2 region may influence infectivity, and reverse genetic experiment showed that V1V2 polymorphisms may influence the infectivity of the clones but did not affect the replication capability at a significant level. pXJDC13 and pXJDC17 displayed strong replication capability and are the first full-length infectious clones of HIV-1 CRF07_BC clade in the world. The availability of CRF07_BC infectious clones provides a useful tool for a wide range of studies, including antiretroviral drug and vaccine research as related to this HIV subtype.

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          High levels of HIV-1 in plasma during all stages of infection determined by competitive PCR.

          Micah B. Hahn, Albert Yang, J Lifson … (1993)
          Quantitative competitive polymerase chain reaction (QC-PCR) methods were used to quantify virion-associated human immunodeficiency virus type-1 (HIV-1) RNA in plasma from 66 patients with Centers for Disease Control stage I to IVC1 infection. HIV-1 RNA, ranging from 100 to nearly 22,000,000 copies per milliliter of plasma (corresponding to 50 to 11,000,000 virions per milliliter), was readily quantified in all subjects, was significantly associated with disease stage and CD4+ T cell counts, and decreased by as much as 235-fold with resolution of primary infection or institution of antiretroviral therapy. Plasma virus levels determined by QC-PCR correlated with, but exceeded by an average of 60,000-fold, virus titers measured by endpoint dilution culture. Quantitation of HIV-1 in plasma by QC-PCR may be useful in assessing the efficacy of antiretroviral agents, especially in early stage disease when conventional viral markers are often negative.
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            A Comprehensive Mapping of HIV-1 Genotypes in Various Risk Groups and Regions across China Based on a Nationwide Molecular Epidemiologic Survey

            Xiang He, Hui-Hui Xing, Yuhua Ruan … (2012)
            Background China is experiencing a dynamic HIV/AIDS epidemic. While serology based surveillance systems have reported the spread of HIV/AIDS, detailed tracking of its transmission in populations and regions is not possible without mapping it at the molecular level. We therefore conducted a nationwide molecular epidemiology survey across the country. Methods HIV-1 genotypes were determined from 1,408 HIV-positive persons newly diagnosed in 2006. The prevalence of each genotype was estimated by weighting the genotype’s prevalence from each province- and risk-specific subpopulation with the number of reported cases in the corresponding subgroups in that year. Results CRF07_BC (35.5%), CRF01_AE (27.6%), CRF08_BC (20.1%), and subtype B' (9.6%) were the four main HIV-1 strains in China. CRF07_BC and CRF08_BC were the primary drivers of infection among injecting drug users in northeastern and southeastern China, respectively, and subtype B' remained dominant among former plasma donors in central China. In contrast, all four strains occurred in significant proportions among heterosexuals nationwide, pointing to an expansion of the HIV-1 epidemic from high-risk populations into the general population. CRF01_AE also replaced subtype B as the principal driver of infection among men-who-have-sex-with-men. Conclusions Our study provides the first comprehensive baseline data on the diversity and characteristics of HIV/AIDS epidemic in China, reflecting unique region- and risk group-specific transmission dynamics. The results provide information critical for designing effective prevention measures against HIV transmission.
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              Structure of the immature retroviral capsid at 8 Å resolution by cryo-electron microscopy.

              Michaela Rumlová, Pavel Ulbrich, Carsten Sachse … (2012)
              The assembly of retroviruses such as HIV-1 is driven by oligomerization of their major structural protein, Gag. Gag is a multidomain polyprotein including three conserved folded domains: MA (matrix), CA (capsid) and NC (nucleocapsid). Assembly of an infectious virion proceeds in two stages. In the first stage, Gag oligomerization into a hexameric protein lattice leads to the formation of an incomplete, roughly spherical protein shell that buds through the plasma membrane of the infected cell to release an enveloped immature virus particle. In the second stage, cleavage of Gag by the viral protease leads to rearrangement of the particle interior, converting the non-infectious immature virus particle into a mature infectious virion. The immature Gag shell acts as the pivotal intermediate in assembly and is a potential target for anti-retroviral drugs both in inhibiting virus assembly and in disrupting virus maturation. However, detailed structural information on the immature Gag shell has not previously been available. For this reason it is unclear what protein conformations and interfaces mediate the interactions between domains and therefore the assembly of retrovirus particles, and what structural transitions are associated with retrovirus maturation. Here we solve the structure of the immature retroviral Gag shell from Mason-Pfizer monkey virus by combining cryo-electron microscopy and tomography. The 8-Å resolution structure permits the derivation of a pseudo-atomic model of CA in the immature retrovirus, which defines the protein interfaces mediating retrovirus assembly. We show that transition of an immature retrovirus into its mature infectious form involves marked rotations and translations of CA domains, that the roles of the amino-terminal and carboxy-terminal domains of CA in assembling the immature and mature hexameric lattices are exchanged, and that the CA interactions that stabilize the immature and mature viruses are almost completely distinct.
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                Author and article information

                Contributors
                Shibo Jiang: Role: Editor
                Journal
                Journal ID (nlm-ta): PLoS One
                Journal ID (iso-abbrev): PLoS ONE
                Journal ID (publisher-id): plos
                Journal ID (pmc): plosone
                Title: PLoS ONE
                Publisher: Public Library of Science (San Francisco, USA )
                ISSN (Electronic): 1932-6203
                Publication date Collection: 2013
                Publication date (Electronic): 18 November 2013
                Volume: 8
                Issue: 11
                Electronic Location Identifier: e79177
                Affiliations
                [1 ]State Key Laboratory for Infectious Disease Prevention and Control, Division of Research of Virology and Immunology, National Center for AIDS/STD Control and Prevention, China CDC, Beijing, China
                [2 ]Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Hangzhou, China
                Shanghai Medical College, Fudan University, China
                Author notes

                Competing Interests: The authors declare that they have no competing interests.

                Conceived and designed the experiments: ZW KH YS. Performed the experiments: ZW JZ LZ. Analyzed the data: ZW KH YS. Contributed reagents/materials/analysis tools: DL LR HL. Wrote the paper: ZW KH YS.

                Article
                Publisher ID: PONE-D-13-22722
                DOI: 10.1371/journal.pone.0079177
                PMC ID: 3850540
                SO-VID: 76ba3234-5cf7-448b-ac2e-fcb3264bff71
                Copyright statement: Copyright @ 2013
                License:

                This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                Date received : 2 June 2013
                Date accepted : 22 September 2013
                Page count
                Pages: 9
                Funding
                This work is partly supported by the National Natural Science Foundation of China (grants 81101281 and 31100126), National Science and Technology Major Project (grant 2012ZX10001-002), and The Project Sponsored by the Yong Scientific Research Foundation of China CDC (grant 2013A102). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Categories
                Subject: Research Article

                ScienceOpen disciplines: Uncategorized
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