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Sample records for hiv-1 pol sequences

  1. Phylogeny and resistance profiles of HIV-1 POL sequences from rectal biopsies and blood

    DEFF Research Database (Denmark)

    Katzenstein, Terese Lea; Petersen, A B; Storgaard, M

    2010-01-01

    The phylogeny and resistance profiles of human immunodeficiency virus type 1 (HIV-1) protease (PR) and reverse transcriptase (RT) sequences were compared among six patients with HIV-1 who had received numerous treatments. RNA and DNA fractions were obtained from concurrent blood and rectal biopsy...

  2. Phylogeny and resistance profiles of HIV-1 POL sequences from rectal biopsies and blood

    DEFF Research Database (Denmark)

    Katzenstein, T L; Petersen, A B; Storgaard, M

    2010-01-01

    The phylogeny and resistance profiles of human immunodeficiency virus type 1 (HIV-1) protease (PR) and reverse transcriptase (RT) sequences were compared among six patients with HIV-1 who had received numerous treatments. RNA and DNA fractions were obtained from concurrent blood and rectal biopsy...... samples. Phylogenetic trees and resistance profiles showed that the rectal mucosa and the peripheral blood mononuclear cells (PBMCs) harbored different HIV-1 strains. The resistance-associated mutations found in each strain corresponded to the treatment history of the patients. The resistance mutations...... acquired during earlier treatment regimens were detected in the sequences obtained from the rectal samples and in the PBMCs in several of the patients. Also, differences in the resistance profiles were observed between anatomical sites and between RNA and DNA fractions. Thus, a single sample probably...

  3. Evaluation of sequence ambiguities of the HIV-1 pol gene as a method to identify recent HIV-1 infection in transmitted drug resistance surveys.

    Science.gov (United States)

    Andersson, Emmi; Shao, Wei; Bontell, Irene; Cham, Fatim; Cuong, Do Duy; Wondwossen, Amogne; Morris, Lynn; Hunt, Gillian; Sönnerborg, Anders; Bertagnolio, Silvia; Maldarelli, Frank; Jordan, Michael R

    2013-08-01

    Identification of recent HIV infection within populations is a public health priority for accurate estimation of HIV incidence rates and transmitted drug resistance at population level. Determining HIV incidence rates by prospective follow-up of HIV-uninfected individuals is challenging and serological assays have important limitations. HIV diversity within an infected host increases with duration of infection. We explore a simple bioinformatics approach to assess viral diversity by determining the percentage of ambiguous base calls in sequences derived from standard genotyping of HIV-1 protease and reverse transcriptase. Sequences from 691 recently infected (≤1 year) and chronically infected (>1 year) individuals from Sweden, Vietnam and Ethiopia were analyzed for ambiguity. A significant difference (p<0.0001) in the proportion of ambiguous bases was observed between sequences from individuals with recent and chronic infection in both HIV-1 subtype B and non-B infection, consistent with previous studies. In our analysis, a cutoff of <0.47% ambiguous base calls identified recent infection with a sensitivity and specificity of 88.8% and 74.6% respectively. 1,728 protease and reverse transcriptase sequences from 36 surveys of transmitted HIV drug resistance performed following World Health Organization guidance were analyzed for ambiguity. The 0.47% ambiguity cutoff was applied and survey sequences were classified as likely derived from recently or chronically infected individuals. 71% of patients were classified as likely to have been infected within one year of genotyping but results varied considerably amongst surveys. This bioinformatics approach may provide supporting population-level information to identify recent infection but its application is limited by infection with more than one viral variant, decreasing viral diversity in advanced disease and technical aspects of population based sequencing. Standardization of sequencing techniques and base calling

  4. Phylogenetic analysis of HIV-1 pol gene: first subgenomic evidence of CRF29-BF among Iranian HIV-1 patients

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    Kazem Baesi

    2014-09-01

    Full Text Available Objective: To identify the dominant subtype among the HIV-1 strains circulation in Iran. Methods: In this cross sectional study 100 HIV positive patients participated. HIV-1 RNA was extracted from plasma. RT nested-PCR was performed and the final products were sequenced and phylogenetically analyzed; reference sequences were downloaded from Los Alamos, aligned with Iranian pol sequences in the study and analyzed by neighbor-joining method. Results: The results of the phylogenetic analysis showed that HIV-1 subtype CRF-35AD was the dominant subtype among HIV-1 infected patients in Iran; this analysis also suggested a new circulating recombinant form that had not previously been identified in Iran: CRF-29BF. Conclusions: The impact of HIV diversity on pathogenesis, transmission and clinical management have been discussed in different studies; therefore, analyses of HIV genetic diversity is required to design effective antiretroviral strategies for different HIV subtypes.

  5. Potent nonnucleoside reverse transcriptase inhibitors target HIV-1 Gag-Pol.

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    Anna Figueiredo

    2006-11-01

    Full Text Available Nonnucleoside reverse transcriptase inhibitors (NNRTIs target HIV-1 reverse transcriptase (RT by binding to a pocket in RT that is close to, but distinct, from the DNA polymerase active site and prevent the synthesis of viral cDNA. NNRTIs, in particular, those that are potent inhibitors of RT polymerase activity, can also act as chemical enhancers of the enzyme's inter-subunit interactions. However, the consequences of this chemical enhancement effect on HIV-1 replication are not understood. Here, we show that the potent NNRTIs efavirenz, TMC120, and TMC125, but not nevirapine or delavirdine, inhibit the late stages of HIV-1 replication. These potent NNRTIs enhanced the intracellular processing of Gag and Gag-Pol polyproteins, and this was associated with a decrease in viral particle production from HIV-1-transfected cells. The increased polyprotein processing is consistent with premature activation of the HIV-1 protease by NNRTI-enhanced Gag-Pol multimerization through the embedded RT sequence. These findings support the view that Gag-Pol multimerization is an important step in viral assembly and demonstrate that regulation of Gag-Pol/Gag-Pol interactions is a novel target for small molecule inhibitors of HIV-1 production. Furthermore, these drugs can serve as useful probes to further understand processes involved in HIV-1 particle assembly and maturation.

  6. HIV-1 pol diversity among female bar and hotel workers in Northern Tanzania.

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    Kiwelu, Ireen E; Novitsky, Vladimir; Kituma, Elimsaada; Margolin, Lauren; Baca, Jeannie; Manongi, Rachel; Sam, Noel; Shao, John; McLane, Mary F; Kapiga, Saidi H; Essex, M

    2014-01-01

    A national ART program was launched in Tanzania in October 2004. Due to the existence of multiple HIV-1 subtypes and recombinant viruses co-circulating in Tanzania, it is important to monitor rates of drug resistance. The present study determined the prevalence of HIV-1 drug resistance mutations among ART-naive female bar and hotel workers, a high-risk population for HIV-1 infection in Moshi, Tanzania. A partial HIV-1 pol gene was analyzed by single-genome amplification and sequencing in 45 subjects (622 pol sequences total; median number of sequences per subject, 13; IQR 5-20) in samples collected in 2005. The prevalence of HIV-1 subtypes A1, C, and D, and inter-subtype recombinant viruses, was 36%, 29%, 9% and 27%, respectively. Thirteen different recombination patterns included D/A1/D, C/A1, A1/C/A1, A1/U/A1, C/U/A1, C/A1, U/D/U, D/A1/D, A1/C, A1/C, A2/C/A2, CRF10_CD/C/CRF10_CD and CRF35_AD/A1/CRF35_AD. CRF35_AD was identified in Tanzania for the first time. All recombinant viruses in this study were unique, suggesting ongoing recombination processes among circulating HIV-1 variants. The prevalence of multiple infections in this population was 16% (n = 7). Primary HIV-1 drug resistance mutations to RT inhibitors were identified in three (7%) subjects (K65R plus Y181C; N60D; and V106M). In some subjects, polymorphisms were observed at the RT positions 41, 69, 75, 98, 101, 179, 190, and 215. Secondary mutations associated with NNRTIs were observed at the RT positions 90 (7%) and 138 (6%). In the protease gene, three subjects (7%) had M46I/L mutations. All subjects in this study had HIV-1 subtype-specific natural polymorphisms at positions 36, 69, 89 and 93 that are associated with drug resistance in HIV-1 subtype B. These results suggested that HIV-1 drug resistance mutations and natural polymorphisms existed in this population before the initiation of the national ART program. With increasing use of ARV, these results highlight the importance of drug

  7. Association of mitochondrial lysyl-tRNA synthetase with HIV-1 GagPol involves catalytic domain of the synthetase and transframe and integrase domains of Pol

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    Shalak V. F.

    2011-10-01

    Full Text Available Aim. Analyze the interaction between Lysyl-tRNA synthetase (LysRS and HIV-1 GagPol to know whether a particular N-terminal sequence of mitochondrial LysRS triggers a specific recognition with GagPol. Methods. Yeast two-hybrid analysis, immunoprecipitation. Results. We have shown that LysRS associates with the Pol domain of GagPol. Conclusions. A model of the assembly of the LysRS:tRNA3Lys:GagPol packaging complex is proposed.

  8. Introducing a frameshift mutation to the POL sequence of HIV-1 provirus and evaluation of the immunogenic characteristics of the mutated virions (RINNL4-3).

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    Zabihollahi, Rezvan; Sadat, Seyed Mehdi; Vahabpour, Rouhollah; Salehi, Mansoor; Azadmanesh, Kayhan; Siadat, Seyed Davar; Saraji, Ali Reza Azizi; Pouriavali, Mohamamd Hassan; Momen, Seyed Bahman; Aghasadeghi, Mohamad Reza

    2012-01-01

    Inactivation of the reverse transcriptase (RT) and integrase (IN) enzymes can abolish the replication of the human immunodeficiency virus (HIV) and, thus, its infectivity. Here, inactivated HIV particles convenient for designing virus-like particle (VLP) based vaccines have been produced. Inactivated HIV-provirus was created by introducing a frame shift mutation. HIV provirus DNA was cut in the pol region by Age I restriction enzyme, followed by filling of sticky ends using the Klenow fragment before ligation. The resulting plasmid was named as pRINNL4-3. HEK-293T cells were used as producer, after being transfected with the modified plasmid. Viral particle production and biological activity were assayed by virus capsid protein (p24) quantification and syncytium formation in MT2 cells, respectively. The immunogenicity of the RINNL4-3 virions was investigated in a mouse model. The mutation was expected to inactivate the virus RT and IN enzymes. The results showed that the VLPs were assembled, as measured by the p24 load of the culture supernatant, and contained functional envelope proteins (Env) as monitored by the syncytium formation. However, these VLPs had no ability to infect target MT2 cells, as well as their VSVG (vesicular stomatitis virus-glycoprotein) pseudotyped counterparts infected HEK-293T cells. A high level of antibody response was observed in immunized mice. Since RINNL4-3 virions are replication incompetent, they are convenient for production and use in biomedical studies. Also, RINNL4-3 is a candidate for a vaccine development due to it contains envelope and structural virus proteins which are crucial for triggering neutralizing antibodies and the cellular immune response.

  9. Deciphering the role of the Gag-Pol ribosomal frameshift signal in HIV-1 RNA genome packaging.

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    Nikolaitchik, Olga A; Hu, Wei-Shau

    2014-04-01

    A key step of retroviral replication is packaging of the viral RNA genome during virus assembly. Specific packaging is mediated by interactions between the viral protein Gag and elements in the viral RNA genome. In HIV-1, similar to most retroviruses, the packaging signal is located within the 5' untranslated region and extends into the gag-coding region. A recent study reported that a region including the Gag-Pol ribosomal frameshift signal plays an important role in HIV-1 RNA packaging; deletions or mutations that affect the RNA structure of this signal lead to drastic decreases (10- to 50-fold) in viral RNA packaging and virus titer. We examined here the role of the ribosomal frameshift signal in HIV-1 RNA packaging by studying the RNA packaging and virus titer in the context of proviruses. Three mutants with altered ribosomal frameshift signal, either through direct deletion of the signal, mutation of the 6U slippery sequence, or alterations of the secondary structure were examined. We found that RNAs from all three mutants were packaged efficiently, and they generate titers similar to that of a virus containing the wild-type ribosomal frameshift signal. We conclude that although the ribosomal frameshift signal plays an important role in regulating the replication cycle, this RNA element is not directly involved in regulating RNA encapsidation. To generate infectious viruses, HIV-1 must package viral RNA genome during virus assembly. The specific HIV-1 genome packaging is mediated by interactions between the structural protein Gag and elements near the 5' end of the viral RNA known as packaging signal. In this study, we examined whether the Gag-Pol ribosomal frameshift signal is important for HIV-1 RNA packaging as recently reported. Our results demonstrated that when Gag/Gag-Pol is supplied in trans, none of the tested ribosomal frameshift signal mutants has defects in RNA packaging or virus titer. These studies provide important information on how HIV-1

  10. Intragenic HIV-1 env sequences that enhance gag expression

    International Nuclear Information System (INIS)

    Suptawiwat, Ornpreya; Sutthent, Ruengpung; Lee, T.-H.; Auewarakul, Prasert

    2003-01-01

    Expression of HIV-1 genes is regulated at multiple levels including the complex RNA splicing and transport mechanisms. Multiple cis-acting elements involved in these regulations have been previously identified in various regions of HIV-1 genome. Here we show that another cis-acting element was present in HIV-1 env region. This element enhanced the expression of Gag when inserted together with Rev response element (RRE) into a truncated HIV-1 genome in the presence of Rev. The enhancing activity was mapped to a 263-bp fragment in the gp41 region downstream to RRE. RNA analysis showed that it might function by promoting RNA stability and Rev-dependent RNA export. The enhancement was specific to Rev-dependent expression, since it did not enhance Gag expression driven by Sam68, a cellular protein that has been shown to be able to substitute for Rev in RNA export function

  11. Improved therapy-success prediction with GSS estimated from clinical HIV-1 sequences.

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    Pironti, Alejandro; Pfeifer, Nico; Kaiser, Rolf; Walter, Hauke; Lengauer, Thomas

    2014-01-01

    Rules-based HIV-1 drug-resistance interpretation (DRI) systems disregard many amino-acid positions of the drug's target protein. The aims of this study are (1) the development of a drug-resistance interpretation system that is based on HIV-1 sequences from clinical practice rather than hard-to-get phenotypes, and (2) the assessment of the benefit of taking all available amino-acid positions into account for DRI. A dataset containing 34,934 therapy-naïve and 30,520 drug-exposed HIV-1 pol sequences with treatment history was extracted from the EuResist database and the Los Alamos National Laboratory database. 2,550 therapy-change-episode baseline sequences (TCEB) were assigned to test set A. Test set B contains 1,084 TCEB from the HIVdb TCE repository. Sequences from patients absent in the test sets were used to train three linear support vector machines to produce scores that predict drug exposure pertaining to each of 20 antiretrovirals: the first one uses the full amino-acid sequences (DEfull), the second one only considers IAS drug-resistance positions (DEonlyIAS), and the third one disregards IAS drug-resistance positions (DEnoIAS). For performance comparison, test sets A and B were evaluated with DEfull, DEnoIAS, DEonlyIAS, geno2pheno[resistance], HIVdb, ANRS, HIV-GRADE, and REGA. Clinically-validated cut-offs were used to convert the continuous output of the first four methods into susceptible-intermediate-resistant (SIR) predictions. With each method, a genetic susceptibility score (GSS) was calculated for each therapy episode in each test set by converting the SIR prediction for its compounds to integer: S=2, I=1, and R=0. The GSS were used to predict therapy success as defined by the EuResist standard datum definition. Statistical significance was assessed using a Wilcoxon signed-rank test. A comparison of the therapy-success prediction performances among the different interpretation systems for test set A can be found in Table 1, while those for test set

  12. Use of four next-generation sequencing platforms to determine HIV-1 coreceptor tropism.

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    Archer, John; Weber, Jan; Henry, Kenneth; Winner, Dane; Gibson, Richard; Lee, Lawrence; Paxinos, Ellen; Arts, Eric J; Robertson, David L; Mimms, Larry; Quiñones-Mateu, Miguel E

    2012-01-01

    HIV-1 coreceptor tropism assays are required to rule out the presence of CXCR4-tropic (non-R5) viruses prior treatment with CCR5 antagonists. Phenotypic (e.g., Trofile™, Monogram Biosciences) and genotypic (e.g., population sequencing linked to bioinformatic algorithms) assays are the most widely used. Although several next-generation sequencing (NGS) platforms are available, to date all published deep sequencing HIV-1 tropism studies have used the 454™ Life Sciences/Roche platform. In this study, HIV-1 co-receptor usage was predicted for twelve patients scheduled to start a maraviroc-based antiretroviral regimen. The V3 region of the HIV-1 env gene was sequenced using four NGS platforms: 454™, PacBio® RS (Pacific Biosciences), Illumina®, and Ion Torrent™ (Life Technologies). Cross-platform variation was evaluated, including number of reads, read length and error rates. HIV-1 tropism was inferred using Geno2Pheno, Web PSSM, and the 11/24/25 rule and compared with Trofile™ and virologic response to antiretroviral therapy. Error rates related to insertions/deletions (indels) and nucleotide substitutions introduced by the four NGS platforms were low compared to the actual HIV-1 sequence variation. Each platform detected all major virus variants within the HIV-1 population with similar frequencies. Identification of non-R5 viruses was comparable among the four platforms, with minor differences attributable to the algorithms used to infer HIV-1 tropism. All NGS platforms showed similar concordance with virologic response to the maraviroc-based regimen (75% to 80% range depending on the algorithm used), compared to Trofile (80%) and population sequencing (70%). In conclusion, all four NGS platforms were able to detect minority non-R5 variants at comparable levels suggesting that any NGS-based method can be used to predict HIV-1 coreceptor usage.

  13. Use of four next-generation sequencing platforms to determine HIV-1 coreceptor tropism.

    Directory of Open Access Journals (Sweden)

    John Archer

    Full Text Available HIV-1 coreceptor tropism assays are required to rule out the presence of CXCR4-tropic (non-R5 viruses prior treatment with CCR5 antagonists. Phenotypic (e.g., Trofile™, Monogram Biosciences and genotypic (e.g., population sequencing linked to bioinformatic algorithms assays are the most widely used. Although several next-generation sequencing (NGS platforms are available, to date all published deep sequencing HIV-1 tropism studies have used the 454™ Life Sciences/Roche platform. In this study, HIV-1 co-receptor usage was predicted for twelve patients scheduled to start a maraviroc-based antiretroviral regimen. The V3 region of the HIV-1 env gene was sequenced using four NGS platforms: 454™, PacBio® RS (Pacific Biosciences, Illumina®, and Ion Torrent™ (Life Technologies. Cross-platform variation was evaluated, including number of reads, read length and error rates. HIV-1 tropism was inferred using Geno2Pheno, Web PSSM, and the 11/24/25 rule and compared with Trofile™ and virologic response to antiretroviral therapy. Error rates related to insertions/deletions (indels and nucleotide substitutions introduced by the four NGS platforms were low compared to the actual HIV-1 sequence variation. Each platform detected all major virus variants within the HIV-1 population with similar frequencies. Identification of non-R5 viruses was comparable among the four platforms, with minor differences attributable to the algorithms used to infer HIV-1 tropism. All NGS platforms showed similar concordance with virologic response to the maraviroc-based regimen (75% to 80% range depending on the algorithm used, compared to Trofile (80% and population sequencing (70%. In conclusion, all four NGS platforms were able to detect minority non-R5 variants at comparable levels suggesting that any NGS-based method can be used to predict HIV-1 coreceptor usage.

  14. nef gene sequence variation among HIV-1-infected African children

    Czech Academy of Sciences Publication Activity Database

    Chakraborty, R.; Reiniš, Milan; Rostron, T.; Philpott, S.; Dong, T.; D'Agostino, A.; Musoke, R.; de Silva, E.; Stumpf, M.; Weiser, B.; Burger, H.; Rowland-Jones, S.L.

    2006-01-01

    Roč. 7, č. 2 (2006), s. 75-84 ISSN 1464-2662 Grant - others:Fogarty International Center, NIH(US) 3D43TW00915; NIH(US) RO1 AI 42555 Institutional research plan: CEZ:AV0Z50520514 Keywords : HIV-1 nef gene * non-clade B * Kenya Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.674, year: 2006

  15. HIV-1 envelope sequence-based diversity measures for identifying recent infections.

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    Alexis Kafando

    Full Text Available Identifying recent HIV-1 infections is crucial for monitoring HIV-1 incidence and optimizing public health prevention efforts. To identify recent HIV-1 infections, we evaluated and compared the performance of 4 sequence-based diversity measures including percent diversity, percent complexity, Shannon entropy and number of haplotypes targeting 13 genetic segments within the env gene of HIV-1. A total of 597 diagnostic samples obtained in 2013 and 2015 from recently and chronically HIV-1 infected individuals were selected. From the selected samples, 249 (134 from recent versus 115 from chronic infections env coding regions, including V1-C5 of gp120 and the gp41 ectodomain of HIV-1, were successfully amplified and sequenced by next generation sequencing (NGS using the Illumina MiSeq platform. The ability of the four sequence-based diversity measures to correctly identify recent HIV infections was evaluated using the frequency distribution curves, median and interquartile range and area under the curve (AUC of the receiver operating characteristic (ROC. Comparing the median and interquartile range and evaluating the frequency distribution curves associated with the 4 sequence-based diversity measures, we observed that the percent diversity, number of haplotypes and Shannon entropy demonstrated significant potential to discriminate recent from chronic infections (p<0.0001. Using the AUC of ROC analysis, only the Shannon entropy measure within three HIV-1 env segments could accurately identify recent infections at a satisfactory level. The env segments were gp120 C2_1 (AUC = 0.806, gp120 C2_3 (AUC = 0.805 and gp120 V3 (AUC = 0.812. Our results clearly indicate that the Shannon entropy measure represents a useful tool for predicting HIV-1 infection recency.

  16. Sieve analysis of breakthrough HIV-1 sequences in HVTN 505 identifies vaccine pressure targeting the CD4 binding site of Env-gp120.

    Science.gov (United States)

    deCamp, Allan C; Rolland, Morgane; Edlefsen, Paul T; Sanders-Buell, Eric; Hall, Breana; Magaret, Craig A; Fiore-Gartland, Andrew J; Juraska, Michal; Carpp, Lindsay N; Karuna, Shelly T; Bose, Meera; LePore, Steven; Miller, Shana; O'Sullivan, Annemarie; Poltavee, Kultida; Bai, Hongjun; Dommaraju, Kalpana; Zhao, Hong; Wong, Kim; Chen, Lennie; Ahmed, Hasan; Goodman, Derrick; Tay, Matthew Z; Gottardo, Raphael; Koup, Richard A; Bailer, Robert; Mascola, John R; Graham, Barney S; Roederer, Mario; O'Connell, Robert J; Michael, Nelson L; Robb, Merlin L; Adams, Elizabeth; D'Souza, Patricia; Kublin, James; Corey, Lawrence; Geraghty, Daniel E; Frahm, Nicole; Tomaras, Georgia D; McElrath, M Juliana; Frenkel, Lisa; Styrchak, Sheila; Tovanabutra, Sodsai; Sobieszczyk, Magdalena E; Hammer, Scott M; Kim, Jerome H; Mullins, James I; Gilbert, Peter B

    2017-01-01

    Although the HVTN 505 DNA/recombinant adenovirus type 5 vector HIV-1 vaccine trial showed no overall efficacy, analysis of breakthrough HIV-1 sequences in participants can help determine whether vaccine-induced immune responses impacted viruses that caused infection. We analyzed 480 HIV-1 genomes sampled from 27 vaccine and 20 placebo recipients and found that intra-host HIV-1 diversity was significantly lower in vaccine recipients (P ≤ 0.04, Q-values ≤ 0.09) in Gag, Pol, Vif and envelope glycoprotein gp120 (Env-gp120). Furthermore, Env-gp120 sequences from vaccine recipients were significantly more distant from the subtype B vaccine insert than sequences from placebo recipients (P = 0.01, Q-value = 0.12). These vaccine effects were associated with signatures mapping to CD4 binding site and CD4-induced monoclonal antibody footprints. These results suggest either (i) no vaccine efficacy to block acquisition of any viral genotype but vaccine-accelerated Env evolution post-acquisition; or (ii) vaccine efficacy against HIV-1s with Env sequences closest to the vaccine insert combined with increased acquisition due to other factors, potentially including the vaccine vector.

  17. Polymorphisms in the HIV-1 gp41 env gene, natural resistance to enfuvirtide (T-20) and pol resistance among pregnant Brazilian women.

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    Reis, Mônica Nogueira da Guarda; de Alcântara, Keila Correa; Cardoso, Ludimila Paula Vaz; Stefani, Mariane Martins Araújo

    2014-01-01

    The selective pressure of antiretroviral drugs (ARVs) targeting HIV-1 pol can promote drug resistance mutations in other genomic regions, such as env. Drug resistance among women should be monitored to avoid horizontal and mother-to-child transmission. To describe natural resistance to T-20 (enfuvirtide), gp41 env polymorphisms, mutations in pol and HIV-1 subtypes, 124 pregnant women were recruited. For 98 patients, the gp41 env, protease (PR) and reverse transcriptase (RT) fragments were sequenced. The patients were ARV naïve (n = 30), taking mother-to-child transmission prophylaxis (n = 50), or being treated with highly active ARV therapy/HAART (n = 18). The Stanford and IAS/USA databases and other sources were used to analyze PR/RT, gp41 env resistance mutations. The HIV-1 genetic diversity was analyzed by REGA/phylogenetic analyses. The patients' median age was 25 years (range, 16-42), 18.4% had AIDS. The frequency of natural resistance to T-20 (N42D, L44M, and R46M-low-impact mutations) was 6.1% (6/98); 20.4% (20/98) had compensatory mutations in HR2. The prevalence of transmitted drug resistance in the pol was 13.3% (4/30), and the prevalence of secondary drug resistance was 33.3% (6/18). Two patients were infected with multidrug resistant/MDR viruses. The analysis of HIV-1 subtypes (PR/RT/gp41) revealed that 61.2% (60/98) were subtype B, 12.2% (12/98) were subtype C, 4.1% (4/98) were subtype F1, and 22.4% (22/98) were possible recombinants (BF1 = 20.4%; BC = 2%). Natural resistance to T-20 was not associated with pol resistance or previous ARV use. The high rate of secondary resistance, including MDR, indicates that the number of women that may need T-20 salvage therapy may be higher than anticipated. © 2013 Wiley Periodicals, Inc.

  18. Detection of viral sequence fragments of HIV-1 subfamilies yet unknown

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    Stanke Mario

    2011-04-01

    Full Text Available Abstract Background Methods of determining whether or not any particular HIV-1 sequence stems - completely or in part - from some unknown HIV-1 subtype are important for the design of vaccines and molecular detection systems, as well as for epidemiological monitoring. Nevertheless, a single algorithm only, the Branching Index (BI, has been developed for this task so far. Moving along the genome of a query sequence in a sliding window, the BI computes a ratio quantifying how closely the query sequence clusters with a subtype clade. In its current version, however, the BI does not provide predicted boundaries of unknown fragments. Results We have developed Unknown Subtype Finder (USF, an algorithm based on a probabilistic model, which automatically determines which parts of an input sequence originate from a subtype yet unknown. The underlying model is based on a simple profile hidden Markov model (pHMM for each known subtype and an additional pHMM for an unknown subtype. The emission probabilities of the latter are estimated using the emission frequencies of the known subtypes by means of a (position-wise probabilistic model for the emergence of new subtypes. We have applied USF to SIV and HIV-1 sequences formerly classified as having emerged from an unknown subtype. Moreover, we have evaluated its performance on artificial HIV-1 recombinants and non-recombinant HIV-1 sequences. The results have been compared with the corresponding results of the BI. Conclusions Our results demonstrate that USF is suitable for detecting segments in HIV-1 sequences stemming from yet unknown subtypes. Comparing USF with the BI shows that our algorithm performs as good as the BI or better.

  19. Molecular epidemiological analysis of paired pol/env sequences from Portuguese HIV type 1 patients.

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    Abecasis, Ana B; Martins, Andreia; Costa, Inês; Carvalho, Ana P; Diogo, Isabel; Gomes, Perpétua; Camacho, Ricardo J

    2011-07-01

    The advent of new therapeutic approaches targeting env and the search for efficient anti-HIV-1 vaccines make it necessary to identify the number of recombinant forms using genomic regions that were previously not frequently sequenced. In this study, we have subtyped paired pol and env sequences from HIV-1 strains infecting 152 patients being clinically followed in Portugal. The percentage of strains in which we found discordant subtypes in pol and env was 25.7%. When the subtype in pol and env was concordant (65.1%), the most prevalent subtypes were subtype B (40.8%), followed by subtype C (17.8%) and subtype G (5.3%). The most prevalent recombinant form was CRF14_BGpol/Genv (7.2%).

  20. Analysis of HIV-1 intersubtype recombination breakpoints suggests region with high pairing probability may be a more fundamental factor than sequence similarity affecting HIV-1 recombination.

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    Jia, Lei; Li, Lin; Gui, Tao; Liu, Siyang; Li, Hanping; Han, Jingwan; Guo, Wei; Liu, Yongjian; Li, Jingyun

    2016-09-21

    With increasing data on HIV-1, a more relevant molecular model describing mechanism details of HIV-1 genetic recombination usually requires upgrades. Currently an incomplete structural understanding of the copy choice mechanism along with several other issues in the field that lack elucidation led us to perform an analysis of the correlation between breakpoint distributions and (1) the probability of base pairing, and (2) intersubtype genetic similarity to further explore structural mechanisms. Near full length sequences of URFs from Asia, Europe, and Africa (one sequence/patient), and representative sequences of worldwide CRFs were retrieved from the Los Alamos HIV database. Their recombination patterns were analyzed by jpHMM in detail. Then the relationships between breakpoint distributions and (1) the probability of base pairing, and (2) intersubtype genetic similarities were investigated. Pearson correlation test showed that all URF groups and the CRF group exhibit the same breakpoint distribution pattern. Additionally, the Wilcoxon two-sample test indicated a significant and inexplicable limitation of recombination in regions with high pairing probability. These regions have been found to be strongly conserved across distinct biological states (i.e., strong intersubtype similarity), and genetic similarity has been determined to be a very important factor promoting recombination. Thus, the results revealed an unexpected disagreement between intersubtype similarity and breakpoint distribution, which were further confirmed by genetic similarity analysis. Our analysis reveals a critical conflict between results from natural HIV-1 isolates and those from HIV-1-based assay vectors in which genetic similarity has been shown to be a very critical factor promoting recombination. These results indicate the region with high-pairing probabilities may be a more fundamental factor affecting HIV-1 recombination than sequence similarity in natural HIV-1 infections. Our

  1. Workup of Human Blood Samples for Deep Sequencing of HIV-1 Genomes

    NARCIS (Netherlands)

    Cornelissen, Marion; Gall, Astrid; van der Kuyl, Antoinette; Wymant, Chris; Blanquart, François; Fraser, Christophe; Berkhout, Ben

    2018-01-01

    We describe a detailed protocol for the manual workup of blood (plasma/serum) samples from individuals infected with the human immunodeficiency virus type 1 (HIV-1) for deep sequence analysis of the viral genome. The study optimizing the assay was performed in the context of the BEEHIVE (Bridging

  2. Nucleic acid amplification of HIV-1 integrase sequence subtypes CRF01_AE and B for development of HIV anti-integrase drug resistance genotyping assay

    Science.gov (United States)

    Adlar, F. R.; Bela, B.

    2017-08-01

    To anticipate the potential use of anti-integrase drugs in Indonesia for treatment of HIV-1 infection, the development of a drug resistance genotyping assay for anti-integrase is crucial in identifying the genetic drug resistance profile of Indonesian HIV-1 strains. This experiment aimed to amplify a target region in the integrase gene of Indonesian HIV-1 subtypes CRF01_AE and B that contain genetic mutations known to confer resistance to anti-integrase drug. Eleven archived plasma samples from individuals living with HIV-1 were obtained from the Virology and Cancer Pathobiology Research Center for Health Service (VCPRC FKUI-RSCM) laboratory. One of the plasma samples contained HIV-1 subtype B, and the remaining plasma samples contained subtype CRF01_AE. The target regions for all samples were amplified through RT-PCR, with an annealing temperature of 55 °C, using the primer pair AE_POL 4086F and AE_POL 5232R that were designed by VCPRC FKUI-RSCM. The results of this experiment show that 18.2% (2/11) of the samples were successfully amplified using the one-step RT-PCR. While the primer pair was effective in amplifying the target region in the integrase gene sequence for subtype B (100%; 1/1), it had a low efficacy (10%, 1/10) for subtype CRF01_AE. In conclusion, the primer pair can be used to amplify the target region in Indonesian HIV-1 strain subtypes CRF01_AE and B. However, optimization of the PCR condition and an increased number of samples would help to determine an accurate representation of the efficacy of the primer pair.

  3. Use of Four Next-Generation Sequencing Platforms to Determine HIV-1 Coreceptor Tropism

    Czech Academy of Sciences Publication Activity Database

    Archer, J.; Weber, Jan; Henry, K.; Winner, D.; Gibson, R.; Lee, L.; Paxinos, E.; Arts, E. J.; Robertson, D. L.; Mimms, L.; Quinones-Mateu, M. E.

    2012-01-01

    Roč. 7, č. 11 (2012), e49602/1-e49602/17 E-ISSN 1932-6203 R&D Projects: GA MŠk(CZ) LK11207 Institutional research plan: CEZ:AV0Z40550506 Keywords : HIV-1 tropism * V3 region * deep sequencing Subject RIV: EE - Microbiology, Virology Impact factor: 3.730, year: 2012 http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0049602

  4. Eliminating HIV-1 Packaging Sequences from Lentiviral Vector Proviruses Enhances Safety and Expedites Gene Transfer for Gene Therapy.

    Science.gov (United States)

    Vink, Conrad A; Counsell, John R; Perocheau, Dany P; Karda, Rajvinder; Buckley, Suzanne M K; Brugman, Martijn H; Galla, Melanie; Schambach, Axel; McKay, Tristan R; Waddington, Simon N; Howe, Steven J

    2017-08-02

    Lentiviral vector genomic RNA requires sequences that partially overlap wild-type HIV-1 gag and env genes for packaging into vector particles. These HIV-1 packaging sequences constitute 19.6% of the wild-type HIV-1 genome and contain functional cis elements that potentially compromise clinical safety. Here, we describe the development of a novel lentiviral vector (LTR1) with a unique genomic structure designed to prevent transfer of HIV-1 packaging sequences to patient cells, thus reducing the total HIV-1 content to just 4.8% of the wild-type genome. This has been achieved by reconfiguring the vector to mediate reverse-transcription with a single strand transfer, instead of the usual two, and in which HIV-1 packaging sequences are not copied. We show that LTR1 vectors offer improved safety in their resistance to remobilization in HIV-1 particles and reduced frequency of splicing into human genes. Following intravenous luciferase vector administration to neonatal mice, LTR1 sustained a higher level of liver transgene expression than an equivalent dose of a standard lentivirus. LTR1 vectors produce reverse-transcription products earlier and start to express transgenes significantly quicker than standard lentiviruses after transduction. Finally, we show that LTR1 is an effective lentiviral gene therapy vector as demonstrated by correction of a mouse hemophilia B model. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  5. High-resolution deep sequencing reveals biodiversity, population structure, and persistence of HIV-1 quasispecies within host ecosystems

    Directory of Open Access Journals (Sweden)

    Yin Li

    2012-12-01

    Full Text Available Abstract Background Deep sequencing provides the basis for analysis of biodiversity of taxonomically similar organisms in an environment. While extensively applied to microbiome studies, population genetics studies of viruses are limited. To define the scope of HIV-1 population biodiversity within infected individuals, a suite of phylogenetic and population genetic algorithms was applied to HIV-1 envelope hypervariable domain 3 (Env V3 within peripheral blood mononuclear cells from a group of perinatally HIV-1 subtype B infected, therapy-naïve children. Results Biodiversity of HIV-1 Env V3 quasispecies ranged from about 70 to 270 unique sequence clusters across individuals. Viral population structure was organized into a limited number of clusters that included the dominant variants combined with multiple clusters of low frequency variants. Next generation viral quasispecies evolved from low frequency variants at earlier time points through multiple non-synonymous changes in lineages within the evolutionary landscape. Minor V3 variants detected as long as four years after infection co-localized in phylogenetic reconstructions with early transmitting viruses or with subsequent plasma virus circulating two years later. Conclusions Deep sequencing defines HIV-1 population complexity and structure, reveals the ebb and flow of dominant and rare viral variants in the host ecosystem, and identifies an evolutionary record of low-frequency cell-associated viral V3 variants that persist for years. Bioinformatics pipeline developed for HIV-1 can be applied for biodiversity studies of virome populations in human, animal, or plant ecosystems.

  6. Cellular specificity of HIV-1 replication can be controlled by LTR sequences

    International Nuclear Information System (INIS)

    Reed-Inderbitzin, Edward; Maury, Wendy

    2003-01-01

    Two well-established determinants of retroviral tropism are envelope sequences that regulate entry and LTR sequences that can regulate viral expression in a cell-specific manner. Studies with human immunodeficiency virus-1 (HIV-1) have demonstrated that tropism of this virus maps primarily to variable envelope sequences. Studies have demonstrated that T cell and macrophage-specific transcription factor binding motifs exist in the upstream region of the LTR U3; however, the ability of the core enhancer/promoter proximal elements (two NF-κB and three Sp1 sites) to function well in macrophages and T cells have led many to conclude that HIV LTR sequences are not primary determinants of HIV tropism. To determine if cellular specificity could be imparted to HIV by the core enhancer elements, the enhancer/promoter proximal region of the HIV LTR was substituted with motifs that control gene expression in a myeloid-specific manner. The enhancer region from equine infectious anemia virus (EIAV) when substituted for the HIV enhancer/promoter proximal region was found to drive expression in a macrophage-specific manner and was responsive to HIV Tat. The addition of a 5' methylation-dependent binding site (MDBP) and a promoter proximal Sp1 motif increased expression without altering cellular specificity. Spacing between the promoter proximal region and the TATA box was also found to influence LTR activity. Infectivity studies using chimeric LTRs within the context of a dual-tropic infectious molecular clone established that these LTRs directed HIV replication and production of infectious virions in macrophages but not primary T cells or T cell lines. This investigation demonstrates that cellular specificity can be imparted onto HIV-1 replication at the level of viral transcription and not entry

  7. HIV-1 transmission patterns in antiretroviral therapy-naive, HIV-infected North Americans based on phylogenetic analysis by population level and ultra-deep DNA sequencing.

    Directory of Open Access Journals (Sweden)

    Lisa L Ross

    Full Text Available Factors that contribute to the transmission of human immunodeficiency virus type 1 (HIV-1, especially drug-resistant HIV-1 variants remain a significant public health concern. In-depth phylogenetic analyses of viral sequences obtained in the screening phase from antiretroviral-naïve HIV-infected patients seeking enrollment in EPZ108859, a large open-label study in the USA, Canada and Puerto Rico (ClinicalTrials.gov NCT00440947 were examined for insights into the roles of drug resistance and epidemiological factors that could impact disease dissemination. Viral transmission clusters (VTCs were initially predicted from a phylogenetic analysis of population level HIV-1 pol sequences obtained from 690 antiretroviral-naïve subjects in 2007. Subsequently, the predicted VTCs were tested for robustness by ultra deep sequencing (UDS using pyrosequencing technology and further phylogenetic analyses. The demographic characteristics of clustered and non-clustered subjects were then compared. From 690 subjects, 69 were assigned to 1 of 30 VTCs, each containing 2 to 5 subjects. Race composition of VTCs were significantly more likely to be white (72% vs. 60%; p = 0.04. VTCs had fewer reverse transcriptase and major PI resistance mutations (9% vs. 24%; p = 0.002 than non-clustered sequences. Both men-who-have-sex-with-men (MSM (68% vs. 48%; p = 0.001 and Canadians (29% vs. 14%; p = 0.03 were significantly more frequent in VTCs than non-clustered sequences. Of the 515 subjects who initiated antiretroviral therapy, 33 experienced confirmed virologic failure through 144 weeks while only 3/33 were from VTCs. Fewer VTCs subjects (as compared to those with non-clustering virus had HIV-1 with resistance-associated mutations or experienced virologic failure during the course of the study. Our analysis shows specific geographical and drug resistance trends that correlate well with transmission clusters defined by HIV sequences of similarity

  8. Timing of the HIV-1 subtype C epidemic in Ethiopia based on early virus strains and subsequent virus diversification

    NARCIS (Netherlands)

    Abebe, A.; Lukashov, V. V.; Pollakis, G.; Kliphuis, A.; Fontanet, A. L.; Goudsmit, J.; Rinke de Wit, T. F.

    2001-01-01

    OBJECTIVE: To trace the introduction of HIV-1 subtype C into Ethiopia based on virus diversification during the epidemic. DESIGN: A set of 474 serum samples obtained in Ethiopia in 1982-1985 was tested for HIV-1. HIV-1 env gp120 V3 and gag or pol regions were sequenced and analysed together with

  9. Generation and Characterization of HIV-1 Transmitted and Founder Virus Consensus Sequence from Intravenous Drug Users in Xinjiang, China.

    Science.gov (United States)

    Li, Fan; Ma, Liying; Feng, Yi; Hu, Jing; Ni, Na; Ruan, Yuhua; Shao, Yiming

    2017-06-01

    HIV-1 transmission in intravenous drug users (IDUs) has been characterized by high genetic multiplicity and suggests a greater challenge for HIV-1 infection blocking. We investigated a total of 749 sequences of full-length gp160 gene obtained by single genome sequencing (SGS) from 22 HIV-1 early infected IDUs in Xinjiang province, northwest China, and generated a transmitted and founder virus (T/F virus) consensus sequence (IDU.CON). The T/F virus was classified as subtype CRF07_BC and predicted to be CCR5-tropic virus. The variable region (V1, V2, and V4 loop) of IDU.CON showed length variation compared with the heterosexual T/F virus consensus sequence (HSX.CON) and homosexual T/F virus consensus sequence (MSM.CON). A total of 26 N-linked glycosylation sites were discovered in the IDU.CON sequence, which is less than that of MSM.CON and HSX.CON. Characterization of T/F virus from IDUs highlights the genetic make-up and complexity of virus near the moment of transmission or in early infection preceding systemic dissemination and is important toward the development of an effective HIV-1 preventive methods, including vaccines.

  10. Production of Mucosally Transmissible SHIV Challenge Stocks from HIV-1 Circulating Recombinant Form 01_AE env Sequences.

    Directory of Open Access Journals (Sweden)

    Lawrence J Tartaglia

    2016-02-01

    Full Text Available Simian-human immunodeficiency virus (SHIV challenge stocks are critical for preclinical testing of vaccines, antibodies, and other interventions aimed to prevent HIV-1. A major unmet need for the field has been the lack of a SHIV challenge stock expressing circulating recombinant form 01_AE (CRF01_AE env sequences. We therefore sought to develop mucosally transmissible SHIV challenge stocks containing HIV-1 CRF01_AE env derived from acutely HIV-1 infected individuals from Thailand. SHIV-AE6, SHIV-AE6RM, and SHIV-AE16 contained env sequences that were >99% identical to the original HIV-1 isolate and did not require in vivo passaging. These viruses exhibited CCR5 tropism and displayed a tier 2 neutralization phenotype. These challenge stocks efficiently infected rhesus monkeys by the intrarectal route, replicated to high levels during acute infection, and established chronic viremia in a subset of animals. SHIV-AE16 was titrated for use in single, high dose as well as repetitive, low dose intrarectal challenge studies. These SHIV challenge stocks should facilitate the preclinical evaluation of vaccines, monoclonal antibodies, and other interventions targeted at preventing HIV-1 CRF01_AE infection.

  11. Study of HIV-1 subtypes in serodiscordant couples attending an integrated counselling and testing centre in Mumbai using heteroduplex mobility analysis and DNA sequencing

    Directory of Open Access Journals (Sweden)

    Mehta P

    2010-01-01

    Full Text Available Aims: To determine the prevalent subtypes of HIV-1 in serodiscordant couples. Setting: Integrated Counselling and Testing Centre (ICTC, Department of Microbiology. Study Design: Prospective pilot study. Participants: Thirty HIV-1 serodiscordant couples. Inclusion Criteria: a Documentation of HIV-1 infection in one partner and seronegative status in the other, current history of continued unprotected sexual activity within the partnership, demonstration that they have been in a partnership for at least 1 year and are not currently on highly active antiretroviral therapy HAART; b willingness of both partners to provide written informed consent including consent to continued couple counselling for 3 months. Materials and Methods: HIV-1 subtyping was carried out by heteroduplex mobility analysis (HMA by amplifying env region; and DNA sequencing by amplifying gag region. Results: HIV-1 env gene was amplified successfully in 10/30 samples; gag gene, in 25/30 samples; and both env and gag gene were amplified successfully in 5/30 samples. HIV-1 subtype C was detected from 21 samples; subtype B, from 7; and subtype A, from 2. Sample from 1 positive partner was detected as subtype C by env HMA and subtype B by gag sequencing. Conclusion: HIV-1 subtype C was found to be the predominant subtype of HIV-1 in serodiscordant couples attending our ICTC, followed by HIV-1 subtype B and HIV-1 subtype A, respectively. DNA sequencing was found to be the most reliable method for determining the subtypes of HIV-1.

  12. High prevalence of HIV-1 transmitted drug-resistance mutations from proviral DNA massively parallel sequencing data of therapy-naïve chronically infected Brazilian blood donors.

    Directory of Open Access Journals (Sweden)

    Rodrigo Pessôa

    Full Text Available An improved understanding of the prevalence of low-abundance transmitted drug-resistance mutations (TDRM in therapy-naïve HIV-1-infected patients may help determine which patients are the best candidates for therapy. In this study, we aimed to obtain a comprehensive picture of the evolving HIV-1 TDRM across the massive parallel sequences (MPS of the viral entire proviral genome in a well-characterized Brazilian blood donor naïve to antiretroviral drugs.The MPS data from 128 samples used in the analysis were sourced from Brazilian blood donors and were previously classified by less-sensitive (LS or "detuned" enzyme immunoassay as non-recent or longstanding HIV-1 infections. The Stanford HIV Resistance Database (HIVDBv 6.2 and IAS-USA mutation lists were used to interpret the pattern of drug resistance. The minority variants with TDRM were identified using a threshold of ≥ 1.0% and ≤ 20% of the reads sequenced. The rate of TDRM in the MPS data of the proviral genome were compared with the corresponding published consensus sequences of their plasma viruses.No TDRM were detected in the integrase or envelope regions. The overall prevalence of TDRM in the protease (PR and reverse transcriptase (RT regions of the HIV-1 pol gene was 44.5% (57/128, including any mutations to the nucleoside analogue reverse transcriptase inhibitors (NRTI and non-nucleoside analogue reverse transcriptase inhibitors (NNRTI. Of the 57 subjects, 43 (75.4% harbored a minority variant containing at least one clinically relevant TDRM. Among the 43 subjects, 33 (76.7% had detectable minority resistant variants to NRTIs, 6 (13.9% to NNRTIs, and 16 (37.2% to PR inhibitors. The comparison of viral sequences in both sources, plasma and cells, would have detected 48 DNA provirus disclosed TDRM by MPS previously missed by plasma bulk analysis.Our findings revealed a high prevalence of TDRM found in this group, as the use of MPS drastically increased the detection of these

  13. The C-terminal sequence of IFITM1 regulates its anti-HIV-1 activity.

    Directory of Open Access Journals (Sweden)

    Rui Jia

    Full Text Available The interferon-inducible transmembrane (IFITM proteins inhibit a wide range of viruses. We previously reported the inhibition of human immunodeficiency virus type 1 (HIV-1 strain BH10 by human IFITM1, 2 and 3. It is unknown whether other HIV-1 strains are similarly inhibited by IFITMs and whether there exists viral countermeasure to overcome IFITM inhibition. We report here that the HIV-1 NL4-3 strain (HIV-1NL4-3 is not restricted by IFITM1 and its viral envelope glycoprotein is partly responsible for this insensitivity. However, HIV-1NL4-3 is profoundly inhibited by an IFITM1 mutant, known as Δ(117-125, which is deleted of 9 amino acids at the C-terminus. In contrast to the wild type IFITM1, which does not affect HIV-1 entry, the Δ(117-125 mutant diminishes HIV-1NL4-3 entry by 3-fold. This inhibition correlates with the predominant localization of Δ(117-125 to the plasma membrane where HIV-1 entry occurs. In spite of strong conservation of IFITM1 among most species, mouse IFITM1 is 19 amino acids shorter at its C-terminus as compared to human IFITM1 and, like the human IFITM1 mutant Δ(117-125, mouse IFITM1 also inhibits HIV-1 entry. This is the first report illustrating the role of viral envelope protein in overcoming IFITM1 restriction. The results also demonstrate the importance of the C-terminal region of IFITM1 in modulating the antiviral function through controlling protein subcellular localization.

  14. Resistance to pyridine-based inhibitor KF116 reveals an unexpected role of integrase in HIV-1 Gag-Pol polyprotein proteolytic processing.

    Science.gov (United States)

    Hoyte, Ashley C; Jamin, Augusta V; Koneru, Pratibha C; Kobe, Matthew J; Larue, Ross C; Fuchs, James R; Engelman, Alan N; Kvaratskhelia, Mamuka

    2017-12-01

    The pyridine-based multimerization selective HIV-1 integrase (IN) inhibitors (MINIs) are a distinct subclass of allosteric IN inhibitors. MINIs potently inhibit HIV-1 replication during virion maturation by inducing hyper- or aberrant IN multimerization but are largely ineffective during the early steps of viral replication. Here, we investigated the mechanism for the evolution of a triple IN substitution (T124N/V165I/T174I) that emerges in cell culture with a representative MINI, KF116. We show that HIV-1 NL4-3(IN T124N/V165I/T174I) confers marked (>2000-fold) resistance to KF116. Two IN substitutions (T124N/T174I) directly weaken inhibitor binding at the dimer interface of the catalytic core domain but at the same time markedly impair HIV-1 replication capacity. Unexpectedly, T124N/T174I IN substitutions inhibited proteolytic processing of HIV-1 polyproteins Gag and Gag-Pol, resulting in immature virions. Strikingly, the addition of the third IN substitution (V165I) restored polyprotein processing, virus particle maturation, and significant levels of replication capacity. These results reveal an unanticipated role of IN for polyprotein proteolytic processing during virion morphogenesis. The complex evolutionary pathway for the emergence of resistant viruses, which includes the need for the compensatory V165I IN substitution, highlights a relatively high genetic barrier exerted by MINI KF116. Additionally, we have solved the X-ray structure of the drug-resistant catalytic core domain protein, which provides means for rational development of second-generation MINIs. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  15. Processing sites in the human immunodeficiency virus type 1 (HIV-1 Gag-Pro-Pol precursor are cleaved by the viral protease at different rates

    Directory of Open Access Journals (Sweden)

    Lindquist Jeffrey N

    2005-11-01

    Full Text Available Abstract We have examined the kinetics of processing of the HIV-1 Gag-Pro-Pol precursor in an in vitro assay with mature protease added in trans. The processing sites were cleaved at different rates to produce distinct intermediates. The initial cleavage occurred at the p2/NC site. Intermediate cleavages occurred at similar rates at the MA/CA and RT/IN sites, and to a lesser extent at sites upstream of RT. Late cleavages occurred at the sites flanking the protease (PR domain, suggesting sequestering of these sites. We observed paired intermediates indicative of half- cleavage of RT/RH site, suggesting that the RT domain in Gag-Pro-Pol was in a dimeric form under these assay conditions. These results clarify our understanding of the processing kinetics of the Gag-Pro-Pol precursor and suggest regulated cleavage. Our results further suggest that early dimerization of the PR and RT domains may serve as a regulatory element to influence the kinetics of processing within the Pol domain.

  16. Processing sites in the human immunodeficiency virus type 1 (HIV-1) Gag-Pro-Pol precursor are cleaved by the viral protease at different rates.

    Science.gov (United States)

    Pettit, Steve C; Lindquist, Jeffrey N; Kaplan, Andrew H; Swanstrom, Ronald

    2005-11-01

    We have examined the kinetics of processing of the HIV-1 Gag-Pro-Pol precursor in an in vitro assay with mature protease added in trans. The processing sites were cleaved at different rates to produce distinct intermediates. The initial cleavage occurred at the p2/NC site. Intermediate cleavages occurred at similar rates at the MA/CA and RT/IN sites, and to a lesser extent at sites upstream of RT. Late cleavages occurred at the sites flanking the protease (PR) domain, suggesting sequestering of these sites. We observed paired intermediates indicative of half- cleavage of RT/RH site, suggesting that the RT domain in Gag-Pro-Pol was in a dimeric form under these assay conditions. These results clarify our understanding of the processing kinetics of the Gag-Pro-Pol precursor and suggest regulated cleavage. Our results further suggest that early dimerization of the PR and RT domains may serve as a regulatory element to influence the kinetics of processing within the Pol domain.

  17. The highly conserved codon following the slippery sequence supports -1 frameshift efficiency at the HIV-1 frameshift site.

    Directory of Open Access Journals (Sweden)

    Suneeth F Mathew

    Full Text Available HIV-1 utilises -1 programmed ribosomal frameshifting to translate structural and enzymatic domains in a defined proportion required for replication. A slippery sequence, U UUU UUA, and a stem-loop are well-defined RNA features modulating -1 frameshifting in HIV-1. The GGG glycine codon immediately following the slippery sequence (the 'intercodon' contributes structurally to the start of the stem-loop but has no defined role in current models of the frameshift mechanism, as slippage is inferred to occur before the intercodon has reached the ribosomal decoding site. This GGG codon is highly conserved in natural isolates of HIV. When the natural intercodon was replaced with a stop codon two different decoding molecules-eRF1 protein or a cognate suppressor tRNA-were able to access and decode the intercodon prior to -1 frameshifting. This implies significant slippage occurs when the intercodon is in the (perhaps distorted ribosomal A site. We accommodate the influence of the intercodon in a model of frame maintenance versus frameshifting in HIV-1.

  18. Clustered epitopes within the Gag-Pol fusion protein DNA vaccine enhance immune responses and protection against challenge with recombinant vaccinia viruses expressing HIV-1 Gag and Pol antigens

    International Nuclear Information System (INIS)

    Bolesta, Elizabeth; Gzyl, Jaroslaw; Wierzbicki, Andrzej; Kmieciak, Dariusz; Kowalczyk, Aleksandra; Kaneko, Yutaro; Srinivasan, Alagarsamy; Kozbor, Danuta

    2005-01-01

    We have generated a codon-optimized hGagp17p24-Polp51 plasmid DNA expressing the human immunodeficiency virus type 1 (HIV-1) Gag-Pol fusion protein that consists of clusters of highly conserved cytotoxic T lymphocyte (CTL) epitopes presented by multiple MHC class I alleles. In the hGagp17p24-Polp51 construct, the ribosomal frameshift site had been deleted together with the potentially immunosuppressive Gag nucleocapsid (p15) as well as Pol protease (p10) and integrase (p31). Analyses of the magnitude and breadth of cellular responses demonstrated that immunization of HLA-A2/K b transgenic mice with the hGagp17p24-Polp51 construct induced 2- to 5-fold higher CD8 + T-cell responses to Gag p17-, p24-, and Pol reverse transcriptase (RT)-specific CTL epitopes than the full-length hGag-PolΔFsΔPr counterpart. The increases were correlated with higher protection against challenge with recombinant vaccinia viruses (rVVs) expressing gag and pol gene products. Consistent with the profile of Gag- and Pol-specific CD8 + T cell responses, an elevated level of type 1 cytokine production was noted in p24- and RT-stimulated splenocyte cultures established from hGagp17p24-Polp51-immunized mice compared to responses induced with the hGag-PolΔFsΔPr vaccine. Sera of mice immunized with the hGagp17p24-Polp51 vaccine also exhibited an increased titer of p24- and RT-specific IgG2 antibody responses. The results from our studies provide insights into approaches for boosting the breadth of Gag- and Pol-specific immune responses

  19. Comprehensive sieve analysis of breakthrough HIV-1 sequences in the RV144 vaccine efficacy trial.

    Science.gov (United States)

    Edlefsen, Paul T; Rolland, Morgane; Hertz, Tomer; Tovanabutra, Sodsai; Gartland, Andrew J; deCamp, Allan C; Magaret, Craig A; Ahmed, Hasan; Gottardo, Raphael; Juraska, Michal; McCoy, Connor; Larsen, Brendan B; Sanders-Buell, Eric; Carrico, Chris; Menis, Sergey; Kijak, Gustavo H; Bose, Meera; Arroyo, Miguel A; O'Connell, Robert J; Nitayaphan, Sorachai; Pitisuttithum, Punnee; Kaewkungwal, Jaranit; Rerks-Ngarm, Supachai; Robb, Merlin L; Kirys, Tatsiana; Georgiev, Ivelin S; Kwong, Peter D; Scheffler, Konrad; Pond, Sergei L Kosakovsky; Carlson, Jonathan M; Michael, Nelson L; Schief, William R; Mullins, James I; Kim, Jerome H; Gilbert, Peter B

    2015-02-01

    The RV144 clinical trial showed the partial efficacy of a vaccine regimen with an estimated vaccine efficacy (VE) of 31% for protecting low-risk Thai volunteers against acquisition of HIV-1. The impact of vaccine-induced immune responses can be investigated through sieve analysis of HIV-1 breakthrough infections (infected vaccine and placebo recipients). A V1/V2-targeted comparison of the genomes of HIV-1 breakthrough viruses identified two V2 amino acid sites that differed between the vaccine and placebo groups. Here we extended the V1/V2 analysis to the entire HIV-1 genome using an array of methods based on individual sites, k-mers and genes/proteins. We identified 56 amino acid sites or "signatures" and 119 k-mers that differed between the vaccine and placebo groups. Of those, 19 sites and 38 k-mers were located in the regions comprising the RV144 vaccine (Env-gp120, Gag, and Pro). The nine signature sites in Env-gp120 were significantly enriched for known antibody-associated sites (p = 0.0021). In particular, site 317 in the third variable loop (V3) overlapped with a hotspot of antibody recognition, and sites 369 and 424 were linked to CD4 binding site neutralization. The identified signature sites significantly covaried with other sites across the genome (mean = 32.1) more than did non-signature sites (mean = 0.9) (p analysis of the breakthrough infections in the RV144 trial, this work describes a set of statistical methods and tools applicable to analysis of breakthrough infection genomes in general vaccine efficacy trials for diverse pathogens.

  20. Murine mammary tumor virus pol-related sequences in human DNA: characterization and sequence comparison with the complete murine mammary tumor virus pol gene

    International Nuclear Information System (INIS)

    Deen, K.C.; Sweet, R.W.

    1986-01-01

    Sequences in the human genome with homology to the murine mammary tumor virus (MMTV) pol gene were isolated from a human phage library. Ten clones with extensive pol homology were shown to define five separate loci. These loci share common sequences immediately adjacent to the pol-like segments and, in addition, contain a related repeat element which bounds this region. This organization is suggestive of a proviral structure. The authors estimate that the human genome contains 30 to 40 copies of these pol-related sequences. The pol region of one of the cloned segments (HM16) and the complete MMTV pol gene were sequenced and compared. The nucleotide homology between these pol sequences is 52% and is concentrated in the terminal regions. The MMTV pol gene contains a single long open reading frame encoding 899 amino acids and is demarcated from the partially overlapping putative gag gene by termination codons and a shift in translational reading frame. The pol sequence of HM16 is multiply terminated but does contain open reading frames which encode 370, 105, and 112 amino acids residues in separate reading frames. The authors deduced a composite pol protein sequence for HM16 by aligning it to the MMTV pol gene and then compared these sequences with other retroviral pol protein sequences. Conserved sequences occur in both the amino and carboxyl regions which lie within the polymerase and endonuclease domains of pol, respectively

  1. HIV-1 molecular epidemiology among newly diagnosed HIV-1 individuals in Hebei, a low HIV prevalence province in China.

    Directory of Open Access Journals (Sweden)

    Xinli Lu

    Full Text Available New human immunodeficiency virus type 1 (HIV-1 diagnoses are increasing rapidly in Hebei. The aim of this study presents the most extensive HIV-1 molecular epidemiology investigation in Hebei province in China thus far. We have carried out the most extensive systematic cross-sectional study based on newly diagnosed HIV-1 positive individuals in 2013, and characterized the molecular epidemiology of HIV-1 based on full length gag-partial pol gene sequences in the whole of Hebei. Nine HIV-1 genotypes based on full length gag-partial pol gene sequence were identified among 610 newly diagnosed naïve individuals. The four main genotypes were circulating recombinant form (CRF01_AE (53.4%, CRF07_BC (23.4%, subtype B (15.9%, and unique recombinant forms URFs (4.9%. Within 1 year, three new genotypes (subtype A1, CRF55_01B, CRF65_cpx, unknown before in Hebei, were first found among men who have sex with men (MSM. All nine genotypes were identified in the sexually contracted HIV-1 population. Among 30 URFs, six recombinant patterns were revealed, including CRF01_AE/BC (40.0%, CRF01_AE/B (23.3%, B/C (16.7%, CRF01_AE/C (13.3%, CRF01_AE/B/A2 (3.3% and CRF01_AE/BC/A2 (3.3%, plus two potential CRFs. This study elucidated the complicated characteristics of HIV-1 molecular epidemiology in a low HIV-1 prevalence northern province of China and revealed the high level of HIV-1 genetic diversity. All nine HIV-1 genotypes circulating in Hebei have spread out of their initial risk groups into the general population through sexual contact, especially through MSM. This highlights the urgency of HIV prevention and control in China.

  2. HIV-1 molecular epidemiology among newly diagnosed HIV-1 individuals in Hebei, a low HIV prevalence province in China.

    Science.gov (United States)

    Lu, Xinli; Kang, Xianjiang; Liu, Yongjian; Cui, Ze; Guo, Wei; Zhao, Cuiying; Li, Yan; Chen, Suliang; Li, Jingyun; Zhang, Yuqi; Zhao, Hongru

    2017-01-01

    New human immunodeficiency virus type 1 (HIV-1) diagnoses are increasing rapidly in Hebei. The aim of this study presents the most extensive HIV-1 molecular epidemiology investigation in Hebei province in China thus far. We have carried out the most extensive systematic cross-sectional study based on newly diagnosed HIV-1 positive individuals in 2013, and characterized the molecular epidemiology of HIV-1 based on full length gag-partial pol gene sequences in the whole of Hebei. Nine HIV-1 genotypes based on full length gag-partial pol gene sequence were identified among 610 newly diagnosed naïve individuals. The four main genotypes were circulating recombinant form (CRF)01_AE (53.4%), CRF07_BC (23.4%), subtype B (15.9%), and unique recombinant forms URFs (4.9%). Within 1 year, three new genotypes (subtype A1, CRF55_01B, CRF65_cpx), unknown before in Hebei, were first found among men who have sex with men (MSM). All nine genotypes were identified in the sexually contracted HIV-1 population. Among 30 URFs, six recombinant patterns were revealed, including CRF01_AE/BC (40.0%), CRF01_AE/B (23.3%), B/C (16.7%), CRF01_AE/C (13.3%), CRF01_AE/B/A2 (3.3%) and CRF01_AE/BC/A2 (3.3%), plus two potential CRFs. This study elucidated the complicated characteristics of HIV-1 molecular epidemiology in a low HIV-1 prevalence northern province of China and revealed the high level of HIV-1 genetic diversity. All nine HIV-1 genotypes circulating in Hebei have spread out of their initial risk groups into the general population through sexual contact, especially through MSM. This highlights the urgency of HIV prevention and control in China.

  3. Comprehensive sieve analysis of breakthrough HIV-1 sequences in the RV144 vaccine efficacy trial.

    Directory of Open Access Journals (Sweden)

    Paul T Edlefsen

    2015-02-01

    Full Text Available The RV144 clinical trial showed the partial efficacy of a vaccine regimen with an estimated vaccine efficacy (VE of 31% for protecting low-risk Thai volunteers against acquisition of HIV-1. The impact of vaccine-induced immune responses can be investigated through sieve analysis of HIV-1 breakthrough infections (infected vaccine and placebo recipients. A V1/V2-targeted comparison of the genomes of HIV-1 breakthrough viruses identified two V2 amino acid sites that differed between the vaccine and placebo groups. Here we extended the V1/V2 analysis to the entire HIV-1 genome using an array of methods based on individual sites, k-mers and genes/proteins. We identified 56 amino acid sites or "signatures" and 119 k-mers that differed between the vaccine and placebo groups. Of those, 19 sites and 38 k-mers were located in the regions comprising the RV144 vaccine (Env-gp120, Gag, and Pro. The nine signature sites in Env-gp120 were significantly enriched for known antibody-associated sites (p = 0.0021. In particular, site 317 in the third variable loop (V3 overlapped with a hotspot of antibody recognition, and sites 369 and 424 were linked to CD4 binding site neutralization. The identified signature sites significantly covaried with other sites across the genome (mean = 32.1 more than did non-signature sites (mean = 0.9 (p < 0.0001, suggesting functional and/or structural relevance of the signature sites. Since signature sites were not preferentially restricted to the vaccine immunogens and because most of the associations were insignificant following correction for multiple testing, we predict that few of the genetic differences are strongly linked to the RV144 vaccine-induced immune pressure. In addition to presenting results of the first complete-genome analysis of the breakthrough infections in the RV144 trial, this work describes a set of statistical methods and tools applicable to analysis of breakthrough infection genomes in general vaccine

  4. A comparison of parallel pyrosequencing and sanger clone-based sequencing and its impact on the characterization of the genetic diversity of HIV-1.

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    Binhua Liang

    Full Text Available BACKGROUND: Pyrosequencing technology has the potential to rapidly sequence HIV-1 viral quasispecies without requiring the traditional approach of cloning. In this study, we investigated the utility of ultra-deep pyrosequencing to characterize genetic diversity of the HIV-1 gag quasispecies and assessed the possible contribution of pyrosequencing technology in studying HIV-1 biology and evolution. METHODOLOGY/PRINCIPAL FINDINGS: HIV-1 gag gene was amplified from 96 patients using nested PCR. The PCR products were cloned and sequenced using capillary based Sanger fluorescent dideoxy termination sequencing. The same PCR products were also directly sequenced using the 454 pyrosequencing technology. The two sequencing methods were evaluated for their ability to characterize quasispecies variation, and to reveal sites under host immune pressure for their putative functional significance. A total of 14,034 variations were identified by 454 pyrosequencing versus 3,632 variations by Sanger clone-based (SCB sequencing. 11,050 of these variations were detected only by pyrosequencing. These undetected variations were located in the HIV-1 Gag region which is known to contain putative cytotoxic T lymphocyte (CTL and neutralizing antibody epitopes, and sites related to virus assembly and packaging. Analysis of the positively selected sites derived by the two sequencing methods identified several differences. All of them were located within the CTL epitope regions. CONCLUSIONS/SIGNIFICANCE: Ultra-deep pyrosequencing has proven to be a powerful tool for characterization of HIV-1 genetic diversity with enhanced sensitivity, efficiency, and accuracy. It also improved reliability of downstream evolutionary and functional analysis of HIV-1 quasispecies.

  5. Sequence requirements of the HIV-1 protease flap region determined by saturation mutagenesis and kinetic analysis of flap mutants

    Science.gov (United States)

    Shao, Wei; Everitt, Lorraine; Manchester, Marianne; Loeb, Daniel D.; Hutchison, Clyde A.; Swanstrom, Ronald

    1997-01-01

    The retroviral proteases (PRs) have a structural feature called the flap, which consists of a short antiparallel β-sheet with a turn. The flap extends over the substrate binding cleft and must be flexible to allow entry and exit of the polypeptide substrates and products. We analyzed the sequence requirements of the amino acids within the flap region (positions 46–56) of the HIV-1 PR. The phenotypes of 131 substitution mutants were determined using a bacterial expression system. Four of the mutant PRs with mutations in different regions of the flap were selected for kinetic analysis. Our phenotypic analysis, considered in the context of published structures of the HIV-1 PR with a bound substrate analogs, shows that: (i) Met-46 and Phe-53 participate in hydrophobic interactions on the solvent-exposed face of the flap; (ii) Ile-47, Ile-54, and Val-56 participate in hydrophobic interactions on the inner face of the flap; (iii) Ile-50 has hydrophobic interactions at the distance of both the δ and γ carbons; (iv) the three glycine residues in the β-turn of the flap are virtually intolerant of substitutions. Among these mutant PRs, we have identified changes in both kcat and Km. These results establish the nature of the side chain requirements at each position in the flap and document a role for the flap in both substrate binding and catalysis. PMID:9122179

  6. Phylogenetic analysis of HIV-1 reverse transcriptase sequences from 382 patients recruited in JJ Hospital of Mumbai, India, between 2002 and 2008.

    Science.gov (United States)

    Deshpande, Alaka; Jauvin, Valerie; Pinson, Patricia; Jeannot, Anne Cecile; Fleury, Herve J

    2009-06-01

    Analysis of reverse transcriptase (RT) sequences of 382 HIV-1 isolates from untreated and treated patients recruited in JJ Hospital (Mumbai, India) between 2002 and 2008 shows that subtype C is largely predominant (98%) and that non-C sequences cluster with A1, B, CRF01_AE, and CRF06_cpx.

  7. Impact of Clinical Parameters in the Intrahost Evolution of HIV-1 Subtype B in Pediatric Patients: A Machine Learning Approach

    Science.gov (United States)

    Rojas Sánchez, Patricia; Cobos, Alberto; Navaro, Marisa; Ramos, José Tomas; Pagán, Israel

    2017-01-01

    Abstract Determining the factors modulating the genetic diversity of HIV-1 populations is essential to understand viral evolution. This study analyzes the relative importance of clinical factors in the intrahost HIV-1 subtype B (HIV-1B) evolution and in the fixation of drug resistance mutations (DRM) during longitudinal pediatric HIV-1 infection. We recovered 162 partial HIV-1B pol sequences (from 3 to 24 per patient) from 24 perinatally infected patients from the Madrid Cohort of HIV-1 infected children and adolescents in a time interval ranging from 2.2 to 20.3 years. We applied machine learning classification methods to analyze the relative importance of 28 clinical/epidemiological/virological factors in the HIV-1B evolution to predict HIV-1B genetic diversity (d), nonsynonymous and synonymous mutations (dN, dS) and DRM presence. Most of the 24 HIV-1B infected pediatric patients were Spanish (91.7%), diagnosed before 2000 (83.3%), and all were antiretroviral therapy experienced. They had from 0.3 to 18.8 years of HIV-1 exposure at sampling time. Most sequences presented DRM. The best-predictor variables for HIV-1B evolutionary parameters were the age of HIV-1 diagnosis for d, the age at first antiretroviral treatment for dN and the year of HIV-1 diagnosis for ds. The year of infection (birth year) and year of sampling seemed to be relevant for fixation of both DRM at large and, considering drug families, to protease inhibitors (PI). This study identifies, for the first time using machine learning, the factors affecting more HIV-1B pol evolution and those affecting DRM fixation in HIV-1B infected pediatric patients. PMID:29044435

  8. Genotypic Resistance Tests Sequences Reveal the Role of Marginalized Populations in HIV-1 Transmission in Switzerland.

    Science.gov (United States)

    Shilaih, Mohaned; Marzel, Alex; Yang, Wan Lin; Scherrer, Alexandra U; Schüpbach, Jörg; Böni, Jürg; Yerly, Sabine; Hirsch, Hans H; Aubert, Vincent; Cavassini, Matthias; Klimkait, Thomas; Vernazza, Pietro L; Bernasconi, Enos; Furrer, Hansjakob; Günthard, Huldrych F; Kouyos, Roger

    2016-06-14

    Targeting hard-to-reach/marginalized populations is essential for preventing HIV-transmission. A unique opportunity to identify such populations in Switzerland is provided by a database of all genotypic-resistance-tests from Switzerland, including both sequences from the Swiss HIV Cohort Study (SHCS) and non-cohort sequences. A phylogenetic tree was built using 11,127 SHCS and 2,875 Swiss non-SHCS sequences. Demographics were imputed for non-SHCS patients using a phylogenetic proximity approach. Factors associated with non-cohort outbreaks were determined using logistic regression. Non-B subtype (univariable odds-ratio (OR): 1.9; 95% confidence interval (CI): 1.8-2.1), female gender (OR: 1.6; 95% CI: 1.4-1.7), black ethnicity (OR: 1.9; 95% CI: 1.7-2.1) and heterosexual transmission group (OR:1.8; 95% CI: 1.6-2.0), were all associated with underrepresentation in the SHCS. We found 344 purely non-SHCS transmission clusters, however, these outbreaks were small (median 2, maximum 7 patients) with a strong overlap with the SHCS'. 65% of non-SHCS sequences were part of clusters composed of >= 50% SHCS sequences. Our data suggests that marginalized-populations are underrepresented in the SHCS. However, the limited size of outbreaks among non-SHCS patients in-care implies that no major HIV outbreak in Switzerland was missed by the SHCS surveillance. This study demonstrates the potential of sequence data to assess and extend the scope of infectious-disease surveillance.

  9. Prediction of HIV-1 coreceptor usage (tropism) by sequence analysis using a genotypic approach.

    Science.gov (United States)

    Sierra, Saleta; Kaiser, Rolf; Lübke, Nadine; Thielen, Alexander; Schuelter, Eugen; Heger, Eva; Däumer, Martin; Reuter, Stefan; Esser, Stefan; Fätkenheuer, Gerd; Pfister, Herbert; Oette, Mark; Lengauer, Thomas

    2011-12-01

    Maraviroc (MVC) is the first licensed antiretroviral drug from the class of coreceptor antagonists. It binds to the host coreceptor CCR5, which is used by the majority of HIV strains in order to infect the human immune cells (Fig. 1). Other HIV isolates use a different coreceptor, the CXCR4. Which receptor is used, is determined in the virus by the Env protein (Fig. 2). Depending on the coreceptor used, the viruses are classified as R5 or X4, respectively. MVC binds to the CCR5 receptor inhibiting the entry of R5 viruses into the target cell. During the course of disease, X4 viruses may emerge and outgrow the R5 viruses. Determination of coreceptor usage (also called tropism) is therefore mandatory prior to administration of MVC, as demanded by EMA and FDA. The studies for MVC efficiency MOTIVATE, MERIT and 1029 have been performed with the Trofile assay from Monogram, San Francisco, U.S.A. This is a high quality assay based on sophisticated recombinant tests. The acceptance for this test for daily routine is rather low outside of the U.S.A., since the European physicians rather tend to work with decentralized expert laboratories, which also provide concomitant resistance testing. These laboratories have undergone several quality assurance evaluations, the last one being presented in 2011. For several years now, we have performed tropism determinations based on sequence analysis from the HIV env-V3 gene region (V3). This region carries enough information to perform a reliable prediction. The genotypic determination of coreceptor usage presents advantages such as: shorter turnover time (equivalent to resistance testing), lower costs, possibility to adapt the results to the patients' needs and possibility of analysing clinical samples with very low or even undetectable viral load (VL), particularly since the number of samples analysed with VL < 1000 copies/μl roughly increased in the last years (Fig. 3). The main steps for tropism testing (Fig. 4) demonstrated in

  10. Structure-Related Roles for the Conservation of the HIV-1 Fusion Peptide Sequence Revealed by Nuclear Magnetic Resonance.

    Science.gov (United States)

    Serrano, Soraya; Huarte, Nerea; Rujas, Edurne; Andreu, David; Nieva, José L; Jiménez, María Angeles

    2017-10-17

    Despite extensive characterization of the human immunodeficiency virus type 1 (HIV-1) hydrophobic fusion peptide (FP), the structure-function relationships underlying its extraordinary degree of conservation remain poorly understood. Specifically, the fact that the tandem repeat of the FLGFLG tripeptide is absolutely conserved suggests that high hydrophobicity may not suffice to unleash FP function. Here, we have compared the nuclear magnetic resonance (NMR) structures adopted in nonpolar media by two FP surrogates, wtFP-tag and scrFP-tag, which had equal hydrophobicity but contained wild-type and scrambled core sequences LFLGFLG and FGLLGFL, respectively. In addition, these peptides were tagged at their C-termini with an epitope sequence that folded independently, thereby allowing Western blot detection without interfering with FP structure. We observed similar α-helical FP conformations for both specimens dissolved in the low-polarity medium 25% (v/v) 1,1,1,3,3,3-hexafluoro-2-propanol (HFIP), but important differences in contact with micelles of the membrane mimetic dodecylphosphocholine (DPC). Thus, whereas wtFP-tag preserved a helix displaying a Gly-rich ridge, the scrambled sequence lost in great part the helical structure upon being solubilized in DPC. Western blot analyses further revealed the capacity of wtFP-tag to assemble trimers in membranes, whereas membrane oligomers were not observed in the case of the scrFP-tag sequence. We conclude that, beyond hydrophobicity, preserving sequence order is an important feature for defining the secondary structures and oligomeric states adopted by the HIV FP in membranes.

  11. New findings on the d(TGGGAG) sequence: Surprising anti-HIV-1 activity.

    Science.gov (United States)

    Romanucci, Valeria; Zarrelli, Armando; Liekens, Sandra; Noppen, Sam; Pannecouque, Christophe; Di Fabio, Giovanni

    2018-02-10

    The biological relevance of tetramolecular G-quadruplexes especially as anti-HIV agents has been extensively reported in the literature over the last years. In the light of our recent results regarding the slow G-quadruplex folding kinetics of ODNs based on d(TGGGAG) sequence, here we report a systematic anti-HIV screening to investigate the impact of the G-quadruplex folding on their anti-HIV activity. In particular, varying the single stranded concentrations of ODNs, it has been tested a pool of ODN sample solutions with different G-quadruplex concentrations. The anti-HIV assays have been designed favouring the limited kinetics involved in the tetramolecular G4-association based on the d(TGGGAG) sequence. Aiming to determine the stoichiometry of G-quadruplex structures in the same experimental conditions of the anti-HIV assays, a native gel electrophoresis was performed. The gel confirmed the G-quadruplex formation for almost all sample solutions while showing the formation of high order G4 structures for the more concentrated ODNs solutions. The most significant result is the discovery of a potent anti-HIV activity of the G-quadruplex formed by the natural d(TGGGAG) sequence (IC 50  = 14 nM) that, until now, has been reported to be completely inactive against HIV infection. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

  12. HIV-1 subtypes D and F are prevalent in Guinea Conakry.

    Science.gov (United States)

    Freimanis, G L; Loua, A; Allain, J P

    2012-04-01

    Limited data is available upon the distribution of different HIV-1/2 genotypes in the blood donor population from Guinea Conakry. To investigate the prevalence of HIV-1/2 subtypes in asymptomatic blood donors in Guinea Conakry, in order to update knowledge of HIV-1/2 epidemiology within this country. Samples from 104 blood donors seropositive for HIV-1/2 were tested for HIV-1 by real-time RT-PCR. Those negative for HIV-1 were tested with HIV-2 nested RT-PCR. Positive samples were further amplified in the HIV-1 gag and pol regions and sequenced. Subtypes were determined by phylogenetic analysis on amplicon sequences. 61 samples were positive by HIV-1 real-time RT-PCR. Of the 43 negative, 2 (4.6%) were positive for HIV-2. 52/61 (85.3%) samples were positive by nested RT-PCR. Of the 52, 43 (70.5%) and 31(59.6%) sequences were obtained in the gag and pol regions, respectively; 23 for both regions. HIV-1 subtype distribution was 1 B (2.1%), 8 F (17%), 8 D (17%) and 28 CRF02_AG (59.6%) with 2 unclassified recombinants (4.3%). Unique clusters for subtype D and F distinguished Guinea from HIV-1 subtype distribution in neighboring countries. Subtype F and subtype D strains, uncommon in West Africa, are a substantial part of HIV-1 epidemiology in Guinea. Copyright © 2011 Elsevier B.V. All rights reserved.

  13. Evaluation of the Roche prototype 454 HIV-1 ultradeep sequencing drug resistance assay in a routine diagnostic laboratory.

    Science.gov (United States)

    Garcia-Diaz, A; Guerrero-Ramos, A; McCormick, A L; Macartney, M; Conibear, T; Johnson, M A; Haque, T; Webster, D P

    2013-10-01

    Studies have shown that low-frequency resistance mutations can influence treatment outcome. However, the lack of a standardized high-throughput assay has precluded their detection in clinical settings. To evaluate the performance of the Roche prototype 454 UDS HIV-1 drug resistance assay (UDS assay) in a routine diagnostic laboratory. 50 plasma samples, previously characterized by population sequencing and that had shown ≥1 resistance associated mutation (RAM), were retrospectively tested by the UDS assay, including 18 B and 32 non-B subtypes; viral loads between 114-1,806,407 cp/ml; drug-naive (n=27) and drug-experienced (n=23) individuals. The UDS assay was successful for 37/50 (74%) samples. It detected all RAMs found by population sequencing at frequencies above 20%. In addition, 39 low-frequency RAMs were exclusively detected by the UDS assay at frequencies below 20% in both drug-naïve (19/26, 73%) and drug-experienced (9/18, 50%) individuals. UDS results would lead to changes from susceptible to resistant to efavirenz (EFV) in one drug-naive individual with suboptimal response to an EFV-containing regimen and from susceptible to resistance to lamivudine (3TC) in one drug naïve subject who subsequently failed a 3TC-containing regimen and in a treatment experienced subject who had failed a 3TC-containing regimen. The UDS assay performed well across a wide range of subtypes and viral loads; it showed perfect agreement with population sequencing for all RAMs analyzed. In addition, the UDS assay detected additional mutations at frequencies below 20% which correlate with patients' treatment history and had in some cases important prognostic implications. Copyright © 2013 Elsevier B.V. All rights reserved.

  14. Stress- and sequence-dependent release into the culture medium of HIV-1 Nef produced in Saccharomyces cerevisiae.

    Science.gov (United States)

    Macreadie, I G; Castelli, L A; Lucantoni, A; Azad, A A

    1995-09-11

    We have produced human immunodeficiency virus type 1 (HIV-1) Nef (a myristylated 206-amino-acid protein) in Saccharomyces cerevisaie and shown that, while Nef is normally found as a predominantly intracellular protein, amounts up to 40 micrograms/ml of Nef are also released into the extracellular medium during stress. By electrophoretic (SDS-PAGE) analysis the extracellular Nef is indistinguishable from intracellular Nef. Conditions of stress that lead to the release of Nef include elevated levels of copper or magnesium ions or growth at elevated temperatures. This release appears to be dependent upon the N-terminal sequences of Nef, including the presence of a myristylation site. Our observations concerning Nef release in yeast suggest new ways in which the behaviour of Nef should be examined in order to gain further insights into the development of AIDS. If the release of Nef is important in the development of AIDS, our work reveals that Nef-associated symptoms may be reduced or delayed by reducing stresses, such as fevers.

  15. Exploration of the effect of sequence variations located inside the binding pocket of HIV-1 and HIV-2 proteases.

    Science.gov (United States)

    Triki, Dhoha; Billot, Telli; Visseaux, Benoit; Descamps, Diane; Flatters, Delphine; Camproux, Anne-Claude; Regad, Leslie

    2018-04-10

    HIV-2 protease (PR2) is naturally resistant to most FDA (Food and Drug Administration)-approved HIV-1 protease inhibitors (PIs), a major antiretroviral class. In this study, we compared the PR1 and PR2 binding pockets extracted from structures complexed with 12 ligands. The comparison of PR1 and PR2 pocket properties showed that bound PR2 pockets were more hydrophobic with more oxygen atoms and fewer nitrogen atoms than PR1 pockets. The structural comparison of PR1 and PR2 pockets highlighted structural changes induced by their sequence variations and that were consistent with these property changes. Specifically, substitutions at residues 31, 46, and 82 induced structural changes in their main-chain atoms that could affect PI binding in PR2. In addition, the modelling of PR1 mutant structures containing V32I and L76M substitutions revealed a cooperative mechanism leading to structural deformation of flap-residue 45 that could modify PR2 flexibility. Our results suggest that substitutions in the PR1 and PR2 pockets can modify PI binding and flap flexibility, which could underlie PR2 resistance against PIs. These results provide new insights concerning the structural changes induced by PR1 and PR2 pocket variation changes, improving the understanding of the atomic mechanism of PR2 resistance to PIs.

  16. Subtype-independent near full-length HIV-1 genome sequencing and assembly to be used in large molecular epidemiological studies and clinical management.

    Science.gov (United States)

    Grossmann, Sebastian; Nowak, Piotr; Neogi, Ujjwal

    2015-01-01

    HIV-1 near full-length genome (HIV-NFLG) sequencing from plasma is an attractive multidimensional tool to apply in large-scale population-based molecular epidemiological studies. It also enables genotypic resistance testing (GRT) for all drug target sites allowing effective intervention strategies for control and prevention in high-risk population groups. Thus, the main objective of this study was to develop a simplified subtype-independent, cost- and labour-efficient HIV-NFLG protocol that can be used in clinical management as well as in molecular epidemiological studies. Plasma samples (n=30) were obtained from HIV-1B (n=10), HIV-1C (n=10), CRF01_AE (n=5) and CRF01_AG (n=5) infected individuals with minimum viral load >1120 copies/ml. The amplification was performed with two large amplicons of 5.5 kb and 3.7 kb, sequenced with 17 primers to obtain HIV-NFLG. GRT was validated against ViroSeq™ HIV-1 Genotyping System. After excluding four plasma samples with low-quality RNA, a total of 26 samples were attempted. Among them, NFLG was obtained from 24 (92%) samples with the lowest viral load being 3000 copies/ml. High (>99%) concordance was observed between HIV-NFLG and ViroSeq™ when determining the drug resistance mutations (DRMs). The N384I connection mutation was additionally detected by NFLG in two samples. Our high efficiency subtype-independent HIV-NFLG is a simple and promising approach to be used in large-scale molecular epidemiological studies. It will facilitate the understanding of the HIV-1 pandemic population dynamics and outline effective intervention strategies. Furthermore, it can potentially be applicable in clinical management of drug resistance by evaluating DRMs against all available antiretrovirals in a single assay.

  17. High level of APOBEC3F/3G editing in HIV-2 DNA vif and pol sequences from antiretroviral-naive patients.

    Science.gov (United States)

    Bertine, Mélanie; Charpentier, Charlotte; Visseaux, Benoit; Storto, Alexandre; Collin, Gilles; Larrouy, Lucile; Damond, Florence; Matheron, Sophie; Brun-Vézinet, Françoise; Descamps, Diane

    2015-04-24

    In HIV-1, hypermutation introduced by APOBEC3F/3G cytidine deaminase activity leads to defective viruses. In-vivo impact of APOBEC3F/3G editing on HIV-2 sequences remains unknown. The objective of this study was to assess the level of APOBEC3F/3G editing in HIV-2-infected antiretroviral-naive patients. Direct sequencing of vif and pol regions was performed on HIV-2 proviral DNA from antiretroviral-naive patients included in the French Agence Nationale de Recherches sur le SIDA et les hépatites virales CO5 HIV-2 cohort. Hypermutated sequences were identified using Hypermut2.0 program. HIV-1 proviral sequences from Genbank were also assessed. Among 82 antiretroviral-naive HIV-2-infected patients assessed, 15 (28.8%) and five (16.7%) displayed Vif proviral defective sequences in HIV-2 groups A and B, respectively. A lower proportion of defective sequences was observed in protease-reverse transcriptase region. A higher median number of G-to-A mutations was observed in HIV-2 group B than in group A, both in Vif and protease-reverse transcriptase regions (P = 0.02 and P = 0.006, respectively). Compared with HIV-1 Vif sequences, a higher number of Vif defective sequences was observed in HIV-2 group A (P = 0.00001) and group B sequences (P = 0.013). We showed for the first time a high level of APOBEC3F/3G editing in HIV-2 sequences from antiretroviral-naive patients. Our study reported a group effect with a significantly higher level of APOBEC3F/3G editing in HIV-2 group B than in group A sequences.

  18. Primary resistance to integrase strand transfer inhibitors in patients infected with diverse HIV-1 subtypes in sub-Saharan Africa

    NARCIS (Netherlands)

    Inzaule, Seth C.; Hamers, Raph L.; Noguera-Julian, Marc; Casadellà, Maria; Parera, Mariona; Rinke de Wit, Tobias F.; Paredes, Roger

    2018-01-01

    To investigate the prevalence and patterns of major and accessory resistance mutations associated with integrase strand transfer inhibitors (INSTIs), across diverse HIV-1 subtypes in sub-Saharan Africa. pol gene sequences were obtained using Illumina next-generation sequencing from 425 INSTI-naive

  19. HIV-1 vaccines

    Science.gov (United States)

    Excler, Jean-Louis; Robb, Merlin L; Kim, Jerome H

    2014-01-01

    The development of a safe and effective preventive HIV-1 vaccine remains a public health priority. Despite scientific difficulties and disappointing results, HIV-1 vaccine clinical development has, for the first time, established proof-of-concept efficacy against HIV-1 acquisition and identified vaccine-associated immune correlates of risk. The correlate of risk analysis showed that IgG antibodies against the gp120 V2 loop correlated with decreased risk of HIV infection, while Env-specific IgA directly correlated with increased risk. The development of vaccine strategies such as improved envelope proteins formulated with potent adjuvants and DNA and vectors expressing mosaics, or conserved sequences, capable of eliciting greater breadth and depth of potentially relevant immune responses including neutralizing and non-neutralizing antibodies, CD4+ and CD8+ cell-mediated immune responses, mucosal immune responses, and immunological memory, is now proceeding quickly. Additional human efficacy trials combined with other prevention modalities along with sustained funding and international collaboration remain key to bring an HIV-1 vaccine to licensure. PMID:24637946

  20. HIV-1 epidemic in Warao Amerindians from Venezuela: spatial phylodynamics and epidemiological patterns.

    Science.gov (United States)

    Villalba, Julian A; Bello, Gonzalo; Maes, Mailis; Sulbaran, Yoneira F; Garzaro, Domingo; Loureiro, Carmen L; Rangel, Hector R; de Waard, Jacobus H; Pujol, Flor H

    2013-07-17

    We previously reported HIV-1 infection in Warao Amerindians from Venezuela. The aim of this study was to evaluate the extent and the dynamic of HIV-1 dissemination in eight Warao communities. HIV-1 infection was evaluated in 576 Warao Amerindians from the Orinoco Delta. Partial HIV-1 pol sequences were analyzed to reconstruct the spatiotemporal and demographic dynamics of the epidemic. HIV-1 antibodies were present in 9.55% of Warao Amerindians, ranging from 0 to 22%. A significantly higher prevalence was found in men (15.6%) compared with women (2.6%), reaching up to 35% in men from one community. All but one isolates were classified as subtype B. Warao's HIV-1 subtype-B epidemic resulted from a single viral introduction at around the early 2000s. After an initial phase of slow growth, the subtype B started to spread at a fast rate (0.8/year) following two major routes of migration within the communities. A dramatic high prevalence was documented in almost all the communities of Warao Amerindians from the Orinoco Delta tested for HIV-1 infection. This epidemic resulted from the dissemination of a single HIV-1 subtype B founder strain introduced about 10 years ago and its size is probably doubling every year, creating a situation that can be devastating for this vulnerable Amerindian group.

  1. HIV-1 epidemiology and circulating subtypes in the countryside of South Brazil

    Directory of Open Access Journals (Sweden)

    Carina Sperotto Librelotto

    2015-06-01

    Full Text Available INTRODUCTION: Human immunodeficiency virus type 1 (HIV-1 has spread worldwide, with several subtypes and circulating recombinant forms. Brazil has an incidence of 20.5 HIV-1/acquired immunodeficiency syndrome (AIDS patients per 100,000 inhabitants; however, the Southernmost State of Rio Grande do Sul (RS has more than twice the number of HIV-1-infected people (41.3/100,000 inhabitants and a different pattern of subtype frequencies, as previously reported in studies conducted in the capital (Porto Alegre and its metropolitan region. This study examined HIV-1/AIDS epidemiological and molecular aspects in the countryside of Rio Grande do Sul. METHODS: Socio-demographic, clinical and risk behavioral characteristics were obtained from HIV-1-positive adult patients using a structured questionnaire. HIV-1 subtypes were determined by nested-polymerase chain reaction (PCR and sequencing of the pol and env genes. RESULTS: The study sample included 149 (55% women patients with a mean age of 41.8 ± 11.9 years. Most (73.8% patients had a low education level and reported heterosexual practices as the most (91.9% probable transmission route. HIV-1 subtypes were detected in 26 patients: 18 (69.2% infected with subtype C, six (23.1% infected with subtype B and two (7.7% infected with BC recombinant forms. CONCLUSIONS: These data highlight the increasing number of HIV-1 subtype C infections in the countryside of South Brazil.

  2. Sequence-specific activation of the DNA sensor cGAS by Y-form DNA structures as found in primary HIV-1 cDNA.

    Science.gov (United States)

    Herzner, Anna-Maria; Hagmann, Cristina Amparo; Goldeck, Marion; Wolter, Steven; Kübler, Kirsten; Wittmann, Sabine; Gramberg, Thomas; Andreeva, Liudmila; Hopfner, Karl-Peter; Mertens, Christina; Zillinger, Thomas; Jin, Tengchuan; Xiao, Tsan Sam; Bartok, Eva; Coch, Christoph; Ackermann, Damian; Hornung, Veit; Ludwig, Janos; Barchet, Winfried; Hartmann, Gunther; Schlee, Martin

    2015-10-01

    Cytosolic DNA that emerges during infection with a retrovirus or DNA virus triggers antiviral type I interferon responses. So far, only double-stranded DNA (dsDNA) over 40 base pairs (bp) in length has been considered immunostimulatory. Here we found that unpaired DNA nucleotides flanking short base-paired DNA stretches, as in stem-loop structures of single-stranded DNA (ssDNA) derived from human immunodeficiency virus type 1 (HIV-1), activated the type I interferon-inducing DNA sensor cGAS in a sequence-dependent manner. DNA structures containing unpaired guanosines flanking short (12- to 20-bp) dsDNA (Y-form DNA) were highly stimulatory and specifically enhanced the enzymatic activity of cGAS. Furthermore, we found that primary HIV-1 reverse transcripts represented the predominant viral cytosolic DNA species during early infection of macrophages and that these ssDNAs were highly immunostimulatory. Collectively, our study identifies unpaired guanosines in Y-form DNA as a highly active, minimal cGAS recognition motif that enables detection of HIV-1 ssDNA.

  3. Phylodynamics of the HIV-1 epidemic in Cuba.

    Science.gov (United States)

    Delatorre, Edson; Bello, Gonzalo

    2013-01-01

    Previous studies have shown that the HIV-1 epidemic in Cuba displayed a complex molecular epidemiologic profile with circulation of several subtypes and circulating recombinant forms (CRF); but the evolutionary and population history of those viral variants remains unknown. HIV-1 pol sequences of the most prevalent Cuban lineages (subtypes B, C and G, CRF18_cpx, CRF19_cpx, and CRFs20/23/24_BG) isolated between 1999 and 2011 were analyzed. Maximum-likelihood analyses revealed multiple introductions of subtype B (n≥66), subtype C (n≥10), subtype G (n≥8) and CRF18_cpx (n≥2) viruses in Cuba. The bulk of HIV-1 infections in this country, however, was caused by dissemination of a few founder strains probably introduced from North America/Europe (clades B(CU-I) and B(CU-II)), east Africa (clade C(CU-I)) and central Africa (clades G(CU), CRF18(CU) and CRF19(CU)), or locally generated (clades CRFs20/23/24_BG). Bayesian-coalescent analyses show that the major HIV-1 founder strains were introduced into Cuba during 1985-1995; whereas the CRFs_BG strains emerged in the second half of the 1990s. Most HIV-1 Cuban clades appear to have experienced an initial period of fast exponential spread during the 1990s and early 2000s, followed by a more recent decline in growth rate. The median initial growth rate of HIV-1 Cuban clades ranged from 0.4 year⁻¹ to 1.6 year⁻¹. Thus, the HIV-1 epidemic in Cuba has been a result of the successful introduction of a few viral strains that began to circulate at a rather late time of the AIDS pandemic, but then were rapidly disseminated through local transmission networks.

  4. Molecular beacon probes-base multiplex NASBA Real-time for detection of HIV-1 and HCV.

    Science.gov (United States)

    Mohammadi-Yeganeh, S; Paryan, M; Mirab Samiee, S; Kia, V; Rezvan, H

    2012-06-01

    Developed in 1991, nucleic acid sequence-based amplification (NASBA) has been introduced as a rapid molecular diagnostic technique, where it has been shown to give quicker results than PCR, and it can also be more sensitive. This paper describes the development of a molecular beacon-based multiplex NASBA assay for simultaneous detection of HIV-1 and HCV in plasma samples. A well-conserved region in the HIV-1 pol gene and 5'-NCR of HCV genome were used for primers and molecular beacon design. The performance features of HCV/HIV-1 multiplex NASBA assay including analytical sensitivity and specificity, clinical sensitivity and clinical specificity were evaluated. The analysis of scalar concentrations of the samples indicated that the limit of quantification of the assay was beacon probes detected all HCV genotypes and all major variants of HIV-1. This method may represent a relatively inexpensive isothermal method for detection of HIV-1/HCV co-infection in monitoring of patients.

  5. In Vitro Transduction and Target-Mutagenesis Efficiency of HIV-1 pol Gene Targeting ZFN and CRISPR/Cas9 Delivered by Various Plasmids and/or Vectors: Toward an HIV Cure.

    Science.gov (United States)

    Okee, Moses; Bayiyana, Alice; Musubika, Carol; Joloba, Moses L; Ashaba-Katabazi, Fred; Bagaya, Bernard; Wayengera, Misaki

    2018-01-01

    Efficiency of artificial restriction enzymes toward curing HIV has only been separately examined, using differing delivery vehicles. We compared the in vitro transduction and target-mutagenesis efficiency of consortium plasmid and adenoviral vector delivered HIV-1 pol gene targeting zinc finger nuclease (ZFN) with CRISPR/Cas, Custom-ZFN, CRISPR-Cas-9, and plasmids and vectors (murCTSD_pZFN, pGS-U-gRNA, pCMV-Cas-D01A, Ad5-RGD); cell lines (TZM-bl and ACH-2/J-Lat cells); and the latency reversing agents prostratin, suberoylanilide hydroxamic acid, and phorbol myristate acetate. Cell lines were grown in either Dulbecco's modified Eagle's medium or Roswell Park Memorial Institute with the antibiotics kanamycin, zeocin, and efavirenz. Efficiency was assayed by GFP/luciferase activity and/or validated by yeast MEL1 reporter assay, CEL1 restriction fragment assay, and quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR). Ad5-RGD vectors had better transduction efficiency than murCTSD and pGS-U-gRNA/pCMV-Cas-D01A plasmids. CRISPR/Cas9 exhibited better target-mutagenesis efficiency relative to ZFN (delivered by either plasmid or Ad5 vector) based on gel electrophoresis of pol gene amplicons within ACH-2 and J-Lat cells. Ad-5-RGD vectors enhanced target mutagenesis of ZFN, relative to murCTSD_pZFN plasmids, to levels of CRISPR/Cas9 plasmids. Similar reduction of luciferase activity among TZM-bl treated with Ad5-ZFN vectors relative to CRISPR/Cas-9 and murCTSD_pZFN plasmids was observed on challenge with HIV-1. qRT-PCR of HIV-1 pol gene transcripts affirmed that Ad5 (RGD) vectors enhanced target mutagenesis of ZFN. Whereas CRISPR/Cas-9 may possess inherent superior target-mutagenesis efficiency; the efficiency of ZFN (off-target toxicity withstanding) can be enhanced by altering delivery vehicle from plasmid to Ad5 (RGD) vectors.

  6. Effects of sequence changes in the HIV-1 gp41 fusion peptide on CCR5 inhibitor resistance

    International Nuclear Information System (INIS)

    Anastassopoulou, Cleo G.; Ketas, Thomas J.; Sanders, Rogier W.; Johan Klasse, Per; Moore, John P.

    2012-01-01

    A rare pathway of HIV-1 resistance to small molecule CCR5 inhibitors such as Vicriviroc (VCV) involves changes solely in the gp41 fusion peptide (FP). Here, we show that the G516V change is critical to VCV resistance in PBMC and TZM-bl cells, although it must be accompanied by either M518V or F519I to have a substantial impact. Modeling VCV inhibition data from the two cell types indicated that G516V allows both double mutants to use VCV-CCR5 complexes for entry. The model further identified F519I as an independent determinant of preference for the unoccupied, high-VCV affinity form of CCR5. From inhibitor-free reversion cultures, we also identified a substitution in the inner domain of gp120, T244A, which appears to counter the resistance phenotype created by the FP substitutions. Examining the interplay of these changes will enhance our understanding of Env complex interactions that influence both HIV-1 entry and resistance to CCR5 inhibitors.

  7. Detailed Molecular Epidemiologic Characterization of HIV-1 Infection in Bulgaria Reveals Broad Diversity and Evolving Phylodynamics

    Science.gov (United States)

    Ivanov, Ivailo Alexiev; Beshkov, Danail; Shankar, Anupama; Hanson, Debra L.; Paraskevis, Dimitrios; Georgieva, Viara; Karamacheva, Lyudmila; Taskov, Hristo; Varleva, Tonka; Elenkov, Ivaylo; Stoicheva, Mariana; Nikolova, Daniela; Switzer, William M.

    2013-01-01

    Limited information is available to describe the molecular epidemiology of HIV-1 in Bulgaria. To better understand the genetic diversity and the epidemiologic dynamics of HIV-1 we analyzed 125 new polymerase (pol) sequences from Bulgarians diagnosed through 2009 and 77 pol sequences available from our previous study from persons infected prior to 2007. Epidemiologic and demographic information was obtained from each participant and phylogenetic analysis was used to infer HIV-1 evolutionary histories. 120 (59.5%) persons were infected with one of five different HIV-1 subtypes (A1, B, C, F1 and H) and 63 (31.2%) persons were infected with one of six different circulating recombinant forms (CRFs; 01_AE, 02_AG, 04_cpx, 05_DF, 14_BG, and 36_cpx). We also for the first time identified infection with two different clusters of unique A-like and F-like sub-subtype variants in 12 persons (5.9%) and seven unique recombinant forms (3.5%), including a novel J/C recombinant. While subtype B was the major genotype identified and was more prevalent in MSM and increased between 2000–2005, most non-B subtypes were present in persons ≥45 years old. CRF01_AE was the most common non-B subtype and was higher in women and IDUs relative to other risk groups combined. Our results show that HIV-1 infection in Bulgaria reflects the shifting distribution of genotypes coincident with the changing epidemiology of the HIV-1 epidemic among different risk groups. Our data support increased public health interventions targeting IDUs and MSM. Furthermore, the substantial and increasing HIV-1 genetic heterogeneity, combined with fluctuating infection dynamics, highlights the importance of sustained and expanded surveillance to prevent and control HIV-1 infection in Bulgaria. PMID:23527245

  8. Genetic Characterization of a Panel of Diverse HIV-1 Isolates at Seven International Sites.

    Directory of Open Access Journals (Sweden)

    Bhavna Hora

    Full Text Available HIV-1 subtypes and drug resistance are routinely tested by many international surveillance groups. However, results from different sites often vary. A systematic comparison of results from multiple sites is needed to determine whether a standardized protocol is required for consistent and accurate data analysis. A panel of well-characterized HIV-1 isolates (N = 50 from the External Quality Assurance Program Oversight Laboratory (EQAPOL was assembled for evaluation at seven international sites. This virus panel included seven subtypes, six circulating recombinant forms (CRFs, nine unique recombinant forms (URFs and three group O viruses. Seven viruses contained 10 major drug resistance mutations (DRMs. HIV-1 isolates were prepared at a concentration of 107 copies/ml and compiled into blinded panels. Subtypes and DRMs were determined with partial or full pol gene sequences by conventional Sanger sequencing and/or Next Generation Sequencing (NGS. Subtype and DRM results were reported and decoded for comparison with full-length genome sequences generated by EQAPOL. The partial pol gene was amplified by RT-PCR and sequenced for 89.4%-100% of group M viruses at six sites. Subtyping results of majority of the viruses (83%-97.9% were correctly determined for the partial pol sequences. All 10 major DRMs in seven isolates were detected at these six sites. The complete pol gene sequence was also obtained by NGS at one site. However, this method missed six group M viruses and sequences contained host chromosome fragments. Three group O viruses were only characterized with additional group O-specific RT-PCR primers employed by one site. These results indicate that PCR protocols and subtyping tools should be standardized to efficiently amplify diverse viruses and more consistently assign virus genotypes, which is critical for accurate global subtype and drug resistance surveillance. Targeted NGS analysis of partial pol sequences can serve as an alternative

  9. Long-term control of HIV-1 in hemophiliacs carrying slow-progressing allele HLA-B*5101.

    Science.gov (United States)

    Kawashima, Yuka; Kuse, Nozomi; Gatanaga, Hiroyuki; Naruto, Takuya; Fujiwara, Mamoru; Dohki, Sachi; Akahoshi, Tomohiro; Maenaka, Katsumi; Goulder, Philip; Oka, Shinichi; Takiguchi, Masafumi

    2010-07-01

    HLA-B*51 alleles are reported to be associated with slow disease progression to AIDS, but the mechanism underlying this association is still unclear. In the present study, we analyzed the effect of HLA-B*5101 on clinical outcome for Japanese hemophiliacs who had been infected with HIV-1 before 1985 and had been recruited in 1998 for this study. HLA-B*5101(+) hemophiliacs exhibited significantly slow progression. The analysis of HLA-B*5101-restricted HIV-1-specific cytotoxic T-lymphocyte (CTL) responses to 4 HLA-B*-restricted epitopes in 10 antiretroviral-therapy (ART)-free HLA-B*5101(+) hemophiliacs showed that the frequency of Pol283-8-specific CD8(+) T cells was inversely correlated with the viral load, whereas the frequencies of CD8(+) T cells specific for 3 other epitopes were positively correlated with the viral load. The HLA-B*5101(+) hemophiliacs whose HIV-1 replication had been controlled for approximately 25 years had HIV-1 possessing the wild-type Pol283-8 sequence or the Pol283-8V mutant, which does not critically affect T-cell recognition, whereas other HLA-B*5101(+) hemophiliacs had HIV-1 with escape mutations in this epitope. The results suggest that the control of HIV-1 over approximately 25 years in HLA-B*5101-positive hemophiliacs is associated with a Pol283-8-specific CD8(+) T-cell response and that lack of control of HIV-1 is associated with the appearance of Pol283-8-specific escape mutants.

  10. Long-Term Control of HIV-1 in Hemophiliacs Carrying Slow-Progressing Allele HLA-B*5101▿ †

    Science.gov (United States)

    Kawashima, Yuka; Kuse, Nozomi; Gatanaga, Hiroyuki; Naruto, Takuya; Fujiwara, Mamoru; Dohki, Sachi; Akahoshi, Tomohiro; Maenaka, Katsumi; Goulder, Philip; Oka, Shinichi; Takiguchi, Masafumi

    2010-01-01

    HLA-B*51 alleles are reported to be associated with slow disease progression to AIDS, but the mechanism underlying this association is still unclear. In the present study, we analyzed the effect of HLA-B*5101 on clinical outcome for Japanese hemophiliacs who had been infected with HIV-1 before 1985 and had been recruited in 1998 for this study. HLA-B*5101+ hemophiliacs exhibited significantly slow progression. The analysis of HLA-B*5101-restricted HIV-1-specific cytotoxic T-lymphocyte (CTL) responses to 4 HLA-B*-restricted epitopes in 10 antiretroviral-therapy (ART)-free HLA-B*5101+ hemophiliacs showed that the frequency of Pol283-8-specific CD8+ T cells was inversely correlated with the viral load, whereas the frequencies of CD8+ T cells specific for 3 other epitopes were positively correlated with the viral load. The HLA-B*5101+ hemophiliacs whose HIV-1 replication had been controlled for approximately 25 years had HIV-1 possessing the wild-type Pol283-8 sequence or the Pol283-8V mutant, which does not critically affect T-cell recognition, whereas other HLA-B*5101+ hemophiliacs had HIV-1 with escape mutations in this epitope. The results suggest that the control of HIV-1 over approximately 25 years in HLA-B*5101-positive hemophiliacs is associated with a Pol283-8-specific CD8+ T-cell response and that lack of control of HIV-1 is associated with the appearance of Pol283-8-specific escape mutants. PMID:20410273

  11. [Molecular epidemiological characteristics of HIV-1 strains isolated from newly diagnosed MSM subjects (2006-2010) in Beijing, China].

    Science.gov (United States)

    Ye, Jing-Rong; Zang, Wan-Chun; Su, Xue-Li; Lu, Hong-Yan; Hao, Ming-Qiang; Xin, Ruo-Lei; Chen, Guo-Min; He, Xiong; Zeng, Yi

    2014-03-01

    This study aims to analyze the molecular epidemiological characteristics of HIV-1 strains prevailing among men who have sex with men (MSM) in Beijing, China. The pol gene fragments from 250 newly diagnosed HIV-1-infected MSM individuals during 2006-2010 in Beijing were amplified by RT-nested PCR, sequenced, and phylogenetically analyzed. HIV-1 pol gene from 189 individuals were amplified and analyzed; 81 (42. 9%), 3 (1. 6%), 2 (1.0%), 88 (46. 6%), and 15 (7.9%) individuals were infected with HIV-1 subtypes B, B', C, CRF01_AE, and CRF07_BC, respectively. The subtypes B and CRF01_AE could both be grouped into two clusters, and CRFO7_BC strains shared high homology and were presumed to originate from a common ancestor. The HIV-1 circulating in MSM in Beijing had a lower genetic diversity than in heterosexuals. The HIV-1 epidemic (2006-2010) in MSM in Beijing was actually a rapid spread of HIV-1 CRF01 AE and B, or rather native strains of the two viruses.

  12. Molecular epidemiological analysis of env and pol sequences in newly diagnosed HIV type 1-infected, untreated patients in Hungary.

    Science.gov (United States)

    Mezei, Mária; Ay, Eva; Koroknai, Anita; Tóth, Renáta; Balázs, Andrea; Bakos, Agnes; Gyori, Zoltán; Bánáti, Ferenc; Marschalkó, Márta; Kárpáti, Sarolta; Minárovits, János

    2011-11-01

    The aim of our study was to monitor the diversity of HIV-1 strains circulating in Hungary and investigate the prevalence of resistance-associated mutations to reverse transcriptase (RT) and protease (PR) inhibitors in newly diagnosed, drug-naive patients. A total of 30 HIV-1-infected patients without prior antiretroviral treatment diagnosed during the period 2008-2010 were included into this study. Viral subtypes and the presence of RT, PR resistance-associated mutations were established by sequencing. Classification of HIV-1 strains showed that 29 (96.6%) patients were infected with subtype B viruses and one patient (3.3%) with subtype A virus. The prevalence of HIV-1 strains with transmitted drug resistance mutations in newly diagnosed individuals was 16.6% (5/30). This study showed that HIV-1 subtype B is still highly predominant in Hungary and documented a relatively high transmission rate of drug resistance in our country.

  13. Development of an HIV-1 Subtype Panel in China: Isolation and Characterization of 30 HIV-1 Primary Strains Circulating in China.

    Directory of Open Access Journals (Sweden)

    Jingwan Han

    Full Text Available The complex epidemic and significant diversity of HIV-1 strains in China pose serious challenges for surveillance and diagnostic assays, vaccine development and clinical management. There is a lack of HIV-1 isolates in current canonical HIV-1 subtype panels that can represent HIV-1 diversity in China; an HIV-1 subtype panel for China is urgently needed.Blood samples were collected from HIV-1 infected patients participating in the drug-resistance surveillance program in China. The samples were isolated, cultured and stored as neat culture supernatant. The HIV-1 isolates were fully characterized. The panel was used to compare 2 viral load assays and 2 p24 assays as the examples of how this panel could be used.An HIV-1 subtype panel for China composed of 30 HIV-1 primary strains of four subtypes (B [including Thai-B], CRF01_AE, CRF07_BC and G was established. The samples were isolated and cultured to a high-titer (10(6-10(9 copies/ml/high-volume (40 ml. The HIV-1 isolates were fully characterized by the final viral load, p24 concentration, gag-pol and envC2V3 sequencing, co-receptor prediction, determination of the four amino acids at the tip of the env V3-loop, glycosylation sites in the V3 loop and the drug-resistance mutations. The comparison of two p24 assays and two viral load assays on the isolates illustrated how this panel may be used for the evaluation of diagnostic assay performance. The Pearson value between p24 assays were 0.938. The viral load results showed excellent concordance and agreement for samples of Thai-B, but lower correlations for samples of CRF01_AE.The current panel of 30 HIV-1 isolates served as a basis for the development of a comprehensive panel of fully characterized viral isolates, which could reflect the current dynamic and complex HIV-1 epidemic in China. This panel will be available to support HIV-1 research, assay evaluation, vaccine and drug development.

  14. Evidence of at Least Two Introductions of HIV-1 in the Amerindian Warao Population from Venezuela

    Science.gov (United States)

    Rangel, Héctor R.; Maes, Mailis; Villalba, Julian; Sulbarán, Yoneira; de Waard, Jacobus H.; Bello, Gonzalo; Pujol, Flor H.

    2012-01-01

    Background The Venezuelan Amerindians were, until recently, free of human immunodeficiency virus (HIV) infection. However, in 2007, HIV-1 infection was detected for the first time in the Warao Amerindian population living in the Eastern part of Venezuela, in the delta of the Orinoco river. The aim of this study was to analyze the genetic diversity of the HIV-1 circulating in this population. Methodology/Principal Findings The pol genomic region was sequenced for 16 HIV-1 isolates and for some of them, sequences from env, vif and nef genomic regions were obtained. All HIV-1 isolates were classified as subtype B, with exception of one that was classified as subtype C. The 15 subtype B isolates exhibited a high degree of genetic similarity and formed a highly supported monophyletic cluster in each genomic region analyzed. Evolutionary analyses of the pol genomic region indicated that the date of the most recent common ancestor of the Waraos subtype B clade dates back to the late 1990s. Conclusions/Significance At least two independent introductions of HIV-1 have occurred in the Warao Amerindians from Venezuela. The HIV-1 subtype B was successfully established and got disseminated in the community, while no evidence of local dissemination of the HIV-1 subtype C was detected in this study. These results warrant further surveys to evaluate the burden of this disease, which can be particularly devastating in this Amerindian population, with a high prevalence of tuberculosis, hepatitis B, among other infectious diseases, and with limited access to primary health care. PMID:22808212

  15. A national study of the molecular epidemiology of HIV-1 in Australia 2005–2012

    Science.gov (United States)

    Castley, Alison; Sawleshwarkar, Shailendra; Varma, Rick; Herring, Belinda; Thapa, Kiran; Dwyer, Dominic; Chibo, Doris; Nguyen, Nam; Hawke, Karen; Ratcliff, Rodney; Garsia, Roger; Kelleher, Anthony; Nolan, David

    2017-01-01

    Introduction Rates of new HIV-1 diagnoses are increasing in Australia, with evidence of an increasing proportion of non-B HIV-1 subtypes reflecting a growing impact of migration and travel. The present study aims to define HIV-1 subtype diversity patterns and investigate possible HIV-1 transmission networks within Australia. Methods The Australian Molecular Epidemiology Network (AMEN) HIV collaborating sites in Western Australia, South Australia, Victoria, Queensland and western Sydney (New South Wales), provided baseline HIV-1 partial pol sequence, age and gender information for 4,873 patients who had genotypes performed during 2005–2012. HIV-1 phylogenetic analyses utilised MEGA V6, with a stringent classification of transmission pairs or clusters (bootstrap ≥98%, genetic distance ≤1.5% from at least one other sequence in the cluster). Results HIV-1 subtype B represented 74.5% of the 4,873 sequences (WA 59%, SA 68.4%, w-Syd 73.8%, Vic 75.6%, Qld 82.1%), with similar proportion of transmission pairs and clusters found in the B and non-B cohorts (23% vs 24.5% of sequences, p = 0.3). Significantly more subtype B clusters were comprised of ≥3 sequences compared with non-B clusters (45.0% vs 24.0%, p = 0.021) and significantly more subtype B pairs and clusters were male-only (88% compared to 53% CRF01_AE and 17% subtype C clusters). Factors associated with being in a cluster of any size included; being sequenced in a more recent time period (p3) was associated with being sequenced in a more recent time period (p = 0.05) and being male (p = 0.008). Conclusion This nationwide HIV-1 study of 4,873 patient sequences highlights the increased diversity of HIV-1 subtypes within the Australian epidemic, as well as differences in transmission networks associated with these HIV-1 subtypes. These findings provide epidemiological insights not readily available using standard surveillance methods and can inform the development of effective public health strategies in the

  16. A national study of the molecular epidemiology of HIV-1 in Australia 2005-2012.

    Directory of Open Access Journals (Sweden)

    Alison Castley

    Full Text Available Rates of new HIV-1 diagnoses are increasing in Australia, with evidence of an increasing proportion of non-B HIV-1 subtypes reflecting a growing impact of migration and travel. The present study aims to define HIV-1 subtype diversity patterns and investigate possible HIV-1 transmission networks within Australia.The Australian Molecular Epidemiology Network (AMEN HIV collaborating sites in Western Australia, South Australia, Victoria, Queensland and western Sydney (New South Wales, provided baseline HIV-1 partial pol sequence, age and gender information for 4,873 patients who had genotypes performed during 2005-2012. HIV-1 phylogenetic analyses utilised MEGA V6, with a stringent classification of transmission pairs or clusters (bootstrap ≥98%, genetic distance ≤1.5% from at least one other sequence in the cluster.HIV-1 subtype B represented 74.5% of the 4,873 sequences (WA 59%, SA 68.4%, w-Syd 73.8%, Vic 75.6%, Qld 82.1%, with similar proportion of transmission pairs and clusters found in the B and non-B cohorts (23% vs 24.5% of sequences, p = 0.3. Significantly more subtype B clusters were comprised of ≥3 sequences compared with non-B clusters (45.0% vs 24.0%, p = 0.021 and significantly more subtype B pairs and clusters were male-only (88% compared to 53% CRF01_AE and 17% subtype C clusters. Factors associated with being in a cluster of any size included; being sequenced in a more recent time period (p3 was associated with being sequenced in a more recent time period (p = 0.05 and being male (p = 0.008.This nationwide HIV-1 study of 4,873 patient sequences highlights the increased diversity of HIV-1 subtypes within the Australian epidemic, as well as differences in transmission networks associated with these HIV-1 subtypes. These findings provide epidemiological insights not readily available using standard surveillance methods and can inform the development of effective public health strategies in the current paradigm of HIV prevention

  17. A national study of the molecular epidemiology of HIV-1 in Australia 2005-2012.

    Science.gov (United States)

    Castley, Alison; Sawleshwarkar, Shailendra; Varma, Rick; Herring, Belinda; Thapa, Kiran; Dwyer, Dominic; Chibo, Doris; Nguyen, Nam; Hawke, Karen; Ratcliff, Rodney; Garsia, Roger; Kelleher, Anthony; Nolan, David

    2017-01-01

    Rates of new HIV-1 diagnoses are increasing in Australia, with evidence of an increasing proportion of non-B HIV-1 subtypes reflecting a growing impact of migration and travel. The present study aims to define HIV-1 subtype diversity patterns and investigate possible HIV-1 transmission networks within Australia. The Australian Molecular Epidemiology Network (AMEN) HIV collaborating sites in Western Australia, South Australia, Victoria, Queensland and western Sydney (New South Wales), provided baseline HIV-1 partial pol sequence, age and gender information for 4,873 patients who had genotypes performed during 2005-2012. HIV-1 phylogenetic analyses utilised MEGA V6, with a stringent classification of transmission pairs or clusters (bootstrap ≥98%, genetic distance ≤1.5% from at least one other sequence in the cluster). HIV-1 subtype B represented 74.5% of the 4,873 sequences (WA 59%, SA 68.4%, w-Syd 73.8%, Vic 75.6%, Qld 82.1%), with similar proportion of transmission pairs and clusters found in the B and non-B cohorts (23% vs 24.5% of sequences, p = 0.3). Significantly more subtype B clusters were comprised of ≥3 sequences compared with non-B clusters (45.0% vs 24.0%, p = 0.021) and significantly more subtype B pairs and clusters were male-only (88% compared to 53% CRF01_AE and 17% subtype C clusters). Factors associated with being in a cluster of any size included; being sequenced in a more recent time period (p3) was associated with being sequenced in a more recent time period (p = 0.05) and being male (p = 0.008). This nationwide HIV-1 study of 4,873 patient sequences highlights the increased diversity of HIV-1 subtypes within the Australian epidemic, as well as differences in transmission networks associated with these HIV-1 subtypes. These findings provide epidemiological insights not readily available using standard surveillance methods and can inform the development of effective public health strategies in the current paradigm of HIV prevention in

  18. Immune defence against HIV-1 infection in HIV-1-exposed seronegative persons.

    Science.gov (United States)

    Schmechel, S C; Russell, N; Hladik, F; Lang, J; Wilson, A; Ha, R; Desbien, A; McElrath, M J

    2001-11-01

    Rare individuals who are repeatedly exposed to HIV-1 through unprotected sexual contact fail to acquire HIV-1 infection. These persons represent a unique study population to evaluate mechanisms by which HIV-1 replication is either prevented or controlled. We followed longitudinally a group of healthy HIV-1 seronegative persons each reporting repeated high-risk sexual activities with their HIV-1-infected partner at enrollment. The volunteers were primarily (90%) male homosexuals, maintaining high risk activities with their known infected partner (45%) or multiple other partners (61%). We evaluated the quantity and specificity of HIV-1-specific T cells in 31 exposed seronegatives (ES) using a IFN-gamma ELISPOT assay to enumerate T cells recognizing epitopes within HIV-1 Env, Gag, Pol and Nef. PBMC from only three of the 31 volunteers demonstrated ex vivo HIV-1-specific IFN-gamma secretion, in contrast to nearly 30% exhibiting cytolytic responses in previous studies. These findings suggest that if T cell responses in ES are induced by HIV-1 exposure, the frequency is at low levels in most of them, and below the level of detection using the ELISPOT assay. Alternative approaches to improve the sensitivity of detection may include use of dendritic cells as antigen-presenting cells in the ex vivo assay and more careful definition of the risk behavior and extent of HIV-1 exposure in conjunction with the evaluation of T cell responses.

  19. HIV-1 transmission linkage in an HIV-1 prevention clinical trial

    Energy Technology Data Exchange (ETDEWEB)

    Leitner, Thomas [Los Alamos National Laboratory; Campbell, Mary S [UNIV OF WASHINGTON; Mullins, James I [UNIV OF WASHINGTON; Hughes, James P [UNIV OF WASHINGTON; Wong, Kim G [UNIV OF WASHINGTON; Raugi, Dana N [UNIV OF WASHINGTON; Scrensen, Stefanie [UNIV OF WASHINGTON

    2009-01-01

    HIV-1 sequencing has been used extensively in epidemiologic and forensic studies to investigate patterns of HIV-1 transmission. However, the criteria for establishing genetic linkage between HIV-1 strains in HIV-1 prevention trials have not been formalized. The Partners in Prevention HSV/HIV Transmission Study (ClinicaITrials.gov NCT00194519) enrolled 3408 HIV-1 serodiscordant heterosexual African couples to determine the efficacy of genital herpes suppression with acyclovir in reducing HIV-1 transmission. The trial analysis required laboratory confirmation of HIV-1 linkage between enrolled partners in couples in which seroconversion occurred. Here we describe the process and results from HIV-1 sequencing studies used to perform transmission linkage determination in this clinical trial. Consensus Sanger sequencing of env (C2-V3-C3) and gag (p17-p24) genes was performed on plasma HIV-1 RNA from both partners within 3 months of seroconversion; env single molecule or pyrosequencing was also performed in some cases. For linkage, we required monophyletic clustering between HIV-1 sequences in the transmitting and seroconverting partners, and developed a Bayesian algorithm using genetic distances to evaluate the posterior probability of linkage of participants sequences. Adjudicators classified transmissions as linked, unlinked, or indeterminate. Among 151 seroconversion events, we found 108 (71.5%) linked, 40 (26.5%) unlinked, and 3 (2.0%) to have indeterminate transmissions. Nine (8.3%) were linked by consensus gag sequencing only and 8 (7.4%) required deep sequencing of env. In this first use of HIV-1 sequencing to establish endpoints in a large clinical trial, more than one-fourth of transmissions were unlinked to the enrolled partner, illustrating the relevance of these methods in the design of future HIV-1 prevention trials in serodiscordant couples. A hierarchy of sequencing techniques, analysis methods, and expert adjudication contributed to the linkage

  20. Deep sequencing analysis of HIV-1 reverse transcriptase at baseline and time of failure in patients receiving rilpivirine in the phase III studies ECHO and THRIVE.

    Science.gov (United States)

    Van Eygen, Veerle; Thys, Kim; Van Hove, Carl; Rimsky, Laurence T; De Meyer, Sandra; Aerssens, Jeroen; Picchio, Gaston; Vingerhoets, Johan

    2016-05-01

    Minority variants (1.0-25.0%) were evaluated by deep sequencing (DS) at baseline and virological failure (VF) in a selection of antiretroviral treatment-naïve, HIV-1-infected patients from the rilpivirine ECHO/THRIVE phase III studies. Linkage between frequently emerging resistance-associated mutations (RAMs) was determined. DS (llIumina®) and population sequencing (PS) results were available at baseline for 47 VFs and time of failure for 48 VFs; and at baseline for 49 responders matched for baseline characteristics. Minority mutations were accurately detected at frequencies down to 1.2% of the HIV-1 quasispecies. No baseline minority rilpivirine RAMs were detected in VFs; one responder carried 1.9% F227C. Baseline minority mutations associated with resistance to other non-nucleoside reverse transcriptase inhibitors (NNRTIs) were detected in 8/47 VFs (17.0%) and 7/49 responders (14.3%). Baseline minority nucleoside/nucleotide reverse transcriptase inhibitor (NRTI) RAMs M184V and L210W were each detected in one VF (none in responders). At failure, two patients without NNRTI RAMs by PS carried minority rilpivirine RAMs K101E and/or E138K; and five additional patients carried other minority NNRTI RAMs V90I, V106I, V179I, V189I, and Y188H. Overall at failure, minority NNRTI RAMs and NRTI RAMs were found in 29/48 (60.4%) and 16/48 VFs (33.3%), respectively. Linkage analysis showed that E138K and K101E were usually not observed on the same viral genome. In conclusion, baseline minority rilpivirine RAMs and other NNRTI/NRTI RAMs were uncommon in the rilpivirine arm of the ECHO and THRIVE studies. DS at failure showed emerging NNRTI resistant minority variants in seven rilpivirine VFs who had no detectable NNRTI RAMs by PS. © 2015 Wiley Periodicals, Inc.

  1. IDEPI: rapid prediction of HIV-1 antibody epitopes and other phenotypic features from sequence data using a flexible machine learning platform.

    Directory of Open Access Journals (Sweden)

    N Lance Hepler

    2014-09-01

    Full Text Available Since its identification in 1983, HIV-1 has been the focus of a research effort unprecedented in scope and difficulty, whose ultimate goals--a cure and a vaccine--remain elusive. One of the fundamental challenges in accomplishing these goals is the tremendous genetic variability of the virus, with some genes differing at as many as 40% of nucleotide positions among circulating strains. Because of this, the genetic bases of many viral phenotypes, most notably the susceptibility to neutralization by a particular antibody, are difficult to identify computationally. Drawing upon open-source general-purpose machine learning algorithms and libraries, we have developed a software package IDEPI (IDentify EPItopes for learning genotype-to-phenotype predictive models from sequences with known phenotypes. IDEPI can apply learned models to classify sequences of unknown phenotypes, and also identify specific sequence features which contribute to a particular phenotype. We demonstrate that IDEPI achieves performance similar to or better than that of previously published approaches on four well-studied problems: finding the epitopes of broadly neutralizing antibodies (bNab, determining coreceptor tropism of the virus, identifying compartment-specific genetic signatures of the virus, and deducing drug-resistance associated mutations. The cross-platform Python source code (released under the GPL 3.0 license, documentation, issue tracking, and a pre-configured virtual machine for IDEPI can be found at https://github.com/veg/idepi.

  2. Reduced evolutionary rates in HIV-1 reveal extensive latency periods among replicating lineages.

    Science.gov (United States)

    Immonen, Taina T; Leitner, Thomas

    2014-10-16

    HIV-1 can persist for the duration of a patient's life due in part to its ability to hide from the immune system, and from antiretroviral drugs, in long-lived latent reservoirs. Latent forms of HIV-1 may also be disproportionally involved in transmission. Thus, it is important to detect and quantify latency in the HIV-1 life cycle. We developed a novel molecular clock-based phylogenetic tool to investigate the prevalence of HIV-1 lineages that have experienced latency. The method removes alternative sources that may affect evolutionary rates, such as hypermutation, recombination, and selection, to reveal the contribution of generation-time effects caused by latency. Our method was able to recover latent lineages with high specificity and sensitivity, and low false discovery rates, even on relatively short branches on simulated phylogenies. Applying the tool to HIV-1 sequences from 26 patients, we show that the majority of phylogenetic lineages have been affected by generation-time effects in every patient type, whether untreated, elite controller, or under effective or failing treatment. Furthermore, we discovered extensive effects of latency in sequence data (gag, pol, and env) from reservoirs as well as in the replicating plasma population. To better understand our phylogenetic findings, we developed a dynamic model of virus-host interactions to investigate the proportion of lineages in the actively replicating population that have ever been latent. Assuming neutral evolution, our dynamic modeling showed that under most parameter conditions, it is possible for a few activated latent viruses to propagate so that in time, most HIV-1 lineages will have been latent at some time in their past. These results suggest that cycling in and out of latency plays a major role in the evolution of HIV-1. Thus, no aspect of HIV-1 evolution can be fully understood without considering latency - including treatment, drug resistance, immune evasion, transmission, and pathogenesis.

  3. Geographic and temporal trends in the molecular epidemiology and genetic mechanisms of transmitted HIV-1 drug resistance: an individual-patient- and sequence-level meta-analysis.

    Science.gov (United States)

    Rhee, Soo-Yon; Blanco, Jose Luis; Jordan, Michael R; Taylor, Jonathan; Lemey, Philippe; Varghese, Vici; Hamers, Raph L; Bertagnolio, Silvia; Rinke de Wit, Tobias F; Aghokeng, Avelin F; Albert, Jan; Avi, Radko; Avila-Rios, Santiago; Bessong, Pascal O; Brooks, James I; Boucher, Charles A B; Brumme, Zabrina L; Busch, Michael P; Bussmann, Hermann; Chaix, Marie-Laure; Chin, Bum Sik; D'Aquin, Toni T; De Gascun, Cillian F; Derache, Anne; Descamps, Diane; Deshpande, Alaka K; Djoko, Cyrille F; Eshleman, Susan H; Fleury, Herve; Frange, Pierre; Fujisaki, Seiichiro; Harrigan, P Richard; Hattori, Junko; Holguin, Africa; Hunt, Gillian M; Ichimura, Hiroshi; Kaleebu, Pontiano; Katzenstein, David; Kiertiburanakul, Sasisopin; Kim, Jerome H; Kim, Sung Soon; Li, Yanpeng; Lutsar, Irja; Morris, Lynn; Ndembi, Nicaise; Ng, Kee Peng; Paranjape, Ramesh S; Peeters, Martine; Poljak, Mario; Price, Matt A; Ragonnet-Cronin, Manon L; Reyes-Terán, Gustavo; Rolland, Morgane; Sirivichayakul, Sunee; Smith, Davey M; Soares, Marcelo A; Soriano, Vincent V; Ssemwanga, Deogratius; Stanojevic, Maja; Stefani, Mariane A; Sugiura, Wataru; Sungkanuparph, Somnuek; Tanuri, Amilcar; Tee, Kok Keng; Truong, Hong-Ha M; van de Vijver, David A M C; Vidal, Nicole; Yang, Chunfu; Yang, Rongge; Yebra, Gonzalo; Ioannidis, John P A; Vandamme, Anne-Mieke; Shafer, Robert W

    2015-04-01

    Regional and subtype-specific mutational patterns of HIV-1 transmitted drug resistance (TDR) are essential for informing first-line antiretroviral (ARV) therapy guidelines and designing diagnostic assays for use in regions where standard genotypic resistance testing is not affordable. We sought to understand the molecular epidemiology of TDR and to identify the HIV-1 drug-resistance mutations responsible for TDR in different regions and virus subtypes. We reviewed all GenBank submissions of HIV-1 reverse transcriptase sequences with or without protease and identified 287 studies published between March 1, 2000, and December 31, 2013, with more than 25 recently or chronically infected ARV-naïve individuals. These studies comprised 50,870 individuals from 111 countries. Each set of study sequences was analyzed for phylogenetic clustering and the presence of 93 surveillance drug-resistance mutations (SDRMs). The median overall TDR prevalence in sub-Saharan Africa (SSA), south/southeast Asia (SSEA), upper-income Asian countries, Latin America/Caribbean, Europe, and North America was 2.8%, 2.9%, 5.6%, 7.6%, 9.4%, and 11.5%, respectively. In SSA, there was a yearly 1.09-fold (95% CI: 1.05-1.14) increase in odds of TDR since national ARV scale-up attributable to an increase in non-nucleoside reverse transcriptase inhibitor (NNRTI) resistance. The odds of NNRTI-associated TDR also increased in Latin America/Caribbean (odds ratio [OR] = 1.16; 95% CI: 1.06-1.25), North America (OR = 1.19; 95% CI: 1.12-1.26), Europe (OR = 1.07; 95% CI: 1.01-1.13), and upper-income Asian countries (OR = 1.33; 95% CI: 1.12-1.55). In SSEA, there was no significant change in the odds of TDR since national ARV scale-up (OR = 0.97; 95% CI: 0.92-1.02). An analysis limited to sequences with mixtures at less than 0.5% of their nucleotide positions—a proxy for recent infection—yielded trends comparable to those obtained using the complete dataset. Four NNRTI SDRMs—K101E, K103N, Y181C, and G190A

  4. HIV-1 transmission between MSM and heterosexuals, and increasing proportions of circulating recombinant forms in the Nordic Countries

    Science.gov (United States)

    Esbjörnsson, Joakim; Mild, Mattias; Audelin, Anne; Fonager, Jannik; Skar, Helena; Bruun Jørgensen, Louise; Liitsola, Kirsi; Björkman, Per; Bratt, Göran; Gisslén, Magnus; Sönnerborg, Anders; Nielsen, Claus; Medstrand, Patrik; Albert, Jan

    2016-01-01

    Increased knowledge about HIV-1 transmission dynamics in different transmission groups and geographical regions is fundamental for assessing and designing prevention efforts against HIV-1 spread. Since the first reported cases of HIV infection during the early 1980s, the HIV-1 epidemic in the Nordic countries has been dominated by HIV-1 subtype B and MSM transmission. HIV-1 pol sequences and clinical data of 51 per cent of all newly diagnosed HIV-1 infections in Sweden, Denmark, and Finland in the period 2000–2012 (N = 3,802) were analysed together with a large reference sequence dataset (N = 4,537) by trend analysis and phylogenetics. Analysis of the eight dominating subtypes and CRFs in the Nordic countries (A, B, C, D, G, CRF01_AE, CRF02_AG, and CRF06_cpx) showed that the subtype B proportion decreased while the CRF proportion increased over the study period. A majority (57 per cent) of the Nordic sequences formed transmission clusters, with evidence of mixing both geographically and between transmission groups. Detailed analyses showed multiple occasions of transmissions from MSM to heterosexuals and that active transmission clusters more often involved single than multiple Nordic countries. The strongest geographical link was between Denmark and Sweden. Finally, Denmark had a larger proportion of heterosexual domestic spread of HIV-1 subtype B (75 per cent) compared with Sweden (49 per cent) and Finland (57 per cent). We describe different HIV-1 transmission patterns between countries and transmission groups in a large geographical region. Our results may have implications for public health interventions in targeting HIV-1 transmission networks and identifying where to introduce such interventions. PMID:27774303

  5. gCUP: rapid GPU-based HIV-1 co-receptor usage prediction for next-generation sequencing.

    Science.gov (United States)

    Olejnik, Michael; Steuwer, Michel; Gorlatch, Sergei; Heider, Dominik

    2014-11-15

    Next-generation sequencing (NGS) has a large potential in HIV diagnostics, and genotypic prediction models have been developed and successfully tested in the recent years. However, albeit being highly accurate, these computational models lack computational efficiency to reach their full potential. In this study, we demonstrate the use of graphics processing units (GPUs) in combination with a computational prediction model for HIV tropism. Our new model named gCUP, parallelized and optimized for GPU, is highly accurate and can classify >175 000 sequences per second on an NVIDIA GeForce GTX 460. The computational efficiency of our new model is the next step to enable NGS technologies to reach clinical significance in HIV diagnostics. Moreover, our approach is not limited to HIV tropism prediction, but can also be easily adapted to other settings, e.g. drug resistance prediction. The source code can be downloaded at http://www.heiderlab.de d.heider@wz-straubing.de. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  6. Dynamics of Preferential Substrate Recognition in HIV-1 Protease: Redefining the Substrate Envelope

    Science.gov (United States)

    Özen, Ayşegül; Haliloğlu, Türkan; Schiffer, Celia A.

    2011-01-01

    HIV-1 protease (PR) permits viral maturation by processing the Gag and Gag-Pro-Pol polyproteins. Though HIV-1 PR inhibitors (PIs) are used in combination antiviral therapy, the emergence of drug resistance has limited their efficacy. The rapid evolution of HIV-1 necessitates the consideration of drug resistance in novel drug-design strategies. Drug-resistant HIV-1 PR variants, while no longer efficiently inhibited, continue to efficiently hydrolyze the natural viral substrates. Though highly diverse in sequence, the HIV-1 PR substrates bind in a conserved three-dimensional shape we defined as the “substrate envelope”. We previously showed that resistance mutations arise where PIs protrude beyond the substrate envelope, as these regions are crucial for drug binding but not for substrate recognition. Here, we extend this model by considering the role of protein dynamics in the interaction of HIV-1 PR with its substrates. Seven molecular dynamics simulations of PR-substrate complexes were performed to estimate the conformational flexibility of substrates in their complexes. Interdependency of the substrate-protease interactions may compensate for the variations in cleavage-site sequences, and explain how a diverse set of sequences can be recognized as substrates by the same enzyme. This diversity may be essential for regulating sequential processing of substrates. We also define a dynamic substrate envelope as a more accurate representation of PR-substrate interactions. This dynamic substrate envelope, described by a probability distribution function, is a powerful tool for drug design efforts targeting ensembles of resistant HIV-1 PR variants with the aim of developing drugs that are less susceptible to resistance. PMID:21762811

  7. An integrated genetic data environment (GDE)-based LINUX interface for analysis of HIV-1 and other microbial sequences.

    Science.gov (United States)

    De Oliveira, T; Miller, R; Tarin, M; Cassol, S

    2003-01-01

    Sequence databases encode a wealth of information needed to develop improved vaccination and treatment strategies for the control of HIV and other important pathogens. To facilitate effective utilization of these datasets, we developed a user-friendly GDE-based LINUX interface that reduces input/output file formatting. GDE was adapted to the Linux operating system, bioinformatics tools were integrated with microbe-specific databases, and up-to-date GDE menus were developed for several clinically important viral, bacterial and parasitic genomes. Each microbial interface was designed for local access and contains Genbank, BLAST-formatted and phylogenetic databases. GDE-Linux is available for research purposes by direct application to the corresponding author. Application-specific menus and support files can be downloaded from (http://www.bioafrica.net).

  8. Differential evolution of a CXCR4-using HIV-1 strain in CCR5wt/wt and CCR5∆32/∆32 hosts revealed by longitudinal deep sequencing and phylogenetic reconstruction.

    Science.gov (United States)

    Le, Anh Q; Taylor, Jeremy; Dong, Winnie; McCloskey, Rosemary; Woods, Conan; Danroth, Ryan; Hayashi, Kanna; Milloy, M-J; Poon, Art F Y; Brumme, Zabrina L

    2015-12-03

    Rare individuals homozygous for a naturally-occurring 32 base pair deletion in the CCR5 gene (CCR5∆32/∆32) are resistant to infection by CCR5-using ("R5") HIV-1 strains but remain susceptible to less common CXCR4-using ("X4") strains. The evolutionary dynamics of X4 infections however, remain incompletely understood. We identified two individuals, one CCR5wt/wt and one CCR5∆32/∆32, within the Vancouver Injection Drug Users Study who were infected with a genetically similar X4 HIV-1 strain. While early-stage plasma viral loads were comparable in the two individuals (~4.5-5 log10 HIV-1 RNA copies/ml), CD4 counts in the CCR5wt/wt individual reached a nadir of 250 cells/mm(3) in the CCR5∆32/∆32 individual. Ancestral phylogenetic reconstructions using longitudinal envelope-V3 deep sequences suggested that both individuals were infected by a single transmitted/founder (T/F) X4 virus that differed at only one V3 site (codon 24). While substantial within-host HIV-1 V3 diversification was observed in plasma and PBMC in both individuals, the CCR5wt/wt individual's HIV-1 population gradually reverted from 100% X4 to ~60% R5 over ~4 years whereas the CCR5∆32/∆32 individual's remained consistently X4. Our observations illuminate early dynamics of X4 HIV-1 infections and underscore the influence of CCR5 genotype on HIV-1 V3 evolution.

  9. Recent evolutionary history of human immunodeficiency virus type 1 subtype B: Reconstruction of epidemic onset based on sequence distances to the common ancestor

    NARCIS (Netherlands)

    Lukashov, Vladimir V.; Goudsmit, Jaap

    2002-01-01

    We obtained and studied HIV-1 sequences with a known sampling year from three outbreaks of the HIV-1 epidemic: 141 env V3 (270 nt) sampled between 1984 and 1992 and 117 pol prot/RT (804 nt) sequences sampled between 1986 and 1999 from Dutch homosexual men and injecting drug users (IDUs), as well as

  10. Molecular epidemiology of HIV-1 in Iceland: Early introductions, transmission dynamics and recent outbreaks among injection drug users.

    Science.gov (United States)

    Sallam, Malik; Esbjörnsson, Joakim; Baldvinsdóttir, Guðrún; Indriðason, Hlynur; Björnsdóttir, Thora Björg; Widell, Anders; Gottfreðsson, Magnús; Löve, Arthur; Medstrand, Patrik

    2017-04-01

    The molecular epidemiology of HIV-1 in Iceland has not been described so far. Detailed analyses of the dynamics of HIV-1 can give insights for prevention of virus spread. The objective of the current study was to characterize the genetic diversity and transmission dynamics of HIV-1 in Iceland. Partial HIV-1 pol (1020bp) sequences were generated from 230 Icelandic samples, representing 77% of all HIV-1 infected individuals reported in the country 1985-2012. Maximum likelihood phylogenies were reconstructed for subtype/CRF assignment and determination of transmission clusters. Timing and demographic growth patterns were determined in BEAST. HIV-1 infection in Iceland was dominated by subtype B (63%, n=145) followed by subtype C (10%, n=23), CRF01_AE (10%, n=22), sub-subtype A1 (7%, n=15) and CRF02_AG (7%, n=15). Trend analysis showed an increase in non-B subtypes/CRFs in Iceland over the study period (p=0.003). The highest proportion of phylogenetic clustering was found among injection drug users (IDUs; 89%), followed by heterosexuals (70%) and men who have sex with men (35%). The time to the most recent common ancestor of the oldest subtype B cluster dated back to 1978 (median estimate, 95% highest posterior density interval: 1974-1981) suggesting an early introduction of HIV-1 into Iceland. A previously reported increase in HIV-1 incidence among IDUs 2009-2011 was revealed to be due to two separate outbreaks. Our study showed that a variety of HIV-1 subtypes and CRFs were prevalent in Iceland 1985-2012, with subtype B being the dominant form both in terms of prevalence and domestic spread. The rapid increase of HIV-1 infections among IDUs following a major economic crisis in Iceland raises questions about casual associations between economic factors, drug use and public health. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. Cascade detection for the extraction of localized sequence features; specificity results for HIV-1 protease and structure-function results for the Schellman loop.

    Science.gov (United States)

    Newell, Nicholas E

    2011-12-15

    The extraction of the set of features most relevant to function from classified biological sequence sets is still a challenging problem. A central issue is the determination of expected counts for higher order features so that artifact features may be screened. Cascade detection (CD), a new algorithm for the extraction of localized features from sequence sets, is introduced. CD is a natural extension of the proportional modeling techniques used in contingency table analysis into the domain of feature detection. The algorithm is successfully tested on synthetic data and then applied to feature detection problems from two different domains to demonstrate its broad utility. An analysis of HIV-1 protease specificity reveals patterns of strong first-order features that group hydrophobic residues by side chain geometry and exhibit substantial symmetry about the cleavage site. Higher order results suggest that favorable cooperativity is weak by comparison and broadly distributed, but indicate possible synergies between negative charge and hydrophobicity in the substrate. Structure-function results for the Schellman loop, a helix-capping motif in proteins, contain strong first-order features and also show statistically significant cooperativities that provide new insights into the design of the motif. These include a new 'hydrophobic staple' and multiple amphipathic and electrostatic pair features. CD should prove useful not only for sequence analysis, but also for the detection of multifactor synergies in cross-classified data from clinical studies or other sources. Windows XP/7 application and data files available at: https://sites.google.com/site/cascadedetect/home. nacnewell@comcast.net Supplementary information is available at Bioinformatics online.

  12. Optimizing HIV-1 protease production in Escherichia coli as fusion protein

    Directory of Open Access Journals (Sweden)

    Piubelli Luciano

    2011-06-01

    Full Text Available Abstract Background Human immunodeficiency virus (HIV is the etiological agent in AIDS and related diseases. The aspartyl protease encoded by the 5' portion of the pol gene is responsible for proteolytic processing of the gag-pol polyprotein precursor to yield the mature capsid protein and the reverse transcriptase and integrase enzymes. The HIV protease (HIV-1Pr is considered an attractive target for designing inhibitors which could be used to tackle AIDS and therefore it is still the object of a number of investigations. Results A recombinant human immunodeficiency virus type 1 protease (HIV-1Pr was overexpressed in Escherichia coli cells as a fusion protein with bacterial periplasmic protein dithiol oxidase (DsbA or glutathione S-transferase (GST, also containing a six-histidine tag sequence. Protein expression was optimized by designing a suitable HIV-1Pr cDNA (for E. coli expression and to avoid autoproteolysis and by screening six different E. coli strains and five growth media. The best expression yields were achieved in E. coli BL21-Codon Plus(DE3-RIL host and in TB or M9 medium to which 1% (w/v glucose was added to minimize basal expression. Among the different parameters assayed, the presence of a buffer system (based on phosphate salts and a growth temperature of 37°C after adding IPTG played the main role in enhancing protease expression (up to 10 mg of chimeric DsbA:HIV-1Pr/L fermentation broth. GST:HIVPr was in part (50% produced as soluble protein while the overexpressed DsbA:HIV-1Pr chimeric protein largely accumulated in inclusion bodies as unprocessed fusion protein. A simple refolding procedure was developed on HiTrap Chelating column that yielded a refolded DsbA:HIV-1Pr with a > 80% recovery. Finally, enterokinase digestion of resolubilized DsbA:HIV-1Pr gave more than 2 mg of HIV-1Pr per liter of fermentation broth with a purity ≤ 80%, while PreScission protease cleavage of soluble GST:HIVPr yielded ~ 0.15 mg of pure HIV-1

  13. Viral linkage in HIV-1 seroconverters and their partners in an HIV-1 prevention clinical trial.

    Directory of Open Access Journals (Sweden)

    Mary S Campbell

    2011-03-01

    Full Text Available Characterization of viruses in HIV-1 transmission pairs will help identify biological determinants of infectiousness and evaluate candidate interventions to reduce transmission. Although HIV-1 sequencing is frequently used to substantiate linkage between newly HIV-1 infected individuals and their sexual partners in epidemiologic and forensic studies, viral sequencing is seldom applied in HIV-1 prevention trials. The Partners in Prevention HSV/HIV Transmission Study (ClinicalTrials.gov #NCT00194519 was a prospective randomized placebo-controlled trial that enrolled serodiscordant heterosexual couples to determine the efficacy of genital herpes suppression in reducing HIV-1 transmission; as part of the study analysis, HIV-1 sequences were examined for genetic linkage between seroconverters and their enrolled partners.We obtained partial consensus HIV-1 env and gag sequences from blood plasma for 151 transmission pairs and performed deep sequencing of env in some cases. We analyzed sequences with phylogenetic techniques and developed a Bayesian algorithm to evaluate the probability of linkage. For linkage, we required monophyletic clustering between enrolled partners' sequences and a Bayesian posterior probability of ≥ 50%. Adjudicators classified each seroconversion, finding 108 (71.5% linked, 40 (26.5% unlinked, and 3 (2.0% indeterminate transmissions, with linkage determined by consensus env sequencing in 91 (84%. Male seroconverters had a higher frequency of unlinked transmissions than female seroconverters. The likelihood of transmission from the enrolled partner was related to time on study, with increasing numbers of unlinked transmissions occurring after longer observation periods. Finally, baseline viral load was found to be significantly higher among linked transmitters.In this first use of HIV-1 sequencing to establish endpoints in a large clinical trial, more than one-fourth of transmissions were unlinked to the enrolled partner

  14. HIV-1 envelope glycoprotein

    Science.gov (United States)

    Caulfield, Michael; Cupo, Albert; Dean, Hansi; Hoffenberg, Simon; King, C. Richter; Klasse, P. J.; Marozsan, Andre; Moore, John P.; Sanders, Rogier W.; Ward, Andrew; Wilson, Ian; Julien, Jean-Philippe

    2017-08-22

    The present application relates to novel HIV-1 envelope glycoproteins, which may be utilized as HIV-1 vaccine immunogens, and antigens for crystallization, electron microscopy and other biophysical, biochemical and immunological studies for the identification of broad neutralizing antibodies. The present invention encompasses the preparation and purification of immunogenic compositions, which are formulated into the vaccines of the present invention.

  15. [Molecular epidemiological analysis of HIV-1 variants circulating in Russia in 1987-2015].

    Science.gov (United States)

    Lapovok, I A; Lopatukhin, A E; Kireev, D E; Kazennova, E V; Lebedev, A V; Bobkova, M R; Kolomeets, A N; Turbina, G I; Shipulin, G A; Ladnaya, N N; Pokrovsky, V V

    To simultaneously analyze HIV-1 samples from all Russian regions to characterize the epidemiology of HIV infection in the country as a whole. The most extensive study was conducted to examine nucleotide sequences of the pol gene of HIV-1 samples isolated from HIV-positive persons in different regions of Russia, with the diagnosis date being fixed during 1987-2015. The nucleotide sequences of the HIV-1 genome were analyzed using computer programs and on-line applications to identify a virus subtype and new recombinant forms. The nucleotide sequences of the pol gene were analyzed in 1697 HIV-1 samples and the findings were that the genetic variant subtype A1 (IDU-A) was dominant throughout the entire territory of Russia (in more than 80% of all infection cases). Other virus variants circulating in Russia were analyzed; the phenomenon of the higher distribution of the recombinant form CRF63/02A in Siberia, which had been previously described in the literature, was also confirmed. Four new recombinant forms generated by the virus subtype A1 (IDU-A) and B and two AG recombinant forms were found. There was a larger genetic distance between the viruses of IDU-A variant circulating among the injecting drug users and those infected through heterosexual contact, as well as a change in the viruses of subtype G that caused the outbreak in the south of the country over time in 1988-1989. The findings demonstrate continuous HIV-1 genetic variability and recombination over time in Russia, as well as increased genetic diversity with higher HIV infection rates in the population.

  16. Identifying HIV-1 dual infections

    Directory of Open Access Journals (Sweden)

    Cornelissen Marion

    2007-09-01

    Full Text Available Abstract Transmission of human immunodeficiency virus (HIV is no exception to the phenomenon that a second, productive infection with another strain of the same virus is feasible. Experiments with RNA viruses have suggested that both coinfections (simultaneous infection with two strains of a virus and superinfections (second infection after a specific immune response to the first infecting strain has developed can result in increased fitness of the viral population. Concerns about dual infections with HIV are increasing. First, the frequent detection of superinfections seems to indicate that it will be difficult to develop a prophylactic vaccine. Second, HIV-1 superinfections have been associated with accelerated disease progression, although this is not true for all persons. In fact, superinfections have even been detected in persons controlling their HIV infections without antiretroviral therapy. Third, dual infections can give rise to recombinant viruses, which are increasingly found in the HIV-1 epidemic. Recombinants could have increased fitness over the parental strains, as in vitro models suggest, and could exhibit increased pathogenicity. Multiple drug resistant (MDR strains could recombine to produce a pan-resistant, transmittable virus. We will describe in this review what is presently known about super- and re-infection among ambient viral infections, as well as the first cases of HIV-1 superinfection, including HIV-1 triple infections. The clinical implications, the impact of the immune system, and the effect of anti-retroviral therapy will be covered, as will as the timing of HIV superinfection. The methods used to detect HIV-1 dual infections will be discussed in detail. To increase the likelihood of detecting a dual HIV-1 infection, pre-selection of patients can be done by serotyping, heteroduplex mobility assays (HMA, counting the degenerate base codes in the HIV-1 genotyping sequence, or surveying unexpected increases in the

  17. [Analysis on HIV-1 genetics and threshold of drug resistance in Dehong prefecture of Yunnan province in 2013].

    Science.gov (United States)

    Ma, Yanling; Wang, Jibao; Xing, Hui; Chen, Min; Yao, Shitang; Chen, Huichao; Yang, Jin; Li, Yanling; Duan, Song; Jia, Manhong

    2015-06-01

    To study the HIV-1 genotypes and transmitted drug resistance (TDR) in Dehong prefecture of Yunnan province in 2013. Referring to the guidelines for HIV drug resistance threshold survey (HIVDR-TS), 54 plasma samples of recently reported HIV-infected individuals, aged between 16 and 25 years, were collected in Dehong prefecture from January to August 2013. Genotyping of partial pol gene was performed by using reverse transcriptional PCR. HIV-1 genotype. Prevalent levels of HIV-1 drug resistance transmission were analyzed. Forty-eight plasma samples were successfully sequenced and analyzed. Among them, 45.8% were Chinese and the rest 54.2% were all Burmese. Based on pol sequences, identified HIV genotypes included subtype C (41.7%), URF (31.3%), CRF01_AE (12.5%), CRF07_BC (10.4%), CRF08_BC (2.1%) and subtype B (2.1%), C subtype appeared dominated in Chinese while URF was dominated in Burmese. One drug resistant mutation to non-nucleoside reverse transcriptase inhibitors (NNRTIs) was detected in one sequence from Burmese. Based on the statistical method of HIVDR-TS, the prevalence of transmitted HIV-1 drug resistance was adjusted as scientific management for people living with HIV/AIDS should be strictly followed. Meanwhile, relevant surveillance, including drug resistance surveillance should also be performed among cross-border migrant population.

  18. Promoter2.0: for the recognition of PolII promoter sequences

    DEFF Research Database (Denmark)

    Knudsen, Steen; Knudsen, Steen

    1999-01-01

    Motivation : A new approach to the prediction of eukaryotic PolII promoters from DNA sequence takesadvantage of a combination of elements similar to neural networks and genetic algorithms to recognize a set ofdiscrete subpatterns with variable separation as one pattern: a promoter. The neural...... of optimization, the algorithm was able todiscriminate between vertebrate promoter and non-promoter sequences in a test set with a correlationcoefficient of 0.63. In addition, all five known transcription start sites on the plus strand of the completeadenovirus genome were within 161 bp of 35 predicted...

  19. The Epidemic History of HIV-1 CRF07_BC in Hetian Prefecture and the Role of It on HIV Spreading in China.

    Science.gov (United States)

    Wu, Jianjun; Guo, Hongxiong; Zhang, Jing; Liu, Xiaoming; Ayoupu, Aideaierli; Shen, Yuelan; Miao, Lifeng; Tang, Jihai; Lei, Yanhua; Su, Bin

    2017-04-01

    CRF07_BC is one of the most prevalent HIV-1 strains in China, and Xinjiang Uygur Autonomous Region has ever been considered to be a second epidemic center after Yunnan Province in previous studies. Here we use HIV-1 pol gene sequences identified from Hetian Prefecture located in Xinjiang Autonomous Region to reconstruct the epidemic history of HIV CRF07_BC strain circulating in this region. We found that CRF07_BC is the predominant HIV-1 form in Hetian Prefecture, and the estimated tMRCA analysis shows that there is no enough evidence supporting Xinjiang Autonomous Region as a second epidemic center of spreading HIV-1. It may imply that every city may be only a point among the HIV spreading network because of the frequent migration of population in the whole country nowadays.

  20. The sequence of the CA-SP1 junction accounts for the differential sensitivity of HIV-1 and SIV to the small molecule maturation inhibitor 3-O-{3',3'-dimethylsuccinyl}-betulinic acid

    Directory of Open Access Journals (Sweden)

    Aiken Christopher

    2004-06-01

    Full Text Available Abstract Background Despite the effectiveness of currently available antiretroviral therapies in the treatment of HIV-1 infection, a continuing need exists for novel compounds that can be used in combination with existing drugs to slow the emergence of drug-resistant viruses. We previously reported that the small molecule 3-O-{3',3'-dimethylsuccinyl}-betulinic acid (DSB specifically inhibits HIV-1 replication by delaying the processing of the CA-SP1 junction in Pr55Gag. By contrast, SIVmac239 replicates efficiently in the presence of high concentrations of DSB. To determine whether sequence differences in the CA-SP1 junction can fully account for the differential sensitivity of HIV-1 and SIV to DSB, we engineered mutations in this region of two viruses and tested their sensitivity to DSB in replication assays using activated human primary CD4+ T cells. Results Substitution of the P2 and P1 residues of HIV-1 by the corresponding amino acids of SIV resulted in strong resistance to DSB, but the mutant virus replicated with reduced efficiency. Conversely, replication of an SIV mutant containing three amino acid substitutions in the CA-SP1 cleavage site was highly sensitive to DSB, and the mutations resulted in delayed cleavage of the CA-SP1 junction in the presence of the drug. Conclusions These results demonstrate that the CA-SP1 junction in Pr55Gag represents the primary viral target of DSB. They further suggest that the therapeutic application of DSB will be accompanied by emergence of mutant viruses that are highly resistant to the drug but which exhibit reduced fitness relative to wild type HIV-1.

  1. HIV-1 Variants and Drug Resistance in Pregnant Women from Bata (Equatorial Guinea): 2012-2013.

    Science.gov (United States)

    Alvarez, Patricia; Fernández McPhee, Carolina; Prieto, Luis; Martín, Leticia; Obiang, Jacinta; Avedillo, Pedro; Vargas, Antonio; Rojo, Pablo; Benito, Agustín; Ramos, José Tomás; Holguín, África

    2016-01-01

    This is the first study describing drug resistance mutations (DRM) and HIV-1 variants among infected pregnant women in Equatorial Guinea (GQ), a country with high (6.2%) and increasing HIV prevalence. Dried blood spots (DBS) were collected from November 2012 to December 2013 from 69 HIV-1 infected women participating in a prevention of mother-to-child transmission program in the Hospital Regional of Bata and Primary Health Care Centre María Rafols, Bata, GQ. The transmitted (TDR) or acquired (ADR) antiretroviral drug resistance mutations at partial pol sequence among naive or antiretroviral therapy (ART)-exposed women were defined following WHO or IAS USA 2015 lists, respectively. HIV-1 variants were identified by phylogenetic analyses. A total of 38 of 69 HIV-1 specimens were successfully amplified and sequenced. Thirty (79%) belonged to ART-experienced women: 15 exposed to nucleoside reverse transcriptase inhibitors (NRTI) monotherapy, and 15 to combined ART (cART) as first regimen including two NRTI and one non-NRTI (NNRTI) or one protease inhibitor (PI). The TDR rate was only found for PI (3.4%). The ADR rate was 37.5% for NNRTI, 8.7% for NRTI and absent for PI or NRTI+NNRTI. HIV-1 group M non-B variants caused most (97.4%) infections, mainly (78.9%) recombinants: CRF02_AG (55.2%), CRF22_A101 (10.5%), subtype C (10.5%), unique recombinants (5.3%), and A3, D, F2, G, CRF06_cpx and CRF11_cpx (2.6% each). The high rate of ADR to retrotranscriptase inhibitors (mainly to NNRTIs) observed among pretreated pregnant women reinforces the importance of systematic DRM monitoring in GQ to reduce HIV-1 resistance transmission and to optimize first and second-line ART regimens when DRM are present.

  2. Reviewing the history of HIV-1: spread of subtype B in the Americas.

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    Dennis Maletich Junqueira

    Full Text Available The dispersal of HIV-1 subtype B (HIV-1B is a reflection of the movement of human populations in response to social, political, and geographical issues. The initial dissemination of HIV-1B outside Africa seems to have included the passive involvement of human populations from the Caribbean in spreading the virus to the United States. However, the exact pathways taken during the establishment of the pandemic in the Americas remain unclear. Here, we propose a geographical scenario for the dissemination of HIV-1B in the Americas, based on phylogenetic and genetic statistical analyses of 313 available sequences of the pol gene from 27 countries. Maximum likelihood and bayesian inference methods were used to explore the phylogenetic relationships between HIV-1B sequences, and molecular variance estimates were analyzed to infer the genetic structure of the viral population. We found that the initial dissemination and subsequent spread of subtype B in the Americas occurred via a single introduction event in the Caribbean around 1964 (1950-1967. Phylogenetic trees present evidence of several primary outbreaks in countries in South America, directly seeded by the Caribbean epidemic. Cuba is an exception insofar as its epidemic seems to have been introduced from South America. One clade comprising isolates from different countries emerged in the most-derived branches, reflecting the intense circulation of the virus throughout the American continents. Statistical analysis supports the genetic compartmentalization of the virus among the Americas, with a close relationship between the South American and Caribbean epidemics. These findings reflect the complex establishment of the HIV-1B pandemic and contribute to our understanding between the migration process of human populations and virus diffusion.

  3. The Genetic Diversity and Evolution of HIV-1 Subtype B Epidemic in Puerto Rico

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    Pablo López

    2015-12-01

    Full Text Available HIV-1 epidemics in Caribbean countries, including Puerto Rico, have been reported to be almost exclusively associated with the subtype B virus (HIV-1B. However, while HIV infections associated with other clades have been only sporadically reported, no organized data exist to accurately assess the prevalence of non-subtype B HIV-1 infection. We analyzed the nucleotide sequence data of the HIV pol gene associated with HIV isolates from Puerto Rican patients. The sequences (n = 945 were obtained from our “HIV Genotyping” test file, which has been generated over a period of 14 years (2001–2014. REGA subtyping tool found the following subtypes: B (90%, B-like (3%, B/D recombinant (6%, and D/B recombinant (0.6%. Though there were fewer cases, the following subtypes were also found (in the given proportions: A1B (0.3%, BF1 (0.2%, subtype A (01-AE (0.1%, subtype A (A2 (0.1%, subtype F (12BF (0.1%, CRF-39 BF-like (0.1%, and others (0.1%. Some of the recombinants were identified as early as 2001. Although the HIV epidemic in Puerto Rico is primarily associated with HIV-1B virus, our analysis uncovered the presence of other subtypes. There was no indication of subtype C, which has been predominantly associated with heterosexual transmission in other parts of the world.

  4. The Genetic Diversity and Evolution of HIV-1 Subtype B Epidemic in Puerto Rico.

    Science.gov (United States)

    López, Pablo; Rivera-Amill, Vanessa; Rodríguez, Nayra; Vargas, Freddie; Yamamura, Yasuhiro

    2015-12-23

    HIV-1 epidemics in Caribbean countries, including Puerto Rico, have been reported to be almost exclusively associated with the subtype B virus (HIV-1B). However, while HIV infections associated with other clades have been only sporadically reported, no organized data exist to accurately assess the prevalence of non-subtype B HIV-1 infection. We analyzed the nucleotide sequence data of the HIV pol gene associated with HIV isolates from Puerto Rican patients. The sequences (n = 945) were obtained from our "HIV Genotyping" test file, which has been generated over a period of 14 years (2001-2014). REGA subtyping tool found the following subtypes: B (90%), B-like (3%), B/D recombinant (6%), and D/B recombinant (0.6%). Though there were fewer cases, the following subtypes were also found (in the given proportions): A1B (0.3%), BF1 (0.2%), subtype A (01-AE) (0.1%), subtype A (A2) (0.1%), subtype F (12BF) (0.1%), CRF-39 BF-like (0.1%), and others (0.1%). Some of the recombinants were identified as early as 2001. Although the HIV epidemic in Puerto Rico is primarily associated with HIV-1B virus, our analysis uncovered the presence of other subtypes. There was no indication of subtype C, which has been predominantly associated with heterosexual transmission in other parts of the world.

  5. Prevalence of drug resistance mutations and non-B subtypes in newly diagnosed HIV-1 patients in Denmark

    DEFF Research Database (Denmark)

    Jørgensen, Louise B; Christensen, Marianne B; Gerstoft, Jan

    2003-01-01

    The aim of this study was to monitor the prevalence of drug resistance mutations in newly diagnosed HIV-1 positive individuals in Denmark. In addition we assessed the prevalence of non-B subtypes based on phylogenetic analysis of the pol gene. Plasma samples from 104 newly diagnosed HIV-1 positive...... patients were obtained in the year 2000. The entire protease gene and 320 amino acids of the reverse transcriptase gene were genotyped. Sequences were obtained from 97 patients. No subjects displayed primary resistance mutations in the protease gene, whereas all carried 1 or more secondary mutations....... Resistance mutations in the RT-gene associated with NRTI-resistance were found in 1 patient, who was infected with zidovudine resistant HIV-1 harbouring the M41L mutation in combination with T215S and L210S. The T215S mutation has been showed to be associated with reversion of zidovudine resistance. The T215...

  6. Geographic and temporal trends in the molecular epidemiology and genetic mechanisms of transmitted HIV-1 drug resistance: an individual-patient- and sequence-level meta-analysis

    NARCIS (Netherlands)

    Rhee, Soo-Yon; Blanco, Jose Luis; Jordan, Michael R.; Taylor, Jonathan; Lemey, Philippe; Varghese, Vici; Hamers, Raph L.; Bertagnolio, Silvia; Rinke de Wit, Tobias F.; Aghokeng, Avelin F.; Albert, Jan; Avi, Radko; Avila-Rios, Santiago; Bessong, Pascal O.; Brooks, James I.; Boucher, Charles A. B.; Brumme, Zabrina L.; Busch, Michael P.; Bussmann, Hermann; Chaix, Marie-Laure; Chin, Bum Sik; D'Aquin, Toni T.; de Gascun, Cillian F.; Derache, Anne; Descamps, Diane; Deshpande, Alaka K.; Djoko, Cyrille F.; Eshleman, Susan H.; Fleury, Herve; Frange, Pierre; Fujisaki, Seiichiro; Harrigan, P. Richard; Hattori, Junko; Holguin, Africa; Hunt, Gillian M.; Ichimura, Hiroshi; Kaleebu, Pontiano; Katzenstein, David; Kiertiburanakul, Sasisopin; Kim, Jerome H.; Kim, Sung Soon; Li, Yanpeng; Lutsar, Irja; Morris, Lynn; Ndembi, Nicaise; Ng, Kee Peng; Paranjape, Ramesh S.; Peeters, Martine; Poljak, Mario; Price, Matt A.; Ragonnet-Cronin, Manon L.; Reyes-Terán, Gustavo; Rolland, Morgane; Sirivichayakul, Sunee; Smith, Davey M.; Soares, Marcelo A.; Soriano, Vincent V.; Ssemwanga, Deogratius; Stanojevic, Maja; Stefani, Mariane A.; Sugiura, Wataru; Sungkanuparph, Somnuek; Tanuri, Amilcar; tee, Kok Keng; Truong, Hong-Ha M.; van de Vijver, David A. M. C.; Vidal, Nicole; Yang, Chunfu; Yang, Rongge; Yebra, Gonzalo; Ioannidis, John P. A.; Vandamme, Anne-Mieke; Shafer, Robert W.

    2015-01-01

    Regional and subtype-specific mutational patterns of HIV-1 transmitted drug resistance (TDR) are essential for informing first-line antiretroviral (ARV) therapy guidelines and designing diagnostic assays for use in regions where standard genotypic resistance testing is not affordable. We sought to

  7. Geographic and Temporal Trends in the Molecular Epidemiology and Genetic Mechanisms of Transmitted HIV-1 Drug Resistance: An Individual-Patient- and Sequence-Level Meta-Analysis

    NARCIS (Netherlands)

    S.Y. Rhee (Soo Yoon); J.L. Blanco (Jose Luis); M.R. Jordan (Michael); J. Taylor (Jonathan); P. Lemey (Philippe); V. Varghese (Vici); R.L. Hamers (Raph); S. Bertagnolio (Silvia); M. De Wit (Meike); A.F. Aghokeng (Avelin); J. Albert (Jan); R. Avi (Radko); S. Avila-Rios (Santiago); P.O. Bessong (Pascal O.); J.I. Brooks (James I.); C.A.B. Boucher (Charles); Z.L. Brumme (Zabrina L.); M.P. Busch (Michael P.); H. Bussmann (Hermann); M.L. Chaix (Marie Laure); B.S. Chin (Bum Sik); T.T. D’Aquin (Toni T.); C. de Gascun (Cillian); A. Derache (Anne); D. Descamps (Diane); A.K. Deshpande (Alaka K.); C.F. Djoko (Cyrille F.); S.H. Eshleman (Susan H.); H. Fleury (Hervé); P. Frange (Pierre); S. Fujisaki (Seiichiro); P. Harrigan (Pr); J. Hattori (Junko); A. Holguin (Africa); G.M. Hunt (Gillian M.); H. Ichimura (Hiroshi); P. Kaleebu (Pontiano); D. Katzenstein (David); S. Kiertiburanakul (Sasisopin); J.H. Kim (Jerome H.); S.S. Kim (Sung Soon); Y. Li (Yanpeng); I. Lutsar (Irja); L. Morris (L.); N. Ndembi (Nicaise); K.P. NG (Kee Peng); R.S. Paranjape (Ramesh S.); M.C. Peeters (Marian); M. Poljak (Mario); M.A. Price (Matt A.); M.L. Ragonnet-Cronin (Manon L.); G. Reyes-Terán (Gustavo); M. Rolland (Morgane); S. Sirivichayakul (Sunee); D.M. Smith (Davey M.); M.A. Soares (Marcelo A.); V. Soriano (Virtudes); D. Ssemwanga (Deogratius); M. Stanojevic (Maja); M.A. Stefani (Mariane A.); W. Sugiura (Wataru); S. Sungkanuparph (Somnuek); A. Tanuri (Amilcar); K.K. Tee (Kok Keng); H.-H.M. Truong (Hong-Ha M.); D.A.M.C. van de Vijver (David); N. Vidal (Nicole); C. Yang (Chunfu); R. Yang (Rongge); G. Yebra (Gonzalo); J.P.A. Ioannidis (John); A.M. Vandamme (Anne Mieke); R.W. Shafer (Robert)

    2015-01-01

    textabstractRegional and subtype-specific mutational patterns of HIV-1 transmitted drug resistance (TDR) are essential for informing first-line antiretroviral (ARV) therapy guidelines and designing diagnostic assays for use in regions where standard genotypic resistance testing is not affordable. We

  8. Sequence similarity between the erythrocyte binding domain 1 of the Plasmodium vivax Duffy binding protein and the V3 loop of HIV-1 strain MN reveals binding residues for the Duffy Antigen Receptor for Chemokines

    OpenAIRE

    Bolton, Michael J; Garry, Robert F

    2011-01-01

    Abstract Background The surface glycoprotein (SU, gp120) of the human immunodeficiency virus (HIV) must bind to a chemokine receptor, CCR5 or CXCR4, to invade CD4+ cells. Plasmodium vivax uses the Duffy Binding Protein (DBP) to bind the Duffy Antigen Receptor for Chemokines (DARC) and invade reticulocytes. Results Variable loop 3 (V3) of HIV-1 SU and domain 1 of the Plasmodium vivax DBP share a sequence similarity. The site of amino acid sequence similarity was necessary, but not sufficient, ...

  9. Molecular epidemiology of HIV-1 among the HIV infected people of Manipur, Northeastern India: Emergence of unique recombinant forms.

    Science.gov (United States)

    Sharma, Adhikarimayum Lakhikumar; Singh, Thiyam Ramsing; Devi, Khuraijam Ranjana; Singh, Lisam Shanjukumar

    2017-06-01

    According to the Joint National Programme on HIV/AIDS (UNAIDS), the northeastern region of India has the highest HIV prevalence in the country. This study was conducted to determine the current HIV-1 molecular epidemiology of Manipur, a state in northeast India. Blood samples from HIV-1 seropositive subjects were collected between June 2011 and February 2014. The partial regions of HIV-1 genes; pol and tat-vpu-env were independently amplified, sequenced, analyzed, and genotyped. Based on all sequences generated from 110 samples using pol and/or tat-vpu-env gene, the overall HIV-1 genotypes distribution of Manipur was as follows: 65.45% (72/110) subtype C, 32.73% (36/110) unique recombinant forms (URFs), and 1.82% (2/110) subtype B. The distribution of HIV-1 genotypes among the risk groups was: heterosexual: 58.33% (35/60) subtype C, 38.33% (23/60) URFs, and 3.34% (2/60) subtype B; intravenous drug users (IDUs): 85.36% (35/41) subtype C, 9.76% (4/41) URFs, and 4.88% (2/41) subtype B; mother to child (MTC): 50% (3/6) URFs and 50% (3/6) subtype C and blood transfusion: 100% (3/3) subtype C. The findings for the first time revealed the emergence of URFs of HIV-1 in Manipur which is predominant among the sexual and MTC risk groups as compared to IDUs. Taking together, this study illustrated that Manipur is the "recombinant hotspot of HIV" of India. The results will provide the clinical importance for continuous monitoring of HIV-infections in order to design appropriate prevention measures to limit the spread of new HIV infections. © 2016 Wiley Periodicals, Inc.

  10. Acyclovir and Transmission of HIV-1 from Persons Infected with HIV-1 and HSV-2

    Science.gov (United States)

    Celum, Connie; Wald, Anna; Lingappa, Jairam R.; Magaret, Amalia S.; Wang, Richard S.; Mugo, Nelly; Mujugira, Andrew; Baeten, Jared M.; Mullins, James I.; Hughes, James P.; Bukusi, Elizabeth A.; Cohen, Craig R.; Katabira, Elly; Ronald, Allan; Kiarie, James; Farquhar, Carey; Stewart, Grace John; Makhema, Joseph; Essex, Myron; Were, Edwin; Fife, Kenneth H.; de Bruyn, Guy; Gray, Glenda E.; McIntyre, James A.; Manongi, Rachel; Kapiga, Saidi; Coetzee, David; Allen, Susan; Inambao, Mubiana; Kayitenkore, Kayitesi; Karita, Etienne; Kanweka, William; Delany, Sinead; Rees, Helen; Vwalika, Bellington; Stevens, Wendy; Campbell, Mary S.; Thomas, Katherine K.; Coombs, Robert W.; Morrow, Rhoda; Whittington, William L.H.; McElrath, M. Juliana; Barnes, Linda; Ridzon, Renee; Corey, Lawrence

    2010-01-01

    BACKGROUND Most persons who are infected with human immunodeficiency virus type 1 (HIV-1) are also infected with herpes simplex virus type 2 (HSV-2), which is frequently reactivated and is associated with increased plasma and genital levels of HIV-1. Therapy to suppress HSV-2 reduces the frequency of reactivation of HSV-2 as well as HIV-1 levels, suggesting that suppression of HSV-2 may reduce the risk of transmission of HIV-1. METHODS We conducted a randomized, placebo-controlled trial of suppressive therapy for HSV-2 (acyclovir at a dose of 400 mg orally twice daily) in couples in which only one of the partners was seropositive for HIV-1 (CD4 count, ≥250 cells per cubic millimeter) and that partner was also infected with HSV-2 and was not taking antiretroviral therapy at the time of enrollment. The primary end point was transmission of HIV-1 to the partner who was not initially infected with HIV-1; linkage of transmissions was assessed by means of genetic sequencing of viruses. RESULTS A total of 3408 couples were enrolled at 14 sites in Africa. Of the partners who were infected with HIV-1, 68% were women, and the baseline median CD4 count was 462 cells per cubic millimeter. Of 132 HIV-1 seroconversions that occurred after randomization (an incidence of 2.7 per 100 person-years), 84 were linked within couples by viral sequencing: 41 in the acyclovir group and 43 in the placebo group (hazard ratio with acyclovir, 0.92, 95% confidence interval [CI], 0.60 to 1.41; P = 0.69). Suppression with acyclovir reduced the mean plasma concentration of HIV-1 by 0.25 log10 copies per milliliter (95% CI, 0.22 to 0.29; P<0.001) and the occurrence of HSV-2–positive genital ulcers by 73% (risk ratio, 0.27; 95% CI, 0.20 to 0.36; P<0.001). A total of 92% of the partners infected with HIV-1 and 84% of the partners not infected with HIV-1 remained in the study for 24 months. The level of adherence to the dispensed study drug was 96%. No serious adverse events related to acyclovir

  11. A HIV-1 heterosexual transmission chain in Guangzhou, China: a molecular epidemiological study.

    Science.gov (United States)

    Han, Zhigang; Leung, Tommy W C; Zhao, Jinkou; Wang, Ming; Fan, Lirui; Li, Kai; Pang, Xinli; Liang, Zhenbo; Lim, Wilina W L; Xu, Huifang

    2009-09-25

    We conducted molecular analyses to confirm four clustering HIV-1 infections (Patient A, B, C & D) in Guangzhou, China. These cases were identified by epidemiological investigation and suspected to acquire the infection through a common heterosexual transmission chain. Env C2V3V4 region, gag p17/p24 junction and partial pol gene of HIV-1 genome from serum specimens of these infected cases were amplified by reverse transcription polymerase chain reaction (RT-PCR) and nucleotide sequenced. Phylogenetic analyses indicated that their viral nucleotide sequences were significantly clustered together (bootstrap value is 99%, 98% and 100% in env, gag and pol tree respectively). Evolutionary distance analysis indicated that their genetic diversities of env, gag and pol genes were significantly lower than non-clustered controls, as measured by unpaired t-test (env gene comparison: p Epidemiological results and molecular analyses consistently illustrated these four cases represented a transmission chain which dispersed in the locality through heterosexual contact involving commercial sex worker.

  12. Short Communication: Reassessing the Origin of the HIV-1 CRF02_AG Lineages Circulating in Brazil.

    Science.gov (United States)

    Delatorre, Edson; Velasco-De-Castro, Carlos A; Pilotto, José H; Couto-Fernandez, José Carlos; Bello, Gonzalo; Morgado, Mariza G

    2015-12-01

    HIV-1 CRF02_AG is responsible for at least 8% of the HIV-1 infections worldwide and is distributed mainly in West Africa. CRF02_AG has recently been reported in countries where it is not native, including Brazil. In a previous study including 10 CRF02_AG Brazilian samples, we found at least four independent introductions and two autochthonous transmission networks of this clade in Brazil. As more CRF02_AG samples have been identified in Brazil, we performed a new phylogeographic analysis using a larger dataset than before. A total of 20 Brazilian (18 from Rio de Janeiro and two from São Paulo) and 1,485 African HIV-1 CRF02_AG pol sequences were analyzed using maximum likelihood (ML). The ML tree showed that the Brazilian sequences were distributed in five different lineages. The Bayesian phylogeographic analysis of the Brazilian and their most closely related African sequences (n = 212) placed the origin of all Brazilian lineages in West Africa, probably Ghana, Senegal, and Nigeria. Two monophyletic clades were identified, comprising only sequences from Rio de Janeiro, and their date of origin was estimated at around 1985 (95% highest posterior density: 1979-1992). These results support the existence of at least five independent introductions of the CRF02_AG lineage from West Africa into Brazil and further indicate that at least two of these lineages have been locally disseminated in the Rio de Janeiro state over the past 30 years.

  13. Mutations Related to Antiretroviral Resistance Identified by Ultra-Deep Sequencing in HIV-1 Infected Children under Structured Interruptions of HAART.

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    Jose Manuel Vazquez-Guillen

    Full Text Available Although Structured Treatment Interruptions (STI are currently not considered an alternative strategy for antiretroviral treatment, their true benefits and limitations have not been fully established. Some studies suggest the possibility of improving the quality of life of patients with this strategy; however, the information that has been obtained corresponds mostly to studies conducted in adults, with a lack of knowledge about its impact on children. Furthermore, mutations associated with antiretroviral resistance could be selected due to sub-therapeutic levels of HAART at each interruption period. Genotyping methods to determine the resistance profiles of the infecting viruses have become increasingly important for the management of patients under STI, thus low-abundance antiretroviral drug-resistant mutations (DRM's at levels under limit of detection of conventional genotyping (<20% of quasispecies could increase the risk of virologic failure. In this work, we analyzed the protease and reverse transcriptase regions of the pol gene by ultra-deep sequencing in pediatric patients under STI with the aim of determining the presence of high- and low-abundance DRM's in the viral rebounds generated by the STI. High-abundance mutations in protease and high- and low-abundance mutations in reverse transcriptase were detected but no one of these are directly associated with resistance to antiretroviral drugs. The results could suggest that the evaluated STI program is virologically safe, but strict and carefully planned studies, with greater numbers of patients and interruption/restart cycles, are still needed to evaluate the selection of DRM's during STI.

  14. Generation and Characterization of a Defective HIV-1 Virus as an Immunogen for a Therapeutic Vaccine

    Science.gov (United States)

    García-Pérez, Javier; García, Felipe; Blanco, Julia; Escribà-García, Laura; Gatell, Jose Maria; Alcamí, Jose; Plana, Montserrat; Sánchez-Palomino, Sonsoles

    2012-01-01

    Background The generation of new immunogens able to elicit strong specific immune responses remains a major challenge in the attempts to obtain a prophylactic or therapeutic vaccine against HIV/AIDS. We designed and constructed a defective recombinant virus based on the HIV-1 genome generating infective but non-replicative virions able to elicit broad and strong cellular immune responses in HIV-1 seropositive individuals. Results Viral particles were generated through transient transfection in producer cells (293-T) of a full length HIV-1 DNA carrying a deletion of 892 base pairs (bp) in the pol gene encompassing the sequence that codes for the reverse transcriptase (NL4-3/ΔRT clone). The viral particles generated were able to enter target cells, but due to the absence of reverse transcriptase no replication was detected. The immunogenic capacity of these particles was assessed by ELISPOT to determine γ-interferon production in a cohort of 69 chronic asymptomatic HIV-1 seropositive individuals. Surprisingly, defective particles produced from NL4-3/ΔRT triggered stronger cellular responses than wild-type HIV-1 viruses inactivated with Aldrithiol-2 (AT-2) and in a larger proportion of individuals (55% versus 23% seropositive individuals tested). Electron microscopy showed that NL4-3/ΔRT virions display immature morphology. Interestingly, wild-type viruses treated with Amprenavir (APV) to induce defective core maturation also induced stronger responses than the same viral particles generated in the absence of protease inhibitors. Conclusions We propose that immature HIV-1 virions generated from NL4-3/ΔRT viral clones may represent new prototypes of immunogens with a safer profile and stronger capacity to induce cellular immune responses than wild-type inactivated viral particles. PMID:23144996

  15. HIV-1 subtypes B and C unique recombinant forms (URFs and transmitted drug resistance identified in the Western Cape Province, South Africa.

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    Graeme Brendon Jacobs

    Full Text Available South Africa has the largest worldwide HIV/AIDS population with 5.6 million people infected and at least 2 million people on antiretroviral therapy. The majority of these infections are caused by HIV-1 subtype C. Using genotyping methods we characterized HIV-1 subtypes of the gag p24 and pol PR and RT fragments, from a cohort of female participants in the Western Cape Province, South Africa. These participants were recruited as part of a study to assess the combined brain and behavioural effects of HIV and early childhood trauma. The partial HIV-1 gag and pol fragments of 84 participants were amplified by PCR and sequenced. Different online tools and manual phylogenetic analysis were used for HIV-1 subtyping. Online tools included: REGA HIV Subtyping tool version 3; Recombinant Identification Program (RIP; Context-based Modeling for Expeditious Typing (COMET; jumping profile Hidden Markov Models (jpHMM webserver; and subtype classification using evolutionary algorithms (SCUEAL. HIV-1 subtype C predominates within the cohort with a prevalence of 93.8%. We also show, for the first time, the presence of circulating BC strains in at least 4.6% of our study cohort. In addition, we detected transmitted resistance associated mutations in 4.6% of analysed sequences. With tourism and migration rates to South Africa currently very high, we are detecting more and more HIV-1 URFs within our study populations. It is still unclear what role these unique strains will play in terms of long term antiretroviral treatment and what challenges they will pose to vaccine development. Nevertheless, it remains vitally important to monitor the HIV-1 diversity in South Africa and worldwide as the face of the epidemic is continually changing.

  16. Proviral HIV-genome-wide and pol-gene specific zinc finger nucleases: usability for targeted HIV gene therapy.

    Science.gov (United States)

    Wayengera, Misaki

    2011-07-22

    Infection with HIV, which culminates in the establishment of a latent proviral reservoir, presents formidable challenges for ultimate cure. Building on the hypothesis that ex-vivo or even in-vivo abolition or disruption of HIV-gene/genome-action by target mutagenesis or excision can irreversibly abrogate HIV's innate fitness to replicate and survive, we previously identified the isoschizomeric bacteria restriction enzymes (REases) AcsI and ApoI as potent cleavers of the HIV-pol gene (11 and 9 times in HIV-1 and 2, respectively). However, both enzymes, along with others found to cleave across the entire HIV-1 genome, slice (SX) at palindromic sequences that are prevalent within the human genome and thereby pose the risk of host genome toxicity. A long-term goal in the field of R-M enzymatic therapeutics has thus been to generate synthetic restriction endonucleases with longer recognition sites limited in specificity to HIV. We aimed (i) to assemble and construct zinc finger arrays and nucleases (ZFN) with either proviral-HIV-pol gene or proviral-HIV-1 whole-genome specificity respectively, and (ii) to advance a model for pre-clinically testing lentiviral vectors (LV) that deliver and transduce either ZFN genotype. First, we computationally generated the consensus sequences of (a) 114 dsDNA-binding zinc finger (Zif) arrays (ZFAs or ZifHIV-pol) and (b) two zinc-finger nucleases (ZFNs) which, unlike the AcsI and ApoI homeodomains, possess specificity to >18 base-pair sequences uniquely present within the HIV-pol gene (ZifHIV-polFN). Another 15 ZFNs targeting >18 bp sequences within the complete HIV-1 proviral genome were constructed (ZifHIV-1FN). Second, a model for constructing lentiviral vectors (LVs) that deliver and transduce a diploid copy of either ZifHIV-polFN or ZifHIV-1FN chimeric genes (termed LV- 2xZifHIV-polFN and LV- 2xZifHIV-1FN, respectively) is proposed. Third, two preclinical models for controlled testing of the safety and efficacy of either of these

  17. Comparison of 454 Ultra-Deep Sequencing and Allele-Specific Real-Time PCR with Regard to the Detection of Emerging Drug-Resistant Minor HIV-1 Variants after Antiretroviral Prophylaxis for Vertical Transmission.

    Directory of Open Access Journals (Sweden)

    Andrea Hauser

    Full Text Available Pregnant HIV-infected women were screened for the development of HIV-1 drug resistance after implementation of a triple-antiretroviral transmission prophylaxis as recommended by the WHO in 2006. The study offered the opportunity to compare amplicon-based 454 ultra-deep sequencing (UDS and allele-specific real-time PCR (ASPCR for the detection of drug-resistant minor variants in the HIV-1 reverse transcriptase (RT.Plasma samples from 34 Tanzanian women were previously analysed by ASPCR for key resistance mutations in the viral RT selected by AZT, 3TC, and NVP (K70R, K103N, Y181C, M184V, T215Y/F. In this study, the RT region of the same samples was investigated by amplicon-based UDS for resistance mutations using the 454 GS FLX System.Drug-resistant HIV-variants were identified in 69% (20/29 of women by UDS and in 45% (13/29 by ASPCR. The absolute number of resistance mutations identified by UDS was twice that identified by ASPCR (45 vs 24. By UDS 14 of 24 ASPCR-detected resistance mutations were identified at the same position. The overall concordance between UDS and ASPCR was 61.0% (25/41. The proportions of variants quantified by UDS were approximately 2-3 times lower than by ASPCR. Amplicon generation from samples with viral loads below 20,000 copies/ml failed more frequently by UDS compared to ASPCR (limit of detection = 650 copies/ml, resulting in missing or insufficient sequence coverage.Both methods can provide useful information about drug-resistant minor HIV-1 variants. ASPCR has a higher sensitivity than UDS, but is restricted to single resistance mutations. In contrast, UDS is limited by its requirement for high viral loads to achieve sufficient sequence coverage, but the sequence information reveals the complete resistance patterns within the genomic region analysed. Improvements to the UDS limit of detection are in progress, and UDS could then facilitate monitoring of drug-resistant minor variants in the HIV-1 quasispecies.

  18. Spatiotemporal dynamics of the HIV-1 subtype G epidemic in West and Central Africa.

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    Delatorre, Edson; Mir, Daiana; Bello, Gonzalo

    2014-01-01

    The human immunodeficiency virus type 1 (HIV-1) subtype G is the second most prevalent HIV-1 clade in West Africa, accounting for nearly 30% of infections in the region. There is no information about the spatiotemporal dynamics of dissemination of this HIV-1 clade in Africa. To this end, we analyzed a total of 305 HIV-1 subtype G pol sequences isolated from 11 different countries from West and Central Africa over a period of 20 years (1992 to 2011). Evolutionary, phylogeographic and demographic parameters were jointly estimated from sequence data using a Bayesian coalescent-based method. Our analyses indicate that subtype G most probably emerged in Central Africa in 1968 (1956-1976). From Central Africa, the virus was disseminated to West and West Central Africa at multiple times from the middle 1970s onwards. Two subtype G strains probably introduced into Nigeria and Togo between the middle and the late 1970s were disseminated locally and to neighboring countries, leading to the origin of two major western African clades (G WA-I and G WA-II). Subtype G clades circulating in western and central African regions displayed an initial phase of exponential growth followed by a decline in growth rate since the early/middle 1990 s; but the mean epidemic growth rate of G WA-I (0.75 year-1) and G WA-II (0.95 year-1) clades was about two times higher than that estimated for central African lineages (0.47 year-1). Notably, the overall evolutionary and demographic history of G WA-I and G WA-II clades was very similar to that estimated for the CRF06_cpx clade circulating in the same region. These results support the notion that the spatiotemporal dissemination dynamics of major HIV-1 clades circulating in western Africa have probably been shaped by the same ecological factors.

  19. Molecular epidemiological study of HIV-1 CRF01_AE transmission in Hong Kong.

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    Chen, J H K; Wong, K H; Li, P; Chan, K C; Lee, M P; Lam, H Y; Cheng, V C C; Yuen, K Y; Yam, W C

    2009-08-15

    The objective of this study was to investigate the transmission history of the HIV-1 CRF01_AE epidemics in Hong Kong between 1994 and 2007. A total of 465 HIV-1 CRF01_AE pol sequences were derived from an in-house or a commercial HIV-1 genotyping system. Phylogenies of CRF01_AE sequences were analyzed by the Bayesian coalescent method. CRF01_AE patient population included 363 males (78.1%) and 102 females (21.9%), whereas 65% (314 of 465) were local Chinese. Major transmission routes were heterosexual contact (63%), followed by intravenous drug use (IDU) (19%) and men having sex with men (MSM) (17%). From phylogenetic analysis, local CRF01_AE strains were from multiple origins with 3 separate transmission clusters identified. Cluster 1 consisted mainly of Chinese male IDUs and heterosexuals. Clusters 2 and 3 included mainly local Chinese MSM and non-Chinese Asian IDUs, respectively. Chinese reference isolates available from China (Fujian, Guangxi, or Liaoning) were clonally related to our transmission clusters, demonstrating the epidemiological linkage of CRF01_AE infections between Hong Kong and China. The 3 individual local transmission clusters were estimated to have initiated since late 1980s and late 1990s, causing subsequent epidemics in the early 2000s. This is the first comprehensive molecular epidemiological study of HIV-1 CRF01_AE in Hong Kong. It revealed that MSM contact is becoming a major route of local CRF01_AE transmission in Hong Kong. Epidemiological linkage of CRF01_AE between Hong Kong and China observed in this study indicates the importance of regular molecular epidemiological surveillance for the HIV-1 epidemic in our region.

  20. [Genetic subtype and epidemiological feature of HIV-1 circulating strains among recently infected patients in Fujian province].

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    Deng, Yongyue; Zhang, Chunyang; Yan, Yansheng; Yan, Pingping; Wu, Shouli

    2014-06-01

    In order to evaluate the distribution of genetic subtypes and epidemiological feature of HIV-1 circulating strains in Fujian province. Blood samples and epidemiological data were collected from 104 newly infected patients who were distinguished by BED-CEIA methodology, during 2011-2012. Viral sequences(n = 81) of HIV-1 gag, env, and pol segments were amplified by nested PCR. Subtypes B and four Circulating Recombinant Forms, (CRF01_AE, CRF07_BC, CRF08_BC and CRF55_01B) were found in the samples, CRF01_AE(45.68%)and CRF07_BC(35.80%) were the two main HIV-1 strains in Fujian province. Compared with previous data, the proportion of CRF07_BC rose significantly while it gradually decreased in CRF01_AE. Heterosexual contact was still the principal transmission route in Fujian province, but the number of infection among men-who-have-sex-with- men grew rapidly. Results from this study suggested that different subtypes of HIV-1 strain existed in Fujian province. The distribution of subtypes and the mode of transmission were changing with the progress of epidemic. Dynamic monitoring of the molecular epidemiology trends of HIV-1 infection should be enhanced.

  1. HIV-1 genetic diversity and its distribution characteristics among newly diagnosed HIV-1 individuals in Hebei province, China.

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    Lu, Xinli; Zhao, Cuiying; Wang, Wei; Nie, Chenxi; Zhang, Yuqi; Zhao, Hongru; Chen, Suliang; Cui, Ze

    2016-01-01

    Since the first HIV-1 case in 1989, Hebei province has presented a clearly rising trend of HIV-1 prevalence, and HIV-1 genetic diversity has become the vital barrier to HIV prevention and control in this area. To obtain detailed information of HIV-1 spread in different populations and in different areas of Hebei, a cross-sectional HIV-1 molecular epidemiological investigation was performed across the province. Blood samples of 154 newly diagnosed HIV-1 individuals were collected from ten prefectures in Hebei using stratified sampling. Partial gag and env genes were amplified and sequenced. HIV-1 genotypes were identified by phylogenetic tree analyses. Among the 139 subjects genotyped, six HIV-1 subtypes were identified successfully, including subtype B (41.0 %), CRF01_AE (40.3 %), CRF07_BC (11.5 %), CRF08_BC (4.3 %), unique recombinant forms (URFs) (1.4 %) and subtype C (1.4 %). Subtype B was identified as the most frequent subtype. Two URF recombination patterns were the same as CRF01_AE/B. HIV-1 genotype distribution showed a significant statistical difference in different demographic characteristics, such as source (P  0.05). The differences in HIV-1 genotype distribution were closely associated with transmission routes. Particularly, all six subtype strains were found in heterosexuals, showing that HIV-1 has spread from the high-risk populations to the general populations in Hebei, China. In addition, CRF01_AE instead of subtype B has become the major strain of HIV-1 infection among homosexuals. Our study revealed HIV-1 evolution and genotype distribution by investigating newly diagnosed HIV-1 individuals in Hebei, China. This study provides important information to enhance the strategic plan for HIV prevention and control in China.

  2. The C-terminal tail of the gp41 transmembrane envelope glycoprotein of HIV-1 clades A, B, C, and D may exist in two conformations: an analysis of sequence, structure, and function

    International Nuclear Information System (INIS)

    Hollier, Mark J.; Dimmock, Nigel J.

    2005-01-01

    In addition to the major ectodomain, the gp41 transmembrane glycoprotein of HIV-1 is now known to have a minor ectodomain that is part of the long C-terminal tail. Both ectodomains are highly antigenic, carry neutralizing and non-neutralizing epitopes, and are involved in virus-mediated fusion activity. However, data have so far been biologically based, and derived solely from T cell line-adapted (TCLA), B clade viruses. Here we have carried out sequence and theoretically based structural analyses of 357 gp41 C-terminal sequences of mainly primary isolates of HIV-1 clades A, B, C, and D. Data show that all these viruses have the potential to form a tail loop structure (the minor ectodomain) supported by three, β-sheet, membrane-spanning domains (MSDs). This means that the first (N-terminal) tyrosine-based sorting signal of the gp41 tail is situated outside the cell membrane and is non-functional, and that gp41 that reaches the cell surface may be recycled back into the cytoplasm through the activity of the second tyrosine-sorting signal. However, we suggest that only a minority of cell-associated gp41 molecules - those destined for incorporation into virions - has 3 MSDs and the minor ectodomain. Most intracellular gp41 has the conventional single MSD, no minor ectodomain, a functional first tyrosine-based sorting signal, and in line with current thinking is degraded intracellularly. The gp41 structural diversity suggested here can be viewed as an evolutionary strategy to minimize HIV-1 envelope glycoprotein expression on the cell surface, and hence possible cytotoxicity and immune attack on the infected cell

  3. Sequence similarity between the erythrocyte binding domain 1 of the Plasmodium vivax Duffy binding protein and the V3 loop of HIV-1 strain MN reveals binding residues for the Duffy Antigen Receptor for Chemokines

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    Garry Robert F

    2011-01-01

    Full Text Available Abstract Background The surface glycoprotein (SU, gp120 of the human immunodeficiency virus (HIV must bind to a chemokine receptor, CCR5 or CXCR4, to invade CD4+ cells. Plasmodium vivax uses the Duffy Binding Protein (DBP to bind the Duffy Antigen Receptor for Chemokines (DARC and invade reticulocytes. Results Variable loop 3 (V3 of HIV-1 SU and domain 1 of the Plasmodium vivax DBP share a sequence similarity. The site of amino acid sequence similarity was necessary, but not sufficient, for DARC binding and contained a consensus heparin binding site essential for DARC binding. Both HIV-1 and P. vivax can be blocked from binding to their chemokine receptors by the chemokine, RANTES and its analog AOP-RANTES. Site directed mutagenesis of the heparin binding motif in members of the DBP family, the P. knowlesi alpha, beta and gamma proteins abrogated their binding to erythrocytes. Positively charged residues within domain 1 are required for binding of P. vivax and P. knowlesi erythrocyte binding proteins. Conclusion A heparin binding site motif in members of the DBP family may form part of a conserved erythrocyte receptor binding pocket.

  4. HIV-1 Genetic Variability in Cuba and Implications for Transmission and Clinical Progression.

    Science.gov (United States)

    Blanco, Madeline; Machado, Liuber Y; Díaz, Héctor; Ruiz, Nancy; Romay, Dania; Silva, Eladio

    2015-10-01

    INTRODUCTION Serological and molecular HIV-1 studies in Cuba have shown very low prevalence of seropositivity, but an increasing genetic diversity attributable to introduction of many HIV-1 variants from different areas, exchange of such variants among HIV-positive people with several coinciding routes of infection and other epidemiologic risk factors in the seropositive population. The high HIV-1 genetic variability observed in Cuba has possible implications for transmission and clinical progression. OBJECTIVE Study genetic variability for the HIV-1 env, gag and pol structural genes in Cuba; determine the prevalence of B and non-B subtypes according to epidemiologic and behavioral variables and determine whether a relationship exists between genetic variability and transmissibility, and between genetic variability and clinical disease progression in people living with HIV/AIDS. METHODS Using two molecular assays (heteroduplex mobility assay and nucleic acid sequencing), structural genes were characterized in 590 people with HIV-1 (480 men and 110 women), accounting for 3.4% of seropositive individuals in Cuba as of December 31, 2013. Nonrandom sampling, proportional to HIV prevalence by province, was conducted. Relationships between molecular results and viral factors, host characteristics, and patients' clinical, epidemiologic and behavioral variables were studied for molecular epidemiology, transmission, and progression analyses. RESULTS Molecular analysis of the three HIV-1 structural genes classified 297 samples as subtype B (50.3%), 269 as non-B subtypes (45.6%) and 24 were not typeable. Subtype B prevailed overall and in men, mainly in those who have sex with men. Non-B subtypes were prevalent in women and heterosexual men, showing multiple circulating variants and recombinant forms. Sexual transmission was the predominant form of infection for all. B and non-B subtypes were encountered throughout Cuba. No association was found between subtypes and

  5. Covariance of charged amino acids at positions 322 and 440 of HIV-1 Env contributes to coreceptor specificity of subtype B viruses, and can be used to improve the performance of V3 sequence-based coreceptor usage prediction algorithms.

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    Kieran Cashin

    Full Text Available The ability to determine coreceptor usage of patient-derived human immunodeficiency virus type 1 (HIV-1 strains is clinically important, particularly for the administration of the CCR5 antagonist maraviroc. The envelope glycoprotein (Env determinants of coreceptor specificity lie primarily within the gp120 V3 loop region, although other Env determinants have been shown to influence gp120-coreceptor interactions. Here, we determined whether conserved amino acid alterations outside the V3 loop that contribute to coreceptor usage exist, and whether these alterations improve the performance of V3 sequence-based coreceptor usage prediction algorithms. We demonstrate a significant covariant association between charged amino acids at position 322 in V3 and position 440 in the C4 Env region that contributes to the specificity of HIV-1 subtype B strains for CCR5 or CXCR4. Specifically, positively charged Lys/Arg at position 322 and negatively charged Asp/Glu at position 440 occurred more frequently in CXCR4-using viruses, whereas negatively charged Asp/Glu at position 322 and positively charged Arg at position 440 occurred more frequently in R5 strains. In the context of CD4-bound gp120, structural models suggest that covariation of amino acids at Env positions 322 and 440 has the potential to alter electrostatic interactions that are formed between gp120 and charged amino acids in the CCR5 N-terminus. We further demonstrate that inclusion of a "440 rule" can improve the sensitivity of several V3 sequence-based genotypic algorithms for predicting coreceptor usage of subtype B HIV-1 strains, without compromising specificity, and significantly improves the AUROC of the geno2pheno algorithm when set to its recommended false positive rate of 5.75%. Together, our results provide further mechanistic insights into the intra-molecular interactions within Env that contribute to coreceptor specificity of subtype B HIV-1 strains, and demonstrate that incorporation

  6. A robust measure of HIV-1 population turnover within chronically infected individuals.

    Science.gov (United States)

    Achaz, G; Palmer, S; Kearney, M; Maldarelli, F; Mellors, J W; Coffin, J M; Wakeley, J

    2004-10-01

    A simple nonparameteric test for population structure was applied to temporally spaced samples of HIV-1 sequences from the gag-pol region within two chronically infected individuals. The results show that temporal structure can be detected for samples separated by about 22 months or more. The performance of the method, which was originally proposed to detect geographic structure, was tested for temporally spaced samples using neutral coalescent simulations. Simulations showed that the method is robust to variation in samples sizes and mutation rates, to the presence/absence of recombination, and that the power to detect temporal structure is high. By comparing levels of temporal structure in simulations to the levels observed in real data, we estimate the effective intra-individual population size of HIV-1 to be between 10(3) and 10(4) viruses, which is in agreement with some previous estimates. Using this estimate and a simple measure of sequence diversity, we estimate an effective neutral mutation rate of about 5 x 10(-6) per site per generation in the gag-pol region. The definition and interpretation of estimates of such "effective" population parameters are discussed.

  7. [Molecular epidemiology and transmission of HIV-1 infection in Zhejiang province, 2015].

    Science.gov (United States)

    Yang, J Z; Chen, W J; Zhang, W J; He, L; Zhang, J F; Pan, X H

    2017-11-10

    Objective: To understand the distribution of HIV-1 subtype diversity and its transmission characteristics in Zhejiang province. Methods: A total of 302 newly diagnosed HIV-1 positive patients were selected through stratified random sampling in Zhejiang in 2015. HIV-1 pol genes were sequenced successfully with reverse transcription PCR/nested PCR and phylogenetic analysis was conducted for 276 patients. Then a molecular epidemiologic study was performed combined with field epidemiological investigation. Results: Of 276 sequence samples analyzed, 122 CRF07_BC strains (44.2%), 103 CRF01_AE strains (37.3%), 17 CRF08_BC strains (6.1%), 9 B strains (3.2%), 6 CRF55_01B strains (2.2%), 5 C strains (1.8%), 1 CRF59_01B strain (0.4%), 1 CRF67_01B strain (0.4%), 1 A1 strain (0.4%), and 11 URFs strains (4.0%) were identified. Phylogenetic analysis revealed 16 clusters with only 15.1% (34/225) sequences involved among CRF07_BC and CRF01_AE strains. The clustered cases in MSM were higher than that in populations with other transmission routes. And clusters existed between the populations with different transmission routes. Conclusion: The major strains of HIV-1 in Zhejiang are CRF07_BC and CRF01_AE. The HIV subtypes showed more complexity in Zhejiang. It is necessary to strengthen the surveillance for HIV subtypes, carry out classified management and conduct effective prevention and control in the population at high risk.

  8. Discovery of novel targets for multi-epitope vaccines: Screening of HIV-1 genomes using association rule mining

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    Piontkivska Helen

    2009-07-01

    Full Text Available Abstract Background Studies have shown that in the genome of human immunodeficiency virus (HIV-1 regions responsible for interactions with the host's immune system, namely, cytotoxic T-lymphocyte (CTL epitopes tend to cluster together in relatively conserved regions. On the other hand, "epitope-less" regions or regions with relatively low density of epitopes tend to be more variable. However, very little is known about relationships among epitopes from different genes, in other words, whether particular epitopes from different genes would occur together in the same viral genome. To identify CTL epitopes in different genes that co-occur in HIV genomes, association rule mining was used. Results Using a set of 189 best-defined HIV-1 CTL/CD8+ epitopes from 9 different protein-coding genes, as described by Frahm, Linde & Brander (2007, we examined the complete genomic sequences of 62 reference HIV sequences (including 13 subtypes and sub-subtypes with approximately 4 representative sequences for each subtype or sub-subtype, and 18 circulating recombinant forms. The results showed that despite inclusion of recombinant sequences that would be expected to break-up associations of epitopes in different genes when two different genomes are recombined, there exist particular combinations of epitopes (epitope associations that occur repeatedly across the world-wide population of HIV-1. For example, Pol epitope LFLDGIDKA is found to be significantly associated with epitopes GHQAAMQML and FLKEKGGL from Gag and Nef, respectively, and this association rule is observed even among circulating recombinant forms. Conclusion We have identified CTL epitope combinations co-occurring in HIV-1 genomes including different subtypes and recombinant forms. Such co-occurrence has important implications for design of complex vaccines (multi-epitope vaccines and/or drugs that would target multiple HIV-1 regions at once and, thus, may be expected to overcome challenges

  9. Evidence of multiple introductions of HIV-1 subtype C in Angola.

    Science.gov (United States)

    Afonso, Joana Morais; Morgado, Mariza G; Bello, Gonzalo

    2012-10-01

    HIV-1 subtype C is the most prevalent group M clade in southern Africa and some eastern African countries. Subtype C is also the most frequent subtype in Angola (southwestern Africa), with an estimated prevalence of 10-20%. In order to better understand the origin of the HIV-1 subtype C strains circulating in Angola, 31 subtype C pol sequences of Angolan origin were compared with 1950 subtype C pol sequences sampled in other African countries. Phylogenetic analyses reveal that the Angolan subtype C sequences were distributed in 16 different lineages that were widely dispersed among other African strains. Ten subtype C Angolan lineages were composed by only one sequence, while the remaining six clades contain between two and seven sequences. Bayesian phylogeographic analysis indicates that most Angolan clades probably originated in different southern African countries with the exception of one lineage that most likely originated in Burundi. Evolutionary analysis suggests that those Angolan subtype C clades composed by ≥ 2 sequences were introduced into the country between the late 1970s and the mid 2000s. The median estimated time frame for the origin of those Angolan lineages coincides with periods of positive migration influx in Angola that were preceded by phases of negative migratory outflow. These results demonstrate that the Angolan subtype C epidemic resulted from multiple introductions of subtype C viruses mainly imported from southern African countries over the last 30years, some of which have been locally disseminated establishing several autochthonous transmission networks. This study also suggests that population mobility between Angola and southern African countries during civil war (1974-2002) may have played a key role in the emergence of the Angolan subtype C epidemic. Copyright © 2012 Elsevier B.V. All rights reserved.

  10. The HIV-1 epidemic in Bolivia is dominated by subtype B and CRF12_BF "family" strains.

    Science.gov (United States)

    Guimarães, Monick L; Velarde-Dunois, Ketty G; Segurondo, David; Morgado, Mariza G

    2012-01-16

    Molecular epidemiological studies of HIV-1 in South America have revealed the occurrence of subtypes B, F1 and BF1 recombinants. Even so, little information concerning the HIV-1 molecular epidemiology in Bolivia is available. In this study we performed phylogenetic analyses from samples collected in Bolivia at two different points in time over a 10 year span. We analyzed these samples to estimate the trends in the HIV subtype and recombinant forms over time. Fifty one HIV-1 positive samples were collected in Bolivia over two distinct periods (1996 and 2005). These samples were genetically characterized based on partial pol protease/reverse transcriptase (pr/rt) and env regions. Alignment and neighbor-joining (NJ) phylogenetic analyses were established from partial env (n = 37) and all pol sequences using Mega 4. The remaining 14 env sequences from 1996 were previously characterized based on HMA-env (Heteroduplex mobility assay). The Simplot v.3.5.1 program was used to verify intragenic recombination, and SplitsTree 4.0 was employed to confirm the phylogenetic relationship of the BF1 recombinant samples. Phylogenetic analysis of both env and pol regions confirmed the predominance of "pure" subtype B (72.5%) samples circulating in Bolivia and revealed a high prevalence of BF1 genotypes (27.5%). Eleven out of 14 BF1 recombinants displayed a mosaic structure identical or similar to that described for the CRF12_BF variant, one sample was classified as CRF17_BF, and two others were F1pol/Benv. No "pure" HIV-1 subtype F1 or B" variant of subtype B was detected in the present study. Of note, samples characterized as CRF12_BF-related were depicted only in 2005. HIV-1 genetic diversity in Bolivia is mostly driven by subtype B followed by BF1 recombinant strains from the CRF12_BF "family". No significant temporal changes were detected between the mid-1990s and the mid-2000s for subtype B (76.2% vs 70.0%) or BF1 recombinant (23.8% vs 30.0%) samples from Bolivia.

  11. The HIV-1 epidemic in Bolivia is dominated by subtype B and CRF12_BF "family" strains

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    Guimarães Monick L

    2012-01-01

    Full Text Available Abstract Background Molecular epidemiological studies of HIV-1 in South America have revealed the occurrence of subtypes B, F1 and BF1 recombinants. Even so, little information concerning the HIV-1 molecular epidemiology in Bolivia is available. In this study we performed phylogenetic analyses from samples collected in Bolivia at two different points in time over a 10 year span. We analyzed these samples to estimate the trends in the HIV subtype and recombinant forms over time. Materials and methods Fifty one HIV-1 positive samples were collected in Bolivia over two distinct periods (1996 and 2005. These samples were genetically characterized based on partial pol protease/reverse transcriptase (pr/rt and env regions. Alignment and neighbor-joining (NJ phylogenetic analyses were established from partial env (n = 37 and all pol sequences using Mega 4. The remaining 14 env sequences from 1996 were previously characterized based on HMA-env (Heteroduplex mobility assay. The Simplot v.3.5.1 program was used to verify intragenic recombination, and SplitsTree 4.0 was employed to confirm the phylogenetic relationship of the BF1 recombinant samples. Results Phylogenetic analysis of both env and pol regions confirmed the predominance of "pure" subtype B (72.5% samples circulating in Bolivia and revealed a high prevalence of BF1 genotypes (27.5%. Eleven out of 14 BF1 recombinants displayed a mosaic structure identical or similar to that described for the CRF12_BF variant, one sample was classified as CRF17_BF, and two others were F1pol/Benv. No "pure" HIV-1 subtype F1 or B" variant of subtype B was detected in the present study. Of note, samples characterized as CRF12_BF-related were depicted only in 2005. Conclusion HIV-1 genetic diversity in Bolivia is mostly driven by subtype B followed by BF1 recombinant strains from the CRF12_BF "family". No significant temporal changes were detected between the mid-1990s and the mid-2000s for subtype B (76.2% vs 70

  12. Modification of a loop sequence between α-helices 6 and 7 of virus capsid (CA protein in a human immunodeficiency virus type 1 (HIV-1 derivative that has simian immunodeficiency virus (SIVmac239 vif and CA α-helices 4 and 5 loop improves replication in cynomolgus monkey cells

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    Adachi Akio

    2009-08-01

    Full Text Available Abstract Background Human immunodeficiency virus type 1 (HIV-1 productively infects only humans and chimpanzees but not cynomolgus or rhesus monkeys while simian immunodeficiency virus isolated from macaque (SIVmac readily establishes infection in those monkeys. Several HIV-1 and SIVmac chimeric viruses have been constructed in order to develop an animal model for HIV-1 infection. Construction of an HIV-1 derivative which contains sequences of a SIVmac239 loop between α-helices 4 and 5 (L4/5 of capsid protein (CA and the entire SIVmac239 vif gene was previously reported. Although this chimeric virus could grow in cynomolgus monkey cells, it did so much more slowly than did SIVmac. It was also reported that intrinsic TRIM5α restricts the post-entry step of HIV-1 replication in rhesus and cynomolgus monkey cells, and we previously demonstrated that a single amino acid in a loop between α-helices 6 and 7 (L6/7 of HIV type 2 (HIV-2 CA determines the susceptibility of HIV-2 to cynomolgus monkey TRIM5α. Results In the study presented here, we replaced L6/7 of HIV-1 CA in addition to L4/5 and vif with the corresponding segments of SIVmac. The resultant HIV-1 derivatives showed enhanced replication capability in established T cell lines as well as in CD8+ cell-depleted primary peripheral blood mononuclear cells from cynomolgus monkey. Compared with the wild type HIV-1 particles, the viral particles produced from a chimeric HIV-1 genome with those two SIVmac loops were less able to saturate the intrinsic restriction in rhesus monkey cells. Conclusion We have succeeded in making the replication of simian-tropic HIV-1 in cynomolgus monkey cells more efficient by introducing into HIV-1 the L6/7 CA loop from SIVmac. It would be of interest to determine whether HIV-1 derivatives with SIVmac CA L4/5 and L6/7 can establish infection of cynomolgus monkeys in vivo.

  13. The connection domain in reverse transcriptase facilitates the in vivo annealing of tRNALys3 to HIV-1 genomic RNA

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    Niu Meijuan

    2004-10-01

    Full Text Available Abstract The primer tRNA for reverse transcription in HIV-1, tRNALys3, is selectively packaged into the virus during its assembly, and annealed to the viral genomic RNA. The ribonucleoprotein complex that is involved in the packaging and annealing of tRNALys into HIV-1 consists of Gag, GagPol, tRNALys, lysyl-tRNA synthetase (LysRS, and viral genomic RNA. Gag targets tRNALys for viral packaging through Gag's interaction with LysRS, a tRNALys-binding protein, while reverse transcriptase (RT sequences within GagPol (the thumb domain bind to tRNALys. The further annealing of tRNALys3 to viral RNA requires nucleocapsid (NC sequences in Gag, but not the NC sequences GagPol. In this report, we further show that while the RT connection domain in GagPol is not required for tRNALys3 packaging into the virus, it is required for tRNALys3 annealing to the viral RNA genome.

  14. Description of HIV-1 Group M Molecular Epidemiology and Drug Resistance Prevalence in Equatorial Guinea from Migrants in Spain

    Science.gov (United States)

    Yebra, Gonzalo; de Mulder, Miguel; Holguín, África

    2013-01-01

    Background The HIV epidemic is increasing in Equatorial Guinea (GQ), West Central Africa, but few studies have reported its HIV molecular epidemiology. We aimed to describe the HIV-1 group M (HIV-1M) variants and drug-resistance mutations in GQ using sequences sampled in this country and in Spain, a frequent destination of Equatoguinean migrants. Methods We collected 195 HIV-1M pol sequences from Equatoguinean subjects attending Spanish clinics during 1997-2011, and 83 additional sequences sampled in GQ in 1997 and 2008 from GenBank. All (n = 278) were re-classified using phylogeny and tested for drug-resistance mutations. To evaluate the origin of CRF02_AG in GQ, we analyzed 2,562 CRF02_AG sequences and applied Bayesian MCMC inference (BEAST program). Results Most Equatoguinean patients recruited in Spain were women (61.1%) or heterosexuals (87.7%). In the 278 sequences, the variants found were CRF02_AG (47.8%), A (13.7%), B (7.2%), C (5.8%), G (5.4%) and others (20.1%). We found 6 CRF02_AG clusters emerged from 1983.9 to 2002.5 with origin in GQ (5.5 sequences/cluster). Transmitted drug-resistance (TDR) rate among naïve patients attended in Spain (n = 144) was 4.7%: 3.4% for PI (all with M46IL), 1.8% for NRTI (all with M184V) and 0.9% for NNRTI (Y188L). Among pre-treated patients, 9/31 (29%) presented any resistance, mainly affecting NNRTI (27.8%). Conclusions We report a low (HIV-1M variant and entered GQ through independent introductions at least since the early 1980s. PMID:23717585

  15. Dynamic HIV-1 genetic recombination and genotypic drug resistance among treatment-experienced adults in northern Ghana.

    Science.gov (United States)

    Nii-Trebi, Nicholas Israel; Brandful, James Ashun Mensah; Ibe, Shiro; Sugiura, Wataru; Barnor, Jacob Samson; Bampoh, Patrick Owiredu; Yamaoka, Shoji; Matano, Tetsuro; Yoshimura, Kazuhisa; Ishikawa, Koichi; Ampofo, William Kwabena

    2017-11-01

    There have been hardly any reports on the human immunodeficiency virus type 1 (HIV-1) drug-resistance profile from northern Ghana since antiretroviral therapy (ART) was introduced over a decade ago. This study investigated prevailing HIV-1 subtypes and examined the occurrence of drug resistance in ART-experienced patients in Tamale, the capital of the Northern Region of Ghana. A cross-sectional study was carried out on HIV-infected adult patients receiving first-line ART. HIV viral load (VL) and CD4 + T-cell counts were measured. The pol gene sequences were analysed for genotypic resistance by an in-house HIV-1 drug-resistance test; the prevailing HIV-1 subtypes were analysed in detail.Results/Key findings. A total of 33 subjects were studied. Participants comprised 11 males (33.3 %) and 22 (66.7 %) females, with a median age of 34.5 years [interquartile range (IQR) 30.0-40.3]. The median duration on ART was 12 months (IQR 8.0-24). Of the 24 subjects successfully genotyped, 10 (41.7 %) viruses possessed at least one mutation conferring resistance to nucleoside or non-nucleoside reverse-transcriptase inhibitors (NRTIs/NNRTIs). Two-class drug resistance to NRTI and NNRTI was mostly detected (25 %, 6/24). The most frequent mutations were lamivudine-resistance M184V and efavirenz/nevirapine-resistance K103N. HIV-1 subtype CRF02_AG was predominant (79.2 %). Other HIV-1 subtypes detected were G (8.3 %), A3 (4.2 %) and importantly two (8.3 %) unique HIV-1 recombinant forms with CRF02_AG/A3 mosaic. HIV-1 shows high genetic diversity and on-going viral genetic recombination in the study region. Nearly 42 % of the patients studied harboured a drug-resistant virus. The study underscores the need for continued surveillance of HIV-1 subtype diversity; and of drug-resistance patterns to guide selection of second-line regimens in northern Ghana.

  16. HIV-1 phylogenetic analysis shows HIV-1 transits through the meninges to brain and peripheral tissues.

    Science.gov (United States)

    Lamers, Susanna L; Gray, Rebecca R; Salemi, Marco; Huysentruyt, Leanne C; McGrath, Michael S

    2011-01-01

    Brain infection by the human immunodeficiency virus type 1 (HIV-1) has been investigated in many reports with a variety of conclusions concerning the time of entry and degree of viral compartmentalization. To address these diverse findings, we sequenced HIV-1 gp120 clones from a wide range of brain, peripheral and meningeal tissues from five patients who died from several HIV-1 associated disease pathologies. High-resolution phylogenetic analysis confirmed previous studies that showed a significant degree of compartmentalization in brain and peripheral tissue subpopulations. Some intermixing between the HIV-1 subpopulations was evident, especially in patients that died from pathologies other than HIV-associated dementia. Interestingly, the major tissue harboring virus from both the brain and peripheral tissues was the meninges. These results show that (1) HIV-1 is clearly capable of migrating out of the brain, (2) the meninges are the most likely primary transport tissues, and (3) infected brain macrophages comprise an important HIV reservoir during highly active antiretroviral therapy. Copyright © 2010 Elsevier B.V. All rights reserved.

  17. [Study on HIV-1 subtype among elderly male clients and female sex workers of low-cost venues in Guangxi Zhuang Autonomous Region, China].

    Science.gov (United States)

    Deng, Y Q; Li, J J; Fang, N Y; Wang, B; Wang, J W; Liang, S S; Shen, Z Y; Lan, G H; Zhang, H M; Wu, X H; Lu, H X; Ge, X M

    2017-03-10

    Objective: To understand HIV-1 subtype characteristics and transmission clusters in elderly male clients and female sex workers (FSWs) of low-cost commercial sex venues in Guangxi Zhuang Autonomous Region, China. Methods: A cross sectional survey was conducted in FSWs and elderly male clients (≥50 years) of low-cost commercial sex venues in 4 cities and 9 counties in Guangxi Zhuang Autonomous Region by convenient sampling in 2012. The blood sample was collected from each case for HIV-1 antibody detection. The pol gene fragments were amplified and sequenced from viral RNA template extracted from plasma samples. The phylogenetic tree was constructed and the subtypes were identified. Results: A total of 4 048 elderly male clients and 784 FSWs were surveyed, and 116 HIV-1 infections were detected, the positive rate was 2.5% (103/4 048) in the clients and 1.7% (13/784) in FSWs. The gene amplification and sequencing of HIV-1 detected in 84 blood samples indicated that 53 pol gene sequences were successfully determined (48 blood samples from elderly male clients and 5 blood samples from FSWs). Among 53 pol sequences, 48(90.6% ), 4(7.5% ), and 1(1.9% ) sequences were identified as CRF01_AE, CRF08_BC, and CRF07_BC, respectively. Two transmission clusters were identified among CRF01_AE, including 4 sub-clusters. One transmission cluster was identified among CRF08_BC. The transmission cluster or sub-cluster were from the infected individuals at same low-cost commercial sex venue, or different low-cost commercial sex venues in the same town, or same place, or adjacent villages and towns. Conclusions: CRF01_AE was the predominant HIV-1 subtype among elderly male clients and FSWs of low-cost commercial sex venues in Guangxi Zhuang Autonomous Region, circulating in same venue or adjacent villages and towns. The HIV-1 positive male clients and FSWs might play an important role in the spread of the strains.

  18. Development of an epitope-based HIV-1 vaccine strategy from HIV-1 lipopeptide to dendritic-based vaccines.

    Science.gov (United States)

    Surenaud, Mathieu; Lacabaratz, Christine; Zurawski, Gérard; Lévy, Yves; Lelièvre, Jean-Daniel

    2017-10-01

    Development of a safe, effective and globally affordable Human Immunodeficiency Virus strain 1 (HIV-1) vaccine offers the best hope for future control of the HIV-1 pandemic. However, with the exception of the recent RV144 trial, which elicited a modest level of protection against infection, no vaccine candidate has shown efficacy in preventing HIV-1 infection or in controlling virus replication in humans. There is also a great need for a successful immunotherapeutic vaccine since combination antiretroviral therapy (cART) does not eliminate the reservoir of HIV-infected cells. But to date, no vaccine candidate has proven to significantly alter the natural history of an individual with HIV-1 infection. Areas covered: For over 25 years, the ANRS (France Recherche Nord&Sud Sida-HIV hépatites) has been committed to an original program combining basic science and clinical research developing an epitope-based vaccine strategy to induce a multiepitopic cellular response against HIV-1. This review describes the evolution of concepts, based on strategies using HIV-1 lipopeptides towards the use of dendritic cell (DC) manipulation. Expert commentary: Understanding the crucial role of DCs in immune responses allowed moving from the non-specific administration of HIV-1 sequences with lipopeptides to DC-based vaccines. These DC-targeting strategies should improve HIV-1 vaccine efficacy.

  19. Hyperthermia stimulates HIV-1 replication.

    Directory of Open Access Journals (Sweden)

    Ferdinand Roesch

    Full Text Available HIV-infected individuals may experience fever episodes. Fever is an elevation of the body temperature accompanied by inflammation. It is usually beneficial for the host through enhancement of immunological defenses. In cultures, transient non-physiological heat shock (42-45°C and Heat Shock Proteins (HSPs modulate HIV-1 replication, through poorly defined mechanisms. The effect of physiological hyperthermia (38-40°C on HIV-1 infection has not been extensively investigated. Here, we show that culturing primary CD4+ T lymphocytes and cell lines at a fever-like temperature (39.5°C increased the efficiency of HIV-1 replication by 2 to 7 fold. Hyperthermia did not facilitate viral entry nor reverse transcription, but increased Tat transactivation of the LTR viral promoter. Hyperthermia also boosted HIV-1 reactivation in a model of latently-infected cells. By imaging HIV-1 transcription, we further show that Hsp90 co-localized with actively transcribing provirus, and this phenomenon was enhanced at 39.5°C. The Hsp90 inhibitor 17-AAG abrogated the increase of HIV-1 replication in hyperthermic cells. Altogether, our results indicate that fever may directly stimulate HIV-1 replication, in a process involving Hsp90 and facilitation of Tat-mediated LTR activity.

  20. Enrichment of intersubtype HIV-1 recombinants in a dual infection system using HIV-1 strain-specific siRNAs

    Science.gov (United States)

    2011-01-01

    Background Intersubtype HIV-1 recombinants in the form of unique or stable circulating recombinants forms (CRFs) are responsible for over 20% of infections in the worldwide epidemic. Mechanisms controlling the generation, selection, and transmission of these intersubtype HIV-1 recombinants still require further investigation. All intersubtype HIV-1 recombinants are generated and evolve from initial dual infections, but are difficult to identify in the human population. In vitro studies provide the most practical system to study mechanisms, but the recombination rates are usually very low in dual infections with primary HIV-1 isolates. This study describes the use of HIV-1 isolate-specific siRNAs to enrich intersubtype HIV-1 recombinants and inhibit the parental HIV-1 isolates from a dual infection. Results Following a dual infection with subtype A and D primary HIV-1 isolates and two rounds of siRNA treatment, nearly 100% of replicative virus was resistant to a siRNA specific for an upstream target sequence in the subtype A envelope (env) gene as well as a siRNA specific for a downstream target sequence in the subtype D env gene. Only 20% (10/50) of the replicating virus had nucleotide substitutions in the siRNA-target sequence whereas the remaining 78% (39/50) harbored a recombination breakpoint that removed both siRNA target sequences, and rendered the intersubtype D/A recombinant virus resistant to the dual siRNA treatment. Since siRNAs target the newly transcribed HIV-1 mRNA, the siRNAs only enrich intersubtype env recombinants and do not influence the recombination process during reverse transcription. Using this system, a strong bias is selected for recombination breakpoints in the C2 region, whereas other HIV-1 env regions, most notably the hypervariable regions, were nearly devoid of intersubtype recombination breakpoints. Sequence conservation plays an important role in selecting for recombination breakpoints, but the lack of breakpoints in many conserved

  1. Sensitive non-radioactive detection of HIV-1

    DEFF Research Database (Denmark)

    Teglbjærg, Lars Stubbe; Nielsen, C; Hansen, J E

    1992-01-01

    This report describes the use of the polymerase chain reaction (PCR) for the non-radioactive detection of HIV-1 proviral genomic sequences in HIV-1 infected cells. We have developed a sensitive assay, using three different sets of nested primers and our results show that this method is superior...... to standard PCR for the detection of HIV-1 DNA. The assay described features the use of a simple and inexpensive sample preparation technique and a non-radioactive hybridization procedure for confirmation of results. To test the suitability of the assay for clinical purposes, we tested cell samples from 76...

  2. The global transmission network of HIV-1.

    Science.gov (United States)

    Wertheim, Joel O; Leigh Brown, Andrew J; Hepler, N Lance; Mehta, Sanjay R; Richman, Douglas D; Smith, Davey M; Kosakovsky Pond, Sergei L

    2014-01-15

    Human immunodeficiency virus type 1 (HIV-1) is pandemic, but its contemporary global transmission network has not been characterized. A better understanding of the properties and dynamics of this network is essential for surveillance, prevention, and eventual eradication of HIV. Here, we apply a simple and computationally efficient network-based approach to all publicly available HIV polymerase sequences in the global database, revealing a contemporary picture of the spread of HIV-1 within and between countries. This approach automatically recovered well-characterized transmission clusters and extended other clusters thought to be contained within a single country across international borders. In addition, previously undescribed transmission clusters were discovered. Together, these clusters represent all known modes of HIV transmission. The extent of international linkage revealed by our comprehensive approach demonstrates the need to consider the global diversity of HIV, even when describing local epidemics. Finally, the speed of this method allows for near-real-time surveillance of the pandemic's progression.

  3. Virtually full-length subtype F and F/D recombinant HIV-1 from Africa and South America

    NARCIS (Netherlands)

    Laukkanen, T.; Carr, J. K.; Janssens, W.; Liitsola, K.; Gotte, D.; McCutchan, F. E.; Op de Coul, E.; Cornelissen, M.; Heyndrickx, L.; van der Groen, G.; Salminen, M. O.

    2000-01-01

    For reliable classification of HIV-1 strains appropriate reference sequences are needed. The HIV-1 genetic subtype F has a wide geographic spread, causing significant epidemics in South America, Africa, and some regions of Europe. Previously only two full-length sequences of each of the HIV-1

  4. Recombination of HIV type 1C (C'/C") in Ethiopia: possible link of EthHIV-1C' to subtype C sequences from the high-prevalence epidemics in India and Southern Africa

    NARCIS (Netherlands)

    Pollakis, Georgios; Abebe, Almaz; Kliphuis, Aletta; Rinke de Wit, Tobias F.; Fisseha, Bitew; Tegbaru, Belete; Tesfaye, Girma; Negassa, Hailu; Mengistu, Yohannes; Fontanet, Arnaud L.; Cornelissen, Marion; Goudsmit, Jaap

    2003-01-01

    The magnitude and complexity of the HIV-1 genetic diversity are major challenges for vaccine development. Investigation of the genotypes circulating in areas of high incidence, as well as their interactions, will be a milestone in the development of an efficacious vaccine. Because HIV-1 subtype C

  5. HIV-1 RNAs are Not Part of the Argonaute 2 Associated RNA Interference Pathway in Macrophages.

    Directory of Open Access Journals (Sweden)

    Valentina Vongrad

    Full Text Available MiRNAs and other small noncoding RNAs (sncRNAs are key players in post-transcriptional gene regulation. HIV-1 derived small noncoding RNAs (sncRNAs have been described in HIV-1 infected cells, but their biological functions still remain to be elucidated. Here, we approached the question whether viral sncRNAs may play a role in the RNA interference (RNAi pathway or whether viral mRNAs are targeted by cellular miRNAs in human monocyte derived macrophages (MDM.The incorporation of viral sncRNAs and/or their target RNAs into RNA-induced silencing complex was investigated using photoactivatable ribonucleoside-induced cross-linking and immunoprecipitation (PAR-CLIP as well as high-throughput sequencing of RNA isolated by cross-linking immunoprecipitation (HITS-CLIP, which capture Argonaute2-bound miRNAs and their target RNAs. HIV-1 infected monocyte-derived macrophages (MDM were chosen as target cells, as they have previously been shown to express HIV-1 sncRNAs. In addition, we applied small RNA deep sequencing to study differential cellular miRNA expression in HIV-1 infected versus non-infected MDMs.PAR-CLIP and HITS-CLIP data demonstrated the absence of HIV-1 RNAs in Ago2-RISC, although the presence of a multitude of HIV-1 sncRNAs in HIV-1 infected MDMs was confirmed by small RNA sequencing. Small RNA sequencing revealed that 1.4% of all sncRNAs were of HIV-1 origin. However, neither HIV-1 derived sncRNAs nor putative HIV-1 target sequences incorporated into Ago2-RISC were identified suggesting that HIV-1 sncRNAs are not involved in the canonical RNAi pathway nor is HIV-1 targeted by this pathway in HIV-1 infected macrophages.

  6. Spatiotemporal dynamics of HIV-1 transmission in France (1999-2014) and impact of targeted prevention strategies.

    Science.gov (United States)

    Chaillon, Antoine; Essat, Asma; Frange, Pierre; Smith, Davey M; Delaugerre, Constance; Barin, Francis; Ghosn, Jade; Pialoux, Gilles; Robineau, Olivier; Rouzioux, Christine; Goujard, Cécile; Meyer, Laurence; Chaix, Marie-Laure

    2017-02-21

    Characterizing HIV-1 transmission networks can be important in understanding the evolutionary patterns and geospatial spread of the epidemic. We reconstructed the broad molecular epidemiology of HIV from individuals with primary HIV-1 infection (PHI) enrolled in France in the ANRS PRIMO C06 cohort over 15 years. Sociodemographic, geographic, clinical, biological and pol sequence data from 1356 patients were collected between 1999 and 2014. Network analysis was performed to infer genetic relationships, i.e. clusters of transmission, between HIV-1 sequences. Bayesian coalescent-based methods were used to examine the temporal and spatial dynamics of identified clusters from different regions in France. We also evaluated the use of network information to target prevention efforts. Participants were mostly Caucasian (85.9%) and men (86.7%) who reported sex with men (MSM, 71.4%). Overall, 387 individuals (28.5%) were involved in clusters: 156 patients (11.5%) in 78 dyads and 231 participants (17%) in 42 larger clusters (median size: 4, range 3-41). Compared to individuals with single PHI (n = 969), those in clusters were more frequently men (95.9 vs 83%, p turnaround time for sample processing, targeting prevention efforts based on phylogenetic monitoring may be an efficient way to deliver prevention interventions but would require near real time targeted interventions on the identified index cases and their partners.

  7. Molecular Basis for Drug Resistance in HIV-1 Protease

    Directory of Open Access Journals (Sweden)

    Celia A. Schiffer

    2010-11-01

    Full Text Available HIV-1 protease is one of the major antiviral targets in the treatment of patients infected with HIV-1. The nine FDA approved HIV-1 protease inhibitors were developed with extensive use of structure-based drug design, thus the atomic details of how the inhibitors bind are well characterized. From this structural understanding the molecular basis for drug resistance in HIV-1 protease can be elucidated. Selected mutations in response to therapy and diversity between clades in HIV-1 protease have altered the shape of the active site, potentially altered the dynamics and even altered the sequence of the cleavage sites in the Gag polyprotein. All of these interdependent changes act in synergy to confer drug resistance while simultaneously maintaining the fitness of the virus. New strategies, such as incorporation of the substrate envelope constraint to design robust inhibitors that incorporate details of HIV-1 protease’s function and decrease the probability of drug resistance, are necessary to continue to effectively target this key protein in HIV-1 life cycle.

  8. Modulation of chromatin structure by the FACT histone chaperone complex regulates HIV-1 integration.

    Science.gov (United States)

    Matysiak, Julien; Lesbats, Paul; Mauro, Eric; Lapaillerie, Delphine; Dupuy, Jean-William; Lopez, Angelica P; Benleulmi, Mohamed Salah; Calmels, Christina; Andreola, Marie-Line; Ruff, Marc; Llano, Manuel; Delelis, Olivier; Lavigne, Marc; Parissi, Vincent

    2017-07-28

    Insertion of retroviral genome DNA occurs in the chromatin of the host cell. This step is modulated by chromatin structure as nucleosomes compaction was shown to prevent HIV-1 integration and chromatin remodeling has been reported to affect integration efficiency. LEDGF/p75-mediated targeting of the integration complex toward RNA polymerase II (polII) transcribed regions ensures optimal access to dynamic regions that are suitable for integration. Consequently, we have investigated the involvement of polII-associated factors in the regulation of HIV-1 integration. Using a pull down approach coupled with mass spectrometry, we have selected the FACT (FAcilitates Chromatin Transcription) complex as a new potential cofactor of HIV-1 integration. FACT is a histone chaperone complex associated with the polII transcription machinery and recently shown to bind LEDGF/p75. We report here that a tripartite complex can be formed between HIV-1 integrase, LEDGF/p75 and FACT in vitro and in cells. Biochemical analyzes show that FACT-dependent nucleosome disassembly promotes HIV-1 integration into chromatinized templates, and generates highly favored nucleosomal structures in vitro. This effect was found to be amplified by LEDGF/p75. Promotion of this FACT-mediated chromatin remodeling in cells both increases chromatin accessibility and stimulates HIV-1 infectivity and integration. Altogether, our data indicate that FACT regulates HIV-1 integration by inducing local nucleosomes dissociation that modulates the functional association between the incoming intasome and the targeted nucleosome.

  9. HLA Class I-Mediated HIV-1 Control in Vietnamese Infected with HIV-1 Subtype A/E.

    Science.gov (United States)

    Chikata, Takayuki; Tran, Giang Van; Murakoshi, Hayato; Akahoshi, Tomohiro; Qi, Ying; Naranbhai, Vivek; Kuse, Nozomi; Tamura, Yoshiko; Koyanagi, Madoka; Sakai, Sachiko; Nguyen, Dung Hoai; Nguyen, Dung Thi; Nguyen, Ha Thu; Nguyen, Trung Vu; Oka, Shinichi; Martin, Maureen P; Carrington, Mary; Sakai, Keiko; Nguyen, Kinh Van; Takiguchi, Masafumi

    2018-03-01

    HIV-1-specific cytotoxic T cells (CTLs) play an important role in the control of HIV-1 subtype B or C infection. However, the role of CTLs in HIV-1 subtype A/E infection still remains unclear. Here we investigated the association of HLA class I alleles with clinical outcomes in treatment-naive Vietnamese infected with subtype A/E virus. We found that HLA-C*12:02 was significantly associated with lower plasma viral loads (pVL) and higher CD4 counts and that the HLA-A*29:01-B*07:05-C*15:05 haplotype was significantly associated with higher pVL and lower CD4 counts than those for individuals without these respective genotypes. Nine Pol and three Nef mutations were associated with at least one HLA allele in the HLA-A*29:01-B*07:05-C*15:05 haplotype, with a strong negative correlation between the number of HLA-associated Pol mutations and CD4 count as well as a positive correlation with pVL for individuals with these HLA alleles. The results suggest that the accumulation of mutations selected by CTLs restricted by these HLA alleles affects HIV control. IMPORTANCE Most previous studies on HLA association with disease progression after HIV-1 infection have been performed on cohorts infected with HIV-1 subtypes B and C, whereas few such population-based studies have been reported for cohorts infected with the Asian subtype A/E virus. In this study, we analyzed the association of HLA class I alleles with clinical outcomes for 536 HIV-1 subtype A/E-infected Vietnamese individuals. We found that HLA-C*12:02 is protective, while the HLA haplotype HLA-A*29:01-B*07:05-C*15:05 is deleterious. The individuals with HIV-1 mutations associated with at least one of the HLA alleles in the deleterious HLA haplotype had higher plasma viral loads and lower CD4 counts than those of individuals without the mutations, suggesting that viral adaptation and escape from HLA-mediated immune control occurred. The present study identifies a protective allele and a deleterious haplotype for HIV-1

  10. A zinc(II)-based two-dimensional MOF for sensitive and selective sensing of HIV-1 ds-DNA sequences

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Hai-Qing; Qiu, Gui-Hua; Liang, Zhen [Guangdong Provincial Key Laboratory of New Drug Screening, School of Pharmaceutical Sciences, Southern Medical University, Guangzhou 510515 (China); Li, Min-Min [The First Affiliated Hospital of Jinan University, Guangzhou 510515 (China); Sun, Bin; Qin, Liang; Yang, Shui-Ping; Chen, Wen-Hua [Guangdong Provincial Key Laboratory of New Drug Screening, School of Pharmaceutical Sciences, Southern Medical University, Guangzhou 510515 (China); Chen, Jin-Xiang, E-mail: jxchen@smu.edu.cn [Guangdong Provincial Key Laboratory of New Drug Screening, School of Pharmaceutical Sciences, Southern Medical University, Guangzhou 510515 (China)

    2016-05-30

    Coordination reaction of a known three-dimensional (3D) polymer precursor {Na_3[Na_9(Cbdcp)_6(H_2O)_1_8]}{sub n} (A, Cbdcp = N-(4-carboxybenzyl)-(3,5-dicarboxyl)pyridinium) with Zn(NO{sub 3}){sub 2}·6H{sub 2}O in H{sub 2}O or H{sub 2}O/DMF at 100 °C and in the presence of aspirin, 5-fluorouracil (5-FU) as modulators, trans-1,2-bis(4-pyridyl)ethylene (bpe) or 1,2-bis(4-pyridyl)ethane (bpea) as ancillary ligands afforded six novel Zn(II)-based metal-organic frameworks (MOFs), that is, {[Zn(Cbdcp)(H_2O)_3]·H_2O}{sub n} (1, 1D zigzag chain), {[Zn(HCbdcp)_2]·H_2O}{sub n} (2, 2D sheet), {[Zn(Cbdcp)(bpe)_1_/_2]·2H_2O}{sub n} (3, 3D polymer), {[Zn(Cbdcp)(bpe)_1_/_2]·2H_2O}{sub n} (4, 2D network), {[Zn(Cbdcp)(bpea)_1_/_2]·2H_2O}{sub n} (5, 3D polymer) and {[Zn(Cbdcp)(bpea)_1_/_2]·2H_2O}{sub n} (6, 2D network). Among them, compound 2 contains aromatic rings, positively charged pyridinium, Zn{sup 2+} cation centers and carboxylic acid groups lined up on the 2D sheet structure with a certain extended surface exposure. The unique structure of 2 facilitates effective association with carboxyfluorescein (FAM) labeled probe single stranded DNA (probe ss-DNA, delineates as P-DNA) to yield a P-DNA@2 system, and leads to fluorescence quenching of FAM via a photoinduced electron transfer process. The P-DNA@2 system is effective and reliable for the detection of human immunodeficiency virus 1 ds-DNA (HIV ds-DNA) sequences and capable of distinguishing complementary HIV ds-DNA from mismatched target sequences with the detection limit as low as 10 pM (S/N = 3). - Graphical abstract: Six water-stable zinc(II) zwitterionic carboxylate compounds with 1D chain, 2D and 3D networks were synthesized. Compound 2 can interact with the probe DNA through noncovalent bonds to form P-DNA@2 system. This system can be used as an effective, fluorescent sensing platform for the detection of HIV ds-DNA with the detection limit as low as 10 pM. - Highlights: • Six water-stable zinc

  11. Detection of Hepatitis B Virus (HBV) Genomes and HBV Drug Resistant Variants by Deep Sequencing Analysis of HBV Genomes in Immune Cell Subsets of HBV Mono-Infected and/or Human Immunodeficiency Virus Type-1 (HIV-1) and HBV Co-Infected Individuals

    Science.gov (United States)

    Lee, Z.; Nishikawa, S.; Gao, S.; Eksteen, J. B.; Czub, M.; Gill, M. J.; Osiowy, C.; van der Meer, F.; van Marle, G.; Coffin, C. S.

    2015-01-01

    The hepatitis B virus (HBV) and the human immunodeficiency virus type 1 (HIV-1) can infect cells of the lymphatic system. It is unknown whether HIV-1 co-infection impacts infection of peripheral blood mononuclear cell (PBMC) subsets by the HBV. Aims To compare the detection of HBV genomes and HBV sequences in unsorted PBMCs and subsets (i.e., CD4+ T, CD8+ T, CD14+ monocytes, CD19+ B, CD56+ NK cells) in HBV mono-infected vs. HBV/HIV-1 co-infected individuals. Methods Total PBMC and subsets isolated from 14 HBV mono-infected (4/14 before and after anti-HBV therapy) and 6 HBV/HIV-1 co-infected individuals (5/6 consistently on dual active anti-HBV/HIV therapy) were tested for HBV genomes, including replication indicative HBV covalently closed circular (ccc)-DNA, by nested PCR/nucleic hybridization and/or quantitative PCR. In CD4+, and/or CD56+ subsets from two HBV monoinfected cases, the HBV polymerase/overlapping surface region was analyzed by next generation sequencing. Results All analyzed whole PBMC from HBV monoinfected and HBV/HIV coinfected individuals were HBV genome positive. Similarly, HBV DNA was detected in all target PBMC subsets regardless of antiviral therapy, but was absent from the CD4+ T cell subset from all HBV/HIV-1 positive cases (PHBV monoinfected cases on tenofovir therapy, mutations at residues associated with drug resistance and/or immune escape (i.e., G145R) were detected in a minor percentage of the population. Summary HBV genomes and drug resistant variants were detectable in PBMC subsets from HBV mono-infected individuals. The HBV replicates in PBMC subsets of HBV/HIV-1 patients except the CD4+ T cell subpopulation. PMID:26390290

  12. Molecular epidemiology of HIV-1 subtype A in former Soviet Union countries.

    Science.gov (United States)

    Aibekova, Lazzat; Foley, Brian; Hortelano, Gonzalo; Raees, Muhammad; Abdraimov, Sabit; Toichuev, Rakhmanbek; Ali, Syed

    2018-01-01

    While in other parts of the world it is on decline, incidence of HIV infection continues to rise in the former Soviet Union (FSU) countries. The present study was conducted to investigate the patterns and modes of HIV transmission in FSU countries. We performed phylogenetic analysis of publicly available 2705 HIV-1 subtype A pol sequences from thirteen FSU countries: Armenia, Azerbaijan, Belarus, Estonia, Georgia, Kazakhstan, Kyrgyzstan, Latvia, Lithuania, Moldova, Russia, Ukraine and Uzbekistan. Our analysis showed that the clusters from FSU countries were intermixed, indicating a possible role of transmigration in HIV transmission. Injection drug use was found to be the most frequent mode of transmission, while the clusters from PWID and heterosexual transmission were intermixed, indicating bridging of HIV infection across populations. To control the expanding HIV epidemic in this region, harm reduction strategies should be focused on three modes of transmission, namely, cross-border migration, injection drug use and heterosexual.

  13. Rare HIV-1 Subtype J Genomes and a New H/U/CRF02_AG Recombinant Genome Suggests an Ancient Origin of HIV-1 in Angola.

    Science.gov (United States)

    Bártolo, Inês; Calado, Rita; Borrego, Pedro; Leitner, Thomas; Taveira, Nuno

    2016-08-01

    Angola has an extremely diverse HIV-1 epidemic fueled in part by the frequent interchange of people with the Democratic Republic of Congo (DRC) and Republic of Congo (RC). Characterization of HIV-1 strains circulating in Angola should help to better understand the origin of HIV-1 subtypes and recombinant forms and their transmission dynamics. In this study we characterize the first near full-length HIV-1 genomic sequences from HIV-1 infected individuals from Angola. Samples were obtained in 1993 from three HIV-1 infected patients living in Cabinda, Angola. Near full-length genomic sequences were obtained from virus isolates. Maximum likelihood phylogenetic tree inference and analyses of potential recombination patterns were performed to evaluate the sequence classifications and origins. Phylogenetic and recombination analyses revealed that one virus was a pure subtype J, another mostly subtype J with a small uncertain region, and the final virus was classified as a H/U/CRF02_AG recombinant. Consistent with their epidemiological data, the subtype J sequences were more closely related to each other than to other J sequences previously published. Based on the env gene, taxa from Angola occur throughout the global subtype J phylogeny. HIV-1 subtypes J and H are present in Angola at low levels since at least 1993. Low transmission efficiency and/or high recombination potential may explain their limited epidemic success in Angola and worldwide. The high diversity of rare subtypes in Angola suggests that Angola was part of the early establishment of the HIV-1 pandemic.

  14. An evolutionary model-based algorithm for accurate phylogenetic breakpoint mapping and subtype prediction in HIV-1.

    Directory of Open Access Journals (Sweden)

    Sergei L Kosakovsky Pond

    2009-11-01

    Full Text Available Genetically diverse pathogens (such as Human Immunodeficiency virus type 1, HIV-1 are frequently stratified into phylogenetically or immunologically defined subtypes for classification purposes. Computational identification of such subtypes is helpful in surveillance, epidemiological analysis and detection of novel variants, e.g., circulating recombinant forms in HIV-1. A number of conceptually and technically different techniques have been proposed for determining the subtype of a query sequence, but there is not a universally optimal approach. We present a model-based phylogenetic method for automatically subtyping an HIV-1 (or other viral or bacterial sequence, mapping the location of breakpoints and assigning parental sequences in recombinant strains as well as computing confidence levels for the inferred quantities. Our Subtype Classification Using Evolutionary ALgorithms (SCUEAL procedure is shown to perform very well in a variety of simulation scenarios, runs in parallel when multiple sequences are being screened, and matches or exceeds the performance of existing approaches on typical empirical cases. We applied SCUEAL to all available polymerase (pol sequences from two large databases, the Stanford Drug Resistance database and the UK HIV Drug Resistance Database. Comparing with subtypes which had previously been assigned revealed that a minor but substantial (approximately 5% fraction of pure subtype sequences may in fact be within- or inter-subtype recombinants. A free implementation of SCUEAL is provided as a module for the HyPhy package and the Datamonkey web server. Our method is especially useful when an accurate automatic classification of an unknown strain is desired, and is positioned to complement and extend faster but less accurate methods. Given the increasingly frequent use of HIV subtype information in studies focusing on the effect of subtype on treatment, clinical outcome, pathogenicity and vaccine design, the importance

  15. Measuring replication competent HIV-1: advances and challenges in defining the latent reservoir.

    Science.gov (United States)

    Wang, Zheng; Simonetti, Francesco R; Siliciano, Robert F; Laird, Gregory M

    2018-02-13

    Antiretroviral therapy cannot cure HIV-1 infection due to the persistence of a small number of latently infected cells harboring replication-competent proviruses. Measuring persistent HIV-1 is challenging, as it consists of a mosaic population of defective and intact proviruses that can shift from a state of latency to active HIV-1 transcription. Due to this complexity, most of the current assays detect multiple categories of persistent HIV-1, leading to an overestimate of the true size of the latent reservoir. Here, we review the development of the viral outgrowth assay, the gold-standard quantification of replication-competent proviruses, and discuss the insights provided by full-length HIV-1 genome sequencing methods, which allowed us to unravel the composition of the proviral landscape. In this review, we provide a dissection of what defines HIV-1 persistence and we examine the unmet needs to measure the efficacy of interventions aimed at eliminating the HIV-1 reservoir.

  16. Dendritic cells exposed to MVA-based HIV-1 vaccine induce highly functional HIV-1-specific CD8(+ T cell responses in HIV-1-infected individuals.

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    Núria Climent

    Full Text Available Currently, MVA virus vectors carrying HIV-1 genes are being developed as HIV-1/AIDS prophylactic/therapeutic vaccines. Nevertheless, little is known about the impact of these vectors on human dendritic cells (DC and their capacity to present HIV-1 antigens to human HIV-specific T cells. This study aimed to characterize the interaction of MVA and MVA expressing the HIV-1 genes Env-Gag-Pol-Nef of clade B (referred to as MVA-B in human monocyte-derived dendritic cells (MDDC and the subsequent processes of HIV-1 antigen presentation and activation of memory HIV-1-specific T lymphocytes. For these purposes, we performed ex vivo assays with MDDC and autologous lymphocytes from asymptomatic HIV-infected patients. Infection of MDDC with MVA-B or MVA, at the optimal dose of 0.3 PFU/MDDC, induced by itself a moderate degree of maturation of MDDC, involving secretion of cytokines and chemokines (IL1-ra, IL-7, TNF-α, IL-6, IL-12, IL-15, IL-8, MCP-1, MIP-1α, MIP-1β, RANTES, IP-10, MIG, and IFN-α. MDDC infected with MVA or MVA-B and following a period of 48 h or 72 h of maturation were able to migrate toward CCL19 or CCL21 chemokine gradients. MVA-B infection induced apoptosis of the infected cells and the resulting apoptotic bodies were engulfed by the uninfected MDDC, which cross-presented HIV-1 antigens to autologous CD8(+ T lymphocytes. MVA-B-infected MDDC co-cultured with autologous T lymphocytes induced a highly functional HIV-specific CD8(+ T cell response including proliferation, secretion of IFN-γ, IL-2, TNF-α, MIP-1β, MIP-1α, RANTES and IL-6, and strong cytotoxic activity against autologous HIV-1-infected CD4(+ T lymphocytes. These results evidence the adjuvant role of the vector itself (MVA and support the clinical development of prophylactic and therapeutic anti-HIV vaccines based on MVA-B.

  17. Clustering patterns of cytotoxic T-lymphocyte epitopes in human immunodeficiency virus type 1 (HIV-1) proteins reveal imprints of immune evasion on HIV-1 global variation

    DEFF Research Database (Denmark)

    Yusim, K.; Kesmir, Can; Gaschen, B.

    2002-01-01

    The human cytotoxic T-lymphocyte (CTL) response to human immunodeficiency virus type 1 (HIV-1) has been intensely studied, and hundreds of CTL epitopes have been experimentally defined, published, and compiled in the HIV Molecular Immunology Database. Maps of CTL epitopes on HIV-1 protein sequenc...

  18. Origin and Population Dynamics of a Novel HIV-1 Subtype G Clade Circulating in Cape Verde and Portugal.

    Science.gov (United States)

    de Pina-Araujo, Isabel Inês M; Delatorre, Edson; Guimarães, Monick L; Morgado, Mariza G; Bello, Gonzalo

    2015-01-01

    The human immunodeficiency virus type 1 (HIV-1) subtype G is the most prevalent and second most prevalent HIV-1 clade in Cape Verde and Portugal, respectively; but there is no information about the origin and spatiotemporal dispersal pattern of this HIV-1 clade circulating in those countries. To this end, we used Maximum Likelihood and Bayesian coalescent-based methods to analyze a collection of 578 HIV-1 subtype G pol sequences sampled throughout Portugal, Cape Verde and 11 other countries from West and Central Africa over a period of 22 years (1992 to 2013). Our analyses indicate that most subtype G sequences from Cape Verde (80%) and Portugal (95%) branched together in a distinct monophyletic cluster (here called G(CV-PT)). The G(CV-PT) clade probably emerged after a single migration of the virus out of Central Africa into Cape Verde between the late 1970s and the middle 1980s, followed by a rapid dissemination to Portugal a couple of years later. Reconstruction of the demographic history of the G(CV-PT) clade circulating in Cape Verde and Portugal indicates that this viral clade displayed an initial phase of exponential growth during the 1980s and 1990s, followed by a decline in growth rate since the early 2000s. Our data also indicate that during the exponential growth phase the G(CV-PT) clade recombined with a preexisting subtype B viral strain circulating in Portugal, originating the CRF14_BG clade that was later disseminated to Spain and Cape Verde. Historical and recent human population movements between Angola, Cape Verde and Portugal probably played a key role in the origin and dispersal of the G(CV-PT )and CRF14_BG clades.

  19. HIV-1 and the macrophage

    NARCIS (Netherlands)

    Bol, Sebastiaan M.; Cobos-Jimenez, Viviana; Kootstra, Neeltje A.; van 't Wout, Angelique B.

    2011-01-01

    Macrophages and CD4(+) T cells are natural target cells for HIV-1, and both cell types contribute to the establishment of the viral reservoir that is responsible for continuous residual virus replication during antiretroviral therapy and viral load rebound upon treatment interruption. Scientific

  20. Developing strategies for HIV-1 eradication

    Science.gov (United States)

    Durand, Christine M.; Blankson, Joel N.; Siliciano, Robert F.

    2014-01-01

    Highly active antiretroviral therapy (HAART) suppresses HIV-1 replication, transforming the outlook for infected patients. However, reservoirs of replication-competent forms of the virus persist during HAART, and when treatment is stopped, high rates of HIV-1 replication return. Recent insights into HIV-1 latency, as well as a report that HIV-1 infection was eradicated in one individual, have renewed interest in finding a cure for HIV-1 infection. Strategies for HIV-1 eradication include gene therapy and hematopoietic stem cell transplantation, stimulating host immunity to control HIV-1 replication, and targeting latent HIV-1 in resting memory CD4+ T cells. Future efforts should aim to provide better understanding of how to reconstitute the CD4+ T cell compartment with genetically engineered cells, exert immune control over HIV-1 replication, and identify and eliminate all viral reservoirs. PMID:22867874

  1. Phylogenetic analysis consistent with a clinical history of sexual transmission of HIV-1 from a single donor reveals transmission of highly distinct variants

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    McClure Myra

    2011-07-01

    Full Text Available Abstract Background To combat the pandemic of human immunodeficiency virus 1 (HIV-1, a successful vaccine will need to cope with the variability of transmissible viruses. Human hosts infected with HIV-1 potentially harbour many viral variants but very little is known about viruses that are likely to be transmitted, or even if there are viral characteristics that predict enhanced transmission in vivo. We show for the first time that genetic divergence consistent with a single transmission event in vivo can represent several years of pre-transmission evolution. Results We describe a highly unusual case consistent with a single donor transmitting highly related but distinct HIV-1 variants to two individuals on the same evening. We confirm that the clustering of viral genetic sequences, present within each recipient, is consistent with the history of a single donor across the viral env, gag and pol genes by maximum likelihood and Bayesian Markov Chain Monte Carlo based phylogenetic analyses. Based on an uncorrelated, lognormal relaxed clock of env gene evolution calibrated with other datasets, the time since the most recent common ancestor is estimated as 2.86 years prior to transmission (95% confidence interval 1.28 to 4.54 years. Conclusion Our results show that an effective design for a preventative vaccine will need to anticipate extensive HIV-1 diversity within an individual donor as well as diversity at the population level.

  2. The HIV-1 transcriptional activator Tat has potent nucleic acid chaperoning activities in vitro.

    Science.gov (United States)

    Kuciak, Monika; Gabus, Caroline; Ivanyi-Nagy, Roland; Semrad, Katharina; Storchak, Roman; Chaloin, Olivier; Muller, Sylviane; Mély, Yves; Darlix, Jean-Luc

    2008-06-01

    The human immunodeficiency virus type 1 (HIV-1) is a primate lentivirus that causes the acquired immunodeficiency syndrome (AIDS). In addition to the virion structural proteins and enzyme precursors, that are Gag, Env and Pol, HIV-1 encodes several regulatory proteins, notably a small nuclear transcriptional activator named Tat. The Tat protein is absolutely required for virus replication since it controls proviral DNA transcription to generate the full-length viral mRNA. Tat can also regulate mRNA capping and splicing and was recently found to interfere with the cellular mi- and siRNA machinery. Because of its extensive interplay with nucleic acids, and its basic and disordered nature we speculated that Tat had nucleic acid-chaperoning properties. This prompted us to examine in vitro the nucleic acid-chaperoning activities of Tat and Tat peptides made by chemical synthesis. Here we report that Tat has potent nucleic acid-chaperoning activities according to the standard DNA annealing, DNA and RNA strand exchange, RNA ribozyme cleavage and trans-splicing assays. The active Tat(44-61) peptide identified here corresponds to the smallest known sequence with DNA/RNA chaperoning properties.

  3. Rational development of radiopharmaceuticals for HIV-1

    International Nuclear Information System (INIS)

    Lau, Chuen-Yen; Maldarelli, Frank; Eckelman, William C.; Neumann, Ronald D.

    2014-01-01

    The global battle against HIV-1 would benefit from a sensitive and specific radiopharmaceutical to localize HIV-infected cells. Ideally, this probe would be able to identify latently infected host cells containing replication competent HIV sequences. Clinical and research applications would include assessment of reservoirs, informing clinical management by facilitating assessment of burden of infection in different compartments, monitoring disease progression and monitoring response to therapy. A “rational” development approach could facilitate efficient identification of an appropriate targeted radiopharmaceutical. Rational development starts with understanding characteristics of the disease that can be effectively targeted and then engineering radiopharmaceuticals to hone in on an appropriate target, which in the case of HIV-1 (HIV) might be an HIV-specific product on or in the host cell, a differentially expressed gene product, an integrated DNA sequence specific enzymatic activity, part of the inflammatory response, or a combination of these. This is different from the current approach that starts with a radiopharmaceutical for a target associated with a disease, mostly from autopsy studies, without a strong rationale for the potential to impact patient care. At present, no targeted therapies are available for HIV latency, although a number of approaches are under study. Here we discuss requirements for a radiopharmaceutical useful in strategies targeting persistently infected cells. The radiopharmaceutical for HIV should be developed based on HIV biology, studied in an animal model and then in humans, and ultimately used in clinical and research settings

  4. HIV-1 subtype D infections among Caucasians from Northwestern Poland--phylogenetic and clinical analysis.

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    Miłosz Parczewski

    Full Text Available BACKGROUND: HIV-1 subtype D infections, which are associated with a faster rate of progression and lymphocyte CD4 decline, cognitive deficit and higher mortality, have rarely been found in native Europeans. In Northwestern Poland, however, infections with this subtype had been identified. This study aimed to analyze the sequence and clinical data for patients with subtype D using molecular phylogeography and identify transmission clusters and ancestry, as well as drug resistance, baseline HIV tropism and antiretroviral treatment efficacy. METHODS: Phylogenetic analyses of local HIV-1 subtype D sequences were performed, with time to the most recent common ancestor inferred using bayesian modeling. Sequence and drug resistance data were linked with the clinical and epidemiological information. RESULTS: Subtype D was found in 24 non-immigrant Caucasian, heterosexually infected patients (75% of females, median age at diagnosis of 49.5 years; IQR: 29-56 years. Partial pol sequences clustered monophyletically with the clades of Ugandan origin and no evidence of transmission from other European countries was found. Time to the most common recent ancestor was 1989.24 (95% HPD: 1968.83-1994.46. Baseline drug resistance to nucleoside reverse transcriptase inhibitors was observed in 54.5% of cases (mutations: M41L, K103N, T215S/D with evidence of clustering, no baseline integrase or protease resistance and infrequent non-R5 tropism (13.6%. Virologic failure was observed in 60% of cases and was associated with poor adherence (p<0.001 and subsequent development of drug resistance (p = 0.008, OR: 20 (95%CI: 1.7-290. CONCLUSIONS: Local subtype D represented an independently transmitted network with probably single index case, high frequency of primary drug resistance and evidence of transmission clusters.

  5. Sieve analysis in HIV-1 vaccine efficacy trials.

    Science.gov (United States)

    Edlefsen, Paul T; Gilbert, Peter B; Rolland, Morgane

    2013-09-01

    The genetic characterization of HIV-1 breakthrough infections in vaccine and placebo recipients offers new ways to assess vaccine efficacy trials. Statistical and sequence analysis methods provide opportunities to mine the mechanisms behind the effect of an HIV vaccine. The release of results from two HIV-1 vaccine efficacy trials, Step/HVTN-502 (HIV Vaccine Trials Network-502) and RV144, led to numerous studies in the last 5 years, including efforts to sequence HIV-1 breakthrough infections and compare viral characteristics between the vaccine and placebo groups. Novel genetic and statistical analysis methods uncovered features that distinguished founder viruses isolated from vaccinees from those isolated from placebo recipients, and identified HIV-1 genetic targets of vaccine-induced immune responses. Studies of HIV-1 breakthrough infections in vaccine efficacy trials can provide an independent confirmation to correlates of risk studies, as they take advantage of vaccine/placebo comparisons, whereas correlates of risk analyses are limited to vaccine recipients. Through the identification of viral determinants impacted by vaccine-mediated host immune responses, sieve analyses can shed light on potential mechanisms of vaccine protection.

  6. Epidemiological trends of HIV-1 infection in blood donors from Catalonia, Spain (2005-2014).

    Science.gov (United States)

    Bes, Marta; Piron, Maria; Casamitjana, Natàlia; Gregori, Josep; Esteban, Juan Ignacio; Ribera, Esteban; Quer, Josep; Puig, Lluís; Sauleda, Sílvia

    2017-09-01

    Human immunodeficiency virus 1 (HIV-1) subtype B is predominant in Spain. However, the recent arrival of immigrant populations has increased the prevalence of non-B subtypes and circulating recombinant forms. The objective of this study was to determine the prevalence of HIV-1 subtypes and transmitted drug-resistance mutations in blood donors from the Catalonian region (northeastern Spain). HIV-1-positive blood donors identified in Catalonia from 2005 to 2014 were included. Demographic variables and risk factors for HIV-1 acquisition were recorded. HIV-1 subtyping was carried out by HIV-1 DNA polymerase region sequencing, and phylogenetic analyses were performed using the neighbor-joining method. During the study period, 2.8 million blood donations were screened, and 214 HIV-1-positive donors were identified, yielding an overall prevalence of 7.7 per 100,000 donations (89% men; mean age, 34 ± 10 years). Most HIV-1-positive donors were native to Spain (81%), and 61% were regular blood donors. When risk factors were known, 62% reportedly were men who had sex with men. HIV-1 subtyping was possible in 176 HIV-1-positive individuals: 143 (81%) had HIV-1 subtype B, and 33 (19%) had non-B subtypes. Most HIV-1 non-B subtypes were circulating recombinant forms (n = 20; 61%). Factors associated with HIV-1 subtype B were male sex (p = 0.007) and men who had sex with men (p HIV-1-positive blood donors in Catalonia. Continuous local epidemiological surveillance is required to implement optimal prevention strategies for controlling transfusion-transmitted HIV and to improve health policies regarding HIV infection. © 2017 AABB.

  7. Conservation patterns of HIV-1 RT connection and RNase H domains: identification of new mutations in NRTI-treated patients.

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    André F A Santos

    Full Text Available BACKGROUND: Although extensive HIV drug resistance information is available for the first 400 amino acids of its reverse transcriptase, the impact of antiretroviral treatment in C-terminal domains of Pol (thumb, connection and RNase H is poorly understood. METHODS AND FINDINGS: We wanted to characterize conserved regions in RT C-terminal domains among HIV-1 group M subtypes and CRF. Additionally, we wished to identify NRTI-related mutations in HIV-1 RT C-terminal domains. We sequenced 118 RNase H domains from clinical viral isolates in Brazil, and analyzed 510 thumb and connection domain and 450 RNase H domain sequences collected from public HIV sequence databases, together with their treatment status and histories. Drug-naïve and NRTI-treated datasets were compared for intra- and inter-group conservation, and differences were determined using Fisher's exact tests. One third of RT C-terminal residues were found to be conserved among group M variants. Three mutations were found exclusively in NRTI-treated isolates. Nine mutations in the connection and 6 mutations in the RNase H were associated with NRTI treatment in subtype B. Some of them lay in or close to amino acid residues which contact nucleic acid or near the RNase H active site. Several of the residues pointed out herein have been recently associated to NRTI exposure or increase drug resistance to NRTI. CONCLUSIONS: This is the first comprehensive genotypic analysis of a large sequence dataset that describes NRTI-related mutations in HIV-1 RT C-terminal domains in vivo. The findings into the conservation of RT C-terminal domains may pave the way to more rational drug design initiatives targeting those regions.

  8. An Efficient Microarray-Based Genotyping Platform for the Identification of Drug-Resistance Mutations in Majority and Minority Subpopulations of HIV-1 Quasispecies.

    Science.gov (United States)

    Martín, Verónica; Perales, Celia; Fernández-Algar, María; Dos Santos, Helena G; Garrido, Patricia; Pernas, María; Parro, Víctor; Moreno, Miguel; García-Pérez, Javier; Alcamí, José; Torán, José Luis; Abia, David; Domingo, Esteban; Briones, Carlos

    2016-01-01

    The response of human immunodeficiency virus type 1 (HIV-1) quasispecies to antiretroviral therapy is influenced by the ensemble of mutants that composes the evolving population. Low-abundance subpopulations within HIV-1 quasispecies may determine the viral response to the administered drug combinations. However, routine sequencing assays available to clinical laboratories do not recognize HIV-1 minority variants representing less than 25% of the population. Although several alternative and more sensitive genotyping techniques have been developed, including next-generation sequencing (NGS) methods, they are usually very time consuming, expensive and require highly trained personnel, thus becoming unrealistic approaches in daily clinical practice. Here we describe the development and testing of a HIV-1 genotyping DNA microarray that detects and quantifies, in majority and minority viral subpopulations, relevant mutations and amino acid insertions in 42 codons of the pol gene associated with drug- and multidrug-resistance to protease (PR) and reverse transcriptase (RT) inhibitors. A customized bioinformatics protocol has been implemented to analyze the microarray hybridization data by including a new normalization procedure and a stepwise filtering algorithm, which resulted in the highly accurate (96.33%) detection of positive/negative signals. This microarray has been tested with 57 subtype B HIV-1 clinical samples extracted from multi-treated patients, showing an overall identification of 95.53% and 89.24% of the queried PR and RT codons, respectively, and enough sensitivity to detect minority subpopulations representing as low as 5-10% of the total quasispecies. The developed genotyping platform represents an efficient diagnostic and prognostic tool useful to personalize antiviral treatments in clinical practice.

  9. Curcumin derivatives as HIV-1 protease inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Sui, Z.; Li, J.; Craik, C.S.; Ortiz de Montellano, P.R. [Univ. of California, San Francisco, CA (United States)

    1993-12-31

    Curcumin, a non-toxic natural compound from Curcuma longa, has been found to be an HIV-1 protease inhibitor. Some of its derivatives were synthesized and their inhibitory activity against the HIV-1 protease was tested. Curcumin analogues containing boron enhanced the inhibitory activity. At least of the the synthesized compounds irreversibly inhibits the HIV-1 protease.

  10. Cyclophilin B enhances HIV-1 infection

    Energy Technology Data Exchange (ETDEWEB)

    DeBoer, Jason; Madson, Christian J. [Department of Medical Microbiology and Immunology, Creighton University, Omaha, NE (United States); Belshan, Michael, E-mail: michaelbelshan@creighton.edu [Department of Medical Microbiology and Immunology, Creighton University, Omaha, NE (United States); The Nebraska Center for Virology, University of Nebraska, Lincoln, NE (United States)

    2016-02-15

    Cyclophilin B (CypB) is a member of the immunophilin family and intracellular chaperone. It predominantly localizes to the ER, but also contains a nuclear localization signal and is secreted from cells. CypB has been shown to interact with the Gag protein of human immunodeficiency type 1 (HIV-1). Several proteomic and genetic studies identified it as a potential factor involved in HIV replication. Herein, we show that over-expression of CypB enhances HIV infection by increasing nuclear import of viral DNA. This enhancement was unaffected by cyclosporine treatment and requires the N-terminus of the protein. The N-terminus contains an ER leader sequence, putative nuclear localization signal, and is required for secretion. Deletion of the N-terminus resulted in mislocalization from the ER and suppression of HIV infection. Passive transfer experiments showed that secreted CypB did not impact HIV infection. Combined, these experiments show that intracellular CypB modulates a pathway of HIV nuclear import. - Highlights: • CypB has been identified in several proteomic studies of HIV-1 infection. • CypB expression is upregulated in activated and infected T-cells. • Over-expression of CypB enhances HIV nuclear import and infection. • The N-terminus of CypB is necessary for these effects.

  11. Cyclophilin B enhances HIV-1 infection

    International Nuclear Information System (INIS)

    DeBoer, Jason; Madson, Christian J.; Belshan, Michael

    2016-01-01

    Cyclophilin B (CypB) is a member of the immunophilin family and intracellular chaperone. It predominantly localizes to the ER, but also contains a nuclear localization signal and is secreted from cells. CypB has been shown to interact with the Gag protein of human immunodeficiency type 1 (HIV-1). Several proteomic and genetic studies identified it as a potential factor involved in HIV replication. Herein, we show that over-expression of CypB enhances HIV infection by increasing nuclear import of viral DNA. This enhancement was unaffected by cyclosporine treatment and requires the N-terminus of the protein. The N-terminus contains an ER leader sequence, putative nuclear localization signal, and is required for secretion. Deletion of the N-terminus resulted in mislocalization from the ER and suppression of HIV infection. Passive transfer experiments showed that secreted CypB did not impact HIV infection. Combined, these experiments show that intracellular CypB modulates a pathway of HIV nuclear import. - Highlights: • CypB has been identified in several proteomic studies of HIV-1 infection. • CypB expression is upregulated in activated and infected T-cells. • Over-expression of CypB enhances HIV nuclear import and infection. • The N-terminus of CypB is necessary for these effects.

  12. Striking HIV-1 Entry by Targeting HIV-1 gp41. But, Where Should We Target?

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    Cátia Teixeira

    Full Text Available HIV-1 gp41 facilitates the viral fusion through a conformational switch involving the association of three C-terminal helices along the conserved hydrophobic grooves of three N-terminal helices coiled-coil. The control of these structural rearrangements is thought to be central to HIV-1 entry and, therefore, different strategies of intervention are being developed. Herewith, we describe a procedure to simulate the folding of an HIV-1 gp41 simplified model. This procedure is based on the construction of plausible conformational pathways, which describe protein transition between non-fusogenic and fusogenic conformations. The calculation of the paths started with 100 molecular dynamics simulations of the non-fusogenic conformation, which were found to converge to different intermediate states. Those presenting defined criteria were selected for separate targeted molecular dynamics simulations, subjected to a force constant imposing a movement towards the gp41 fusogenic conformation. Despite significant diversity, a preferred sequence of events emerged when the simulations were analyzed in terms of the formation, breakage and evolution of the contacts. We pointed out 29 residues as the most relevant for the movement of gp41; also, 2696 possible interactions were reduced to only 48 major interactions, which reveals the efficiency of the method. The analysis of the evolution of the main interactions lead to the detection of four main behaviors for those contacts: stable, increasing, decreasing and repulsive interactions. Altogether, these results suggest a specific small cavity of the HIV-1 gp41 hydrophobic groove as the preferred target to small molecules.

  13. Recognition of HIV-1 peptides by host CTL is related to HIV-1 similarity to human proteins.

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    Morgane Rolland

    Full Text Available BACKGROUND: While human immunodeficiency virus type 1 (HIV-1-specific cytotoxic T lymphocytes preferentially target specific regions of the viral proteome, HIV-1 features that contribute to immune recognition are not well understood. One hypothesis is that similarities between HIV and human proteins influence the host immune response, i.e., resemblance between viral and host peptides could preclude reactivity against certain HIV epitopes. METHODOLOGY/PRINCIPAL FINDINGS: We analyzed the extent of similarity between HIV-1 and the human proteome. Proteins from the HIV-1 B consensus sequence from 2001 were dissected into overlapping k-mers, which were then probed against a non-redundant database of the human proteome in order to identify segments of high similarity. We tested the relationship between HIV-1 similarity to host encoded peptides and immune recognition in HIV-infected individuals, and found that HIV immunogenicity could be partially modulated by the sequence similarity to the host proteome. ELISpot responses to peptides spanning the entire viral proteome evaluated in 314 individuals showed a trend indicating an inverse relationship between the similarity to the host proteome and the frequency of recognition. In addition, analysis of responses by a group of 30 HIV-infected individuals against 944 overlapping peptides representing a broad range of individual HIV-1B Nef variants, affirmed that the degree of similarity to the host was significantly lower for peptides with reactive epitopes than for those that were not recognized. CONCLUSIONS/SIGNIFICANCE: Our results suggest that antigenic motifs that are scarcely represented in human proteins might represent more immunogenic CTL targets not selected against in the host. This observation could provide guidance in the design of more effective HIV immunogens, as sequences devoid of host-like features might afford superior immune reactivity.

  14. Use of competitive polymerase chain reaction to determine HIV-1 levels in response to antiviral treatments

    NARCIS (Netherlands)

    Bruisten, S. M.; Koppelman, M. H.; Roos, M. T.; Loeliger, A. E.; Reiss, P.; Boucher, C. A.; Huisman, H. G.

    1993-01-01

    OBJECTIVE: To develop a competitive polymerase chain reaction technique with which to evaluate the usefulness of HIV-1 level as a marker of response to antiviral treatment. DESIGN: HIV-1 sequences were assessed by competitive polymerase chain reaction in four subjects participating in a double-blind

  15. HIV-1 drug resistance prevalence, drug susceptibility and variant characterization in the Jacobi Medical Center paediatric cohort, Bronx, NY, USA.

    Science.gov (United States)

    de Mulder, M; York, V A; Wiznia, A A; Michaud, H A; Nixon, D F; Holguin, A; Rosenberg, M G

    2014-03-01

    With the advent of combined antiretroviral therapy (cART), perinatally HIV-infected children are surviving into adolescence and beyond. However, drug resistance mutations (DRMs) compromise viral control, affecting the long-term effectiveness of ART. The aims of this study were to detect and identify DRMs in a HIV-1 infected paediatric cohort. Paired plasma and dried blood spots (DBSs) specimens were obtained from HIV-1 perinatally infected patients attending the Jacobi Medical Center, New York, USA. Clinical, virological and immunological data for these patients were analysed. HIV-1 pol sequences were generated from samples to identify DRMs according to the International AIDS Society (IAS) 2011 list. Forty-seven perinatally infected patients were selected, with a median age of 17.7 years, of whom 97.4% were carrying subtype B. They had a mean viral load of 3143 HIV-1 RNA copies/mL and a mean CD4 count of 486 cells/μL at the time of sampling. Nineteen patients (40.4%) had achieved undetectable viraemia (40.5% had a CD4 count of > 500 cells/μL. Most of the patients (97.9%) had received cART, including protease inhibitor (PI)-based regimens in 59.6% of cases. The DRM prevalence was 54.1, 27.6 and 27.0% for nucleoside reverse transcriptase inhibitors (NRTIs), PIs and nonnucleoside reverse transcriptase inhibitors (NNRTIs), respectively. Almost two-thirds (64.9%) of the patients harboured DRMs to at least one drug class and 5.4% were triple resistant. The mean nucleotide similarity between plasma and DBS sequences was 97.9%. Identical DRM profiles were present in 60% of plasma-DBS paired sequences. A total of 30 DRMs were detected in plasma and 26 in DBSs, with 23 present in both. Although more perinatally HIV-1-infected children are reaching adulthood as a result of advances in cART, our study cohort presented a high prevalence of resistant viruses, especially viruses resistant to NRTIs. DBS specimens can be used for DRM detection. © 2013 British HIV Association.

  16. Characterization of natural polymorphic sites of the HIV-1 integrase before the introduction of HIV-1 integrase inhibitors in Germany

    Science.gov (United States)

    Meixenberger, Karolin; Pouran Yousef, Kaveh; Somogyi, Sybille; Fiedler, Stefan; Bartmeyer, Barbara; von Kleist, Max; Kücherer, Claudia

    2014-01-01

    Introduction The aim of our study was to analyze the occurrence and evolution of HIV-1 integrase polymorphisms during the HIV-1 epidemic in Germany prior to the introduction of the first integrase inhibitor raltegravir in 2007. Materials and Methods Plasma samples from drug-naïve HIV-1 infected individuals newly diagnosed between 1986 and 2006 were used to determine PCR-based population sequences of the HIV-1 integrase (amino acids 1–278). The HIV-1 subtype was determined using the REGA HIV-1 subtyping tool. We calculated the frequency of amino acids at each position of the HIV-1 integrase in 337 subtype B strains for the time periods 1986–1989, 1991–1994, 1995–1998, 1999–2002, and 2003–2006. Positions were defined as polymorphic if amino acid variation was >1% in any period. Logistic regression was used to identify trends in amino acid variation over time. Resistance-associated mutations were identified according to the IAS 2013 list and the HIVdb, ANRS and GRADE algorithms. Results Overall, 56.8% (158/278) amino acid positions were polymorphic and 15.8% (25/158) of these positions exhibited a significant trend in amino acid variation over time. Proportionately, most polymorphic positions (63.3%, 31/49) were detected in the N-terminal zinc finger domain of the HIV-1 integrase. Motifs and residues essential for HIV-1 integrase activity were little polymorphic, but within the minimal non-specific DNA binding region I220-D270 up to 18.1% amino acid variation was noticed, including four positions with significant amino acid variation over time (S230, D232, D256, A265). No major resistance mutations were identified, and minor resistance mutations were rarely observed without trend over time. E157Q considered by HIVdb, ANRS, and GRADE algorithms was the most frequent resistance-associated polymorphism with an overall prevalence of 2.4%. Conclusions Detailed knowledge of the evolutionary variation of HIV-1 integrase polymorphisms is important to understand

  17. Cellular and molecular mechanisms of HIV-1 integration targeting.

    Science.gov (United States)

    Engelman, Alan N; Singh, Parmit K

    2018-07-01

    Integration is central to HIV-1 replication and helps mold the reservoir of cells that persists in AIDS patients. HIV-1 interacts with specific cellular factors to target integration to interior regions of transcriptionally active genes within gene-dense regions of chromatin. The viral capsid interacts with several proteins that are additionally implicated in virus nuclear import, including cleavage and polyadenylation specificity factor 6, to suppress integration into heterochromatin. The viral integrase protein interacts with transcriptional co-activator lens epithelium-derived growth factor p75 to principally position integration within gene bodies. The integrase additionally senses target DNA distortion and nucleotide sequence to help fine-tune the specific phosphodiester bonds that are cleaved at integration sites. Research into virus-host interactions that underlie HIV-1 integration targeting has aided the development of a novel class of integrase inhibitors and may help to improve the safety of viral-based gene therapy vectors.

  18. Phylodynamics of HIV-1 subtype F1 in Angola, Brazil and Romania.

    Science.gov (United States)

    Bello, Gonzalo; Afonso, Joana Morais; Morgado, Mariza G

    2012-07-01

    The HIV-1 subtype F1 is exceptionally prevalent in Angola, Brazil and Romania. The epidemiological context in which the spread of HIV occurred was highly variable from one country to another, mainly due to the existence of a long-term civil war in Angola and the contamination of a large number of children in Romania. Here we apply phylogenetic and Bayesian coalescent-based methods to reconstruct the phylodynamic patterns of HIV-1 subtype F1 in such different epidemiological settings. The phylogenetic analyses of HIV-1 subtype F1 pol sequences sampled worldwide confirmed that most sequences from Angola, Brazil and Romania segregated in country-specific monophyletic groups, while most subtype F1 sequences from Romanian children branched as a monophyletic sub-cluster (Romania-CH) nested within sequences from adults. The inferred time of the most recent common ancestor of the different subtype F1 clades were as follow: Angola=1983 (1978-1989), Brazil=1977 (1972-1981), Romania adults=1980 (1973-1987), and Romania-CH=1985 (1978-1989). All subtype F1 clades showed a demographic history best explained by a model of logistic population growth. Although the expansion phase of subtype F1 epidemic in Angola (mid 1980s to early 2000s) overlaps with the civil war period (1975-2002), the mean estimated growth rate of the Angolan F1 clade (0.49 year(-1)) was not exceptionally high, but quite similar to that estimated for the Brazilian (0.69 year(-1)) and Romanian adult (0.36 year(-1)) subtype F1 clades. The Romania-CH subtype F1 lineage, by contrast, displayed a short and explosive dissemination phase, with a median growth rate (2.47 year(-1)) much higher than that estimated for adult populations. This result supports the idea that the AIDS epidemic that affected the Romanian children was mainly caused by the spread of the HIV through highly efficient parenteral transmission networks, unlike adult populations where HIV is predominantly transmitted through sexual route. Copyright

  19. Herpes viruses and HIV-1 drug resistance mutations influence the virologic and immunologic milieu of the male genital tract.

    Science.gov (United States)

    Gianella, Sara; Morris, Sheldon R; Anderson, Christy; Spina, Celsa A; Vargas, Milenka V; Young, Jason A; Richman, Douglas D; Little, Susan J; Smith, Davey M

    2013-01-02

    To further understand the role that chronic viral infections of the male genital tract play on HIV-1 dynamics and replication. Retrospective, observational study including 236 paired semen and blood samples collected from 115 recently HIV-1 infected antiretroviral naive men who have sex with men. In this study, we evaluated the association of seminal HIV-1 shedding to coinfections with seven herpes viruses, blood plasma HIV-1 RNA levels, CD4 T-cell counts, presence of transmitted drug resistance mutations (DRMs) in HIV-1 pol, participants' age and stage of HIV-infection using multivariate generalized estimating equation methods. Associations between herpes virus shedding, seminal HIV-1 levels, number and immune activation of seminal T-cells was also investigated (Mann-Whitney). Seminal herpes virus shedding was observed in 75.7% of individuals. Blood HIV-1 RNA levels (P herpes virus (HHV)-8 levels (P herpes viruses seminal shedding in our cohort. Shedding of CMV, EBV and HHV-8 and absence of DRM were associated with increased frequency of HIV-1 shedding and/or higher levels of HIV-1 RNA in semen, which are likely important cofactors for HIV-1 transmission.

  20. Structure of the HIV-1 reverse transcriptase Q151M mutant: insights into the inhibitor resistance of HIV-1 reverse transcriptase and the structure of the nucleotide-binding pocket of Hepatitis B virus polymerase

    International Nuclear Information System (INIS)

    Nakamura, Akiyoshi; Tamura, Noriko; Yasutake, Yoshiaki

    2015-01-01

    The structure of the HIV-1 reverse transcriptase Q151M mutant was determined at a resolution of 2.6 Å in space group P321. Hepatitis B virus polymerase (HBV Pol) is an important target for anti-HBV drug development; however, its low solubility and stability in vitro has hindered detailed structural studies. Certain nucleotide reverse transcriptase (RT) inhibitors (NRTIs) such as tenofovir and lamivudine can inhibit both HBV Pol and Human immunodeficiency virus 1 (HIV-1) RT, leading to speculation on structural and mechanistic analogies between the deoxynucleotide triphosphate (dNTP)-binding sites of these enzymes. The Q151M mutation in HIV-1 RT, located at the dNTP-binding site, confers resistance to various NRTIs, while maintaining sensitivity to tenofovir and lamivudine. The residue corresponding to Gln151 is strictly conserved as a methionine in HBV Pol. Therefore, the structure of the dNTP-binding pocket of the HIV-1 RT Q151M mutant may reflect that of HBV Pol. Here, the crystal structure of HIV-1 RT Q151M, determined at 2.6 Å resolution, in a new crystal form with space group P321 is presented. Although the structure of HIV-1 RT Q151M superimposes well onto that of HIV-1 RT in a closed conformation, a slight movement of the β-strands (β2–β3) that partially create the dNTP-binding pocket was observed. This movement might be caused by the introduction of the bulky thioether group of Met151. The structure also highlighted the possibility that the hydrogen-bonding network among amino acids and NRTIs is rearranged by the Q151M mutation, leading to a difference in the affinity of NRTIs for HIV-1 RT and HBV Pol

  1. Structure of the HIV-1 reverse transcriptase Q151M mutant: insights into the inhibitor resistance of HIV-1 reverse transcriptase and the structure of the nucleotide-binding pocket of Hepatitis B virus polymerase

    Energy Technology Data Exchange (ETDEWEB)

    Nakamura, Akiyoshi; Tamura, Noriko; Yasutake, Yoshiaki, E-mail: y-yasutake@aist.go.jp [National Institute of Advanced Industrial Science and Technology (AIST), 2-17-2-1 Tsukisamu-Higashi, Toyohira, Sapporo, Hokkaido 062-8517 (Japan)

    2015-10-23

    The structure of the HIV-1 reverse transcriptase Q151M mutant was determined at a resolution of 2.6 Å in space group P321. Hepatitis B virus polymerase (HBV Pol) is an important target for anti-HBV drug development; however, its low solubility and stability in vitro has hindered detailed structural studies. Certain nucleotide reverse transcriptase (RT) inhibitors (NRTIs) such as tenofovir and lamivudine can inhibit both HBV Pol and Human immunodeficiency virus 1 (HIV-1) RT, leading to speculation on structural and mechanistic analogies between the deoxynucleotide triphosphate (dNTP)-binding sites of these enzymes. The Q151M mutation in HIV-1 RT, located at the dNTP-binding site, confers resistance to various NRTIs, while maintaining sensitivity to tenofovir and lamivudine. The residue corresponding to Gln151 is strictly conserved as a methionine in HBV Pol. Therefore, the structure of the dNTP-binding pocket of the HIV-1 RT Q151M mutant may reflect that of HBV Pol. Here, the crystal structure of HIV-1 RT Q151M, determined at 2.6 Å resolution, in a new crystal form with space group P321 is presented. Although the structure of HIV-1 RT Q151M superimposes well onto that of HIV-1 RT in a closed conformation, a slight movement of the β-strands (β2–β3) that partially create the dNTP-binding pocket was observed. This movement might be caused by the introduction of the bulky thioether group of Met151. The structure also highlighted the possibility that the hydrogen-bonding network among amino acids and NRTIs is rearranged by the Q151M mutation, leading to a difference in the affinity of NRTIs for HIV-1 RT and HBV Pol.

  2. HIV-1 protease-substrate coevolution in nelfinavir resistance.

    Science.gov (United States)

    Kolli, Madhavi; Ozen, Ayşegül; Kurt-Yilmaz, Nese; Schiffer, Celia A

    2014-07-01

    Resistance to various human immunodeficiency virus type 1 (HIV-1) protease inhibitors (PIs) challenges the effectiveness of therapies in treating HIV-1-infected individuals and AIDS patients. The virus accumulates mutations within the protease (PR) that render the PIs less potent. Occasionally, Gag sequences also coevolve with mutations at PR cleavage sites contributing to drug resistance. In this study, we investigated the structural basis of coevolution of the p1-p6 cleavage site with the nelfinavir (NFV) resistance D30N/N88D protease mutations by determining crystal structures of wild-type and NFV-resistant HIV-1 protease in complex with p1-p6 substrate peptide variants with L449F and/or S451N. Alterations of residue 30's interaction with the substrate are compensated by the coevolving L449F and S451N cleavage site mutations. This interdependency in the PR-p1-p6 interactions enhances intermolecular contacts and reinforces the overall fit of the substrate within the substrate envelope, likely enabling coevolution to sustain substrate recognition and cleavage in the presence of PR resistance mutations. Resistance to human immunodeficiency virus type 1 (HIV-1) protease inhibitors challenges the effectiveness of therapies in treating HIV-1-infected individuals and AIDS patients. Mutations in HIV-1 protease selected under the pressure of protease inhibitors render the inhibitors less potent. Occasionally, Gag sequences also mutate and coevolve with protease, contributing to maintenance of viral fitness and to drug resistance. In this study, we investigated the structural basis of coevolution at the Gag p1-p6 cleavage site with the nelfinavir (NFV) resistance D30N/N88D protease mutations. Our structural analysis reveals the interdependency of protease-substrate interactions and how coevolution may restore substrate recognition and cleavage in the presence of protease drug resistance mutations. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  3. A European multicientre study on the comparison of HIV-1 viral loads between VERIS HIV-1 Assay and Roche COBAS® TAQMAN® HIV-1 test, Abbott RealTime HIV-1 Assay, and Siemens VERSANT HIV-1 Assay.

    Science.gov (United States)

    Braun, Patrick; Delgado, Rafael; Drago, Monica; Fanti, Diana; Fleury, Hervé; Hofmann, Jörg; Izopet, Jacques; Kühn, Sebastian; Lombardi, Alessandra; Mancon, Alessandro; Marcos, Mª Angeles; Mileto, Davide; Sauné, Karine; O'Shea, Siobhan; Pérez-Rivilla, Alfredo; Ramble, John; Trimoulet, Pascale; Vila, Jordi; Whittaker, Duncan; Artus, Alain; Rhodes, Daniel

    2017-07-01

    Viral load monitoring is essential for patients under treatment for HIV. Beckman Coulter has developed the VERIS HIV-1 Assay for use on the novel, automated DxN VERIS Molecular Diagnostics System. ¥ OBJECTIVES: Evaluation of the clinical performance of the new quantitative VERIS HIV-1 Assay at multiple EU laboratories. Method comparison with the VERIS HIV-1 Assay was performed with 415 specimens at 5 sites tested with COBAS ® AmpliPrep/COBAS ® TaqMan ® HIV-1 Test, v2.0, 169 specimens at 3 sites tested with RealTime HIV-1 Assay, and 202 specimens from 2 sites tested with VERSANT HIV-1 Assay. Patient monitoring sample results from 4 sites were also compared. Bland-Altman analysis showed the average bias between VERIS HIV-1 Assay and COBAS HIV-1 Test, RealTime HIV-1 Assay, and VERSANT HIV-1 Assay to be 0.28, 0.39, and 0.61 log 10 cp/mL, respectively. Bias at low end levels below 1000cp/mL showed predicted bias to be <0.3 log 10 cp/mL for VERIS HIV-1 Assay versus COBAS HIV-1 Test and RealTime HIV-1 Assay, and <0.5 log 10 cp/mL versus VERSANT HIV-1 Assay. Analysis on 174 specimens tested with the 0.175mL volume VERIS HIV-1 Assay and COBAS HIV-1 Test showed average bias of 0.39 log 10 cp/mL. Patient monitoring results using VERIS HIV-1 Assay demonstrated similar viral load trends over time to all comparators. The VERIS HIV-1 Assay for use on the DxN VERIS System demonstrated comparable clinical performance to COBAS ® HIV-1 Test, RealTime HIV-1 Assay, and VERSANT HIV-1 Assay. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Proviral HIV-genome-wide and pol-gene specific Zinc Finger Nucleases: Usability for targeted HIV gene therapy

    Directory of Open Access Journals (Sweden)

    Wayengera Misaki

    2011-07-01

    Full Text Available Abstract Background Infection with HIV, which culminates in the establishment of a latent proviral reservoir, presents formidable challenges for ultimate cure. Building on the hypothesis that ex-vivo or even in-vivo abolition or disruption of HIV-gene/genome-action by target mutagenesis or excision can irreversibly abrogate HIV's innate fitness to replicate and survive, we previously identified the isoschizomeric bacteria restriction enzymes (REases AcsI and ApoI as potent cleavers of the HIV-pol gene (11 and 9 times in HIV-1 and 2, respectively. However, both enzymes, along with others found to cleave across the entire HIV-1 genome, slice (SX at palindromic sequences that are prevalent within the human genome and thereby pose the risk of host genome toxicity. A long-term goal in the field of R-M enzymatic therapeutics has thus been to generate synthetic restriction endonucleases with longer recognition sites limited in specificity to HIV. We aimed (i to assemble and construct zinc finger arrays and nucleases (ZFN with either proviral-HIV-pol gene or proviral-HIV-1 whole-genome specificity respectively, and (ii to advance a model for pre-clinically testing lentiviral vectors (LV that deliver and transduce either ZFN genotype. Methods and Results First, we computationally generated the consensus sequences of (a 114 dsDNA-binding zinc finger (Zif arrays (ZFAs or ZifHIV-pol and (b two zinc-finger nucleases (ZFNs which, unlike the AcsI and ApoI homeodomains, possess specificity to >18 base-pair sequences uniquely present within the HIV-pol gene (ZifHIV-polFN. Another 15 ZFNs targeting >18 bp sequences within the complete HIV-1 proviral genome were constructed (ZifHIV-1FN. Second, a model for constructing lentiviral vectors (LVs that deliver and transduce a diploid copy of either ZifHIV-polFN or ZifHIV-1FN chimeric genes (termed LV- 2xZifHIV-polFN and LV- 2xZifHIV-1FN, respectively is proposed. Third, two preclinical models for controlled testing of

  5. Frequent intra-subtype recombination among HIV-1 circulating in Tanzania.

    Directory of Open Access Journals (Sweden)

    Ireen E Kiwelu

    Full Text Available The study estimated the prevalence of HIV-1 intra-subtype recombinant variants among female bar and hotel workers in Tanzania. While intra-subtype recombination occurs in HIV-1, it is generally underestimated. HIV-1 env gp120 V1-C5 quasispecies from 45 subjects were generated by single-genome amplification and sequencing (median (IQR of 38 (28-50 sequences per subject. Recombination analysis was performed using seven methods implemented within the recombination detection program version 3, RDP3. HIV-1 sequences were considered recombinant if recombination signals were detected by at least three methods with p-values of ≤0.05 after Bonferroni correction for multiple comparisons. HIV-1 in 38 (84% subjects showed evidence for intra-subtype recombination including 22 with HIV-1 subtype A1, 13 with HIV-1 subtype C, and 3 with HIV-1 subtype D. The distribution of intra-patient recombination breakpoints suggested ongoing recombination and showed selective enrichment of recombinant variants in 23 (60% subjects. The number of subjects with evidence of intra-subtype recombination increased from 29 (69% to 36 (82% over one year of follow-up, although the increase did not reach statistical significance. Adjustment for intra-subtype recombination is important for the analysis of multiplicity of HIV infection. This is the first report of high prevalence of intra-subtype recombination in the HIV/AIDS epidemic in Tanzania, a region where multiple HIV-1 subtypes co-circulate. HIV-1 intra-subtype recombination increases viral diversity and presents additional challenges for HIV-1 vaccine design.

  6. Compartmentalization of the gut viral reservoir in HIV-1 infected patients

    Directory of Open Access Journals (Sweden)

    Grant Tannika

    2007-12-01

    Full Text Available Abstract Background Recently there has been an increasing interest and appreciation for the gut as both a viral reservoir as well as an important host-pathogen interface in human immunodefiency virus type 1 (HIV-1 infection. The gut associated lymphoid tissue (GALT is the largest lymphoid organ infected by HIV-1. In this study we examined if different HIV-1 quasispecies are found in different parts of the gut of HIV-1 infected individuals. Results Gut biopsies (esophagus, stomach, duodenum and colorectum were obtained from eight HIV-1 infected preHAART (highly active antiretroviral therapy patients. HIV-1 Nef and Reverse transcriptase (RT encoding sequences were obtained through nested PCR amplification from DNA isolated from the gut biopsy tissues. The PCR fragments were cloned and sequenced. The resulting sequences were subjected to various phylogenetic analyses. Expression of the nef gene and viral RNA in the different gut tissues was determined using real-time RT-PCR. Phylogenetic analysis of the Nef protein-encoding region revealed compartmentalization of viral replication in the gut within patients. Viral diversity in both the Nef and RT encoding region varied in different parts of the gut. Moreover, increased nef gene expression (p Conclusion Our results indicated that different HIV-1 quasispecies populate different parts of the gut, and that viral replication in the gut is compartmentalized. These observations underscore the importance of the gut as a host-pathogen interface in HIV-1 infection.

  7. Spatio-Temporal History of HIV-1 CRF35_AD in Afghanistan and Iran.

    Science.gov (United States)

    Eybpoosh, Sana; Bahrampour, Abbas; Karamouzian, Mohammad; Azadmanesh, Kayhan; Jahanbakhsh, Fatemeh; Mostafavi, Ehsan; Zolala, Farzaneh; Haghdoost, Ali Akbar

    2016-01-01

    HIV-1 Circulating Recombinant Form 35_AD (CRF35_AD) has an important position in the epidemiological profile of Afghanistan and Iran. Despite the presence of this clade in Afghanistan and Iran for over a decade, our understanding of its origin and dissemination patterns is limited. In this study, we performed a Bayesian phylogeographic analysis to reconstruct the spatio-temporal dispersion pattern of this clade using eligible CRF35_AD gag and pol sequences available in the Los Alamos HIV database (432 sequences available from Iran, 16 sequences available from Afghanistan, and a single CRF35_AD-like pol sequence available from USA). Bayesian Markov Chain Monte Carlo algorithm was implemented in BEAST v1.8.1. Between-country dispersion rates were tested with Bayesian stochastic search variable selection method and were considered significant where Bayes factor values were greater than three. The findings suggested that CRF35_AD sequences were genetically similar to parental sequences from Kenya and Uganda, and to a set of subtype A1 sequences available from Afghan refugees living in Pakistan. Our results also showed that across all phylogenies, Afghan and Iranian CRF35_AD sequences formed a monophyletic cluster (posterior clade credibility> 0.7). The divergence date of this cluster was estimated to be between 1990 and 1992. Within this cluster, a bidirectional dispersion of the virus was observed across Afghanistan and Iran. We could not clearly identify if Afghanistan or Iran first established or received this epidemic, as the root location of this cluster could not be robustly estimated. Three CRF35_AD sequences from Afghan refugees living in Pakistan nested among Afghan and Iranian CRF35_AD branches. However, the CRF35_AD-like sequence available from USA diverged independently from Kenyan subtype A1 sequences, suggesting it not to be a true CRF35_AD lineage. Potential factors contributing to viral exchange between Afghanistan and Iran could be injection drug

  8. Curaxin CBL0100 Blocks HIV-1 Replication and Reactivation through Inhibition of Viral Transcriptional Elongation

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    Maxime J. Jean

    2017-10-01

    Full Text Available Despite combination antiretroviral therapy (cART, acquired immunodeficiency syndrome (AIDS, predominantly caused by the human immunodeficiency virus type 1 (HIV-1, remains incurable. The barrier to a cure lies in the virus' ability to establish a latent infection in HIV/AIDS patients. Unsurprisingly, efforts for a sterilizing cure have focused on the “shock and kill” strategy using latency-reversing agents (LRAs to complement cART in order to eliminate these latent reservoirs. However, this method faces numerous challenges. Recently, the “block and lock” strategy has been proposed. It aims to reinforce a deep state of latency and prevent sporadic reactivation (“blip” of HIV-1 using latency-promoting agents (LPAs for a functional cure. Our studies of curaxin 100 (CBL0100, a small-molecule targeting the facilitates chromatin transcription (FACT complex, show that it blocks both HIV-1 replication and reactivation in in vitro and ex vivo models of HIV-1. Mechanistic investigation elucidated that CBL0100 preferentially targets HIV-1 transcriptional elongation and decreases the occupancy of RNA Polymerase II (Pol II and FACT at the HIV-1 promoter region. In conclusion, CBL0100 is a newly identified inhibitor of HIV-1 transcription that can be used as an LPA in the “block and lock” cure strategy.

  9. Identification of dual-tropic HIV-1 using evolved neural networks.

    Science.gov (United States)

    Fogel, Gary B; Lamers, Susanna L; Liu, Enoch S; Salemi, Marco; McGrath, Michael S

    2015-11-01

    Blocking the binding of the envelope HIV-1 protein to immune cells is a popular concept for development of anti-HIV therapeutics. R5 HIV-1 binds CCR5, X4 HIV-1 binds CXCR4, and dual-tropic HIV-1 can bind either coreceptor for cellular entry. R5 viruses are associated with early infection and over time can evolve to X4 viruses that are associated with immune failure. Dual-tropic HIV-1 is less studied; however, it represents functional antigenic intermediates during the transition of R5 to X4 viruses. Viral tropism is linked partly to the HIV-1 envelope V3 domain, where the amino acid sequence helps dictate the receptor a particular virus will target; however, using V3 sequence information to identify dual-tropic HIV-1 isolates has remained difficult. Our goal in this study was to elucidate features of dual-tropic HIV-1 isolates that assist in the biological understanding of dual-tropism and develop an approach for their detection. Over 1559 HIV-1 subtype B sequences with known tropisms were analyzed. Each sequence was represented by 73 structural, biochemical and regional features. These features were provided to an evolved neural network classifier and evaluated using balanced and unbalanced data sets. The study resolved R5X4 viruses from R5 with an accuracy of 81.8% and from X4 with an accuracy of 78.8%. The approach also identified a set of V3 features (hydrophobicity, structural and polarity) that are associated with tropism transitions. The ability to distinguish R5X4 isolates will improve computational tropism decisions for R5 vs. X4 and assist in HIV-1 research and drug development efforts. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  10. Rising prevalence of non-B HIV-1 subtypes in North Carolina and evidence for local onward transmission.

    Science.gov (United States)

    Dennis, Ann M; Hué, Stephane; Learner, Emily; Sebastian, Joseph; Miller, William C; Eron, Joseph J

    2017-01-01

    HIV-1 diversity is increasing in North American and European cohorts which may have public health implications. However, little is known about non-B subtype diversity in the southern United States, despite the region being the epicenter of the nation's epidemic. We characterized HIV-1 diversity and transmission clusters to identify the extent to which non-B strains are transmitted locally. We conducted cross-sectional analyses of HIV-1 partial pol sequences collected from 1997 to 2014 from adults accessing routine clinical care in North Carolina (NC). Subtypes were evaluated using COMET and phylogenetic analysis. Putative transmission clusters were identified using maximum-likelihood trees. Clusters involving non-B strains were confirmed and their dates of origin were estimated using Bayesian phylogenetics. Data were combined with demographic information collected at the time of sample collection and country of origin for a subset of patients. Among 24,972 sequences from 15,246 persons, the non-B subtype prevalence increased from 0% to 3.46% over the study period. Of 325 persons with non-B subtypes, diversity was high with over 15 pure subtypes and recombinants; subtype C (28.9%) and CRF02_AG (24.0%) were most common. While identification of transmission clusters was lower for persons with non-B versus B subtypes, several local transmission clusters (≥3 persons) involving non-B subtypes were identified and all were presumably due to heterosexual transmission. Prevalence of non-B subtype diversity remains low in NC but a statistically significant rise was identified over time which likely reflects multiple importation. However, the combined phylogenetic clustering analysis reveals evidence for local onward transmission. Detection of these non-B clusters suggests heterosexual transmission and may guide diagnostic and prevention interventions.

  11. Phylogenetic investigation of a statewide HIV-1 epidemic reveals ongoing and active transmission networks among men who have sex with men

    Science.gov (United States)

    Chan, Philip A.; Hogan, Joseph W.; Huang, Austin; DeLong, Allison; Salemi, Marco; Mayer, Kenneth H.; Kantor, Rami

    2015-01-01

    Background Molecular epidemiologic evaluation of HIV-1 transmission networks can elucidate behavioral components of transmission that can be targets for intervention. Methods We combined phylogenetic and statistical approaches using pol sequences from patients diagnosed 2004-2011 at a large HIV center in Rhode Island, following 75% of the state’s HIV population. Phylogenetic trees were constructed using maximum likelihood and putative transmission clusters were evaluated using latent class analyses (LCA) to determine association of cluster size with underlying demographic/behavioral characteristics. A logistic growth model was used to assess intra-cluster dynamics over time and predict “active” clusters that were more likely to harbor undiagnosed infections. Results Of 1,166 HIV-1 subtype B sequences, 31% were distributed among 114 statistically-supported, monophyletic clusters (range: 2-15 sequences/cluster). Sequences from men who have sex with men (MSM) formed 52% of clusters. LCA demonstrated that sequences from recently diagnosed (2008-2011) MSM with primary HIV infection (PHI) and other sexually transmitted infections (STIs) were more likely to form larger clusters (Odds Ratio 1.62-11.25, ppornographic stores. Four large clusters with 38 sequences (100% male, 89% MSM) had a high-probability of harboring undiagnosed infections and included younger MSM with PHI and STIs. Conclusions In this first large-scale molecular epidemiologic investigation of HIV-1 transmission in New England, sexual networks among recently diagnosed MSM with PHI and concomitant STIs contributed to ongoing transmission. Characterization of transmission dynamics revealed actively growing clusters which may be targets for intervention. PMID:26258569

  12. Genetic architecture of HIV-1 genes circulating in north India & their functional implications.

    Science.gov (United States)

    Neogi, Ujjwal; Sood, Vikas; Ronsard, Larence; Singh, Jyotsna; Lata, Sneh; Ramachandran, V G; Das, S; Wanchu, Ajay; Banerjea, Akhil C

    2011-12-01

    This review presents data on genetic and functional analysis of some of the HIV-1 genes derived from HIV-1 infected individuals from north India (Delhi, Punjab and Chandigarh). We found evidence of novel B/C recombinants in HIV-1 LTR region showing relatedness to China/Myanmar with 3 copies of Nfκb sites; B/C/D mosaic genomes for HIV-1 Vpr and novel B/C Tat. We reported appearance of a complex recombinant form CRF_02AG of HIV-1 envelope sequences which is predominantly found in Central/Western Africa. Also one Indian HIV-1 envelope subtype C sequence suggested exclusive CXCR4 co-receptor usage. This extensive recombination, which is observed in about 10 per cent HIV-1 infected individuals in the Vpr genes, resulted in remarkably altered functions when compared with prototype subtype B Vpr. The Vpu C was found to be more potent in causing apoptosis when compared with Vpu B when analyzed for subG1 DNA content. The functional implications of these changes as well as in other genes of HIV-1 are discussed in detail with possible implications for subtype-specific pathogenesis highlighted.

  13. Dynamics of HIV-1 RNA Near the Plasma Membrane during Virus Assembly.

    Science.gov (United States)

    Sardo, Luca; Hatch, Steven C; Chen, Jianbo; Nikolaitchik, Olga; Burdick, Ryan C; Chen, De; Westlake, Christopher J; Lockett, Stephen; Pathak, Vinay K; Hu, Wei-Shau

    2015-11-01

    To increase our understanding of the events that lead to HIV-1 genome packaging, we examined the dynamics of viral RNA and Gag-RNA interactions near the plasma membrane by using total internal reflection fluorescence microscopy. We labeled HIV-1 RNA with a photoconvertible Eos protein via an RNA-binding protein that recognizes stem-loop sequences engineered into the viral genome. Near-UV light exposure causes an irreversible structural change in Eos and alters its emitted fluorescence from green to red. We studied the dynamics of HIV-1 RNA by photoconverting Eos near the plasma membrane, and we monitored the population of photoconverted red-Eos-labeled RNA signals over time. We found that in the absence of Gag, most of the HIV-1 RNAs stayed near the plasma membrane transiently, for a few minutes. The presence of Gag significantly increased the time that RNAs stayed near the plasma membrane: most of the RNAs were still detected after 30 min. We then quantified the proportion of HIV-1 RNAs near the plasma membrane that were packaged into assembling viral complexes. By tagging Gag with blue fluorescent protein, we observed that only a portion, ∼13 to 34%, of the HIV-1 RNAs that reached the membrane were recruited into assembling particles in an hour, and the frequency of HIV-1 RNA packaging varied with the Gag expression level. Our studies reveal the HIV-1 RNA dynamics on the plasma membrane and the efficiency of RNA recruitment and provide insights into the events leading to the generation of infectious HIV-1 virions. Nascent HIV-1 particles assemble on plasma membranes. During the assembly process, HIV-1 RNA genomes must be encapsidated into viral complexes to generate infectious particles. To gain insights into the RNA packaging and virus assembly mechanisms, we labeled and monitored the HIV-1 RNA signals near the plasma membrane. Our results showed that most of the HIV-1 RNAs stayed near the plasma membrane for only a few minutes in the absence of Gag, whereas

  14. Neuroinflammation and Behavior in HIV-1 Transgenic Rats Exposed to Chronic Adolescent Stress.

    Science.gov (United States)

    Rowson, Sydney A; Harrell, Constance S; Bekhbat, Mandakh; Gangavelli, Apoorva; Wu, Matthew J; Kelly, Sean D; Reddy, Renuka; Neigh, Gretchen N

    2016-01-01

    Highly active antiretroviral therapy (HAART) has improved prognosis for people living with HIV (PLWH) and dramatically reduced the incidence of AIDS. However, even when viral load is controlled, PLWH develop psychiatric and neurological disorders more frequently than those living without HIV. Adolescents with HIV are particularly susceptible to the development of psychiatric illnesses and neurocognitive impairments. While both psychiatric and neurocognitive disorders have been found to be exacerbated by stress, the extent to which chronic stress and HIV-1 viral proteins interact to impact behavior and relevant neuroinflammatory processes is unknown. Determination of the individual contributions of stress and HIV to neuropsychiatric disorders is heavily confounded in humans. In order to isolate the influence of HIV-1 proteins and chronic stress on behavior and neuroinflammation, we employed the HIV-1 transgenic (Tg) rat model, which expresses HIV-1 proteins with a gag and pol deletion, allowing for viral protein expression without viral replication. This Tg line has been characterized as a model of HAART-controlled HIV-1 infection due to the lack of viral replication but continued presence of HIV-1 proteins. We exposed male and female adolescent HIV-1 Tg rats to a mixed-modality chronic stress paradigm consisting of isolation, social defeat and restraint, and assessed behavior, cerebral vascularization, and neuroinflammatory endpoints. Stress, sex, and presence of the HIV-1 transgene impacted weight gain in adolescent rats. Female HIV-1 Tg rats showed decreases in central tendency during the light cycle in the open field regardless of stress exposure. Both male and female HIV-1 Tg rats exhibited decreased investigative behavior in the novel object recognition task, but no memory impairments. Adolescent stress had no effect on the tested behaviors. Microglia in female HIV-1 Tg rats exhibited a hyper-ramified structure, and gene expression of complement factor B was

  15. Epidemiological study of phylogenetic transmission clusters in a local HIV-1 epidemic reveals distinct differences between subtype B and non-B infections.

    Science.gov (United States)

    Chalmet, Kristen; Staelens, Delfien; Blot, Stijn; Dinakis, Sylvie; Pelgrom, Jolanda; Plum, Jean; Vogelaers, Dirk; Vandekerckhove, Linos; Verhofstede, Chris

    2010-09-07

    The number of HIV-1 infected individuals in the Western world continues to rise. More in-depth understanding of regional HIV-1 epidemics is necessary for the optimal design and adequate use of future prevention strategies. The use of a combination of phylogenetic analysis of HIV sequences, with data on patients' demographics, infection route, clinical information and laboratory results, will allow a better characterization of individuals responsible for local transmission. Baseline HIV-1 pol sequences, obtained through routine drug-resistance testing, from 506 patients, newly diagnosed between 2001 and 2009, were used to construct phylogenetic trees and identify transmission-clusters. Patients' demographics, laboratory and clinical data, were retrieved anonymously. Statistical analysis was performed to identify subtype-specific and transmission-cluster-specific characteristics. Multivariate analysis showed significant differences between the 59.7% of individuals with subtype B infection and the 40.3% non-B infected individuals, with regard to route of transmission, origin, infection with Chlamydia (p = 0.01) and infection with Hepatitis C virus (p = 0.017). More and larger transmission-clusters were identified among the subtype B infections (p HIV (p = 0.017). Combination of phylogenetics with demographic information, laboratory and clinical data, revealed that HIV-1 subtype B infected Caucasian men-who-have-sex-with-men with high prevalence of sexually transmitted diseases, account for the majority of local HIV-transmissions. This finding elucidates observed epidemiological trends through molecular analysis, and justifies sustained focus in prevention on this high risk group.

  16. HIV-1 infection and pregnancy in young women in Brazil: socioeconomic and drug resistance profiles in a cross-sectional study.

    Science.gov (United States)

    Lima, Yanna Andressa Ramos; Reis, Mônica Nogueira Guarda; Cardoso, Ludimila Paula Vaz; Stefani, Mariane Martins Araújo

    2016-07-05

    To describe socioeconomic and antiretroviral (ARV) drug resistance profiles among young pregnant women infected with HIV-1. A public health antenatal programme responsible for screening ∼90 000 pregnant women per year for nine different infectious diseases in Central Western Brazil. 96 young pregnant women (15-24 years) infected with HIV-1. Standard interviews and blood samples were taken at the time of recruitment, at the first medical appointment after confirmation of diagnosis of HIV-1 infection, and before ARV prophylaxis initiation. Clinical and laboratory data were retrieved from medical files. HIV-1 pol gene sequences (entire protease/PR, partial reverse transcriptase/RT) were obtained from plasma RNA. ARV resistance mutations (CPR/Stanford HIV-1; International AIDS Society-USA databases) were identified. The median age was 21 years; most reported pregnancies. Possible heterosexual transmission by an HIV-1 infected partner (17%) and commercial sex work (2%) were reported. The median of CD4 cell count was 526 cells/mm(3); the median viral load was: 10 056 copies/mL in ARV-naïve (48/96) patients and 5881 copies/mL in ARV-exposed (48/96) patients. Two probable seroconversion cases during pregnancy were identified in adolescents. One mother-to-child transmission case (1.0%) was observed. Transmitted drug resistance among ARV-naïve patients was 9.3% (CI 95% 3.3% to 19.6%); secondary drug resistance among ARV-exposed patients was 12.5% (CI 95% 4.7% to 25.6%). Despite high access to antenatal care, the low socioeconomic-educational profiles seen in these young HIV-1-infected women highlight the necessity of improved public health educational and preventive strategies regarding HIV infection and early unplanned pregnancy. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/

  17. W-curve alignments for HIV-1 genomic comparisons.

    Directory of Open Access Journals (Sweden)

    Douglas J Cork

    2010-06-01

    Full Text Available The W-curve was originally developed as a graphical visualization technique for viewing DNA and RNA sequences. Its ability to render features of DNA also makes it suitable for computational studies. Its main advantage in this area is utilizing a single-pass algorithm for comparing the sequences. Avoiding recursion during sequence alignments offers advantages for speed and in-process resources. The graphical technique also allows for multiple models of comparison to be used depending on the nucleotide patterns embedded in similar whole genomic sequences. The W-curve approach allows us to compare large numbers of samples quickly.We are currently tuning the algorithm to accommodate quirks specific to HIV-1 genomic sequences so that it can be used to aid in diagnostic and vaccine efforts. Tracking the molecular evolution of the virus has been greatly hampered by gap associated problems predominantly embedded within the envelope gene of the virus. Gaps and hypermutation of the virus slow conventional string based alignments of the whole genome. This paper describes the W-curve algorithm itself, and how we have adapted it for comparison of similar HIV-1 genomes. A treebuilding method is developed with the W-curve that utilizes a novel Cylindrical Coordinate distance method and gap analysis method. HIV-1 C2-V5 env sequence regions from a Mother/Infant cohort study are used in the comparison.The output distance matrix and neighbor results produced by the W-curve are functionally equivalent to those from Clustal for C2-V5 sequences in the mother/infant pairs infected with CRF01_AE.Significant potential exists for utilizing this method in place of conventional string based alignment of HIV-1 genomes, such as Clustal X. With W-curve heuristic alignment, it may be possible to obtain clinically useful results in a short time-short enough to affect clinical choices for acute treatment. A description of the W-curve generation process, including a comparison

  18. W-curve alignments for HIV-1 genomic comparisons.

    Science.gov (United States)

    Cork, Douglas J; Lembark, Steven; Tovanabutra, Sodsai; Robb, Merlin L; Kim, Jerome H

    2010-06-01

    The W-curve was originally developed as a graphical visualization technique for viewing DNA and RNA sequences. Its ability to render features of DNA also makes it suitable for computational studies. Its main advantage in this area is utilizing a single-pass algorithm for comparing the sequences. Avoiding recursion during sequence alignments offers advantages for speed and in-process resources. The graphical technique also allows for multiple models of comparison to be used depending on the nucleotide patterns embedded in similar whole genomic sequences. The W-curve approach allows us to compare large numbers of samples quickly. We are currently tuning the algorithm to accommodate quirks specific to HIV-1 genomic sequences so that it can be used to aid in diagnostic and vaccine efforts. Tracking the molecular evolution of the virus has been greatly hampered by gap associated problems predominantly embedded within the envelope gene of the virus. Gaps and hypermutation of the virus slow conventional string based alignments of the whole genome. This paper describes the W-curve algorithm itself, and how we have adapted it for comparison of similar HIV-1 genomes. A treebuilding method is developed with the W-curve that utilizes a novel Cylindrical Coordinate distance method and gap analysis method. HIV-1 C2-V5 env sequence regions from a Mother/Infant cohort study are used in the comparison. The output distance matrix and neighbor results produced by the W-curve are functionally equivalent to those from Clustal for C2-V5 sequences in the mother/infant pairs infected with CRF01_AE. Significant potential exists for utilizing this method in place of conventional string based alignment of HIV-1 genomes, such as Clustal X. With W-curve heuristic alignment, it may be possible to obtain clinically useful results in a short time-short enough to affect clinical choices for acute treatment. A description of the W-curve generation process, including a comparison technique of

  19. HIV epidemiology. The early spread and epidemic ignition of HIV-1 in human populations.

    Science.gov (United States)

    Faria, Nuno R; Rambaut, Andrew; Suchard, Marc A; Baele, Guy; Bedford, Trevor; Ward, Melissa J; Tatem, Andrew J; Sousa, João D; Arinaminpathy, Nimalan; Pépin, Jacques; Posada, David; Peeters, Martine; Pybus, Oliver G; Lemey, Philippe

    2014-10-03

    Thirty years after the discovery of HIV-1, the early transmission, dissemination, and establishment of the virus in human populations remain unclear. Using statistical approaches applied to HIV-1 sequence data from central Africa, we show that from the 1920s Kinshasa (in what is now the Democratic Republic of Congo) was the focus of early transmission and the source of pre-1960 pandemic viruses elsewhere. Location and dating estimates were validated using the earliest HIV-1 archival sample, also from Kinshasa. The epidemic histories of HIV-1 group M and nonpandemic group O were similar until ~1960, after which group M underwent an epidemiological transition and outpaced regional population growth. Our results reconstruct the early dynamics of HIV-1 and emphasize the role of social changes and transport networks in the establishment of this virus in human populations. Copyright © 2014, American Association for the Advancement of Science.

  20. HIV-1 Latency in Monocytes/Macrophages

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    Amit Kumar

    2014-04-01

    Full Text Available Human immunodeficiency virus type 1 (HIV-1 targets CD4+ T cells and cells of the monocyte/macrophage lineage. HIV pathogenesis is characterized by the depletion of T lymphocytes and by the presence of a population of cells in which latency has been established called the HIV-1 reservoir. Highly active antiretroviral therapy (HAART has significantly improved the life of HIV-1 infected patients. However, complete eradication of HIV-1 from infected individuals is not possible without targeting latent sources of infection. HIV-1 establishes latent infection in resting CD4+ T cells and findings indicate that latency can also be established in the cells of monocyte/macrophage lineage. Monocyte/macrophage lineage includes among others, monocytes, macrophages and brain resident macrophages. These cells are relatively more resistant to apoptosis induced by HIV-1, thus are important stable hideouts of the virus. Much effort has been made in the direction of eliminating HIV-1 resting CD4+ T-cell reservoirs. However, it is impossible to achieve a cure for HIV-1 without considering these neglected latent reservoirs, the cells of monocyte/macrophage lineage. In this review we will describe our current understanding of the mechanism of latency in monocyte/macrophage lineage and how such cells can be specifically eliminated from the infected host.

  1. First report of an HIV-1 triple recombinant of subtypes B, C and F in Buenos Aires, Argentina

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    Weissenbacher Mercedes

    2006-09-01

    Full Text Available Abstract We describe the genetic diversity of currently transmitted strains of HIV-1 in men who have sex with men (MSM in Buenos Aires, Argentina between 2000 and 2004. Nearly full-length sequence analysis of 10 samples showed that 6 were subtype B, 3 were BF recombinant and 1 was a triple recombinant of subtypes B, C and F. The 3 BF recombinants were 3 different unique recombinant forms. Full genome analysis of one strain that was subtype F when sequenced in pol was found to be a triple recombinant. Gag and pol were predominantly subtype F, while gp120 was subtype B; there were regions of subtype C interspersed throughout. The young man infected with this strain reported multiple sexual partners and sero-converted between May and November of 2004. This study reported for the first time the full genome analysis of a triple recombinant between subtypes B, C and F, that combines in one virus the three most common subtypes in South America.

  2. Differential expression and interaction of host factors augment HIV-1 gene expression in neonatal mononuclear cells

    International Nuclear Information System (INIS)

    Sundaravaradan, Vasudha; Mehta, Roshni; Harris, David T.; Zack, Jerome A.; Ahmad, Nafees

    2010-01-01

    We have previously shown a higher level of HIV-1 replication and gene expression in neonatal (cord) blood mononuclear cells (CBMC) compared with adult blood cells (PBMC), which could be due to differential expression of host factors. We performed the gene expression profile of CBMC and PBMC and found that 8013 genes were expressed at higher levels in CBMC than PBMC and 8028 genes in PBMC than CBMC, including 1181 and 1414 genes upregulated after HIV-1 infection in CBMC and PBMC, respectively. Several transcription factors (NF-κB, E2F, HAT-1, TFIIE, Cdk9, Cyclin T1), signal transducers (STAT3, STAT5A) and cytokines (IL-1β, IL-6, IL-10) were upregulated in CBMC than PBMC, which are known to influence HIV-1 replication. In addition, a repressor of HIV-1 transcription, YY1, was down regulated in CBMC than PBMC and several matrix metalloproteinase (MMP-7, -12, -14) were significantly upregulated in HIV-1 infected CBMC than PBMC. Furthermore, we show that CBMC nuclear extracts interacted with a higher extent to HIV-1 LTR cis-acting sequences, including NF-κB, NFAT, AP1 and NF-IL6 compared with PBMC nuclear extracts and retroviral based short hairpin RNA (shRNA) for STAT3 and IL-6 down regulated their own and HIV-1 gene expression, signifying that these factors influenced differential HIV-1 gene expression in CBMC than PBMC.

  3. Prevalence of genotypic HIV-1 drug resistance in Thailand, 2002

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    Watitpun Chotip

    2003-03-01

    Full Text Available Abstract Background The prices of reverse transcriptase (RT inhibitors in Thailand have been reduced since December 1, 2001. It is expected that reduction in the price of these inhibitors may influence the drug resistance mutation pattern of HIV-1 among infected people. This study reports the frequency of HIV-1 genetic mutation associated with drug resistance in antiretroviral-treated patients from Thailand. Methods Genotypic resistance testing was performed on samples collected in 2002 from 88 HIV-1 infected individuals. Automated DNA sequencing was used to genotype the HIV-1 polymerase gene isolated from patients' plasma. Results Resistance to protease inhibitors, nucleoside and non-nucleoside reverse transcriptase inhibitors were found in 10 (12%, 42 (48% and 19 (21% patients, respectively. The most common drug resistance mutations in the protease gene were at codon 82 (8%, 90 (7% and 54 (6%, whereas resistant mutations at codon 215 (45%, 67 (40%, 41 (38% and 184 (27% were commonly found in the RT gene. This finding indicates that genotypic resistance to nucleoside reverse transcriptase inhibitors was prevalent in 2002. The frequency of resistant mutations corresponding to non-nucleoside reverse transcriptase inhibitors was three times higher-, while resistant mutation corresponding to protease inhibitors was two times lower than those frequencies determined in 2001. Conclusion This study shows that the frequencies of RT inhibitor resistance mutations have been increased after the reduction in the price of RT inhibitors since December 2001. We believe that this was an important factor that influenced the mutation patterns of HIV-1 protease and RT genes in Thailand.

  4. Molecular epidemiology of HIV-1 infection in Europe: An overview.

    Science.gov (United States)

    Beloukas, Apostolos; Psarris, Alexandros; Giannelou, Polina; Kostaki, Evangelia; Hatzakis, Angelos; Paraskevis, Dimitrios

    2016-12-01

    Human Immunodeficiency Virus type 1 (HIV-1) is characterised by vast genetic diversity. Globally circulating HIV-1 viruses are classified into distinct phylogenetic strains (subtypes, sub-subtypes) and several recombinant forms. Here we describe the characteristics and evolution of European HIV-1 epidemic over time through a review of published literature and updated queries of existing HIV-1 sequence databases. HIV-1 in Western and Central Europe was introduced in the early-1980s in the form of subtype B, which is still the predominant clade. However, in Eastern Europe (Former Soviet Union (FSU) countries and Russia) the predominant strain, introduced into Ukraine in the mid-1990s, is subtype A (A FSU ) with transmission mostly occurring in People Who Inject Drugs (PWID). In recent years, the epidemic is evolving towards a complex tapestry with an increase in the prevalence of non-B subtypes and recombinants in Western and Central Europe. Non-B epidemics are mainly associated with immigrants, heterosexuals and females but more recently, non-B clades have also spread amongst groups where non-B strains were previously absent - non-immigrant European populations and amongst men having sex with men (MSM). In some countries, non-B clades have spread amongst the native population, for example subtype G in Portugal and subtype A in Greece, Albania and Cyprus. Romania provides a unique case where sub-subtype F1 has predominated throughout the epidemic. In contrast, HIV-1 epidemic in FSU countries remains more homogeneous with A FSU clade predominating in all countries. The differences between the evolution of the Western epidemic and the Eastern epidemic may be attributable to differences in transmission risk behaviours, lifestyle and the patterns of human mobility. The study of HIV-1 epidemic diversity provides a useful tool by which we can understand the history of the pandemic in addition to allowing us to monitor the spread and growth of the epidemic over time

  5. Cytoplasmic Dynein Promotes HIV-1 Uncoating

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    Paulina Pawlica

    2014-11-01

    Full Text Available Retroviral capsid (CA cores undergo uncoating during their retrograde transport (toward the nucleus, and/or after reaching the nuclear membrane. However, whether HIV-1 CA core uncoating is dependent upon its transport is not understood. There is some evidence that HIV-1 cores retrograde transport involves cytoplasmic dynein complexes translocating on microtubules. Here we investigate the role of dynein-dependent transport in HIV-1 uncoating. To interfere with dynein function, we depleted dynein heavy chain (DHC using RNA interference, and we over-expressed p50/dynamitin. In immunofluorescence microscopy experiments, DHC depletion caused an accumulation of CA foci in HIV-1 infected cells. Using a biochemical assay to monitor HIV-1 CA core disassembly in infected cells, we observed an increase in amounts of intact (pelletable CA cores upon DHC depletion or p50 over-expression. Results from these two complementary assays suggest that inhibiting dynein-mediated transport interferes with HIV-1 uncoating in infected cells, indicating the existence of a functional link between HIV-1 transport and uncoating.

  6. Cyclophilin B enhances HIV-1 infection.

    Science.gov (United States)

    DeBoer, Jason; Madson, Christian J; Belshan, Michael

    2016-02-01

    Cyclophilin B (CypB) is a member of the immunophilin family and intracellular chaperone. It predominantly localizes to the ER, but also contains a nuclear localization signal and is secreted from cells. CypB has been shown to interact with the Gag protein of human immunodeficiency type 1 (HIV-1). Several proteomic and genetic studies identified it as a potential factor involved in HIV replication. Herein, we show that over-expression of CypB enhances HIV infection by increasing nuclear import of viral DNA. This enhancement was unaffected by cyclosporine treatment and requires the N-terminus of the protein. The N-terminus contains an ER leader sequence, putative nuclear localization signal, and is required for secretion. Deletion of the N-terminus resulted in mislocalization from the ER and suppression of HIV infection. Passive transfer experiments showed that secreted CypB did not impact HIV infection. Combined, these experiments show that intracellular CypB modulates a pathway of HIV nuclear import. Copyright © 2015 Elsevier Inc. All rights reserved.

  7. RNA interactions in the 5' region of the HIV-1 genome

    DEFF Research Database (Denmark)

    Damgaard, Christian Kroun; Andersen, Ebbe Sloth; Knudsen, Bjarne

    2004-01-01

    The untranslated leader of the dimeric HIV-1 RNA genome is folded into a complex structure that plays multiple and essential roles in the viral replication cycle. Here, we have investigated secondary and tertiary structural elements within the 5' 744 nucleotides of the HIV-1 genome using...... a combination of bioinformatics, enzymatic probing, native gel electrophoresis, and UV-crosslinking experiments. We used a recently developed RNA folding algorithm (Pfold) to predict the common secondary structure of an alignment of 20 divergent HIV-1 sequences. Combining this analysis with biochemical data, we...

  8. Forensic application of phylogenetic analyses - Exploration of suspected HIV-1 transmission case.

    Science.gov (United States)

    Siljic, Marina; Salemovic, Dubravka; Cirkovic, Valentina; Pesic-Pavlovic, Ivana; Ranin, Jovan; Todorovic, Marija; Nikolic, Slobodan; Jevtovic, Djordje; Stanojevic, Maja

    2017-03-01

    Transmission of human immunodeficiency virus (HIV) between individuals may have important legal implications and therefore may come to require forensic investigation based upon phylogenetic analysis. In criminal trials results of phylogenetic analyses have been used as evidence of responsibility for HIV transmission. In Serbia, as in many countries worldwide, exposure and deliberate transmission of HIV are criminalized. We present the results of applying state of the art phylogenetic analyses, based on pol and env genetic sequences, in exploration of suspected HIV transmission among three subjects: a man and two women, with presumed assumption of transmission direction from one woman to a man. Phylogenetic methods included relevant neighbor-joining (NJ), maximum likelihood (ML) and Bayesian methods of phylogenetic trees reconstruction and hypothesis testing, that has been shown to be the most sensitive for the reconstruction of epidemiological links mostly from sexually infected individuals. End-point limiting-dilution PCR (EPLD-PCR) assay, generating the minimum of 10 sequences per genetic region per subject, was performed to assess HIV quasispecies distribution and to explore the direction of HIV transmission between three subjects. Phylogenetic analysis revealed that the viral sequences from the three subjects were more genetically related to each other than to other strains circulating in the same area with the similar epidemiological profile, forming strongly supported transmission chain, which could be in favour of a priori hypothesis of one of the women infecting the man. However, in the EPLD based phylogenetic trees for both pol and env genetic region, viral sequences of one subject (man) were paraphyletic to those of two other subjects (women), implying the direction of transmission opposite to the a priori assumption. The dated tree in our analysis confirmed the clustering pattern of query sequences. Still, in the context of unsampled sequences and

  9. Predicting Bevirimat resistance of HIV-1 from genotype

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    Hoffmann Daniel

    2010-01-01

    Full Text Available Abstract Background Maturation inhibitors are a new class of antiretroviral drugs. Bevirimat (BVM was the first substance in this class of inhibitors entering clinical trials. While the inhibitory function of BVM is well established, the molecular mechanisms of action and resistance are not well understood. It is known that mutations in the regions CS p24/p2 and p2 can cause phenotypic resistance to BVM. We have investigated a set of p24/p2 sequences of HIV-1 of known phenotypic resistance to BVM to test whether BVM resistance can be predicted from sequence, and to identify possible molecular mechanisms of BVM resistance in HIV-1. Results We used artificial neural networks and random forests with different descriptors for the prediction of BVM resistance. Random forests with hydrophobicity as descriptor performed best and classified the sequences with an area under the Receiver Operating Characteristics (ROC curve of 0.93 ± 0.001. For the collected data we find that p2 sequence positions 369 to 376 have the highest impact on resistance, with positions 370 and 372 being particularly important. These findings are in partial agreement with other recent studies. Apart from the complex machine learning models we derived a number of simple rules that predict BVM resistance from sequence with surprising accuracy. According to computational predictions based on the data set used, cleavage sites are usually not shifted by resistance mutations. However, we found that resistance mutations could shorten and weaken the α-helix in p2, which hints at a possible resistance mechanism. Conclusions We found that BVM resistance of HIV-1 can be predicted well from the sequence of the p2 peptide, which may prove useful for personalized therapy if maturation inhibitors reach clinical practice. Results of secondary structure analysis are compatible with a possible route to BVM resistance in which mutations weaken a six-helix bundle discovered in recent experiments

  10. Molecular mimicry of human tRNALys anti-codon domain by HIV-1 RNA genome facilitates tRNA primer annealing.

    Science.gov (United States)

    Jones, Christopher P; Saadatmand, Jenan; Kleiman, Lawrence; Musier-Forsyth, Karin

    2013-02-01

    The primer for initiating reverse transcription in human immunodeficiency virus type 1 (HIV-1) is tRNA(Lys3). Host cell tRNA(Lys) is selectively packaged into HIV-1 through a specific interaction between the major tRNA(Lys)-binding protein, human lysyl-tRNA synthetase (hLysRS), and the viral proteins Gag and GagPol. Annealing of the tRNA primer onto the complementary primer-binding site (PBS) in viral RNA is mediated by the nucleocapsid domain of Gag. The mechanism by which tRNA(Lys3) is targeted to the PBS and released from hLysRS prior to annealing is unknown. Here, we show that hLysRS specifically binds to a tRNA anti-codon-like element (TLE) in the HIV-1 genome, which mimics the anti-codon loop of tRNA(Lys) and is located proximal to the PBS. Mutation of the U-rich sequence within the TLE attenuates binding of hLysRS in vitro and reduces the amount of annealed tRNA(Lys3) in virions. Thus, LysRS binds specifically to the TLE, which is part of a larger LysRS binding domain in the viral RNA that includes elements of the Psi packaging signal. Our results suggest that HIV-1 uses molecular mimicry of the anti-codon of tRNA(Lys) to increase the efficiency of tRNA(Lys3) annealing to viral RNA.

  11. Clinical, epidemiological and treatment failure data among HIV-1 non-B-infected patients in the Spanish AIDS Research Network Cohort.

    Science.gov (United States)

    Torrecilla García, Esther; Yebra Sanz, Gonzalo; Llácer-Delicado, Teresa; Rubio García, Rafael; González-García, Juan; García García, Federico; López-Aldeguer, José; Asensi Álvarez, Víctor; Holguín Fernández, África

    2016-01-01

    The prevalence of HIV-1 non-B variants is increasing in Spain, showing a higher number of transmitted drug resistance mutations (TDR) since 2002. This study presents the features of non-B-infected patients enrolled in the cohort of antiretroviral treatment (ART) naïve HIV-infected patients included in the Research Network on HIV/AIDS (CoRIS). The study includes a selected group of HIV-1 non-B-infected subjects from 670 subjects with pol sequences collected from 2004 to 2008 in the CoRIS cohort. Epidemiological-clinical-virological data were analyzed since cohort entry until October 2011, considering the presence or absence of treatment failure (TF). Eighty two non-B infected subjects with known HIV-1 variants were selected from 2004 to 2008 in the CoRIS cohort, being mainly female, immigrants, infected by recombinant viruses, and by heterosexual route. They had an intermediate TDR rate (9.4%), a high rate of TF (25.6%), of losses to follow-up (35%), of coinfections (32.9%), and baseline CD4+ counts ≥350cells/mm(3) (61.8%). Non-B subjects with TF showed higher rates of heterosexual infection (85.7% vs. 69.5%, pHIV-1 non-B-infected patients in Spain had a particular epidemiological and clinical profile that should be considered during their clinical management. Copyright © 2015 Elsevier España, S.L.U. and Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

  12. HIV-1 Tat regulates the expression of the dcw operon and stimulates the proliferation of bacteria.

    Science.gov (United States)

    Wei, Jinsong; Zhang, Yumin; Knapp, Pamela E; Zhao, Tianyong

    2016-01-01

    Infections of pathogenic bacteria are very common in acquired immunodeficiency syndrome (AIDS) patients. However, the biological effects of HIV-1 Tat on bacteria are incompletely understood. In this study, HIV-1 Tat was expressed in Escherichia coli and Pseudomonas aeruginosa (PA01) to investigate its biological effects on bacteria. Bacterial cells expressing either HIV-1 Tat1-86 (Tat1-86) or HIV-1 Tat1-72 (Tat1-72) grow significantly faster than those with either only an empty vector or an unrelated control (GFP or Rluc). Supplementation of purified HIV-1 Tat1-86 or Tat1-101 protein into bacterial culture medium stimulated the growth of both E. coli and PA01. The expression profile of certain cell division-associated genes, such as those in the division cell wall (dcw) operon (ftsA, ftsQ, ftsW and ftsZ), yafO and zipA, was altered in HIV-1 Tat1-86 expressing E. coli BL21(DE3). Furthermore, the expression of firefly luciferase (Fluc) reporter gene, when engineered for control by the dcw promoter and terminator, was enhanced by HIV-1 Tat in E. coli, confirming that HIV-1 Tat transcriptionally regulates the expression of the dcw operon. The finding that HIV-1 Tat stimulates bacterial growth whether it is produced intracellularly or applied extracellularly may have relevance for HIV patients who are highly susceptible to opportunistic bacterial infections. Contents category: Viruses -Retroviruses. The GenBank accession number for the sequence of HIV-1 Tat1-86 is AF324439.1. Copyright © 2015 Elsevier Ltd. All rights reserved.

  13. Low-cost HIV-1 diagnosis and quantification in dried blood spots by real time PCR.

    Science.gov (United States)

    Mehta, Nishaki; Trzmielina, Sonia; Nonyane, Bareng A S; Eliot, Melissa N; Lin, Rongheng; Foulkes, Andrea S; McNeal, Kristina; Ammann, Arthur; Eulalievyolo, Vindu; Sullivan, John L; Luzuriaga, Katherine; Somasundaran, Mohan

    2009-06-05

    Rapid and cost-effective methods for HIV-1 diagnosis and viral load monitoring would greatly enhance the clinical management of HIV-1 infected adults and children in limited-resource settings. Recent recommendations to treat perinatally infected infants within the first year of life are feasible only if early diagnosis is routinely available. Dried blood spots (DBS) on filter paper are an easy and convenient way to collect and transport blood samples. A rapid and cost effective method to diagnose and quantify HIV-1 from DBS is urgently needed to facilitate early diagnosis of HIV-1 infection and monitoring of antiretroviral therapy. We have developed a real-time LightCycler (rtLC) PCR assay to detect and quantify HIV-1 from DBS. HIV-1 RNA extracted from DBS was amplified in a one-step, single-tube system using primers specific for long-terminal repeat sequences that are conserved across all HIV-1 clades. SYBR Green dye was used to quantify PCR amplicons and HIV-1 RNA copy numbers were determined from a standard curve generated using serially diluted known copies of HIV-1 RNA. This assay detected samples across clades, has a dynamic range of 5 log(10), and %CV real-time systems demonstrated similar performance. The accuracy, reliability, genotype inclusivity and affordability, along with the small volumes of blood required for the assay suggest that the rtLC DBS assay will be useful for early diagnosis and monitoring of pediatric HIV-1 infection in resource-limited settings.

  14. Origin and dynamics of HIV-1 subtype C infection in India.

    Directory of Open Access Journals (Sweden)

    Chengli Shen

    Full Text Available To investigate the geographical origin and evolution dynamics of HIV-1 subtype C infection in India.Ninety HIV-1 subtype C env gp120 subtype C sequences from India were compared with 312 env gp120 reference subtype C sequences from 27 different countries obtained from Los Alamos HIV database. All the HIV-1 subtype C env gp120 sequences from India were used for the geographical origin analysis and 61 subtype C env gp120 sequences with known sampling year (from 1991 to 2008 were employed to determine the origin of HIV infection in India.Phylogenetic analysis of HIV-1 env sequences was used to investigate the geographical origin and tMRCA of Indian HIV-1 subtype C. Evolutionary parameters including origin date and demographic growth patterns of Indian subtype C were estimated using a Bayesian coalescent-based approach under relaxed molecular clock models.The majority of the analyzed Indian and South African HIV-1 subtype C sequences formed a single monophyletic cluster. The most recent common ancestor date was calculated to be 1975.56 (95% HPD, 1968.78-1981.52. Reconstruction of the effective population size revealed three phases of epidemic growth: an initial slow growth, followed by exponential growth, and then a plateau phase approaching present time. Stabilization of the epidemic growth phase correlated with the foundation of National AIDS Control Organization in India.Indian subtype C originated from a single South African lineage in the middle of 1970s. The current study emphasizes not only the utility of HIV-1 sequence data for epidemiological studies but more notably highlights the effectiveness of community or government intervention strategies in controlling the trend of the epidemic.

  15. Six Highly Conserved Targets of RNAi Revealed in HIV-1-Infected Patients from Russia Are Also Present in Many HIV-1 Strains Worldwide

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    Olga V. Kretova

    2017-09-01

    Full Text Available RNAi has been suggested for use in gene therapy of HIV/AIDS, but the main problem is that HIV-1 is highly variable and could escape attack from the small interfering RNAs (siRNAs due to even single nucleotide substitutions in the potential targets. To exhaustively check the variability in selected RNA targets of HIV-1, we used ultra-deep sequencing of six regions of HIV-1 from the plasma of two independent cohorts of patients from Russia. Six RNAi targets were found that are invariable in 82%–97% of viruses in both cohorts and are located inside the domains specifying reverse transcriptase (RT, integrase, vpu, gp120, and p17. The analysis of mutation frequencies and their characteristics inside the targets suggests a likely role for APOBEC3G (apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like 3G, A3G in G-to-A mutations and a predominant effect of RT biases in the detected variability of the virus. The lowest frequency of mutations was detected in the central part of all six targets. We also discovered that the identical RNAi targets are present in many HIV-1 strains from many countries and from all continents. The data are important for both the understanding of the patterns of HIV-1 mutability and properties of RT and for the development of gene therapy approaches using RNAi for the treatment of HIV/AIDS. Keywords: HIV-1, RNAi targets, gene therapy, ultra-deep sequencing, conserved HIV-1 sequences

  16. Lack of mutational hot spots during decitabine-mediated HIV-1 mutagenesis.

    Science.gov (United States)

    Rawson, Jonathan M O; Landman, Sean R; Reilly, Cavan S; Bonnac, Laurent; Patterson, Steven E; Mansky, Louis M

    2015-11-01

    Decitabine has previously been shown to induce lethal mutagenesis of human immunodeficiency virus type 1 (HIV-1). However, the factors that determine the susceptibilities of individual sequence positions in HIV-1 to decitabine have not yet been defined. To investigate this, we performed Illumina high-throughput sequencing of multiple amplicons prepared from proviral DNA that was recovered from decitabine-treated cells infected with HIV-1. We found that decitabine induced an ≈4.1-fold increase in the total mutation frequency of HIV-1, primarily due to a striking ≈155-fold increase in the G-to-C transversion frequency. Intriguingly, decitabine also led to an ≈29-fold increase in the C-to-G transversion frequency. G-to-C frequencies varied substantially (up to ≈80-fold) depending upon sequence position, but surprisingly, mutational hot spots (defined as upper outliers within the mutation frequency distribution) were not observed. We further found that every single guanine position examined was significantly susceptible to the mutagenic effects of decitabine. Taken together, these observations demonstrate for the first time that decitabine-mediated HIV-1 mutagenesis is promiscuous and occurs in the absence of a clear bias for mutational hot spots. These data imply that decitabine-mediated G-to-C mutagenesis is a highly effective antiviral mechanism for extinguishing HIV-1 infectivity. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  17. An efficient procedure for the expression and purification of HIV-1 protease from inclusion bodies.

    Science.gov (United States)

    Nguyen, Hong-Loan Thi; Nguyen, Thuy Thi; Vu, Quy Thi; Le, Hang Thi; Pham, Yen; Trinh, Phuong Le; Bui, Thuan Phuong; Phan, Tuan-Nghia

    2015-12-01

    Several studies have focused on HIV-1 protease for developing drugs for treating AIDS. Recombinant HIV-1 protease is used to screen new drugs from synthetic compounds or natural substances. However, large-scale expression and purification of this enzyme is difficult mainly because of its low expression and solubility. In this study, we constructed 9 recombinant plasmids containing a sequence encoding HIV-1 protease along with different fusion tags and examined the expression of the enzyme from these plasmids. Of the 9 plasmids, pET32a(+) plasmid containing the HIV-1 protease-encoding sequence along with sequences encoding an autocleavage site GTVSFNF at the N-terminus and TEV plus 6× His tag at the C-terminus showed the highest expression of the enzyme and was selected for further analysis. The recombinant protein was isolated from inclusion bodies by using 2 tandem Q- and Ni-Sepharose columns. SDS-PAGE of the obtained HIV-1 protease produced a single band of approximately 13 kDa. The enzyme was recovered efficiently (4 mg protein/L of cell culture) and had high specific activity of 1190 nmol min(-1) mg(-1) at an optimal pH of 4.7 and optimal temperature of 37 °C. This procedure for expressing and purifying HIV-1 protease is now being scaled up to produce the enzyme on a large scale for its application. Copyright © 2015 Elsevier Inc. All rights reserved.

  18. Continued Follow-Up of Phambili Phase 2b Randomized HIV-1 Vaccine Trial Participants Supports Increased HIV-1 Acquisition among Vaccinated Men.

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    Zoe Moodie

    Full Text Available The Phase 2b double-blinded, randomized Phambili/HVTN 503 trial evaluated safety and efficacy of the MRK Ad5 gag/pol/nef subtype B HIV-1 preventive vaccine vs placebo in sexually active HIV-1 seronegative participants in South Africa. Enrollment and vaccinations stopped and participants were unblinded but continued follow-up when the Step study evaluating the same vaccine in the Americas, Caribbean, and Australia was unblinded for non-efficacy. Final Phambili analyses found more HIV-1 infections amongst vaccine than placebo recipients, impelling the HVTN 503-S recall study.HVTN 503-S sought to enroll all 695 HIV-1 uninfected Phambili participants, provide HIV testing, risk reduction counseling, physical examination, risk behavior assessment and treatment assignment recall. After adding HVTN 503-S data, HIV-1 infection hazard ratios (HR vaccine vs. placebo were estimated by Cox models.Of the 695 eligible, 465 (67% enrolled with 230 from the vaccine group and 235 from the placebo group. 38% of the 184 Phambili dropouts were enrolled. Enrollment did not differ by treatment group, gender, or baseline HSV-2. With the additional 1286 person years of 503-S follow-up, the estimated HR over Phambili and HVTN 503-S follow-up was 1.52 (95% CI 1.08-2.15, p = 0.02, 82 vaccine/54 placebo infections. The HR was significant for men (HR = 2.75, 95% CI 1.49, 5.06, p = 0.001 but not for women (HR = 1.12, 95% CI 0.73, 1.72, p = 0.62.The additional follow-up from HVTN 503-S supported the Phambili finding of increased HIV-1 acquisition among vaccinated men and strengthened the evidence of lack of vaccine effect among women.clinicaltrials.gov NCT00413725 SA National Health Research Database DOH-27-0207-1539.

  19. NFAT5 regulates HIV-1 in primary monocytes via a highly conserved long terminal repeat site.

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    Shahin Ranjbar

    2006-12-01

    Full Text Available To replicate, HIV-1 capitalizes on endogenous cellular activation pathways resulting in recruitment of key host transcription factors to its viral enhancer. RNA interference has been a powerful tool for blocking key checkpoints in HIV-1 entry into cells. Here we apply RNA interference to HIV-1 transcription in primary macrophages, a major reservoir of the virus, and specifically target the transcription factor NFAT5 (nuclear factor of activated T cells 5, which is the most evolutionarily divergent NFAT protein. By molecularly cloning and sequencing isolates from multiple viral subtypes, and performing DNase I footprinting, electrophoretic mobility shift, and promoter mutagenesis transfection assays, we demonstrate that NFAT5 functionally interacts with a specific enhancer binding site conserved in HIV-1, HIV-2, and multiple simian immunodeficiency viruses. Using small interfering RNA to ablate expression of endogenous NFAT5 protein, we show that the replication of three major HIV-1 viral subtypes (B, C, and E is dependent upon NFAT5 in human primary differentiated macrophages. Our results define a novel host factor-viral enhancer interaction that reveals a new regulatory role for NFAT5 and defines a functional DNA motif conserved across HIV-1 subtypes and representative simian immunodeficiency viruses. Inhibition of the NFAT5-LTR interaction may thus present a novel therapeutic target to suppress HIV-1 replication and progression of AIDS.

  20. Characterization and frequency of a newly identified HIV-1 BF1 intersubtype circulating recombinant form in São Paulo, Brazil

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    Neto Walter

    2010-04-01

    Full Text Available Abstract Background HIV circulating recombinant forms (CRFs play an important role in the global and regional HIV epidemics, particularly in regions where multiple subtypes are circulating. To date, several (>40 CRFs are recognized worldwide with five currently circulating in Brazil. Here, we report the characterization of near full-length genome sequences (NFLG of six phylogenetically related HIV-1 BF1 intersubtype recombinants (five from this study and one from other published sequences representing CRF46_BF1. Methods Initially, we selected 36 samples from 888 adult patients residing in São Paulo who had previously been diagnosed as being infected with subclade F1 based on pol subgenomic fragment sequencing. Proviral DNA integrated in peripheral blood mononuclear cells (PBMC was amplified from the purified genomic DNA of all 36-blood samples by five overlapping PCR fragments followed by direct sequencing. Sequence data were obtained from the five fragments that showed identical genomic structure and phylogenetic trees were constructed and compared with previously published sequences. Genuine subclade F1 sequences and any other sequences that exhibited unique mosaic structures were omitted from further analysis Results Of the 36 samples analyzed, only six sequences, inferred from the pol region as subclade F1, displayed BF1 identical mosaic genomes with a single intersubtype breakpoint identified at the nef-U3 overlap (HXB2 position 9347-9365; LTR region. Five of these isolates formed a rigid cluster in phylogentic trees from different subclade F1 fragment regions, which we can now designate as CRF46_BF1. According to our estimate, the new CRF accounts for 0.56% of the HIV-1 circulating strains in São Paulo. Comparison with previously published sequences revealed an additional five isolates that share an identical mosaic structure with those reported in our study. Despite sharing a similar recombinant structure, only one sequence appeared to

  1. [Molecular epidemiological analysis of HIV-1 in Kazakhstan in 2009-2013].

    Science.gov (United States)

    Lapovok, I A; Laga, V Y; Kazennova, E V; Vasilyev, A V; Dzissyuk, N V; Utegenova, A K; Abishev, A T; Tukeev, M S; Bobkova, M R

    2015-01-01

    In this study pol gene analysis of 205 HIV-1 samples collected in Kazakhstan in 2009 and 2012-2013 was carried out. CRF02_AG variant is dominating in Almaty and actively circulates in East Kazakhstan Province. IDU-A variant is dominating in the rest of Kazakhstan. The data on low prevalence (3%) of HIV drug resistance mutations in native patients were obtained.

  2. Assessment of phylogenetic sensitivity for reconstructing HIV-1 epidemiological relationships.

    Science.gov (United States)

    Beloukas, Apostolos; Magiorkinis, Emmanouil; Magiorkinis, Gkikas; Zavitsanou, Asimina; Karamitros, Timokratis; Hatzakis, Angelos; Paraskevis, Dimitrios

    2012-06-01

    Phylogenetic analysis has been extensively used as a tool for the reconstruction of epidemiological relations for research or for forensic purposes. It was our objective to assess the sensitivity of different phylogenetic methods and various phylogenetic programs to reconstruct epidemiological links among HIV-1 infected patients that is the probability to reveal a true transmission relationship. Multiple datasets (90) were prepared consisting of HIV-1 sequences in protease (PR) and partial reverse transcriptase (RT) sampled from patients with documented epidemiological relationship (target population), and from unrelated individuals (control population) belonging to the same HIV-1 subtype as the target population. Each dataset varied regarding the number, the geographic origin and the transmission risk groups of the sequences among the control population. Phylogenetic trees were inferred by neighbor-joining (NJ), maximum likelihood heuristics (hML) and Bayesian methods. All clusters of sequences belonging to the target population were correctly reconstructed by NJ and Bayesian methods receiving high bootstrap and posterior probability (PP) support, respectively. On the other hand, TreePuzzle failed to reconstruct or provide significant support for several clusters; high puzzling step support was associated with the inclusion of control sequences from the same geographic area as the target population. In contrary, all clusters were correctly reconstructed by hML as implemented in PhyML 3.0 receiving high bootstrap support. We report that under the conditions of our study, hML using PhyML, NJ and Bayesian methods were the most sensitive for the reconstruction of epidemiological links mostly from sexually infected individuals. Copyright © 2012 Elsevier B.V. All rights reserved.

  3. A small set of succinct signature patterns distinguishes Chinese and non-Chinese HIV-1 genomes.

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    Yan Wang

    Full Text Available The epidemiology of HIV-1 in China has unique features that may have led to unique viral strains. We therefore tested the hypothesis that it is possible to find distinctive patterns in HIV-1 genomes sampled in China. Using a rule inference algorithm we could indeed extract from sequences of the third variable loop (V3 of HIV-1 gp120 a set of 14 signature patterns that with 89% accuracy distinguished Chinese from non-Chinese sequences. These patterns were found to be specific to HIV-1 subtype, i.e. sequences complying with pattern 1 were of subtype B, pattern 2 almost exclusively covered sequences of subtype 01_AE, etc. We then analyzed the first of these signature patterns in depth, namely that L and W at two V3 positions are specifically occurring in Chinese sequences of subtype B/B' (3% false positives. This pattern was found to be in agreement with the phylogeny of HIV-1 of subtype B inside and outside of China. We could neither reject nor convincingly confirm that the pattern is stabilized by immune escape. For further interpretation of the signature pattern we used the recently developed measure of Direct Information, and in this way discovered evidence for physical interactions between V2 and V3. We conclude by a discussion of limitations of signature patterns, and the applicability of the approach to other genomic regions and other countries.

  4. Lack of detection of XMRV in seminal plasma from HIV-1 infected men in The Netherlands.

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    Marion Cornelissen

    Full Text Available BACKGROUND: Xenotropic murine leukaemia virus-related virus (XMRV is a recently discovered human gammaretrovirus with yet unknown prevalence and transmission route(s. Its presence in prostate stromal fibroblasts and prostatic secretions suggests that XMRV might be sexually transmitted. We chose to study a compartment closely connected to the prostate, a location where XMRV was detected in independent studies. Seminal plasma samples from HIV-1 infected men were examined as they have an increased probability of acquiring sexually transmitted pathogens. METHODOLOGY/PRINCIPAL FINDINGS: We studied the prevalence of XMRV in 93 seminal plasma samples of 54 HIV-1 infected men living in The Netherlands with a nested PCR amplification specifically targeting the XMRV gag gene. As a control for the presence and integrity of retrovirus particles, HIV-1 was amplified from the same samples with a PCR amplification targeting the env gene of the virus, or HIV-1 was quantified with a real-time PCR amplifying part of the pol gene. CONCLUSIONS/SIGNIFICANCE: Although HIV-1 was amplified from 25% of the seminal plasma samples, no XMRV was detected, suggesting that either the prevalence of XMRV is very low in The Netherlands, or that XMRV is not naturally present in the seminal plasma.

  5. Resistance mutations and CTL epitopes in archived HIV-1 DNA of patients on antiviral treatment: toward a new concept of vaccine.

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    Jennifer Papuchon

    Full Text Available Eleven patients responding successfully to first-line antiretroviral therapy (ART were investigated for proviral drug resistance mutations (DRMs in RT by ultra-deep pyrosequencing (UDPS. After molecular typing of the class I alleles A and B, the CTL epitopes in the Gag, Nef and Pol regions of the provirus were sequenced and compared to the reference HXB2 HIV-1 epitopes. They were then matched with the HLA alleles with determination of theoretical affinity (TA. For 3 patients, the results could be compared with an RNA sample of the circulating virus at initiation of therapy. Five out of 11 patients exhibited DRMs by UDPS. The issue is whether a therapeutic switch is relevant in these patients by taking into account the identity of the archived resistance mutations. When the archived CTL epitopes were determined on the basis of the HLA alleles, different patterns were observed. Some epitopes were identical to those reported for the reference with the same TA, while others were mutated with a decrease in TA. In 2 cases, an epitope was observed as a combination of subpopulations at entry and was retrieved as a single population with lower TA at success. With regard to immunological stimulation and given the variability of the archived CTL epitopes, we propose a new concept of curative vaccine based on identification of HIV-1 CTL epitopes after prior sequencing of proviral DNA and matching with HLA class I alleles.

  6. Positron emission tomography in patients suffering from HIV-1 infection

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    Sathekge, Mike [University Hospital of Pretoria, Department of Nuclear Medicine, Pretoria (South Africa); Goethals, Ingeborg; Wiele, Christophe van de [University Hospital Ghent, Department of Nuclear Medicine, Ghent (Belgium); Maes, Alex [AZ Groening, Department of Nuclear Medicine, Kortrijk (Belgium)

    2009-07-15

    This paper reviews currently available PET studies performed either to improve our understanding of the pathogenesis of HIV-1 infection or to assess the value of PET imaging in the clinical decision making of patients infected with HIV-1 presenting with AIDS-related opportunistic infections and malignancies. FDG PET has shown that HIV-1 infection progresses by distinct anatomical steps, with involvement of the upper torso preceding involvement of the lower part of the torso, and that the degree of FDG uptake relates to viral load. The former finding suggests that lymphoid tissues are engaged in a predictable sequence and that diffusible mediators of activation might be important targets for vaccine or therapeutic intervention strategies. In lipodystrophic HIV-infected patients, limited available data support the hypothesis that stavudine-related lipodystrophy is associated with increased glucose uptake by adipose tissue as a result of the metabolic stress of adipose tissue in response to highly active antiretroviral treatment (HAART). Finally, in early AIDS-related dementia complex (ADC), striatal hypermetabolism is observed, whereas progressive ADC is characterized by a decrease in subcortical and cortical metabolism. In the clinical setting, PET has been shown to allow the differentiation of AIDS-related opportunistic infections and malignancies, and to allow monitoring of side effects of HAART. However, in patients suffering from HIV infection and presenting with extracerebral lymphoma or other human malignancies, knowledge of viraemia is essential when interpreting FDG PET imaging. (orig.)

  7. Positron emission tomography in patients suffering from HIV-1 infection

    International Nuclear Information System (INIS)

    Sathekge, Mike; Goethals, Ingeborg; Wiele, Christophe van de; Maes, Alex

    2009-01-01

    This paper reviews currently available PET studies performed either to improve our understanding of the pathogenesis of HIV-1 infection or to assess the value of PET imaging in the clinical decision making of patients infected with HIV-1 presenting with AIDS-related opportunistic infections and malignancies. FDG PET has shown that HIV-1 infection progresses by distinct anatomical steps, with involvement of the upper torso preceding involvement of the lower part of the torso, and that the degree of FDG uptake relates to viral load. The former finding suggests that lymphoid tissues are engaged in a predictable sequence and that diffusible mediators of activation might be important targets for vaccine or therapeutic intervention strategies. In lipodystrophic HIV-infected patients, limited available data support the hypothesis that stavudine-related lipodystrophy is associated with increased glucose uptake by adipose tissue as a result of the metabolic stress of adipose tissue in response to highly active antiretroviral treatment (HAART). Finally, in early AIDS-related dementia complex (ADC), striatal hypermetabolism is observed, whereas progressive ADC is characterized by a decrease in subcortical and cortical metabolism. In the clinical setting, PET has been shown to allow the differentiation of AIDS-related opportunistic infections and malignancies, and to allow monitoring of side effects of HAART. However, in patients suffering from HIV infection and presenting with extracerebral lymphoma or other human malignancies, knowledge of viraemia is essential when interpreting FDG PET imaging. (orig.)

  8. Assessing the HIV-1 Epidemic in Brazilian Drug Users: A Molecular Epidemiology Approach.

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    Monick Lindenmeyer Guimarães

    Full Text Available Person who inject illicit substances have an important role in HIV-1 blood and sexual transmission and together with person who uses heavy non-injecting drugs may have less than optimal adherence to anti-retroviral treatment and eventually could transmit resistant HIV variants. Unfortunately, molecular biology data on such key population remain fragmentary in most low and middle-income countries. The aim of the present study was to assess HIV infection rates, evaluate HIV-1 genetic diversity, drug resistance, and to identify HIV transmission clusters in heavy drug users (DUs. For this purpose, DUs were recruited in the context of a Respondent-Driven Sampling (RDS study in different Brazilian cities during 2009. Overall, 2,812 individuals were tested for HIV, and 168 (6% of them were positive, of which 19 (11.3% were classified as recent seroconverters, corresponding to an estimated incidence rate of 1.58%/year (95% CI 0.92-2.43%. Neighbor joining phylogenetic trees from env and pol regions and bootscan analyses were employed to subtype the virus from132 HIV-1-infected individuals. HIV-1 subtype B was prevalent in most of the cities under analysis, followed by BF recombinants (9%-35%. HIV-1 subtype C was the most prevalent in Curitiba (46% and Itajaí (86% and was also detected in Brasília (9% and Campo Grande (20%. Pure HIV-1F infections were detected in Rio de Janeiro (9%, Recife (6%, Salvador (6% and Brasília (9%. Clusters of HIV transmission were assessed by Maximum likelihood analyses and were cross-compared with the RDS network structure. Drug resistance mutations were verified in 12.2% of DUs. Our findings reinforce the importance of the permanent HIV-1 surveillance in distinct Brazilian cities due to viral resistance and increasing subtype heterogeneity all over Brazil, with relevant implications in terms of treatment monitoring, prophylaxis and vaccine development.

  9. Semen Bacterial Concentrations and HIV-1 RNA Shedding Among HIV-1–Seropositive Kenyan Men

    Science.gov (United States)

    Srinivasan, Sujatha; Huang, Dandi; Ko, Daisy L.; Sanders, Eduard J.; Peshu, Norbert M.; Krieger, John N.; Muller, Charles H.; Coombs, Robert W.; Fredricks, David N.; Graham, Susan M.

    2017-01-01

    Introduction: HIV-1 is transmitted through semen from men to their sexual partners. Genital infections can increase HIV-1 RNA shedding in semen, but shedding also occurs in the absence of typical pathogens. We hypothesized that higher bacterial concentrations in semen would be associated with higher HIV-1 RNA levels. Methods: We analyzed semen samples from 42 HIV-1–seropositive Kenyan men using quantitative polymerase chain reaction (PCR) to assess bacterial concentrations and real-time PCR to measure HIV-1 RNA levels. Generalized estimation equations were used to evaluate associations between these 2 measures. Broad-range 16S rRNA gene PCR with pyrosequencing was performed on a subset of 13 samples to assess bacterial community composition. Results: Bacteria were detected in 96.6% of 88 samples by quantitative PCR. Semen bacterial concentration and HIV-1 RNA levels were correlated 0.30 (P = 0.01). The association between bacterial concentration and HIV-1 RNA detection was not significant after adjustment for antiretroviral therapy (ART) (adjusted odds ratio: 1.27, 95% CI: 0.84 to 1.91). Factors associated with semen bacterial concentration included insertive anal sex (adjusted beta 0.92, 95% CI: 0.12 to 1.73) and ART use (adjusted beta: −0.77, 95% CI: −1.50 to 0.04). Among 13 samples with pyrosequencing data, Corynebacterium spp., Staphylococcus spp., and Streptococcus spp. were most frequently detected. Conclusion: Most of these HIV-1–infected men had bacteria in their semen. ART use was associated with undetectable semen HIV-1 RNA and lower semen bacterial concentrations, whereas insertive anal sex was associated with higher bacterial concentrations. Additional studies evaluating the relationship between semen bacteria, inflammation, mucosal immunity, and HIV-1 shedding are needed to understand implications for HIV-1 transmission. PMID:27861240

  10. Semen Bacterial Concentrations and HIV-1 RNA Shedding Among HIV-1-Seropositive Kenyan Men.

    Science.gov (United States)

    Korhonen, Christine J; Srinivasan, Sujatha; Huang, Dandi; Ko, Daisy L; Sanders, Eduard J; Peshu, Norbert M; Krieger, John N; Muller, Charles H; Coombs, Robert W; Fredricks, David N; Graham, Susan M

    2017-03-01

    HIV-1 is transmitted through semen from men to their sexual partners. Genital infections can increase HIV-1 RNA shedding in semen, but shedding also occurs in the absence of typical pathogens. We hypothesized that higher bacterial concentrations in semen would be associated with higher HIV-1 RNA levels. We analyzed semen samples from 42 HIV-1-seropositive Kenyan men using quantitative polymerase chain reaction (PCR) to assess bacterial concentrations and real-time PCR to measure HIV-1 RNA levels. Generalized estimation equations were used to evaluate associations between these 2 measures. Broad-range 16S rRNA gene PCR with pyrosequencing was performed on a subset of 13 samples to assess bacterial community composition. Bacteria were detected in 96.6% of 88 samples by quantitative PCR. Semen bacterial concentration and HIV-1 RNA levels were correlated 0.30 (P = 0.01). The association between bacterial concentration and HIV-1 RNA detection was not significant after adjustment for antiretroviral therapy (ART) (adjusted odds ratio: 1.27, 95% CI: 0.84 to 1.91). Factors associated with semen bacterial concentration included insertive anal sex (adjusted beta 0.92, 95% CI: 0.12 to 1.73) and ART use (adjusted beta: -0.77, 95% CI: -1.50 to 0.04). Among 13 samples with pyrosequencing data, Corynebacterium spp., Staphylococcus spp., and Streptococcus spp. were most frequently detected. Most of these HIV-1-infected men had bacteria in their semen. ART use was associated with undetectable semen HIV-1 RNA and lower semen bacterial concentrations, whereas insertive anal sex was associated with higher bacterial concentrations. Additional studies evaluating the relationship between semen bacteria, inflammation, mucosal immunity, and HIV-1 shedding are needed to understand implications for HIV-1 transmission.

  11. Tenofovir-based regimens associated with less drug resistance in HIV-1-infected Nigerians failing first-line antiretroviral therapy.

    Science.gov (United States)

    Etiebet, Mary-Ann A; Shepherd, James; Nowak, Rebecca G; Charurat, Man; Chang, Harry; Ajayi, Samuel; Elegba, Olufunmilayo; Ndembi, Nicaise; Abimiku, Alashle; Carr, Jean K; Eyzaguirre, Lindsay M; Blattner, William A

    2013-02-20

    In resource-limited settings, HIV-1 drug resistance testing to guide antiretroviral therapy (ART) selection is unavailable. We retrospectively conducted genotypic analysis on archived samples from Nigerian patients who received targeted viral load testing to confirm treatment failure and report their drug resistance mutation patterns. Stored plasma from 349 adult patients on non-nucleoside reverse transcriptase inhibitor (NNRTI) regimens was assayed for HIV-1 RNA viral load, and samples with more than 1000 copies/ml were sequenced in the pol gene. Analysis for resistance mutations utilized the IAS-US 2011 Drug Resistance Mutation list. One hundred and seventy-five samples were genotyped; the majority of the subtypes were G (42.9%) and CRF02_AG (33.7%). Patients were on ART for a median of 27 months. 90% had the M184V/I mutation, 62% had at least one thymidine analog mutation, and 14% had the K65R mutation. 97% had an NNRTI resistance mutation and 47% had at least two etravirine-associated mutations. In multivariate analysis tenofovir-based regimens were less likely to have at least three nucleoside reverse transcriptase inhibitor (NRTI) mutations after adjusting for subtype, previous ART, CD4, and HIV viral load [P < 0.001, odds ratio (OR) 0.04]. 70% of patients on tenofovir-based regimens had at least two susceptible NRTIs to include in a second-line regimen compared with 40% on zidovudine-based regimens (P = 0.04, OR = 3.4). At recognition of treatment failure, patients on tenofovir-based first-line regimens had fewer NRTI drug-resistant mutations and more active NRTI drugs available for second-line regimens. These findings can inform strategies for ART regimen sequencing to optimize long-term HIV treatment outcomes in low-resource settings.

  12. μ-opioid modulation of HIV-1 coreceptor expressionand HIV-1 replication

    International Nuclear Information System (INIS)

    Steele, Amber D.; Henderson, Earl E.; Rogers, Thomas J.

    2003-01-01

    A substantial proportion of HIV-1-infected individuals are intravenous drug users (IVDUs) who abuse opiates. Opioids induce a number of immunomodulatory effects that may directly influence HIV-1 disease progression. In the present report, we have investigated the effect of opioids on the expression of the major HIV-1 coreceptors CXCR4 and CCR5. For these studies we have focused on opiates which are ligands for the μ-opioid receptor. Our results show that DAMGO, a selective μ-opioid agonist, increases CXCR4 and CCR5 expression in both CD3 + lymphoblasts and CD14 + monocytes three- to fivefold. Furthermore, DAMGO-induced elevation of HIV-1 coreceptor expression translates into enhanced replication of both X4 and R5 viral strains of HIV-1. We have confirmed the role of the μ-opioid receptor based on the ability of a μ-opioid receptor-selective antagonist to block the effects of DAMGO. We have also found that morphine enhances CXCR4 and CCR5 expression and subsequently increases both X4 and R5 HIV-1 infection. We suggest that the capacity of μ-opioids to increase HIV-1 coreceptor expression and replication may promote viral binding, trafficking of HIV-1-infected cells, and enhanced disease progression

  13. Influenza vaccination of HIV-1-positive and HIV-1-negative former intravenous drug users.

    Science.gov (United States)

    Amendola, A; Boschini, A; Colzani, D; Anselmi, G; Oltolina, A; Zucconi, R; Begnini, M; Besana, S; Tanzi, E; Zanetti, A R

    2001-12-01

    The immunogenicity of an anti-influenza vaccine was assessed in 409 former intravenous drug user volunteers and its effect on the levels of HIV-1 RNA, proviral DNA and on CD4+ lymphocyte counts in a subset HIV-1-positive subjects was measured. HIV-1-positive individuals (n = 72) were divided into three groups on the basis of their CD4+ lymphocyte counts, while the 337 HIV-1-negative participants were allocated into group four. Haemagglutination inhibiting (HI) responses varied from 45.8 to 70% in the HIV-1-positive subjects and were significantly higher in group four (80.7% responses to the H1N1 strain, 81.6% to the H3N2 strain, and 83% to the B strain). The percentage of subjects with HI protective antibody titres (> or = 1:40) increased significantly after vaccination, especially in HIV-1 uninfected subjects. Immunization caused no significant changes in CD4+ counts and in neither plasma HIV-1 RNA nor proviral DNA levels. Therefore, vaccination against influenza may benefit persons infected by HIV-1. Copyright 2001 Wiley-Liss, Inc.

  14. HIV-1 subtype A gag variability and epitope evolution.

    Science.gov (United States)

    Abidi, Syed Hani; Kalish, Marcia L; Abbas, Farhat; Rowland-Jones, Sarah; Ali, Syed

    2014-01-01

    The aim of this study was to examine the course of time-dependent evolution of HIV-1 subtype A on a global level, especially with respect to the dynamics of immunogenic HIV gag epitopes. We used a total of 1,893 HIV-1 subtype A gag sequences representing a timeline from 1985 through 2010, and 19 different countries in Africa, Europe and Asia. The phylogenetic relationship of subtype A gag and its epidemic dynamics was analysed through a Maximum Likelihood tree and Bayesian Skyline plot, genomic variability was measured in terms of G → A substitutions and Shannon entropy, and the time-dependent evolution of HIV subtype A gag epitopes was examined. Finally, to confirm observations on globally reported HIV subtype A sequences, we analysed the gag epitope data from our Kenyan, Pakistani, and Afghan cohorts, where both cohort-specific gene epitope variability and HLA restriction profiles of gag epitopes were examined. The most recent common ancestor of the HIV subtype A epidemic was estimated to be 1956 ± 1. A period of exponential growth began about 1980 and lasted for approximately 7 years, stabilized for 15 years, declined for 2-3 years, then stabilized again from about 2004. During the course of evolution, a gradual increase in genomic variability was observed that peaked in 2005-2010. We observed that the number of point mutations and novel epitopes in gag also peaked concurrently during 2005-2010. It appears that as the HIV subtype A epidemic spread globally, changing population immunogenetic pressures may have played a role in steering immune-evolution of this subtype in new directions. This trend is apparent in the genomic variability and epitope diversity of HIV-1 subtype A gag sequences.

  15. HIV-1 subtype A gag variability and epitope evolution.

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    Syed Hani Abidi

    Full Text Available OBJECTIVE: The aim of this study was to examine the course of time-dependent evolution of HIV-1 subtype A on a global level, especially with respect to the dynamics of immunogenic HIV gag epitopes. METHODS: We used a total of 1,893 HIV-1 subtype A gag sequences representing a timeline from 1985 through 2010, and 19 different countries in Africa, Europe and Asia. The phylogenetic relationship of subtype A gag and its epidemic dynamics was analysed through a Maximum Likelihood tree and Bayesian Skyline plot, genomic variability was measured in terms of G → A substitutions and Shannon entropy, and the time-dependent evolution of HIV subtype A gag epitopes was examined. Finally, to confirm observations on globally reported HIV subtype A sequences, we analysed the gag epitope data from our Kenyan, Pakistani, and Afghan cohorts, where both cohort-specific gene epitope variability and HLA restriction profiles of gag epitopes were examined. RESULTS: The most recent common ancestor of the HIV subtype A epidemic was estimated to be 1956 ± 1. A period of exponential growth began about 1980 and lasted for approximately 7 years, stabilized for 15 years, declined for 2-3 years, then stabilized again from about 2004. During the course of evolution, a gradual increase in genomic variability was observed that peaked in 2005-2010. We observed that the number of point mutations and novel epitopes in gag also peaked concurrently during 2005-2010. CONCLUSION: It appears that as the HIV subtype A epidemic spread globally, changing population immunogenetic pressures may have played a role in steering immune-evolution of this subtype in new directions. This trend is apparent in the genomic variability and epitope diversity of HIV-1 subtype A gag sequences.

  16. Etravirine and Rilpivirine Drug Resistance Among HIV-1 Subtype C Infected Children Failing Non-Nucleoside Reverse Transcriptase Inhibitor-Based Regimens in South India.

    Science.gov (United States)

    Saravanan, Shanmugam; Kausalya, Bagavathi; Gomathi, Selvamurthi; Sivamalar, Sathasivam; Pachamuthu, Balakrishnan; Selvamuthu, Poongulali; Pradeep, Amrose; Sunil, Solomon; Mothi, Sarvode N; Smith, Davey M; Kantor, Rami

    2017-06-01

    We have analyzed reverse transcriptase (RT) region of HIV-1 pol gene from 97 HIV-infected children who were identified as failing first-line therapy that included first-generation non-nucleoside RT inhibitors (Nevirapine and Efavirenz) for at least 6 months. We found that 54% and 65% of the children had genotypically predicted resistance to second-generation non-nucleoside RT inhibitors drugs Etravirine (ETR) and Rilpivirine, respectively. These cross-resistance mutations may compromise future NNRTI-based regimens, especially in resource-limited settings. To complement these investigations, we also analyzed the sequences in Stanford database, Monogram weighted score, and DUET weighted score algorithms for ETR susceptibility and found almost perfect agreement between the three algorithms in predicting ETR susceptibility from genotypic data.

  17. HIV-1 Reservoir Association with Immune Activation

    Directory of Open Access Journals (Sweden)

    Alejandro Vallejo

    2015-09-01

    Full Text Available In this issue of EBioMedicine, Ruggiero and colleagues describe immune activation biomarkers associated with the size of the HIV reservoir in a carefully designed cross-sectional study. The cohort consists of a homogeneous sample of HIV-1-infected patients with long-term plasma HIV-1 RNA suppression under antiretroviral treatment (ART. It is crucial to explore the potential utility of biomarkers that are easier (less labor intensive, less expensive to measure than integrated HIV DNA load, in order to quickly and accurately quantify cellular reservoirs of HIV.

  18. MAS NMR of HIV-1 protein assemblies

    Science.gov (United States)

    Suiter, Christopher L.; Quinn, Caitlin M.; Lu, Manman; Hou, Guangjin; Zhang, Huilan; Polenova, Tatyana

    2015-04-01

    The negative global impact of the AIDS pandemic is well known. In this perspective article, the utility of magic angle spinning (MAS) NMR spectroscopy to answer pressing questions related to the structure and dynamics of HIV-1 protein assemblies is examined. In recent years, MAS NMR has undergone major technological developments enabling studies of large viral assemblies. We discuss some of these evolving methods and technologies and provide a perspective on the current state of MAS NMR as applied to the investigations into structure and dynamics of HIV-1 assemblies of CA capsid protein and of Gag maturation intermediates.

  19. Alterations of HIV-1 envelope phenotype and antibody-mediated neutralization by signal peptide mutations.

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    Chitra Upadhyay

    2018-01-01

    Full Text Available HIV-1 envelope glycoprotein (Env mediates virus attachment and entry into the host cells. Like other membrane-bound and secreted proteins, HIV-1 Env contains at its N terminus a signal peptide (SP that directs the nascent Env to the endoplasmic reticulum (ER where Env synthesis and post-translational modifications take place. SP is cleaved during Env biosynthesis but potentially influences the phenotypic traits of the Env protein. The Env SP sequences of HIV-1 isolates display high sequence variability, and the significance of such variability is unclear. We postulate that changes in the Env SP influence Env transport through the ER-Golgi secretory pathway and Env folding and/or glycosylation that impact on Env incorporation into virions, receptor binding and antibody recognition. We first evaluated the consequences of mutating the charged residues in the Env SP in the context of infectious molecular clone HIV-1 REJO.c/2864. Results show that three different mutations affecting histidine at position 12 affected Env incorporation into virions that correlated with reduction of virus infectivity and DC-SIGN-mediated virus capture and transmission. Mutations at positions 8, 12, and 15 also rendered the virus more resistant to neutralization by monoclonal antibodies against the Env V1V2 region. These mutations affected the oligosaccharide composition of N-glycans as shown by changes in Env reactivity with specific lectins and by mass spectrometry. Increased neutralization resistance and N-glycan composition changes were also observed when analogous mutations were introduced to another HIV-1 strain, JRFL. To the best of our knowledge, this is the first study showing that certain residues in the HIV-1 Env SP can affect virus neutralization sensitivity by modulating oligosaccharide moieties on the Env N-glycans. The HIV-1 Env SP sequences thus may be under selective pressure to balance virus infectiousness with virus resistance to the host antibody

  20. Complexity and dynamics of HIV-1 chemokine receptor usage in a multidrug-resistant adolescent.

    Science.gov (United States)

    Cavarelli, Mariangela; Mainetti, Lara; Pignataro, Angela Rosa; Bigoloni, Alba; Tolazzi, Monica; Galli, Andrea; Nozza, Silvia; Castagna, Antonella; Sampaolo, Michela; Boeri, Enzo; Scarlatti, Gabriella

    2014-12-01

    Maraviroc (MVC) is licensed in clinical practice for patients with R5 virus and virological failure; however, in anecdotal reports, dual/mixed viruses were also inhibited. We retrospectively evaluated the evolution of HIV-1 coreceptor tropism in plasma and peripheral blood mononuclear cells (PBMCs) of an infected adolescent with a CCR5/CXCR4 Trofile profile who experienced an important but temporary immunological and virological response during a 16-month period of MVC-based therapy. Coreceptor usage of biological viral clones isolated from PBMCs was investigated in U87.CD4 cells expressing wild-type or chimeric CCR5 and CXCR4. Plasma and PBMC-derived viral clones were sequenced to predict coreceptor tropism using the geno2pheno algorithm from the V3 envelope sequence and pol gene-resistant mutations. From start to 8.5 months of MVC treatment only R5X4 viral clones were observed, whereas at 16 months the phenotype enlarged to also include R5 and X4 clones. Chimeric receptor usage suggested the preferential usage of the CXCR4 coreceptor by the R5X4 biological clones. According to phenotypic data, R5 viruses were susceptible, whereas R5X4 and X4 viruses were resistant to RANTES and MVC in vitro. Clones at 16 months, but not at baseline, showed an amino acidic resistance pattern in protease and reverse transcription genes, which, however, did not drive their tropisms. The geno2pheno algorithm predicted at baseline R5 viruses in plasma, and from 5.5 months throughout follow-up only CXCR4-using viruses. An extended methodological approach is needed to unravel the complexity of the phenotype and variation of viruses resident in the different compartments of an infected individual. The accurate evaluation of the proportion of residual R5 viruses may guide therapeutic intervention in highly experienced patients with limited therapeutic options.

  1. Novel tetra-peptide insertion in Gag-p6 ALIX-binding motif in HIV-1 subtype C associated with protease inhibitor failure in Indian patients.

    Science.gov (United States)

    Neogi, Ujjwal; Rao, Shwetha D; Bontell, Irene; Verheyen, Jens; Rao, Vasudev R; Gore, Sagar C; Soni, Neelesh; Shet, Anita; Schülter, Eugen; Ekstrand, Maria L; Wondwossen, Amogne; Kaiser, Rolf; Madhusudhan, Mallur S; Prasad, Vinayaka R; Sonnerborg, Anders

    2014-09-24

    A novel tetra-peptide insertion was identified in Gag-p6 ALIX-binding region, which appeared in protease inhibitor failure Indian HIV-1C sequences (odds ratio=17.1, P < 0.001) but was naturally present in half of untreated Ethiopian HIV-1C sequences. The insertion is predicted to restore ALIX-mediated virus release pathway, which is lacking in HIV-1C. The clinical importance of the insertion needs to be evaluated in HIV-1C dominating regions wherein the use of protease inhibitor drugs are being scaled up.

  2. Prediction of HIV-1 sensitivity to broadly neutralizing antibodies shows a trend towards resistance over time.

    Science.gov (United States)

    Hake, Anna; Pfeifer, Nico

    2017-10-01

    Treatment with broadly neutralizing antibodies (bNAbs) has proven effective against HIV-1 infections in humanized mice, non-human primates, and humans. Due to the high mutation rate of HIV-1, resistance testing of the patient's viral strains to the bNAbs is still inevitable. So far, bNAb resistance can only be tested in expensive and time-consuming neutralization experiments. Here, we introduce well-performing computational models that predict the neutralization response of HIV-1 to bNAbs given only the envelope sequence of the virus. Using non-linear support vector machines based on a string kernel, the models learnt even the important binding sites of bNAbs with more complex epitopes, i.e., the CD4 binding site targeting bNAbs, proving thereby the biological relevance of the models. To increase the interpretability of the models, we additionally provide a new kind of motif logo for each query sequence, visualizing those residues of the test sequence that influenced the prediction outcome the most. Moreover, we predicted the neutralization sensitivity of around 34,000 HIV-1 samples from different time points to a broad range of bNAbs, enabling the first analysis of HIV resistance to bNAbs on a global scale. The analysis showed for many of the bNAbs a trend towards antibody resistance over time, which had previously only been discovered for a small non-representative subset of the global HIV-1 population.

  3. CRISPR/Cas9 Inhibits Multiple Steps of HIV-1 Infection.

    Science.gov (United States)

    Yin, Lijuan; Hu, Siqi; Mei, Shan; Sun, Hong; Xu, Fengwen; Li, Jian; Zhu, Weijun; Liu, Xiaoman; Zhao, Fei; Zhang, Di; Cen, Shan; Liang, Chen; Guo, Fei

    2018-05-09

    CRISPR/Cas9 is an adaptive immune system where bacteria and archaea have evolved to resist the invading viruses and plasmid DNA by creating site-specific double-strand breaks in DNA. This study tested this gene editing system in inhibiting human immunodeficiency virus type 1 (HIV-1) infection by targeting the viral long terminal repeat and the gene coding sequences. Strong inhibition of HIV-1 infection by Cas9/gRNA was observed, which resulted not only from insertions and deletions (indels) that were introduced into viral DNA due to Cas9 cleavage, but also from the marked decrease in the levels of the late viral DNA products and the integrated viral DNA. This latter defect might have reflected the degradation of viral DNA that has not been immediately repaired after Cas9 cleavage. It was further observed that Cas9, when solely located in the cytoplasm, inhibits HIV-1 as strongly as the nuclear Cas9, except that the cytoplasmic Cas9 does not act on the integrated HIV-1 DNA and thus cannot be used to excise the latent provirus. Together, the results suggest that Cas9/gRNA is able to target and edit HIV-1 DNA both in the cytoplasm and in the nucleus. The inhibitory effect of Cas9 on HIV-1 is attributed to both the indels in viral DNA and the reduction in the levels of viral DNA.

  4. Correlation of Naturally Occurring HIV-1 Resistance to DEB025 with Capsid Amino Acid Polymorphisms

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    Brigitte Rosenwirth

    2013-03-01

    Full Text Available DEB025 (alisporivir is a synthetic cyclosporine with inhibitory activity against human immunodeficiency virus type-1 (HIV-1 and hepatitis C virus (HCV. It binds to cyclophilin A (CypA and blocks essential functions of CypA in the viral replication cycles of both viruses. DEB025 inhibits clinical HIV-1 isolates in vitro and decreases HIV-1 virus load in the majority of patients. HIV-1 isolates being naturally resistant to DEB025 have been detected in vitro and in nonresponder patients. By sequence analysis of their capsid protein (CA region, two amino acid polymorphisms that correlated with DEB025 resistance were identified: H87Q and I91N, both located in the CypA-binding loop of the CA protein of HIV-1. The H87Q change was by far more abundant than I91N. Additional polymorphisms in the CypA-binding loop (positions 86, 91 and 96, as well as in the N-terminal loop of CA were detected in resistant isolates and are assumed to contribute to the degree of resistance. These amino acid changes may modulate the conformation of the CypA-binding loop of CA in such a way that binding and/or isomerase function of CypA are no longer necessary for virus replication. The resistant HIV-1 isolates thus are CypA-independent.

  5. Molecular mechanisms of HIV-1 associated neurodegeneration

    Indian Academy of Sciences (India)

    Since identification of the human immunodeficiency virus-1 (HIV-1), numerous studies suggest a link between neurological impairments, in particular dementia, with acquired immunodeficiency syndrome (AIDS) with alarming occurrence worldwide. Approximately, 60% of HIV-infected people show some form of neurological ...

  6. Epidemiology of HIV-1 and emerging problems

    NARCIS (Netherlands)

    Lukashov, V. V.; de Ronde, A.; de Jong, J. J.; Goudsmit, J.

    2000-01-01

    Broad use of antiretroviral drugs is becoming a factor that is important to consider for understanding the HIV-1 epidemiology. Since 1993, we observe that a proportion of new infections within major risk groups in Amsterdam is caused by azidothymidine (AZT)-resistant viruses. After the introduction

  7. Six Highly Conserved Targets of RNAi Revealed in HIV-1-Infected Patients from Russia Are Also Present in Many HIV-1 Strains Worldwide.

    Science.gov (United States)

    Kretova, Olga V; Fedoseeva, Daria M; Gorbacheva, Maria A; Gashnikova, Natalya M; Gashnikova, Maria P; Melnikova, Nataliya V; Chechetkin, Vladimir R; Kravatsky, Yuri V; Tchurikov, Nickolai A

    2017-09-15

    RNAi has been suggested for use in gene therapy of HIV/AIDS, but the main problem is that HIV-1 is highly variable and could escape attack from the small interfering RNAs (siRNAs) due to even single nucleotide substitutions in the potential targets. To exhaustively check the variability in selected RNA targets of HIV-1, we used ultra-deep sequencing of six regions of HIV-1 from the plasma of two independent cohorts of patients from Russia. Six RNAi targets were found that are invariable in 82%-97% of viruses in both cohorts and are located inside the domains specifying reverse transcriptase (RT), integrase, vpu, gp120, and p17. The analysis of mutation frequencies and their characteristics inside the targets suggests a likely role for APOBEC3G (apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like 3G, A3G) in G-to-A mutations and a predominant effect of RT biases in the detected variability of the virus. The lowest frequency of mutations was detected in the central part of all six targets. We also discovered that the identical RNAi targets are present in many HIV-1 strains from many countries and from all continents. The data are important for both the understanding of the patterns of HIV-1 mutability and properties of RT and for the development of gene therapy approaches using RNAi for the treatment of HIV/AIDS. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  8. Phase 1 safety and immunogenicity evaluation of ADMVA, a multigenic, modified vaccinia Ankara-HIV-1 B'/C candidate vaccine.

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    Sandhya Vasan

    Full Text Available BACKGROUND: We conducted a Phase I dose-escalation trial of ADMVA, a Clade-B'/C-based HIV-1 candidate vaccine expressing env, gag, pol, nef, and tat in a modified vaccinia Ankara viral vector. Sequences were derived from a prevalent circulating HIV-1 recombinant form in Yunnan, China, an area of high HIV incidence. The objective was to evaluate the safety and immunogenicity of ADMVA in human volunteers. METHODOLOGY/PRINCIPAL FINDINGS: ADMVA or placebo was administered intramuscularly at months 0, 1 and 6 to 50 healthy adult volunteers not at high risk for HIV-1. In each dosage group [1x10(7 (low, 5x10(7 (mid, or 2.5x10(8 pfu (high] volunteers were randomized in a 3:1 ratio to receive ADMVA or placebo in a double-blinded design. Subjects were followed for local and systemic reactogenicity, adverse events including cardiac adverse events, and clinical laboratory parameters. Study follow up was 18 months. Humoral immunogenicity was evaluated by anti-gp120 binding ELISA, immunoflourescent staining, and HIV-1 neutralization. Cellular immunogenicity was assessed by a validated IFNgamma ELISpot assay and intracellular cytokine staining. Anti-vaccinia binding titers were measured by ELISA. ADMVA was generally well-tolerated, with no vaccine-related serious adverse events or cardiac adverse events. Local or systemic reactogenicity events were reported by 77% and 78% of volunteers, respectively. The majority of events were of mild intensity. The IFNgamma ELISpot response rate to any HIV antigen was 0/12 (0% in the placebo group, 3/12 (25% in the low dosage group, 6/12 (50% in the mid dosage group, and 8/13 (62% in the high dosage group. Responses were often multigenic and occasionally persisted up to one year post vaccination. Antibodies to gp120 were detected in 0/12 (0%, 8/13 (62%, 6/12 (50% and 10/13 (77% in the placebo, low, mid, and high dosage groups, respectively. Antibodies persisted up to 12 months after vaccination, with a trend toward agreement

  9. Regulatory T cells expanded from HIV-1-infected individuals maintain phenotype, TCR repertoire and suppressive capacity.

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    Mathieu Angin

    Full Text Available While modulation of regulatory T cell (Treg function and adoptive Treg transfer are being explored as therapeutic modalities in the context of autoimmune diseases, transplantation and cancer, their role in HIV-1 pathogenesis remains less well defined. Controversy persists regarding their beneficial or detrimental effects in HIV-1 disease, which warrants further detailed exploration. Our objectives were to investigate if functional CD4(+ Tregs can be isolated and expanded from HIV-1-infected individuals for experimental or potential future therapeutic use and to determine phenotype and suppressive capacity of expanded Tregs from HIV-1 positive blood and tissue. Tregs and conventional T cell controls were isolated from blood and gut-associated lymphoid tissue of individuals with HIV-1 infection and healthy donors using flow-based cell-sorting. The phenotype of expanded Tregs was assessed by flow-cytometry and quantitative PCR. T-cell receptor ß-chain (TCR-β repertoire diversity was investigated by deep sequencing. Flow-based T-cell proliferation and chromium release cytotoxicity assays were used to determine Treg suppressive function. Tregs from HIV-1 positive individuals, including infants, were successfully expanded from PBMC and GALT. Expanded Tregs expressed high levels of FOXP3, CTLA4, CD39 and HELIOS and exhibited a highly demethylated TSDR (Treg-specific demethylated region, characteristic of Treg lineage. The TCRß repertoire was maintained following Treg expansion and expanded Tregs remained highly suppressive in vitro. Our data demonstrate that Tregs can be expanded from blood and tissue compartments of HIV-1+ donors with preservation of Treg phenotype, function and TCR repertoire. These results are highly relevant for the investigation of potential future therapeutic use, as currently investigated for other disease states and hold great promise for detailed studies on the role of Tregs in HIV-1 infection.

  10. Near Full-Length Identification of a Novel HIV-1 CRF01_AE/B/C Recombinant in Northern Myanmar.

    Science.gov (United States)

    Zhou, Yan-Heng; Chen, Xin; Liang, Yue-Bo; Pang, Wei; Qin, Wei-Hong; Zhang, Chiyu; Zheng, Yong-Tang

    2015-08-01

    The Myanmar-China border appears to be the "hot spot" region for the occurrence of HIV-1 recombination. The majority of the previous analyses of HIV-1 recombination were based on partial genomic sequences, which obviously cannot reflect the reality of the genetic diversity of HIV-1 in this area well. Here, we present a near full-length characterization of a novel HIV-1 CRF01_AE/B/C recombinant isolated from a long-distance truck driver in Northern Myanmar. It is the first description of a near full-length genomic sequence in Myanmar since 2003, and might be one of the most complicated HIV-1 chimeras ever detected in Myanmar, containing four CRF01_AE, six B segments, and five C segments separated by 14 breakpoints throughout its genome. The discovery and characterization of this new CRF01_AE/B/C recombinant indicate that intersubtype recombination is ongoing in Myanmar, continuously generating new forms of HIV-1. More work based on near full-length sequence analyses is urgently needed to better understand the genetic diversity of HIV-1 in these regions.

  11. Molecular and epidemiological characterization of HIV-1 subtypes among Libyan patients.

    Science.gov (United States)

    Daw, Mohamed A; El-Bouzedi, Abdallah; Ahmed, Mohamed O; Dau, Aghnyia A

    2017-04-28

    The epidemiological and clinical aspects of human immunodeficiency virus subtypes are of great interest worldwide. These subtypes are rarely studied in North African countries. Libya is a large country with the longest coast on the Mediterranean Sea, facing the Southern European countries. Studies on the characterization of HIV-1 subtypes are limited in Libya. This study aimed to determine the magnitude of the HIV problem among the Libyan population and to better understand the genetic diversity and the epidemiologic dynamics of HIV 1, as well as to correlate that with the risk factors involved. A total of 159 HIV-1 strains were collected from 814 HIV positive patients from the four Libyan regions during a 16-year period (1995-2010). To determine the HIV-1 subtypes, genetic analysis and molecular sequencing were carried out using provirus polygene. Epidemiologic and demographic information was obtained from each participant and correlated with HIV-1 subtypes using logistic regression. The overall prevalence of HIV among Libyans ranged from 5 to 10 per 100,000 during the study period. It was higher among intravenous drug users (IVDUs) (53.9%), blood recipients (25.9%) and heterosexuals (17.6%) than by vertical transmission (2.6%). Prevalence was higher among males aged 20-40 years (M:F 1:6, P > 0.001). Among the 159 strains of HIV-1 available for typing, 117 strains (73.6%) were subtype B, 29 (18.2%) were CRF02_AG, and 13 (8.2%) were subtype A. HIV-1 subtype B was the most prevalent all over the country, and it was more prevalent in the Northern region, particularly among IVDUs (P HIV-1 infection is emerging in Libya with a shifting prevalence of subtypes associated with the changing epidemiology of HIV-1 among risk groups. A genetic analysis of HIV-1 strains demonstrated low subtype heterogeneity with the evolution of subtype B, and CRF_20 AG, as well as HIV-1 subtype A. Our study highlights the importance of expanded surveillance programs to control HIV

  12. Host-specific adaptation of HIV-1 subtype B in the Japanese population.

    Science.gov (United States)

    Chikata, Takayuki; Carlson, Jonathan M; Tamura, Yoshiko; Borghan, Mohamed Ali; Naruto, Takuya; Hashimoto, Masao; Murakoshi, Hayato; Le, Anh Q; Mallal, Simon; John, Mina; Gatanaga, Hiroyuki; Oka, Shinichi; Brumme, Zabrina L; Takiguchi, Masafumi

    2014-05-01

    The extent to which HIV-1 clade B strains exhibit population-specific adaptations to host HLA alleles remains incompletely known, in part due to incomplete characterization of HLA-associated HIV-1 polymorphisms (HLA-APs) in different global populations. Moreover, it remains unknown to what extent the same HLA alleles may drive significantly different escape pathways across populations. As the Japanese population exhibits distinctive HLA class I allele distributions, comparative analysis of HLA-APs between HIV-1 clade B-infected Japanese and non-Asian cohorts could shed light on these questions. However, HLA-APs remain incompletely mapped in Japan. In a cohort of 430 treatment-naive Japanese with chronic HIV-1 clade B infection, we identified 284 HLA-APs in Gag, Pol, and Nef using phylogenetically corrected methods. The number of HLA-associated substitutions in Pol, notably those restricted by HLA-B*52:01, was weakly inversely correlated with the plasma viral load (pVL), suggesting that the transmission and persistence of B*52:01-driven Pol mutations could modulate the pVL. Differential selection of HLA-APs between HLA subtype members, including those differing only with respect to substitutions outside the peptide-binding groove, was observed, meriting further investigation as to their mechanisms of selection. Notably, two-thirds of HLA-APs identified in Japan had not been reported in previous studies of predominantly Caucasian cohorts and were attributable to HLA alleles unique to, or enriched in, Japan. We also identified 71 cases where the same HLA allele drove significantly different escape pathways in Japan versus predominantly Caucasian cohorts. Our results underscore the distinct global evolution of HIV-1 clade B as a result of host population-specific cellular immune pressures. Cytotoxic T lymphocyte (CTL) escape mutations in HIV-1 are broadly predictable based on the HLA class I alleles expressed by the host. Because HLA allele distributions differ among

  13. Safety and immunogenicity of HIV-1 Tat toxoid in immunocompromised HIV-1-infected patients.

    Science.gov (United States)

    Gringeri, A; Santagostino, E; Muça-Perja, M; Mannucci, P M; Zagury, J F; Bizzini, B; Lachgar, A; Carcagno, M; Rappaport, J; Criscuolo, M; Blattner, W; Burny, A; Gallo, R C; Zagury, D

    1998-01-01

    To antagonize the deleterious effects of the HIV-1 toxin extracellular Tat on uninfected immune cells, we developed a new strategy of anti-HIV-1 vaccine using an inactivated but immunogenic Tat (Tat toxoid). Tat toxoid has been assayed for safety and immunogenicity in seropositive patients. The phase I vaccine clinical trial testing Tat toxoid preparation in Seppic Isa 51 oil adjuvant was performed on 14 HIV-1-infected asymptomatic although biologically immunocompromised individuals (500-200 CD4+ cells/mm3). Following as many as 8 injections, no clinical defects were observed. All patients exhibited an antibody (Ab) response to Tat, and some had cell-mediated immunity (CMI) as evaluated by skin test in vivo and T-cell proliferation in vitro. These results provide initial evidence of safety and potency of Tat toxoid vaccination in HIV-1-infected individuals.

  14. Lack of integrase inhibitors associated resistance mutations among HIV-1C isolates.

    Science.gov (United States)

    Mulu, Andargachew; Maier, Melanie; Liebert, Uwe Gerd

    2015-12-01

    Although biochemical analysis of HIV-1 integrase enzyme suggested the use of integrase inhibitors (INIs) against HIV-1C, different viral subtypes may favor different mutational pathways potentially leading to varying levels of drug resistance. Thus, the aim of this study was to search for the occurrence and natural evolution of integrase polymorphisms and/or resistance mutations in HIV-1C Ethiopian clinical isolates prior to the introduction of INIs. Plasma samples from chronically infected drug naïve patients (N = 45), of whom the PR and RT sequence was determined previously, were used to generate population based sequences of HIV-1 integrase. HIV-1 subtype was determined using the REGA HIV-1 subtyping tool. Resistance mutations were interpreted according to the Stanford HIV drug resistance database ( http://hivdb.stanford.edu ) and the updated International Antiviral Society (IAS)-USA mutation lists. Moreover, rates of polymorphisms in the current isolates were compared with South African and global HIV-1C isolates. All subjects were infected with HIV-1C concordant to the protease (PR) and reverse transcriptase (RT) regions. Neither major resistance-associated IN mutations (T66I/A/K, E92Q/G, T97A, Y143HCR, S147G, Q148H/R/K, and N155H) nor silent mutations known to change the genetic barrier were observed. Moreover, the DDE-catalytic motif (D64G/D116G/E152 K) and signature HHCC zinc-binding motifs at codon 12, 16, 40 and 43 were found to be highly conserved. However, compared to other South African subtype C isolates, the rate of polymorphism was variable at various positions. Although the sample size is small, the findings suggest that this drug class could be effective in Ethiopia and other southern African countries where HIV-1C is predominantly circulating. The data will contribute to define the importance of integrase polymorphism and to improve resistance interpretation algorithms in HIV-1C isolates.

  15. Sequence similarity between the erythrocyte binding domain of the Plasmodium vivax Duffy binding protein and the V3 loop of HIV-1 strain MN reveals a functional heparin binding motif involved in binding to the Duffy antigen receptor for chemokines

    OpenAIRE

    Bolton, Michael J; Garry, Robert F

    2011-01-01

    Abstract Background The HIV surface glycoprotein gp120 (SU, gp120) and the Plasmodium vivax Duffy binding protein (PvDBP) bind to chemokine receptors during infection and have a site of amino acid sequence similarity in their binding domains that often includes a heparin binding motif (HBM). Infection by either pathogen has been found to be inhibited by polyanions. Results Specific polyanions that inhibit HIV infection and bind to the V3 loop of X4 strains also inhibited DBP-mediated infectio...

  16. Molecular cloning and anti-HIV-1 activities of APOBEC3s from northern pig-tailed macaques (Macaca leonina

    Directory of Open Access Journals (Sweden)

    Xiao-Liang ZHANG

    2016-07-01

    Full Text Available Northern pig-tailed macaques (NPMs, Macaca leonina are susceptible to HIV-1 infection largely due to the loss of HIV-1-restricting factor TRIM5α. However, great impediments still exist in the persistent replication of HIV-1 in vivo, suggesting some viral restriction factors are reserved in this host. The APOBEC3 proteins have demonstrated a capacity to restrict HIV-1 replication, but their inhibitory effects in NPMs remain elusive. In this study, we cloned the NPM A3A-A3H genes, and determined by BLAST searching that their coding sequences (CDSs showed 99% identity to the corresponding counterparts from rhesus and southern pig-tailed macaques. We further analyzed the anti-HIV-1 activities of the A3A-A3H genes, and found that A3G and A3F had the greatest anti-HIV-1 activity compared with that of other members. The results of this study indicate that A3G and A3F might play critical roles in limiting HIV-1 replication in NPMs in vivo. Furthermore, this research provides valuable information for the optimization of monkey models of HIV-1 infection.

  17. Molecular detection of HIV-1 subtype B, CRF01_AE, CRF33_01B, and newly emerging recombinant lineages in Malaysia.

    Science.gov (United States)

    Chook, Jack Bee; Ong, Lai Yee; Takebe, Yutaka; Chan, Kok Gan; Choo, Martin; Kamarulzaman, Adeeba; Tee, Kok Keng

    2015-03-01

    A molecular genotyping assay for human immunodeficiency virus type 1 (HIV-1) circulating in Southeast Asia is difficult to design because of the high level of genetic diversity. We developed a multiplex real-time polymerase chain reaction (PCR) assay to detect subtype B, CRF01_AE, CRF33_01B, and three newly described circulating recombinant forms, (CRFs) (CRF53_01B, CRF54_01B, and CRF58_01B). A total of 785 reference genomes were used for subtype-specific primers and TaqMan probes design targeting the gag, pol, and env genes. The performance of this assay was compared and evaluated with direct sequencing and phylogenetic analysis. A total of 180 HIV-infected subjects from Kuala Lumpur, Malaysia were screened and 171 samples were successfully genotyped, in agreement with the phylogenetic data. The HIV-1 genotype distribution was as follows: subtype B (16.7%); CRF01_AE (52.8%); CRF33_01B (24.4%); CRF53_01B (1.1%); CRF54_01B (0.6%); and CRF01_AE/B unique recombinant forms (4.4%). The overall accuracy of the genotyping assay was over 95.0%, in which the sensitivities for subtype B, CRF01_AE, and CRF33_01B detection were 100%, 100%, and 97.7%, respectively. The specificity of genotyping was 100%, inter-subtype specificities were > 95% and the limit of detection of 10(3) copies/mL for plasma. The newly developed real-time PCR assay offers a rapid and cost-effective alternative for large-scale molecular epidemiological surveillance for HIV-1. © The American Society of Tropical Medicine and Hygiene.

  18. Picomolar dichotomous activity of gnidimacrin against HIV-1.

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    Li Huang

    Full Text Available Highly active antiretroviral therapy (HAART has offered a promising approach for controlling HIV-1 replication in infected individuals. However, with HARRT, HIV-1 is suppressed rather than eradicated due to persistence of HIV-1 in latent viral reservoirs. Thus, purging the virus from latent reservoirs is an important strategy toward eradicating HIV-1 infection. In this study, we discovered that the daphnane diterpene gnidimacrin, which was previously reported to have potent anti-cancer cell activity, activated HIV-1 replication and killed persistently-infected cells at picomolar concentrations. In addition to its potential to purge HIV-1 from latently infected cells, gnidimacrin potently inhibited a panel of HIV-1 R5 virus infection of peripheral blood mononuclear cells (PBMCs at an average concentration lower than 10 pM. In contrast, gnidimacrin only partially inhibited HIV-1 ×4 virus infection of PBMCs. The strong anti-HIV-1 R5 virus activity of gnidimacrin was correlated with its effect on down-regulation of the HIV-1 coreceptor CCR5. The anti-R5 virus activity of gnidimacrin was completely abrogated by a selective protein kinase C beta inhibitor enzastaurin, which suggests that protein kinase C beta plays a key role in the potent anti-HIV-1 activity of gnidimacrin in PBMCs. In summary, these results suggest that gnidimacrin could activate latent HIV-1, specifically kill HIV-1 persistently infected cells, and inhibit R5 viruses at picomolar concentrations.

  19. Variability of HIV-1 genomes among children and adolescents from Sao Paulo, Brazil.

    Directory of Open Access Journals (Sweden)

    Sabri Saeed Sanabani

    Full Text Available BACKGROUND: Genetic variability is a major feature of the human immunodeficiency virus type 1 (HIV-1 and considered the key factor to frustrating efforts to halt the virus epidemic. In this study, we aimed to investigate the genetic variability of HIV-1 strains among children and adolescents born from 1992 to 2009 in the state of Sao Paulo, Brazil. METHODOLOGY: Plasma and peripheral blood mononuclear cells (PBMC were collected from 51 HIV-1-positive children and adolescents on ART followed between September 1992 and July 2009. After extraction, the genetic materials were used in a polymerase chain reaction (PCR to amplify the viral near full length genomes (NFLGs from 5 overlapped fragments. NFLGs and partial amplicons were directly sequenced and data were phylogenetically inferred. RESULTS: Of the 51 samples studied, the NFLGs and partial fragments of HIV-1 from 42 PBMCs and 25 plasma were successfully subtyped. Results based on proviral DNA revealed that 22 (52.4% patients were infected with subtype B, 16 (38.1% were infected with BF1 mosaic variants and 4 (9.5% were infected with sub-subtype F1. All the BF1 recombinants were unique and distinct from any previously identified unique or circulating recombinant forms in South America. Evidence of dual infections was detected in 3 patients coinfected with the same or distinct HIV-1 subtypes. Ten of the 31 (32.2% and 12 of the 21 (57.1% subjects with recovered proviral and plasma, respectively, protease sequences were infected with major mutants resistant to protease inhibitors. The V3 sequences of 14 patients with available sequences from PBMC/or plasma were predicted to be R5-tropic virus except for two patients who harbored an X4 strain. CONCLUSIONS: The high proportion of HIV-1 BF1 recombinant, coinfection rate and vertical transmission in Brazil merits urgent attention and effective measures to reduce the transmission of HIV among spouses and sex partners.

  20. Minority drug-resistant HIV-1 variants in treatment naïve East-African and Caucasian patients detected by allele-specific real-time PCR.

    Directory of Open Access Journals (Sweden)

    Halime Ekici

    Full Text Available To assess the presence of two major non-nucleoside reverse transcriptase inhibitors (NNRTI drug resistance mutations (DRMs, Y181C and K103N, in minor viral quasispecies of treatment naïve HIV-1 infected East-African and Swedish patients by allele-specific polymerase chain reaction (AS-PCR.Treatment naïve adults (n=191 with three epidemiological backgrounds were included: 92 Ethiopians living in Ethiopia; 55 East-Africans who had migrated to Sweden; and 44 Caucasians living in Sweden. The pol gene was analysed by standard population sequencing and by AS-PCR for the detection of Y181C and K103N.The Y181C was detected in the minority quasispecies of six Ethiopians (6.5%, in two Caucasians (4.5%, and in one East-African (1.8%. The K103N was detected in one East- African (1.8%, by both methods. The proportion of mutants ranged from 0.25% to 17.5%. Additional DRMs were found in all three treatment naïve patient groups by population sequencing.Major NNRTI mutations can be found by AS-PCR in minor quasispecies of treatment naïve HIV-1 infected Ethiopians living in Ethiopia, in East-African and Caucasian patients living in Sweden in whom population sequencing reveal wild-type virus only. Surveys with standard sequencing are likely to underestimate transmitted drug resistance and the presence of resistant minor quasispecies in treatment naïve patients should be topic for future large scale studies.

  1. The combination of phylogenetic analysis with epidemiological and serological data to track HIV-1 transmission in a sexual transmission case.

    Directory of Open Access Journals (Sweden)

    Min Chen

    Full Text Available To investigate the linkage of HIV transmission from a man to a woman through unprotected sexual contact without disclosing his HIV-positive status.Combined with epidemiological information and serological tests, phylogenetic analysis was used to test the a priori hypothesis of HIV transmission from the man to the woman. Control subjects, infected with HIV through heterosexual intercourse, from the same location were also sampled. Phylogenetic analyses were performed using the consensus gag, pol and env sequences obtained from blood samples of the man, the woman and the local control subjects. The env quasispecies of the man, the woman, and two controls were also obtained using single genome amplification and sequencing (SGA/S to explore the paraphyletic relationship by phylogenetic analysis.Epidemiological information and serological tests indicated that the man was infected with HIV-1 earlier than the woman. Phylogenetic analyses of the consensus sequences showed a monophyletic cluster for the man and woman in all three genomic regions. Furthermore, gag sequences of the man and woman shared a unique recombination pattern from subtype B and C, which was different from those of CRF07_BC or CRF08_BC observed in the local samples. These indicated that the viral sequences from the two subjects display a high level of similarity. Further, viral quasispecies from the man exhibited a paraphyletic relationship with those from the woman in the Bayesian and maximum-likelihood (ML phylogenetic trees of the env region, which supported the transmission direction from the man to the woman.In the context of epidemiological and serological evidence, the results of phylogenetic analyses support the transmission from the man to the woman.

  2. Sequence similarity between the erythrocyte binding domain of the Plasmodium vivax Duffy binding protein and the V3 loop of HIV-1 strain MN reveals a functional heparin binding motif involved in binding to the Duffy antigen receptor for chemokines

    Directory of Open Access Journals (Sweden)

    Bolton Michael J

    2011-11-01

    Full Text Available Abstract Background The HIV surface glycoprotein gp120 (SU, gp120 and the Plasmodium vivax Duffy binding protein (PvDBP bind to chemokine receptors during infection and have a site of amino acid sequence similarity in their binding domains that often includes a heparin binding motif (HBM. Infection by either pathogen has been found to be inhibited by polyanions. Results Specific polyanions that inhibit HIV infection and bind to the V3 loop of X4 strains also inhibited DBP-mediated infection of erythrocytes and DBP binding to the Duffy Antigen Receptor for Chemokines (DARC. A peptide including the HBM of PvDBP had similar affinity for heparin as RANTES and V3 loop peptides, and could be specifically inhibited from heparin binding by the same polyanions that inhibit DBP binding to DARC. However, some V3 peptides can competitively inhibit RANTES binding to heparin, but not the PvDBP HBM peptide. Three other members of the DBP family have an HBM sequence that is necessary for erythrocyte binding, however only the protein which binds to DARC, the P. knowlesi alpha protein, is inhibited by heparin from binding to erythrocytes. Heparitinase digestion does not affect the binding of DBP to erythrocytes. Conclusion The HBMs of DBPs that bind to DARC have similar heparin binding affinities as some V3 loop peptides and chemokines, are responsible for specific sulfated polysaccharide inhibition of parasite binding and invasion of red blood cells, and are more likely to bind to negative charges on the receptor than cell surface glycosaminoglycans.

  3. Membrane topology analysis of HIV-1 envelope glycoprotein gp41

    Directory of Open Access Journals (Sweden)

    Xiao Dan

    2010-11-01

    Full Text Available Abstract Background The gp41 subunit of the HIV-1 envelope glycoprotein (Env has been widely regarded as a type I transmembrane protein with a single membrane-spanning domain (MSD. An alternative topology model suggested multiple MSDs. The major discrepancy between the two models is that the cytoplasmic Kennedy sequence in the single MSD model is assigned as the extracellular loop accessible to neutralizing antibodies in the other model. We examined the membrane topology of the gp41 subunit in both prokaryotic and mammalian systems. We attached topological markers to the C-termini of serially truncated gp41. In the prokaryotic system, we utilized a green fluorescent protein (GFP that is only active in the cytoplasm. The tag protein (HaloTag and a membrane-impermeable ligand specific to HaloTag was used in the mammalian system. Results In the absence of membrane fusion, both the prokaryotic and mammalian systems (293FT cells supported the single MSD model. In the presence of membrane fusion in mammalian cells (293CD4 cells, the data obtained seem to support the multiple MSD model. However, the region predicted to be a potential MSD is the highly hydrophilic Kennedy sequence and is least likely to become a MSD based on several algorithms. Further analysis revealed the induction of membrane permeability during membrane fusion, allowing the membrane-impermeable ligand and antibodies to cross the membrane. Therefore, we cannot completely rule out the possible artifacts. Addition of membrane fusion inhibitors or alterations of the MSD sequence decreased the induction of membrane permeability. Conclusions It is likely that a single MSD model for HIV-1 gp41 holds true even in the presence of membrane fusion. The degree of the augmentation of membrane permeability we observed was dependent on the membrane fusion and sequence of the MSD.

  4. A study of vaccine-induced immune pressure on breakthrough infections in the Phambili phase 2b HIV-1 vaccine efficacy trial

    Science.gov (United States)

    Rolland, M.; Magaret, C.A.; Rademeyer, C.; Fiore-Gartland, A.; Edlefsen, P.T.; DeCamp, A.; Ahmed, H.; Ngandu, N.; Larsen, B.B.; Frahm, N.; Marais, J.; Thebus, R.; Geraghty, D.; Hural, J.; Corey, L.; Kublin, J.; Gray, G.; McElrath, M.J.; Mullins, J.I.; Gilbert, P.B.; Williamson, C.

    2016-01-01

    Introduction The Merck Adenovirus-5 Gag/Pol/Nef HIV-1 subtype-B vaccine evaluated in predominately subtype B epidemic regions (Step Study), while not preventing infection, exerted vaccine-induced immune pressure on HIV-1 breakthrough infections. Here we investigated if the same vaccine exerted immune pressure when tested in the Phambili Phase 2b study in a subtype C epidemic. Materials and methods A sieve analysis, which compares breakthrough viruses from placebo and vaccine arms, was performed on 277 near full-length genomes generated from 23 vaccine and 20 placebo recipients. Vaccine coverage was estimated by computing the percentage of 9-mers that were exact matches to the vaccine insert. Results There was significantly greater protein distances from the vaccine immunogen sequence in Gag (p = 0.045) and Nef (p = 0.021) in viruses infecting vaccine recipients compared to placebo recipients. Twenty-seven putative sites of vaccine-induced pressure were identified (p sieve effect in Step was driven by HLA A*02:01; an allele which was found in low frequency in Phambili participants compared to Step participants. Furthermore, the coverage of the vaccine against subtype C Phambili viruses was 31%, 46% and 14% for Gag, Pol and Nef, respectively, compared to subtype B Step virus coverage of 56%, 61% and 26%, respectively. Discussion This study presents evidence of sieve effects in Gag and Nef; however could not confirm effects on specific amino acid sites. We propose that this weaker signal of vaccine immune pressure detected in the Phambili study compared to the Step study may have been influenced by differences in host genetics (HLA allele frequency) and reduced impact of vaccine-induced immune responses due to mismatch between the viral subtype in the vaccine and infecting subtypes. PMID:27756485

  5. Identification of human microRNA-like sequences embedded within the protein-encoding genes of the human immunodeficiency virus.

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    Bryan Holland

    Full Text Available BACKGROUND: MicroRNAs (miRNAs are highly conserved, short (18-22 nts, non-coding RNA molecules that regulate gene expression by binding to the 3' untranslated regions (3'UTRs of mRNAs. While numerous cellular microRNAs have been associated with the progression of various diseases including cancer, miRNAs associated with retroviruses have not been well characterized. Herein we report identification of microRNA-like sequences in coding regions of several HIV-1 genomes. RESULTS: Based on our earlier proteomics and bioinformatics studies, we have identified 8 cellular miRNAs that are predicted to bind to the mRNAs of multiple proteins that are dysregulated during HIV-infection of CD4+ T-cells in vitro. In silico analysis of the full length and mature sequences of these 8 miRNAs and comparisons with all the genomic and subgenomic sequences of HIV-1 strains in global databases revealed that the first 18/18 sequences of the mature hsa-miR-195 sequence (including the short seed sequence, matched perfectly (100%, or with one nucleotide mismatch, within the envelope (env genes of five HIV-1 genomes from Africa. In addition, we have identified 4 other miRNA-like sequences (hsa-miR-30d, hsa-miR-30e, hsa-miR-374a and hsa-miR-424 within the env and the gag-pol encoding regions of several HIV-1 strains, albeit with reduced homology. Mapping of the miRNA-homologues of env within HIV-1 genomes localized these sequence to the functionally significant variable regions of the env glycoprotein gp120 designated V1, V2, V4 and V5. CONCLUSIONS: We conclude that microRNA-like sequences are embedded within the protein-encoding regions of several HIV-1 genomes. Given that the V1 to V5 regions of HIV-1 envelopes contain specific, well-characterized domains that are critical for immune responses, virus neutralization and disease progression, we propose that the newly discovered miRNA-like sequences within the HIV-1 genomes may have evolved to self-regulate survival of the

  6. Reliable reconstruction of HIV-1 whole genome haplotypes reveals clonal interference and genetic hitchhiking among immune escape variants

    Science.gov (United States)

    2014-01-01

    Background Following transmission, HIV-1 evolves into a diverse population, and next generation sequencing enables us to detect variants occurring at low frequencies. Studying viral evolution at the level of whole genomes was hitherto not possible because next generation sequencing delivers relatively short reads. Results We here provide a proof of principle that whole HIV-1 genomes can be reliably reconstructed from short reads, and use this to study the selection of immune escape mutations at the level of whole genome haplotypes. Using realistically simulated HIV-1 populations, we demonstrate that reconstruction of complete genome haplotypes is feasible with high fidelity. We do not reconstruct all genetically distinct genomes, but each reconstructed haplotype represents one or more of the quasispecies in the HIV-1 population. We then reconstruct 30 whole genome haplotypes from published short sequence reads sampled longitudinally from a single HIV-1 infected patient. We confirm the reliability of the reconstruction by validating our predicted haplotype genes with single genome amplification sequences, and by comparing haplotype frequencies with observed epitope escape frequencies. Conclusions Phylogenetic analysis shows that the HIV-1 population undergoes selection driven evolution, with successive replacement of the viral population by novel dominant strains. We demonstrate that immune escape mutants evolve in a dependent manner with various mutations hitchhiking along with others. As a consequence of this clonal interference, selection coefficients have to be estimated for complete haplotypes and not for individual immune escapes. PMID:24996694

  7. Morphogenesis of the infectious HIV-1 virion

    Directory of Open Access Journals (Sweden)

    Jun-Ichi eSakuragi

    2011-12-01

    Full Text Available The virion of HIV-1 is spherical and viral glycoprotein spikes (gp120, gp41 protrude from its envelope. The characteristic cone-shaped core exists within the virion, caging the ribonucleoprotein (RNP complex, which is comprised of viral RNA, nucleocapsid (NC and viral enzymes. The HIV-1 virion is budded and released from the infected cell as an immature donut-shaped particle. During or immediately after release, viral protease (PR is activated and subsequently processes the viral structural protein Gag. Through this maturation process, virions acquire infectivity, but its mechanism and transition of morphology largely remain unclear. Recent technological advances in experimental devices and techniques have made it possible to closely dissect the viral production site on the cell, the exterior – or even the interior – of an individual virion, and many new aspects on virion morphology and maturation. In this manuscript, I review the morphogenesis of HIV-1 virions. I focus on several studies, including some of our recent findings, which examined virion formation and/or maturation processes. The story of novel compound, which inhibits virion maturation, and the importance of maturation research are also discussed.

  8. HIV-1 Eradication Strategies: Design and Assessment

    Science.gov (United States)

    Siliciano, Robert F.

    2014-01-01

    Purpose of review Recent developments have generated renewed interest in the possibility of curing HIV-1 infection. This review describes some of the practical challenges that will need to be overcome if curative strategies are to be successful. Recent findings The latent reservoir for HIV-1 in resting memory CD4+ T cells is the major barrier to curing the infection. The most widely discussed approach to curing the infection involves finding agents that reverse latency in resting CD4+ T cells, with the assumption that the cells will then die from viral cytopathic effects or be lysed by host cytolytic T lymphocytes (CTL). A major challenge is the development of in vitro models that can be used to explore mechanisms and identify latency reversing agents (LRA). Although several models have been developed, including primary cell models, none of them may fully capture the quiescent state of the cells that harbor latent HIV-1 in vivo. An additional problem is that LRA that do not cause T cell activation may not lead to the death of infected cells. Finally, measuring the effects of LRAs in vivo is complicated by the lack of correlation between different assays for the latent reservoir. Summary Progress on these practical issues is essential to finding a cure. PMID:23698561

  9. A newly emerging HIV-1 recombinant lineage (CRF58_01B) disseminating among people who inject drugs in Malaysia.

    Science.gov (United States)

    Chow, Wei Zhen; Takebe, Yutaka; Syafina, Nur Ezreen; Prakasa, Malarvelli Soorya; Chan, Kok Gan; Al-Darraji, Haider Abdulrazzaq Abed; Koh, Clayton; Kamarulzaman, Adeeba; Tee, Kok Keng

    2014-01-01

    The HIV epidemic is primarily characterised by the circulation of HIV-1 group M (main) comprising of 11 subtypes and sub-subtypes (A1, A2, B-D, F1, F2, G, H, J, and K) and to date 55 circulating recombinant forms (CRFs). In Southeast Asia, active inter-subtype recombination involving three main circulating genotypes--subtype B (including subtype B', the Thai variant of subtype B), CRF01_AE, and CRF33_01B--have contributed to the emergence of novel unique recombinant forms. In the present study, we conducted the molecular epidemiological surveillance of HIV-1 gag-RT genes among 258 people who inject drugs (PWIDs) in Kuala Lumpur, Malaysia, between 2009 and 2011 whereby a novel CRF candidate was recently identified. The near full-length genome sequences obtained from six epidemiologically unlinked individuals showed identical mosaic structures consisting of subtype B' and CRF01_AE, with six unique recombination breakpoints in the gag-RT, pol, and env regions. Among the high-risk population of PWIDs in Malaysia, which was predominantly infected by CRF33_01B (>70%), CRF58_01B circulated at a low but significant prevalence (2.3%, 6/258). Interestingly, the CRF58_01B shared two unique recombination breakpoints with other established CRFs in the region: CRF33_01B, CRF48_01B, and CRF53_01B in the gag gene, and CRF15_01B (from Thailand) in the env gene. Extended Bayesian Markov chain Monte Carlo sampling analysis showed that CRF58_01B and other recently discovered CRFs were most likely to have originated in Malaysia, and that the recent spread of recombinant lineages in the country had little influence from neighbouring countries. The isolation, genetic characterization, and evolutionary features of CRF58_01B among PWIDs in Malaysia signify the increasingly complex HIV-1 diversity in Southeast Asia that may hold an implication on disease treatment, control, and prevention.

  10. A newly emerging HIV-1 recombinant lineage (CRF58_01B disseminating among people who inject drugs in Malaysia.

    Directory of Open Access Journals (Sweden)

    Wei Zhen Chow

    Full Text Available The HIV epidemic is primarily characterised by the circulation of HIV-1 group M (main comprising of 11 subtypes and sub-subtypes (A1, A2, B-D, F1, F2, G, H, J, and K and to date 55 circulating recombinant forms (CRFs. In Southeast Asia, active inter-subtype recombination involving three main circulating genotypes--subtype B (including subtype B', the Thai variant of subtype B, CRF01_AE, and CRF33_01B--have contributed to the emergence of novel unique recombinant forms. In the present study, we conducted the molecular epidemiological surveillance of HIV-1 gag-RT genes among 258 people who inject drugs (PWIDs in Kuala Lumpur, Malaysia, between 2009 and 2011 whereby a novel CRF candidate was recently identified. The near full-length genome sequences obtained from six epidemiologically unlinked individuals showed identical mosaic structures consisting of subtype B' and CRF01_AE, with six unique recombination breakpoints in the gag-RT, pol, and env regions. Among the high-risk population of PWIDs in Malaysia, which was predominantly infected by CRF33_01B (>70%, CRF58_01B circulated at a low but significant prevalence (2.3%, 6/258. Interestingly, the CRF58_01B shared two unique recombination breakpoints with other established CRFs in the region: CRF33_01B, CRF48_01B, and CRF53_01B in the gag gene, and CRF15_01B (from Thailand in the env gene. Extended Bayesian Markov chain Monte Carlo sampling analysis showed that CRF58_01B and other recently discovered CRFs were most likely to have originated in Malaysia, and that the recent spread of recombinant lineages in the country had little influence from neighbouring countries. The isolation, genetic characterization, and evolutionary features of CRF58_01B among PWIDs in Malaysia signify the increasingly complex HIV-1 diversity in Southeast Asia that may hold an implication on disease treatment, control, and prevention.

  11. Inhibition of HIV-1 in vitro by C-5 propyne phosphorothioate antisense to rev

    DEFF Research Database (Denmark)

    Lund, O S; Nielsen, Jens Ole; Hansen, J E

    1995-01-01

    in a sequence-dependent manner. The antiviral effect was obtained by lipofection or simple addition of 0.2-1 microM modified oligodeoxynucleotide to the culture medium of H9 cells chronically infected with the HIV-1LAI isolate of human immunodeficiency virus type 1. We conclude that C-5 propyne...

  12. CRISPR-Cas9 Can Inhibit HIV-1 Replication but NHEJ Repair Facilitates Virus Escape

    NARCIS (Netherlands)

    Wang, Gang; Zhao, Na; Berkhout, Ben; Das, Atze T.

    2016-01-01

    Several recent studies demonstrated that the clustered regularly interspaced short palindromic repeats (CRISPR)-associated endonuclease Cas9 can be used for guide RNA (gRNA)-directed, sequence-specific cleavage of HIV proviral DNA in infected cells. We here demonstrate profound inhibition of HIV-1

  13. Frequency and site mapping of HIV-1/SIVcpz, HIV- 2/SIVsmm and ...

    African Journals Online (AJOL)

    out to analyze the effects of various restriction enzymes on the HIV genome. A computer simulated model using Web cutter Version 2.0, and cytogenetic analysis. 339 restriction enzymes from Promega database, 10 HIV-1/SIVcpz genes, 10 HIV-2/SIVsmm genes and 10 other SIV genes. Gene sequences were fed into Web ...

  14. Transmitted drug resistance and type of infection in newly diagnosed HIV-1 individuals in Honduras.

    Science.gov (United States)

    Murillo, Wendy; Paz-Bailey, Gabriela; Morales, Sonia; Monterroso, Edgar; Paredes, Mayte; Dobbs, Trudy; Parekh, Bharat S; Albert, Jan; Rivera, Ivette Lorenzana de

    2010-12-01

    Transmitted drug resistance (TDR) reduces the efficacy of antiretroviral treatment and is a public health concern. To gain insight in the epidemiology of TDR in Honduras by evaluating the amount of TDR in a representative sample of newly diagnosed individuals and by determining whether these are recent or established infections. Two hundred treatment-naïve, newly diagnosed HIV-positive individuals representing different population groups (general population, Garifunas ethnic group, female sex workers and men who have sex with men) and different geographic regions were enrolled during April 2004-April 2007. The HIV-1 pol gene was sequenced to identify drug-resistant mutations and TDR was scored as recommended by the WHO. Specimens were classified as recent or established infections using the BED assay. Among 200 samples analyzed from Honduran patients the prevalence of TDR was 7% (95% CI: 3.9-11.5%), 5% for non-nucleoside reverse transcriptase inhibitors (NNRTIs), 3% for nucleoside reverse transcriptase inhibitors (NRTIs) and 0.5% for protease inhibitors (PIs). Testing of these samples with the BED assay revealed that 12% of the specimens were associated with recent infections. TDR was significantly more common in specimens with recent infection (21%) than established infection (5%) (p=0.016). The prevalence of TDR in Honduras was moderate (7%). The percentage of specimens who were recently infected was low (12%), suggesting that late HIV diagnosis is common. The TDR prevalence was higher in recent than in established infections, which may indicate that TDR is increasing over time. The higher prevalence of NNRTI and NRTI mutations as compared to PI mutations is probably due to a broader and longer use of these drugs in Honduras. Copyright © 2010 Elsevier B.V. All rights reserved.

  15. Large-scale functional purification of recombinant HIV-1 capsid.

    Directory of Open Access Journals (Sweden)

    Magdeleine Hung

    Full Text Available During human immunodeficiency virus type-1 (HIV-1 virion maturation, capsid proteins undergo a major rearrangement to form a conical core that protects the viral nucleoprotein complexes. Mutations in the capsid sequence that alter the stability of the capsid core are deleterious to viral infectivity and replication. Recently, capsid assembly has become an attractive target for the development of a new generation of anti-retroviral agents. Drug screening efforts and subsequent structural and mechanistic studies require gram quantities of active, homogeneous and pure protein. Conventional means of laboratory purification of Escherichia coli expressed recombinant capsid protein rely on column chromatography steps that are not amenable to large-scale production. Here we present a function-based purification of wild-type and quadruple mutant capsid proteins, which relies on the inherent propensity of capsid protein to polymerize and depolymerize. This method does not require the packing of sizable chromatography columns and can generate double-digit gram quantities of functionally and biochemically well-behaved proteins with greater than 98% purity. We have used the purified capsid protein to characterize two known assembly inhibitors in our in-house developed polymerization assay and to measure their binding affinities. Our capsid purification procedure provides a robust method for purifying large quantities of a key protein in the HIV-1 life cycle, facilitating identification of the next generation anti-HIV agents.

  16. The evolutionary rate dynamically tracks changes in HIV-1 epidemics

    Energy Technology Data Exchange (ETDEWEB)

    Maljkovic-berry, Irina [Los Alamos National Laboratory; Athreya, Gayathri [Los Alamos National Laboratory; Daniels, Marcus [Los Alamos National Laboratory; Bruno, William [Los Alamos National Laboratory; Korber, Bette [Los Alamos National Laboratory; Kuiken, Carla [Los Alamos National Laboratory; Ribeiro, Ruy M [Los Alamos National Laboratory

    2009-01-01

    Large-sequence datasets provide an opportunity to investigate the dynamics of pathogen epidemics. Thus, a fast method to estimate the evolutionary rate from large and numerous phylogenetic trees becomes necessary. Based on minimizing tip height variances, we optimize the root in a given phylogenetic tree to estimate the most homogenous evolutionary rate between samples from at least two different time points. Simulations showed that the method had no bias in the estimation of evolutionary rates and that it was robust to tree rooting and topological errors. We show that the evolutionary rates of HIV-1 subtype B and C epidemics have changed over time, with the rate of evolution inversely correlated to the rate of virus spread. For subtype B, the evolutionary rate slowed down and tracked the start of the HAART era in 1996. Subtype C in Ethiopia showed an increase in the evolutionary rate when the prevalence increase markedly slowed down in 1995. Thus, we show that the evolutionary rate of HIV-1 on the population level dynamically tracks epidemic events.

  17. Nup153 and Nup98 bind the HIV-1 core and contribute to the early steps of HIV-1 replication

    International Nuclear Information System (INIS)

    Di Nunzio, Francesca; Fricke, Thomas; Miccio, Annarita; Valle-Casuso, Jose Carlos; Perez, Patricio; Souque, Philippe; Rizzi, Ermanno; Severgnini, Marco; Mavilio, Fulvio; Charneau, Pierre; Diaz-Griffero, Felipe

    2013-01-01

    The early steps of HIV-1 replication involve the entry of HIV-1 into the nucleus, which is characterized by viral interactions with nuclear pore components. HIV-1 developed an evolutionary strategy to usurp the nuclear pore machinery and chromatin in order to integrate and efficiently express viral genes. In the current work, we studied the role of nucleoporins 153 and 98 (Nup153 and Nup98) in infection of human Jurkat lymphocytes by HIV-1. We showed that Nup153-depleted cells exhibited a defect in nuclear import, while depletion of Nup 98 caused a slight defect in HIV integration. To explore the biochemical viral determinants for the requirement of Nup153 and Nup98 during HIV-1 infection, we tested the ability of these nucleoporins to interact with HIV-1 cores. Our findings showed that both nucleoporins bind HIV-1 cores suggesting that this interaction is important for HIV-1 nuclear import and/or integration. Distribution analysis of integration sites in Nup153-depleted cells revealed a reduced tendency of HIV-1 to integrate in intragenic sites, which in part could account for the large infectivity defect observed in Nup153-depleted cells. Our work strongly supports a role for Nup153 in HIV-1 nuclear import and integration. - Highlights: ► We studied the role of Nup98 and Nup153 in HIV-1 infection. ► Nup98 binds the HIV-1 core and is involved in HIV-1 integration. ► Nup153 binds the HIV-1 core and is involved in HIV-1 nuclear import. ► Depletion of Nup153 decreased the integration of HIV-1 in transcriptionally active sites

  18. Nup153 and Nup98 bind the HIV-1 core and contribute to the early steps of HIV-1 replication

    Energy Technology Data Exchange (ETDEWEB)

    Di Nunzio, Francesca, E-mail: francesca.di-nunzio@pasteur.fr [Molecular Virology and Vaccinology unit, CNRS URA 3015, Department of Virology, Institut Pasteur, 25-28 rue du Dr. Roux, 75015 Paris (France); Fricke, Thomas [Department of Microbiology and Immunology, Albert Einstein College of Medicine Bronx, NY 10461 (United States); Miccio, Annarita [University of Modena e Reggio Emilia, Centro di Medicina Rigenerativa, Modena (Italy); Valle-Casuso, Jose Carlos; Perez, Patricio [Department of Microbiology and Immunology, Albert Einstein College of Medicine Bronx, NY 10461 (United States); Souque, Philippe [Molecular Virology and Vaccinology unit, CNRS URA 3015, Department of Virology, Institut Pasteur, 25-28 rue du Dr. Roux, 75015 Paris (France); Rizzi, Ermanno; Severgnini, Marco [Institute of Biomedical Technologies, CNR, Milano (Italy); Mavilio, Fulvio [University of Modena e Reggio Emilia, Centro di Medicina Rigenerativa, Modena (Italy); Genethon, Evry (France); Charneau, Pierre [Molecular Virology and Vaccinology unit, CNRS URA 3015, Department of Virology, Institut Pasteur, 25-28 rue du Dr. Roux, 75015 Paris (France); Diaz-Griffero, Felipe, E-mail: felipe.diaz-griffero@einstein.yu.edu [Department of Microbiology and Immunology, Albert Einstein College of Medicine Bronx, NY 10461 (United States)

    2013-05-25

    The early steps of HIV-1 replication involve the entry of HIV-1 into the nucleus, which is characterized by viral interactions with nuclear pore components. HIV-1 developed an evolutionary strategy to usurp the nuclear pore machinery and chromatin in order to integrate and efficiently express viral genes. In the current work, we studied the role of nucleoporins 153 and 98 (Nup153 and Nup98) in infection of human Jurkat lymphocytes by HIV-1. We showed that Nup153-depleted cells exhibited a defect in nuclear import, while depletion of Nup 98 caused a slight defect in HIV integration. To explore the biochemical viral determinants for the requirement of Nup153 and Nup98 during HIV-1 infection, we tested the ability of these nucleoporins to interact with HIV-1 cores. Our findings showed that both nucleoporins bind HIV-1 cores suggesting that this interaction is important for HIV-1 nuclear import and/or integration. Distribution analysis of integration sites in Nup153-depleted cells revealed a reduced tendency of HIV-1 to integrate in intragenic sites, which in part could account for the large infectivity defect observed in Nup153-depleted cells. Our work strongly supports a role for Nup153 in HIV-1 nuclear import and integration. - Highlights: ► We studied the role of Nup98 and Nup153 in HIV-1 infection. ► Nup98 binds the HIV-1 core and is involved in HIV-1 integration. ► Nup153 binds the HIV-1 core and is involved in HIV-1 nuclear import. ► Depletion of Nup153 decreased the integration of HIV-1 in transcriptionally active sites.

  19. Human endogenous retrovirus K Gag coassembles with HIV-1 Gag and reduces the release efficiency and infectivity of HIV-1.

    Science.gov (United States)

    Monde, Kazuaki; Contreras-Galindo, Rafael; Kaplan, Mark H; Markovitz, David M; Ono, Akira

    2012-10-01

    Human endogenous retroviruses (HERVs), which are remnants of ancestral retroviruses integrated into the human genome, are defective in viral replication. Because activation of HERV-K and coexpression of this virus with HIV-1 have been observed during HIV-1 infection, it is conceivable that HERV-K could affect HIV-1 replication, either by competition or by cooperation, in cells expressing both viruses. In this study, we found that the release efficiency of HIV-1 Gag was 3-fold reduced upon overexpression of HERV-K(CON) Gag. In addition, we observed that in cells expressing Gag proteins of both viruses, HERV-K(CON) Gag colocalized with HIV-1 Gag at the plasma membrane. Furthermore, HERV-K(CON) Gag was found to coassemble with HIV-1 Gag, as demonstrated by (i) processing of HERV-K(CON) Gag by HIV-1 protease in virions, (ii) coimmunoprecipitation of virion-associated HERV-K(CON) Gag with HIV-1 Gag, and (iii) rescue of a late-domain-defective HERV-K(CON) Gag by wild-type (WT) HIV-1 Gag. Myristylation-deficient HERV-K(CON) Gag localized to nuclei, suggesting cryptic nuclear trafficking of HERV-K Gag. Notably, unlike WT HERV-K(CON) Gag, HIV-1 Gag failed to rescue myristylation-deficient HERV-K(CON) Gag to the plasma membrane. Efficient colocalization and coassembly of HIV-1 Gag and HERV-K Gag also required nucleocapsid (NC). These results provide evidence that HIV-1 Gag heteromultimerizes with HERV-K Gag at the plasma membrane, presumably through NC-RNA interaction. Intriguingly, HERV-K Gag overexpression reduced not only HIV-1 release efficiency but also HIV-1 infectivity in a myristylation- and NC-dependent manner. Altogether, these results indicate that Gag proteins of endogenous retroviruses can coassemble with HIV-1 Gag and modulate the late phase of HIV-1 replication.

  20. Excision of HIV-1 proviral DNA by recombinant cell permeable tre-recombinase.

    Directory of Open Access Journals (Sweden)

    Lakshmikanth Mariyanna

    Full Text Available Over the previous years, comprehensive studies on antiretroviral drugs resulted in the successful introduction of highly active antiretroviral therapy (HAART into clinical practice for treatment of HIV/AIDS. However, there is still need for new therapeutic approaches, since HAART cannot eradicate HIV-1 from the infected organism and, unfortunately, can be associated with long-term toxicity and the development of drug resistance. In contrast, novel gene therapy strategies may have the potential to reverse the infection by eradicating HIV-1. For example, expression of long terminal repeat (LTR-specific recombinase (Tre-recombinase has been shown to result in chromosomal excision of proviral DNA and, in consequence, in the eradication of HIV-1 from infected cell cultures. However, the delivery of Tre-recombinase currently depends on the genetic manipulation of target cells, a process that is complicating such therapeutic approaches and, thus, might be undesirable in a clinical setting. In this report we demonstrate that E.coli expressed Tre-recombinases, tagged either with the protein transduction domain (PTD from the HIV-1 Tat trans-activator or the translocation motif (TLM of the Hepatitis B virus PreS2 protein, were able to translocate efficiently into cells and showed significant recombination activity on HIV-1 LTR sequences. Tre activity was observed using episomal and stable integrated reporter constructs in transfected HeLa cells. Furthermore, the TLM-tagged enzyme was able to excise the full-length proviral DNA from chromosomal integration sites of HIV-1-infected HeLa and CEM-SS cells. The presented data confirm Tre-recombinase activity on integrated HIV-1 and provide the basis for the non-genetic transient application of engineered recombinases, which may be a valuable component of future HIV eradication strategies.

  1. Origin, imports and exports of HIV-1 subtype C in South Africa: A historical perspective.

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    Wilkinson, Eduan; Rasmussen, David; Ratmann, Oliver; Stadler, Tanja; Engelbrecht, Susan; de Oliveira, Tulio

    2016-12-01

    While the HIV epidemic in South Africa had a later onset than epidemics in other southern African countries, prevalence grew rapidly during the 1990's when the country was going through socio-political changes with the end of Apartheid. South Africa currently has the largest number of people living with HIV in the world and the epidemic is dominated by a unique subtype, HIV-1 subtype C. This large epidemic is also characterized by high level of genetic diversity. We hypothesize that this diversity is due to multiple introductions of the virus during the period of change. In this paper, we apply novel phylogeographic methods to estimate the number of viral imports and exports from the start of the epidemic to the present. We assembled 11,289 unique subtype C pol sequences from southern Africa. These represent one of the largest sequence datasets ever analyzed in the region. Sequences were stratified based on country of sampling and levels of genetic diversity were estimated for each country. Sequences were aligned and a maximum-likelihood evolutionary tree was inferred. Least-Squares Dating was then used to obtain a dated phylogeny from which we estimated the number of introductions into and exports out of South Africa using parsimony-based ancestral location reconstructions. Our results identified 189 viral introductions into South Africa with the largest number of introductions attributed to Zambia (n=109), Botswana (n=32), Malawi (n=26) and Zimbabwe (n=13). South Africa also exported many viral lineages to its neighbours. The bulk viral imports and exports appear to have occurred between 1985 and 2000, coincident with the period of socio-political transition. The high level of subtype C genetic diversity in South Africa is related to multiple introductions of the virus to the country. While the number of viral imports and exports we identified was highly sensitive to the number of samples included from each country, they mostly clustered around the period of

  2. Population dynamics of HIV-2 in rural West Africa: comparison with HIV-1 and ongoing transmission at the heart of the epidemic

    NARCIS (Netherlands)

    de Silva, Thushan I.; van Tienen, Carla; Onyango, Clayton; Jabang, Abdoulie; Vincent, Tim; Loeff, Maarten F. Schim van der; Coutinho, Roel A.; Jaye, Assan; Rowland-Jones, Sarah; Whittle, Hilton; Cotten, Matthew; Hué, Stéphane

    2013-01-01

    To compare the population dynamics of HIV-2 and HIV-1, and to characterize ongoing HIV-2 transmission in rural Guinea-Bissau. Phylogenetic and phylodynamic analyses using HIV-2 gag and env, and HIV-1 env sequences, combined with epidemiological data from a community cohort. Samples were obtained

  3. Molecular Diversity of HIV-1 among People Who Inject Drugs in Kuala Lumpur, Malaysia: Massive Expansion of Circulating Recombinant Form (CRF) 33_01B and Emergence of Multiple Unique Recombinant Clusters

    Science.gov (United States)

    Chow, Wei Zhen; Ng, Kim Tien; Yong, Yean Kong; Azmel, Azureen; Takebe, Yutaka; Al-Darraji, Haider Abdulrazzaq Abed; Kamarulzaman, Adeeba; Tee, Kok Keng

    2013-01-01

    Since the discovery of HIV-1 circulating recombinant form (CRF) 33_01B in Malaysia in the early 2000 s, continuous genetic diversification and active recombination involving CRF33_01B and other circulating genotypes in the region including CRF01_AE and subtype B′ of Thai origin, have led to the emergence of novel CRFs and unique recombinant forms. The history and magnitude of CRF33_01B transmission among various risk groups including people who inject drugs (PWID) however have not been investigated despite the high epidemiological impact of CRF33_01B in the region. We update the most recent molecular epidemiology of HIV-1 among PWIDs recruited in Malaysia between 2010 and 2011 by population sequencing and phylogenetic analysis of 128 gag-pol sequences. HIV-1 CRF33_01B was circulating among 71% of PWIDs whilst a lower prevalence of other previously dominant HIV-1 genotypes [subtype B′ (11%) and CRF01_AE (5%)] and CRF01_AE/B′ unique recombinants (13%) were detected, indicating a significant shift in genotype replacement in this population. Three clusters of CRF01_AE/B′ recombinants displaying divergent yet phylogenetically-related mosaic genomes to CRF33_01B were identified and characterized, suggestive of an abrupt emergence of multiple novel CRF clades. Using rigorous maximum likelihood approach and the Bayesian Markov chain Monte Carlo (MCMC) sampling of CRF33_01Bpol sequences to elucidate the past population dynamics, we found that the founder lineages of CRF33_01B were likely to have first emerged among PWIDs in the early 1990 s before spreading exponentially to various high and low-risk populations (including children who acquired infections from their mothers) and later on became endemic around the early 2000 s. Taken together, our findings provide notable genetic evidence indicating the widespread expansion of CRF33_01B among PWIDs and into the general population. The emergence of numerous previously unknown recombinant clades highlights the

  4. Molecular diversity of HIV-1 among people who inject drugs in Kuala Lumpur, Malaysia: massive expansion of circulating recombinant form (CRF) 33_01B and emergence of multiple unique recombinant clusters.

    Science.gov (United States)

    Chow, Wei Zhen; Ong, Lai Yee; Razak, Siti Humaira; Lee, Yeat Mei; Ng, Kim Tien; Yong, Yean Kong; Azmel, Azureen; Takebe, Yutaka; Al-Darraji, Haider Abdulrazzaq Abed; Kamarulzaman, Adeeba; Tee, Kok Keng

    2013-01-01

    Since the discovery of HIV-1 circulating recombinant form (CRF) 33_01B in Malaysia in the early 2000 s, continuous genetic diversification and active recombination involving CRF33_01B and other circulating genotypes in the region including CRF01_AE and subtype B' of Thai origin, have led to the emergence of novel CRFs and unique recombinant forms. The history and magnitude of CRF33_01B transmission among various risk groups including people who inject drugs (PWID) however have not been investigated despite the high epidemiological impact of CRF33_01B in the region. We update the most recent molecular epidemiology of HIV-1 among PWIDs recruited in Malaysia between 2010 and 2011 by population sequencing and phylogenetic analysis of 128 gag-pol sequences. HIV-1 CRF33_01B was circulating among 71% of PWIDs whilst a lower prevalence of other previously dominant HIV-1 genotypes [subtype B' (11%) and CRF01_AE (5%)] and CRF01_AE/B' unique recombinants (13%) were detected, indicating a significant shift in genotype replacement in this population. Three clusters of CRF01_AE/B' recombinants displaying divergent yet phylogenetically-related mosaic genomes to CRF33_01B were identified and characterized, suggestive of an abrupt emergence of multiple novel CRF clades. Using rigorous maximum likelihood approach and the Bayesian Markov chain Monte Carlo (MCMC) sampling of CRF33_01Bpol sequences to elucidate the past population dynamics, we found that the founder lineages of CRF33_01B were likely to have first emerged among PWIDs in the early 1990 s before spreading exponentially to various high and low-risk populations (including children who acquired infections from their mothers) and later on became endemic around the early 2000 s. Taken together, our findings provide notable genetic evidence indicating the widespread expansion of CRF33_01B among PWIDs and into the general population. The emergence of numerous previously unknown recombinant clades highlights the escalating

  5. Molecular diversity of HIV-1 among people who inject drugs in Kuala Lumpur, Malaysia: massive expansion of circulating recombinant form (CRF 33_01B and emergence of multiple unique recombinant clusters.

    Directory of Open Access Journals (Sweden)

    Wei Zhen Chow

    Full Text Available Since the discovery of HIV-1 circulating recombinant form (CRF 33_01B in Malaysia in the early 2000 s, continuous genetic diversification and active recombination involving CRF33_01B and other circulating genotypes in the region including CRF01_AE and subtype B' of Thai origin, have led to the emergence of novel CRFs and unique recombinant forms. The history and magnitude of CRF33_01B transmission among various risk groups including people who inject drugs (PWID however have not been investigated despite the high epidemiological impact of CRF33_01B in the region. We update the most recent molecular epidemiology of HIV-1 among PWIDs recruited in Malaysia between 2010 and 2011 by population sequencing and phylogenetic analysis of 128 gag-pol sequences. HIV-1 CRF33_01B was circulating among 71% of PWIDs whilst a lower prevalence of other previously dominant HIV-1 genotypes [subtype B' (11% and CRF01_AE (5%] and CRF01_AE/B' unique recombinants (13% were detected, indicating a significant shift in genotype replacement in this population. Three clusters of CRF01_AE/B' recombinants displaying divergent yet phylogenetically-related mosaic genomes to CRF33_01B were identified and characterized, suggestive of an abrupt emergence of multiple novel CRF clades. Using rigorous maximum likelihood approach and the Bayesian Markov chain Monte Carlo (MCMC sampling of CRF33_01Bpol sequences to elucidate the past population dynamics, we found that the founder lineages of CRF33_01B were likely to have first emerged among PWIDs in the early 1990 s before spreading exponentially to various high and low-risk populations (including children who acquired infections from their mothers and later on became endemic around the early 2000 s. Taken together, our findings provide notable genetic evidence indicating the widespread expansion of CRF33_01B among PWIDs and into the general population. The emergence of numerous previously unknown recombinant clades highlights the

  6. Co-evolution of a broadly neutralizing HIV-1 antibody and founder virus

    Science.gov (United States)

    Liao, Hua-Xin; Lynch, Rebecca; Zhou, Tongqing; Gao, Feng; Alam, S. Munir; Boyd, Scott D.; Fire, Andrew Z.; Roskin, Krishna M.; Schramm, Chaim A.; Zhang, Zhenhai; Zhu, Jiang; Shapiro, Lawrence; Mullikin, James C.; Gnanakaran, S.; Hraber, Peter; Wiehe, Kevin; Kelsoe, Garnett; Yang, Guang; Xia, Shi-Mao; Montefiori, David C.; Parks, Robert; Lloyd, Krissey E.; Scearce, Richard M.; Soderberg, Kelly A.; Cohen, Myron; Kaminga, Gift; Louder, Mark K.; Tran, Lillan M.; Chen, Yue; Cai, Fangping; Chen, Sheri; Moquin, Stephanie; Du, Xiulian; Joyce, Gordon M.; Srivatsan, Sanjay; Zhang, Baoshan; Zheng, Anqi; Shaw, George M.; Hahn, Beatrice H.; Kepler, Thomas B.; Korber, Bette T.M.; Kwong, Peter D.; Mascola, John R.; Haynes, Barton F.

    2013-01-01

    Current HIV-1 vaccines elicit strain-specific neutralizing antibodies. However, cross-reactive neutralizing antibodies arise in ~20% of HIV-1-infected individuals, and details of their generation could provide a roadmap for effective vaccination. Here we report the isolation, evolution and structure of a broadly neutralizing antibody from an African donor followed from time of infection. The mature antibody, CH103, neutralized ~55% of HIV-1 isolates, and its co-crystal structure with gp120 revealed a novel loop-based mechanism of CD4-binding site recognition. Virus and antibody gene sequencing revealed concomitant virus evolution and antibody maturation. Notably, the CH103-lineage unmutated common ancestor avidly bound the transmitted/founder HIV-1 envelope glycoprotein, and evolution of antibody neutralization breadth was preceded by extensive viral diversification in and near the CH103 epitope. These data elucidate the viral and antibody evolution leading to induction of a lineage of HIV-1 broadly neutralizing antibodies and provide insights into strategies to elicit similar antibodies via vaccination. PMID:23552890

  7. Correlation between HIV-1 genotype and clinical progression in HIV/AIDS patients in Surabaya, Indonesia

    Science.gov (United States)

    Rachman, B. E.; Khairunisa, S. Q.; Witaningrum, A. M.; Yunifiar, M. Q.; Nasronudin

    2018-03-01

    Several factors such as host and viral factors can affect the progression of HIV/AIDS. This study aims to identify the correlation viral factors, especially the HIV-1 subtype with HIV/AIDS progression. Inpatient HIV/AIDS during the period March to September 2017 and willing to participate are included in the study. Historical data of disease and treatment was taken by medical record. Blood samples were amplified, sequenced and undergone phylogenetic analysis. Linear regression analysis was used to estimate beta coefficient (β) and 95%CI of HIV/AIDS progression (measured by the CD4 change rate, ΔCD4 cell count/time span in months).This study has 17 samples. The HIV-1 subtype was dominated by CRF01_AE (81.8%) followed by subtype B (18.2%). There was significant correlation between subtype HIV-1 (p = 0.04) and body mass index (p = 0.038) with HIV/AIDS clinical stage. Many factors were assumed to be correlated with increased rate of CD4, but we only subtype HIV-1 had a significant correlation (p = 0.024) with it. From multivariate analysis, we also found that subtype HIV-1 had a significant correlation (β = 0.788, 95%CI: 17.5-38.6, p = 0.004).

  8. Differential trends in the codon usage patterns in HIV-1 genes.

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    Aridaman Pandit

    Full Text Available Host-pathogen interactions underlie one of the most complex evolutionary phenomena resulting in continual adaptive genetic changes, where pathogens exploit the host's molecular resources for growth and survival, while hosts try to eliminate the pathogen. Deciphering the molecular basis of host-pathogen interactions is useful in understanding the factors governing pathogen evolution and disease propagation. In host-pathogen context, a balance between mutation, selection, and genetic drift is known to maintain codon bias in both organisms. Studies revealing determinants of the bias and its dynamics are central to the understanding of host-pathogen evolution. We considered the Human Immunodeficiency Virus (HIV type 1 and its human host to search for evolutionary signatures in the viral genome. Positive selection is known to dominate intra-host evolution of HIV-1, whereas high genetic variability underlies the belief that neutral processes drive inter-host differences. In this study, we analyze the codon usage patterns of HIV-1 genomes across all subtypes and clades sequenced over a period of 23 years. We show presence of unique temporal correlations in the codon bias of three HIV-1 genes illustrating differential adaptation of the HIV-1 genes towards the host preferred codons. Our results point towards gene-specific translational selection to be an important force driving the evolution of HIV-1 at the population level.

  9. Synthetic AAV/CRISPR vectors for blocking HIV-1 expression in persistently infected astrocytes.

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    Kunze, Christine; Börner, Kathleen; Kienle, Eike; Orschmann, Tanja; Rusha, Ejona; Schneider, Martha; Radivojkov-Blagojevic, Milena; Drukker, Micha; Desbordes, Sabrina; Grimm, Dirk; Brack-Werner, Ruth

    2018-02-01

    Astrocytes, the most abundant cells in the mammalian brain, perform key functions and are involved in several neurodegenerative diseases. The human immunodeficiency virus (HIV) can persist in astrocytes, contributing to the HIV burden and neurological dysfunctions in infected individuals. While a comprehensive approach to HIV cure must include the targeting of HIV-1 in astrocytes, dedicated tools for this purpose are still lacking. Here we report a novel Adeno-associated virus-based vector (AAV9P1) with a synthetic surface peptide for transduction of astrocytes. Analysis of AAV9P1 transduction efficiencies with single brain cell populations, including primary human brain cells, as well as human brain organoids demonstrated that AAV9P1 targeted terminally differentiated human astrocytes much more efficiently than neurons. We then investigated whether AAV9P1 can be used to deliver HIV-inhibitory genes to astrocytes. To this end we generated AAV9P1 vectors containing genes for HIV-1 proviral editing by CRISPR/Cas9. Latently HIV-1 infected astrocytes transduced with these vectors showed significantly diminished reactivation of proviruses, compared with untransduced cultures. Sequence analysis identified mutations/deletions in key HIV-1 transcriptional control regions. We conclude that AAV9P1 is a promising tool for gene delivery to astrocytes and may facilitate inactivation/destruction of persisting HIV-1 proviruses in astrocyte reservoirs. © 2017 Wiley Periodicals, Inc.

  10. Association between HIV-1 coreceptor usage and resistance to broadly neutralizing antibodies.

    Science.gov (United States)

    Pfeifer, Nico; Walter, Hauke; Lengauer, Thomas

    2014-10-01

    Recently discovered broadly neutralizing antibodies have revitalized hopes of developing a universal vaccine against HIV-1. Mainly responsible for new infections are variants only using CCR5 for cell entry, whereas CXCR4-using variants can become dominant in later infection stages. We performed a statistical analysis on two different previously published data sets. The first data set was a panel of 199 diverse HIV-1 isolates for which IC50 neutralization titers were determined for the broadly neutralizing antibodies VRC01, VRC-PG04, PG9, and PG16. The second data set contained env sequences of viral variants extracted from HIV-1-infected humanized mice treated with the antibody PGT128 and from untreated control mice. For the panel of 199 diverse HIV-1 isolates, we found a statistically significant association between viral resistance to PG9 and PG16 and CXCR4 coreceptor usage (P = 0.0011 and P = 0.0010, respectively). Our analysis of viral variants from HIV-1-infected humanized mice under treatment with the broadly neutralizing antibody PGT128 indicated that certain antibodies might drive a viral population toward developing CXCR4 coreceptor usage capability (P = 0.0011 for the comparison between PGT128 and control measurement). These analyses highlight the importance of accounting for a possible coreceptor usage bias pertaining to the effectiveness of an HIV vaccine and to passive antibody transfer as therapeutic approach.

  11. Increased Risk of HIV-1 Transmission in Pregnancy: A Prospective Study among African HIV-1 Serodiscordant Couples

    Science.gov (United States)

    MUGO, Nelly R.; HEFFRON, Renee; DONNELL, Deborah; WALD, Anna; WERE, Edwin O.; REES, Helen; CELUM, Connie; KIARIE, James N.; COHEN, Craig R.; KAYINTEKORE, Kayitesi; BAETEN, Jared M.

    2011-01-01

    Background Physiologic and behavioral changes during pregnancy may alter HIV-1 susceptibility and infectiousness. Prospective studies exploring pregnancy and HIV-1 acquisition risk in women have found inconsistent results. No study has explored the effect of pregnancy on HIV-1 transmission risk from HIV-1 infected women to male partners. Methods In a prospective study of African HIV-1 serodiscordant couples, we evaluated the relationship between pregnancy and the risk of 1) HIV-1 acquisition among women and 2) HIV-1 transmission from women to men. Results 3321 HIV-1 serodiscordant couples were enrolled, 1085 (32.7%) with HIV-1 susceptible female partners and 2236 (67.3%) with susceptible male partners. HIV-1 incidence in women was 7.35 versus 3.01 per 100 person-years during pregnant and non-pregnant periods (hazard ratio [HR] 2.34, 95% confidence interval [CI] 1.33–4.09). This effect was attenuated and not statistically significant after adjusting for sexual behavior and other confounding factors (adjusted HR 1.71, 95% CI 0.93–3.12). HIV-1 incidence in male partners of infected women was 3.46 versus 1.58 per 100 person-years when their partners were pregnant versus not pregnant (HR 2.31, 95% CI 1.22–4.39). This effect was not attenuated in adjusted analysis (adjusted HR 2.47, 95% CI 1.26–4.85). Conclusions HIV-1 risk increased two-fold during pregnancy. Elevated risk of HIV-1 acquisition in pregnant women appeared in part to be explained by behavioral and other factors. This is the first study to show pregnancy increased the risk of female-to-male HIV-1 transmission, which may reflect biological changes of pregnancy that could increase HIV-1 infectiousness. PMID:21785321

  12. Early low-titer neutralizing antibodies impede HIV-1 replication and select for virus escape.

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    Katharine J Bar

    Full Text Available Single genome sequencing of early HIV-1 genomes provides a sensitive, dynamic assessment of virus evolution and insight into the earliest anti-viral immune responses in vivo. By using this approach, together with deep sequencing, site-directed mutagenesis, antibody adsorptions and virus-entry assays, we found evidence in three subjects of neutralizing antibody (Nab responses as early as 2 weeks post-seroconversion, with Nab titers as low as 1∶20 to 1∶50 (IC(50 selecting for virus escape. In each of the subjects, Nabs targeted different regions of the HIV-1 envelope (Env in a strain-specific, conformationally sensitive manner. In subject CH40, virus escape was first mediated by mutations in the V1 region of the Env, followed by V3. HIV-1 specific monoclonal antibodies from this subject mapped to an immunodominant region at the base of V3 and exhibited neutralizing patterns indistinguishable from polyclonal antibody responses, indicating V1-V3 interactions within the Env trimer. In subject CH77, escape mutations mapped to the V2 region of Env, several of which selected for alterations of glycosylation. And in subject CH58, escape mutations mapped to the Env outer domain. In all three subjects, initial Nab recognition was followed by sequential rounds of virus escape and Nab elicitation, with Nab escape variants exhibiting variable costs to replication fitness. Although delayed in comparison with autologous CD8 T-cell responses, our findings show that Nabs appear earlier in HIV-1 infection than previously recognized, target diverse sites on HIV-1 Env, and impede virus replication at surprisingly low titers. The unexpected in vivo sensitivity of early transmitted/founder virus to Nabs raises the possibility that similarly low concentrations of vaccine-induced Nabs could impair virus acquisition in natural HIV-1 transmission, where the risk of infection is low and the number of viruses responsible for transmission and productive clinical

  13. [Study on the molecular-epidemiological characteristics of HIV-1 in Shenzhen, 1992-2008].

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    Zhao, Guang-lu; Yu, Wei; Zhang, Juan-juan; Chen, Lin; Feng, Tie-jian; Wang, Feng; Hong, Fu-chang; Wang, Xiao-hui; Li, Qing

    2012-01-01

    To investigate the epidemiological characteristics of HIV-1 subtype in Shenzhen from 1992 to 2008. 489 HIV-1 positive plasma samples were collected from 1992 to 2008 in Shenzhen. HIV-1 env genes were amplified by nested-PCR from RNA. Phylogenetic analysis was performed on data regarding the nucleotide sequence. A total of 464 sequences were amplified and genotyped. Data from this study revealed that CRF01_AE was a predominant HIV-1 subtype in Shenzhen (64.4%, 299/464), followed by subtypes CRF_BC (17.5%, 81/464), B' (14.7%, 68/464) and B (2.4%, 11/464). Subtype C (0.4%, 2/464), A1 (0.2%, 1/464), CRF02_AG (0.2%, 1/464) and CRF06_cpx (0.2%, 1/464) were also prevalent in Shenzhen. CRF01_AE and CRF_BC were predominant among heterosexuals, homosexuals and injection drug users, while B' was predominant among blood donors. Results from phylogenetic tree analysis showed that some of the HIV-1 clusters had been defined in CRF01_AE strains at different time or groups with different transmission routes. Cross-infections were also seen. CRF01_AE was the predominant HIV-1 subtype in Shenzhen while CRF_BC, B, B', C, A1, CRF02_AG and a small amount of CRF06_cpx or recombinant subtypes were prevalent in this city. Different subtypes showed great variation in the process of epidemics.

  14. Zinc finger nuclease: a new approach for excising HIV-1 proviral DNA from infected human T cells.

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    Qu, Xiying; Wang, Pengfei; Ding, Donglin; Wang, Xiaohui; Zhang, Gongmin; Zhou, Xin; Liu, Lin; Zhu, Xiaoli; Zeng, Hanxian; Zhu, Huanzhang

    2014-09-01

    A major reason that Acquired Immune Deficiency Syndrome (AIDS) cannot be completely cured is the human immunodeficiency virus 1 (HIV-1) provirus integrated into the human genome. Though existing therapies can inhibit replication of HIV-1, they cannot eradicate it. A molecular therapy gains popularity due to its specifically targeting to HIV-1 infected cells and effectively removing the HIV-1, regardless of viral genes being active or dormant. Now, we propose a new method which can excellently delete the HIV provirus from the infected human T cell genome. First, we designed zinc-finger nucleases (ZFNs) that target a sequence within the long terminal repeat (LTR) U3 region that is highly conserved in whole clade. Then, we screened out one pair of ZFN and named it as ZFN-U3. We discovered that ZFN-U3 can exactly target and eliminate the full-length HIV-1 proviral DNA after the infected human cell lines treated with it, and the frequency of its excision was about 30 % without cytotoxicity. These results prove that ZFN-U3 can efficiently excise integrated HIV-1 from the human genome in infected cells. This method to delete full length HIV-1 in human genome can therefore provide a novel approach to cure HIV-infected individuals in the future.

  15. A real-time view of the TAR:Tat:P-TEFb complex at HIV-1 transcription sites

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    Knezevich Anna

    2007-05-01

    Full Text Available Abstract HIV-1 transcription is tightly regulated: silent in long-term latency and highly active in acutely-infected cells. Transcription is activated by the viral protein Tat, which recruits the elongation factor P-TEFb by binding the TAR sequence present in nascent HIV-1 RNAs. In this study, we analyzed the dynamic of the TAR:Tat:P-TEFb complex in living cells, by performing FRAP experiments at HIV-1 transcription sites. Our results indicate that a large fraction of Tat present at these sites is recruited by Cyclin T1. We found that in the presence of Tat, Cdk9 remained bound to nascent HIV-1 RNAs for 71s. In contrast, when transcription was activated by PMA/ionomycin, in the absence of Tat, Cdk9 turned-over rapidly and resided on the HIV-1 promoter for only 11s. Thus, the mechanism of trans-activation determines the residency time of P-TEFb at the HIV-1 gene, possibly explaining why Tat is such a potent transcriptional activator. In addition, we observed that Tat occupied HIV-1 transcription sites for 55s, suggesting that the TAR:Tat:P-TEFb complex dissociates from the polymerase following transcription initiation, and undergoes subsequent cycles of association/dissociation.

  16. Human CNS cultures exposed to HIV-1 gp120 reproduce dendritic injuries of HIV-1-associated dementia

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    Hammond Robert R

    2004-05-01

    Full Text Available Abstract HIV-1-associated dementia remains a common subacute to chronic central nervous system degeneration in adult and pediatric HIV-1 infected populations. A number of viral and host factors have been implicated including the HIV-1 120 kDa envelope glycoprotein (gp120. In human post-mortem studies using confocal scanning laser microscopy for microtubule-associated protein 2 and synaptophysin, neuronal dendritic pathology correlated with dementia. In the present study, primary human CNS cultures exposed to HIV-1 gp120 at 4 weeks in vitro suffered gliosis and dendritic damage analogous to that described in association with HIV-1-associated dementia.

  17. Genetic Signatures of HIV-1 Envelope-mediated Bystander Apoptosis

    Science.gov (United States)

    Joshi, Anjali; Lee, Raphael T. C.; Mohl, Jonathan; Sedano, Melina; Khong, Wei Xin; Ng, Oon Tek; Maurer-Stroh, Sebastian; Garg, Himanshu

    2014-01-01

    The envelope (Env) glycoprotein of HIV is an important determinant of viral pathogenesis. Several lines of evidence support the role of HIV-1 Env in inducing bystander apoptosis that may be a contributing factor in CD4+ T cell loss. However, most of the studies testing this phenomenon have been conducted with laboratory-adapted HIV-1 isolates. This raises the question of whether primary Envs derived from HIV-infected patients are capable of inducing bystander apoptosis and whether specific Env signatures are associated with this phenomenon. We developed a high throughput assay to determine the bystander apoptosis inducing activity of a panel of primary Envs. We tested 38 different Envs for bystander apoptosis, virion infectivity, neutralizing antibody sensitivity, and putative N-linked glycosylation sites along with a comprehensive sequence analysis to determine if specific sequence signatures within the viral Env are associated with bystander apoptosis. Our studies show that primary Envs vary considerably in their bystander apoptosis-inducing potential, a phenomenon that correlates inversely with putative N-linked glycosylation sites and positively with virion infectivity. By use of a novel phylogenetic analysis that avoids subtype bias coupled with structural considerations, we found specific residues like Arg-476 and Asn-425 that were associated with differences in bystander apoptosis induction. A specific role of these residues was also confirmed experimentally. These data demonstrate for the first time the potential of primary R5 Envs to mediate bystander apoptosis in CD4+ T cells. Furthermore, we identify specific genetic signatures within the Env that may be associated with the bystander apoptosis-inducing phenotype. PMID:24265318

  18. Somatic populations of PGT135-137 HIV-1-neutralizing antibodies identified by 454 pyrosequencing and bioinformatics

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    Jiang eZhu

    2012-09-01

    Full Text Available Select HIV-1-infected individuals develop sera capable of neutralizing diverse viral strains. The molecular basis of this neutralization is currently being deciphered by the isolation of HIV-1-neutralizing antibodies. In one infected donor, three neutralizing antibodies, PGT135-137, were identified by assessment of neutralization from individually sorted B cells and found to recognize an epitope containing an N-linked glycan at residue 332 on HIV-1 gp120. Here we use deep sequencing and bioinformatics methods to interrogate the B cell record of this donor to gain a more complete understanding of the humoral immune response. PGT135-137-gene family-specific primers were used to amplify heavy and light chain-variable domain sequences. 454 pyrosequencing produced 141,298 heavy-chain sequences of IGHV4-39 origin and 87,229 light-chain sequences of IGKV3-15 origin. A number of heavy and light chain sequences of ~90% identity to PGT137, several to PGT136, and none of high identity to PGT135 were identified. After expansion of these sequences to include close phylogenetic relatives, a total of 202 heavy-chain sequences and 72 light-chain sequences were identified. These sequences were clustered into populations of 95% identity comprising 15 for heavy chain and 10 for light chain, and a select sequence from each population was synthesized and reconstituted with a PGT137-partner chain. Reconstituted antibodies showed varied neutralization phenotypes for HIV-1 clade A and D isolates. Sequence diversity of the antibody population represented by these tested sequences was notably higher than observed with a 454 pyrosequencing-control analysis on 10 antibodies of defined sequence, suggesting that this diversity results primarily from somatic maturation. Our results thus provide an example of how pathogens like HIV-1 are opposed by a varied humoral immune response, derived from intrinsic mechanisms of antibody development, and embodied by somatic populations

  19. Importance of Viral Sequence Length and Number of Variable and Informative Sites in Analysis of HIV Clustering.

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    Novitsky, Vlad; Moyo, Sikhulile; Lei, Quanhong; DeGruttola, Victor; Essex, M

    2015-05-01

    To improve the methodology of HIV cluster analysis, we addressed how analysis of HIV clustering is associated with parameters that can affect the outcome of viral clustering. The extent of HIV clustering and tree certainty was compared between 401 HIV-1C near full-length genome sequences and subgenomic regions retrieved from the LANL HIV Database. Sliding window analysis was based on 99 windows of 1,000 bp and 45 windows of 2,000 bp. Potential associations between the extent of HIV clustering and sequence length and the number of variable and informative sites were evaluated. The near full-length genome HIV sequences showed the highest extent of HIV clustering and the highest tree certainty. At the bootstrap threshold of 0.80 in maximum likelihood (ML) analysis, 58.9% of near full-length HIV-1C sequences but only 15.5% of partial pol sequences (ViroSeq) were found in clusters. Among HIV-1 structural genes, pol showed the highest extent of clustering (38.9% at a bootstrap threshold of 0.80), although it was significantly lower than in the near full-length genome sequences. The extent of HIV clustering was significantly higher for sliding windows of 2,000 bp than 1,000 bp. We found a strong association between the sequence length and proportion of HIV sequences in clusters, and a moderate association between the number of variable and informative sites and the proportion of HIV sequences in clusters. In HIV cluster analysis, the extent of detectable HIV clustering is directly associated with the length of viral sequences used, as well as the number of variable and informative sites. Near full-length genome sequences could provide the most informative HIV cluster analysis. Selected subgenomic regions with a high extent of HIV clustering and high tree certainty could also be considered as a second choice.

  20. HIV-1 adaptation studies reveal a novel Env-mediated homeostasis mechanism for evading lethal hypermutation by APOBEC3G.

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    Terumasa Ikeda

    2018-04-01

    Full Text Available HIV-1 replication normally requires Vif-mediated neutralization of APOBEC3 antiviral enzymes. Viruses lacking Vif succumb to deamination-dependent and -independent restriction processes. Here, HIV-1 adaptation studies were leveraged to ask whether viruses with an irreparable vif deletion could develop resistance to restrictive levels of APOBEC3G. Several resistant viruses were recovered with multiple amino acid substitutions in Env, and these changes alone are sufficient to protect Vif-null viruses from APOBEC3G-dependent restriction in T cell lines. Env adaptations cause decreased fusogenicity, which results in higher levels of Gag-Pol packaging. Increased concentrations of packaged Pol in turn enable faster virus DNA replication and protection from APOBEC3G-mediated hypermutation of viral replication intermediates. Taken together, these studies reveal that a moderate decrease in one essential viral activity, namely Env-mediated fusogenicity, enables the virus to change other activities, here, Gag-Pol packaging during particle production, and thereby escape restriction by the antiviral factor APOBEC3G. We propose a new paradigm in which alterations in viral homeostasis, through compensatory small changes, constitute a general mechanism used by HIV-1 and other viral pathogens to escape innate antiviral responses and other inhibitions including antiviral drugs.

  1. Methamphetamine inhibits HIV-1 replication in CD4+ T cells by modulating anti-HIV-1 miRNA expression.

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    Mantri, Chinmay K; Mantri, Jyoti V; Pandhare, Jui; Dash, Chandravanu

    2014-01-01

    Methamphetamine is the second most frequently used illicit drug in the United States. Methamphetamine abuse is associated with increased risk of HIV-1 acquisition, higher viral loads, and enhanced HIV-1 pathogenesis. Although a direct link between methamphetamine abuse and HIV-1 pathogenesis remains to be established in patients, methamphetamine has been shown to increase HIV-1 replication in macrophages, dendritic cells, and cells of HIV transgenic mice. Intriguingly, the effects of methamphetamine on HIV-1 replication in human CD4(+) T cells that serve as the primary targets of infection in vivo are not clearly understood. Therefore, we examined HIV-1 replication in primary CD4(+) T cells in the presence of methamphetamine in a dose-dependent manner. Our results demonstrate that methamphetamine had a minimal effect on HIV-1 replication at concentrations of 1 to 50 μmol/L. However, at concentrations >100 μmol/L, it inhibited HIV-1 replication in a dose-dependent manner. We also discovered that methamphetamine up-regulated the cellular anti-HIV-1 microRNAs (miR-125b, miR-150, and miR-28-5p) in CD4(+) T cells. Knockdown experiments illustrated that up-regulation of the anti-HIV miRNAs inhibited HIV-1 replication. These results are contrary to the paradigm that methamphetamine accentuates HIV-1 pathogenesis by increasing HIV-1 replication. Therefore, our findings underline the complex interaction between drug use and HIV-1 and necessitate comprehensive understanding of the effects of methamphetamine on HIV-1 pathogenesis. Copyright © 2014 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  2. HIV-1 proteins dysregulate motivational processes and dopamine circuitry.

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    Bertrand, Sarah J; Mactutus, Charles F; Harrod, Steven B; Moran, Landhing M; Booze, Rosemarie M

    2018-05-18

    Motivational alterations, such as apathy, in HIV-1+ individuals are associated with decreased performance on tasks involving frontal-subcortical circuitry. We used the HIV-1 transgenic (Tg) rat to assess effect of long-term HIV-1 protein exposure on motivated behavior using sucrose (1-30%, w/v) and cocaine (0.01-1.0 mg/kg/infusion) maintained responding with fixed-ratio (FR) and progressive-ratio (PR) schedules of reinforcement. For sucrose-reinforced responding, HIV-1 Tg rats displayed no change in EC 50 relative to controls, suggesting no change in sucrose reinforcement but had a downward shifted concentration-response curves, suggesting a decrease in response vigor. Cocaine-maintained responding was attenuated in HIV-1 Tg rats (FR1 0.33 mg/kg/infusion and PR 1.0 mg/kg/infusion). Dose-response tests (PR) revealed that HIV-1 Tg animals responded significantly less than F344 control rats and failed to earn significantly more infusions of cocaine as the unit dose increased. When choosing between cocaine and sucrose, control rats initially chose sucrose but with time shifted to a cocaine preference. In contrast, HIV-1 disrupted choice behaviors. DAT function was altered in the striatum of HIV-1 Tg rats; however, prior cocaine self-administration produced a unique effect on dopamine homeostasis in the HIV-1 Tg striatum. These findings of altered goal directed behaviors may determine neurobiological mechanisms of apathy in HIV-1+ patients.

  3. Interferon-α Subtypes in an Ex Vivo Model of Acute HIV-1 Infection: Expression, Potency and Effector Mechanisms.

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    Michael S Harper

    Full Text Available HIV-1 is transmitted primarily across mucosal surfaces and rapidly spreads within the intestinal mucosa during acute infection. The type I interferons (IFNs likely serve as a first line of defense, but the relative expression and antiviral properties of the 12 IFNα subtypes against HIV-1 infection of mucosal tissues remain unknown. Here, we evaluated the expression of all IFNα subtypes in HIV-1-exposed plasmacytoid dendritic cells by next-generation sequencing. We then determined the relative antiviral potency of each IFNα subtype ex vivo using the human intestinal Lamina Propria Aggregate Culture model. IFNα subtype transcripts from the centromeric half of the IFNA gene complex were highly expressed in pDCs following HIV-1 exposure. There was an inverse relationship between IFNA subtype expression and potency. IFNα8, IFNα6 and IFNα14 were the most potent in restricting HIV-1 infection. IFNα2, the clinically-approved subtype, and IFNα1 were both highly expressed but exhibited relatively weak antiviral activity. The relative potencies correlated with binding affinity to the type I IFN receptor and the induction levels of HIV-1 restriction factors Mx2 and Tetherin/BST-2 but not APOBEC3G, F and D. However, despite the lack of APOBEC3 transcriptional induction, the higher relative potency of IFNα8 and IFNα14 correlated with stronger inhibition of virion infectivity, which is linked to deaminase-independent APOBEC3 restriction activity. By contrast, both potent (IFNα8 and weak (IFNα1 subtypes significantly induced HIV-1 GG-to-AG hypermutation. The results unravel non-redundant functions of the IFNα subtypes against HIV-1 infection, with strong implications for HIV-1 mucosal immunity, viral evolution and IFNα-based functional cure strategies.

  4. Comprehensive Characterization of HIV-1 Molecular Epidemiology and Demographic History in the Brazilian Region Most Heavily Affected by AIDS.

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    Gräf, Tiago; Machado Fritsch, Hegger; de Medeiros, Rúbia Marília; Maletich Junqueira, Dennis; Esteves de Matos Almeida, Sabrina; Pinto, Aguinaldo Roberto

    2016-09-15

    The high incidence of AIDS cases and the dominance of HIV-1 subtype C infections are two features that distinguish the HIV-1 epidemic in the two southernmost Brazilian states (Rio Grande do Sul [RS] and Santa Catarina [SC]) from the epidemic in other parts of the country. Nevertheless, previous studies on HIV molecular epidemiology were conducted mainly in capital cities, and a more comprehensive understanding of factors driving this unique epidemic in Brazil is necessary. Blood samples were collected from individuals in 13 municipalities in the Brazilian southern region. HIV-1 env and pol genes were submitted to phylogenetic analyses for assignment of subtype, and viral population phylodynamics were reconstructed by applying Skygrid and logistic coalescent models in a Bayesian analysis. A high prevalence of subtype C was observed in all sampled locations; however, an increased frequency of recombinant strains was found in RS, with evidence for new circulating forms (CRFs). In the SC state, subtype B and C epidemics were associated with distinct exposure groups. Although logistic models estimated similar growth rates for HIV-1 subtype C (HIV-1C) and HIV-1B, a Skygrid plot reveals that the former epidemic has been expanding for a longer time. Our results highlight a consistent expansion of HIV-1C in south Brazil, and we also discuss how heterosexual and men who have sex with men (MSM) transmission chains might have impacted the current prevalence of HIV-1 subtypes in this region. The AIDS epidemic in south Brazil is expanding rapidly, but the circumstances driving this condition are not well known. A high prevalence of HIV-1 subtype C was reported in the capital cities of this region, in contrast to the subtype B dominance in the rest of the country. This study sought to comparatively investigate the HIV-1 subtype B and C epidemics by sampling individuals from several cities in the two states with the highest AIDS incidences in Brazil. Our analyses showed distinct

  5. Bacterial vaginosis associated with increased risk of female-to-male HIV-1 transmission: a prospective cohort analysis among African couples.

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    Craig R Cohen

    Full Text Available Bacterial vaginosis (BV, a disruption of the normal vaginal flora, has been associated with a 60% increased risk of HIV-1 acquisition in women and higher concentration of HIV-1 RNA in the genital tract of HIV-1-infected women. However, whether BV, which is present in up to half of African HIV-1-infected women, is associated with an increase in HIV-1 transmission to male partners has not been assessed in previous studies.We assessed the association between BV on female-to-male HIV-1 transmission risk in a prospective study of 2,236 HIV-1-seropositive women and their HIV-1 uninfected male partners from seven African countries from a randomized placebo-controlled trial that enrolled heterosexual African adults who were seropositive for both HIV-1 and herpes simplex virus (HSV-2, and their HIV-1-seronegative partners. Participants were followed for up to 24 months; every three months, vaginal swabs were obtained from female partners for Gram stain and male partners were tested for HIV-1. BV and normal vaginal flora were defined as a Nugent score of 7-10 and 0-3, respectively. To reduce misclassification, HIV-1 sequence analysis of viruses from seroconverters and their partners was performed to determine linkage of HIV-1 transmissions. Overall, 50 incident HIV-1 infections occurred in men in which the HIV-1-infected female partner had an evaluable vaginal Gram stain. HIV-1 incidence in men whose HIV-1-infected female partners had BV was 2.91 versus 0.76 per 100 person-years in men whose female partners had normal vaginal flora (hazard ratio 3.62, 95% CI 1.74-7.52. After controlling for sociodemographic factors, sexual behavior, male circumcision, sexually transmitted infections, pregnancy, and plasma HIV-1 RNA levels in female partners, BV was associated with a greater than 3-fold increased risk of female-to-male HIV-1 transmission (adjusted hazard ratio 3.17, 95% CI 1.37-7.33.This study identified an association between BV and increased risk of HIV

  6. Sensitivity of the ViroSeq HIV-1 Genotyping System for Detection of the K103N Resistance Mutation in HIV-1 Subtypes A, C, and D

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    Church, Jessica D.; Jones, Dana; Flys, Tamara; Hoover, Donald; Marlowe, Natalia; Chen, Shu; Shi, Chanjuan; Eshleman, James R.; Guay, Laura A.; Jackson, J. Brooks; Kumwenda, Newton; Taha, Taha E.; Eshleman, Susan H.

    2006-01-01

    The US Food and Drug Administration-cleared ViroSeq HIV-1 Genotyping System (ViroSeq) and other population sequencing-based human immunodeficiency virus type 1 (HIV-1) genotyping methods detect antiretroviral drug resistance mutations present in the major viral population of a test sample. These assays also detect some mutations in viral variants that are present as mixtures. We compared detection of the K103N nevirapine resistance mutation using ViroSeq and a sensitive, quantitative point mutation assay, LigAmp. The LigAmp assay measured the percentage of K103N-containing variants in the viral population (percentage of K103N). We analyzed 305 samples with HIV-1 subtypes A, C, and D collected from African women after nevirapine administration. ViroSeq detected K103N in 100% of samples with >20% K103N, 77.8% of samples with 10 to 20% K103N, 71.4% of samples with 5 to 10% K103N, and 16.9% of samples with 1 to 5% K103N. The sensitivity of ViroSeq for detection of K103N was similar for subtypes A, C, and D. These data indicate that the ViroSeq system reliably detects the K103N mutation at levels above 20% and frequently detects the mutation at lower levels. Further studies are needed to compare the sensitivity of different assays for detection of HIV-1 drug resistance mutations and to determine the clinical relevance of HIV-1 minority variants. PMID:16931582

  7. Antibodyomics: bioinformatics technologies for understanding B-cell immunity to HIV-1.

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    Kwong, Peter D; Chuang, Gwo-Yu; DeKosky, Brandon J; Gindin, Tatyana; Georgiev, Ivelin S; Lemmin, Thomas; Schramm, Chaim A; Sheng, Zizhang; Soto, Cinque; Yang, An-Suei; Mascola, John R; Shapiro, Lawrence

    2017-01-01

    Numerous antibodies have been identified from HIV-1-infected donors that neutralize diverse strains of HIV-1. These antibodies may provide the basis for a B cell-mediated HIV-1 vaccine. However, it has been unclear how to elicit similar antibodies by vaccination. To address this issue, we have undertaken an informatics-based approach to understand the genetic and immunologic processes controlling the development of HIV-1-neutralizing antibodies. As DNA sequencing comprises the fastest growing database of biological information, we focused on incorporating next-generation sequencing of B-cell transcripts to determine the origin, maturation pathway, and prevalence of broadly neutralizing antibody lineages (Antibodyomics1, 2, 4, and 6). We also incorporated large-scale robotic analyses of serum neutralization to identify and quantify neutralizing antibodies in donor cohorts (Antibodyomics3). Statistical analyses furnish another layer of insight (Antibodyomics5), with physical characteristics of antibodies and their targets through molecular dynamics simulations (Antibodyomics7) and free energy perturbation analyses (Antibodyomics8) providing information-rich output. Functional interrogation of individual antibodies (Antibodyomics9) and synthetic antibody libraries (Antibodyomics10) also yields multi-dimensional data by which to understand and improve antibodies. Antibodyomics, described here, thus comprise resolution-enhancing tools, which collectively embody an information-driven discovery engine aimed toward the development of effective B cell-based vaccines. © 2017 The Authors. Immunological Reviews published by John Wiley & Sons Ltd.

  8. A Directed Molecular Evolution Approach to Improved Immunogenicity of the HIV-1 Envelope Glycoprotein

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    Du, Sean X.; Xu, Li; Zhang, Wenge; Tang, Susan; Boenig, Rebecca I.; Chen, Helen; Mariano, Ellaine B.; Zwick, Michael B.; Parren, Paul W. H. I.; Burton, Dennis R.; Wrin, Terri; Petropoulos, Christos J.; Ballantyne, John A.; Chambers, Michael; Whalen, Robert G.

    2011-01-01

    A prophylactic vaccine is needed to slow the spread of HIV-1 infection. Optimization of the wild-type envelope glycoproteins to create immunogens that can elicit effective neutralizing antibodies is a high priority. Starting with ten genes encoding subtype B HIV-1 gp120 envelope glycoproteins and using in vitro homologous DNA recombination, we created chimeric gp120 variants that were screened for their ability to bind neutralizing monoclonal antibodies. Hundreds of variants were identified with novel antigenic phenotypes that exhibit considerable sequence diversity. Immunization of rabbits with these gp120 variants demonstrated that the majority can induce neutralizing antibodies to HIV-1. One novel variant, called ST-008, induced significantly improved neutralizing antibody responses when assayed against a large panel of primary HIV-1 isolates. Further study of various deletion constructs of ST-008 showed that the enhanced immunogenicity results from a combination of effective DNA priming, an enhanced V3-based response, and an improved response to the constant backbone sequences. PMID:21738594

  9. Rapid turnover of 2-LTR HIV-1 DNA during early stage of highly active antiretroviral therapy.

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    Weijun Zhu

    Full Text Available BACKGROUND: Despite prolonged treatment with highly active antiretroviral therapy (HAART, the infectious HIV-1 continues to replicate and resides latently in the resting memory CD4+ T lymphocytes, which blocks the eradication of HIV-1. The viral persistence of HIV-1 is mainly caused by its proviral DNA being either linear nonintegrated, circular nonintegrated, or integrated. Previous reports have largely focused on the dynamics of HIV-1 DNA from the samples collected with relatively long time intervals during the process of disease and HAART treatment, which may have missed the intricate changes during the intervals in early treatment. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we investigated the dynamics of HIV-1 DNA in patients during the early phase of HARRT treatment. Using optimized real time PCR, we observed significant changes in 2-LTR during the first 12-week of treatment, while total and integrated HIV-1 DNA remained stable. The doubling time and half-life of 2-LTR were not correlated with the baseline and the rate of changes in plasma viral load and various CD4+ T-cell populations. Longitudinal analyses on 2-LTR sequences and plasma lipopolysaccharide (LPS levels did not reveal any significant changes in the same treatment period. CONCLUSIONS/SIGNIFICANCE: Our study revealed the rapid changes in 2-LTR concentration in a relatively large number of patients during the early HAART treatment. The rapid changes indicate the rapid infusion and clearance of cells bearing 2-LTR in the peripheral blood. Those changes are not expected to be caused by the blocking of viral integration, as our study did not include the integrase inhibitor raltegravir. Our study helps better understand the dynamics of HIV-DNA and its potential role as a biomarker for the diseases and for the treatment efficacy of HAART.

  10. Phylogenetic characteristics of HIV-1 among travelers entering China from Myanmar: A retrospective study.

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    Zhang, Li; Wang, Binhui; Liang, Yaobo; Feng, Yue; Dong, Shuwei; Wang, Yajuan; Li, Yaping; Zhang, A-Mei; Liu, Li; Qin, Weihong; Xia, Xueshan

    2017-08-01

    Due to the open policy of the Chinese government, a large number of Burmese individuals enter China at land ports in Yunnan province for travel or business. However, the situation of HIV-1 infection and its phylogenetic characteristics among these travelers remains unclear, which is a potential threat to public health. From January 2003 to December 2012, a total of 1,961 travelers were detected to be positive for HIV-1 infection at land ports between Myanmar and Yunnan province, China. From 1153 (58.8%) Burmese of them, we randomly collected 489 serum samples for HIV-1 subtype/recombinant analysis. Based on successfully obtained 223 gag-RT sequences, 187 of them were genotyped as 2 subtypes and 3 CRFs. CRF01_AE was showed to be the most prevalent genotype (54.3%), followed by subtypes C (13.5%) and B (10.8%). Notably, CRF07_BC (1.3%) and CRF08_BC (4.0%) were mainly distributed in travelers from Shan state and Kachin (91.7%, 11/12), but was not found in travelers from the capital city of Yangon (0/16). Additionally, there were 36 samples (16.1%) were preliminary determined as unique recombinant forms (URFs). The higher HIV-1 infection among entering travelers from Myanmar and its diverse and complex genotypes distribution suggest this bridge population may facilitate the transmission of HIV-1. It is necessary to have the strict monitoring on this population for prevention of HIV-1 cross-border transmission. © 2017 Wiley Periodicals, Inc.

  11. Interactions between the HIV-1 Unspliced mRNA and Host mRNA Decay Machineries

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    Daniela Toro-Ascuy

    2016-11-01

    Full Text Available The human immunodeficiency virus type-1 (HIV-1 unspliced transcript is used both as mRNA for the synthesis of structural proteins and as the packaged genome. Given the presence of retained introns and instability AU-rich sequences, this viral transcript is normally retained and degraded in the nucleus of host cells unless the viral protein REV is present. As such, the stability of the HIV-1 unspliced mRNA must be particularly controlled in the nucleus and the cytoplasm in order to ensure proper levels of this viral mRNA for translation and viral particle formation. During its journey, the HIV-1 unspliced mRNA assembles into highly specific messenger ribonucleoproteins (mRNPs containing many different host proteins, amongst which are well-known regulators of cytoplasmic mRNA decay pathways such as up-frameshift suppressor 1 homolog (UPF1, Staufen double-stranded RNA binding protein 1/2 (STAU1/2, or components of miRNA-induced silencing complex (miRISC and processing bodies (PBs. More recently, the HIV-1 unspliced mRNA was shown to contain N6-methyladenosine (m6A, allowing the recruitment of YTH N6-methyladenosine RNA binding protein 2 (YTHDF2, an m6A reader host protein involved in mRNA decay. Interestingly, these host proteins involved in mRNA decay were shown to play positive roles in viral gene expression and viral particle assembly, suggesting that HIV-1 interacts with mRNA decay components to successfully accomplish viral replication. This review summarizes the state of the art in terms of the interactions between HIV-1 unspliced mRNA and components of different host mRNA decay machineries.

  12. Clonal expansion of genome-intact HIV-1 in functionally polarized Th1 CD4+ T cells.

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    Lee, Guinevere Q; Orlova-Fink, Nina; Einkauf, Kevin; Chowdhury, Fatema Z; Sun, Xiaoming; Harrington, Sean; Kuo, Hsiao-Hsuan; Hua, Stephane; Chen, Hsiao-Rong; Ouyang, Zhengyu; Reddy, Kavidha; Dong, Krista; Ndung'u, Thumbi; Walker, Bruce D; Rosenberg, Eric S; Yu, Xu G; Lichterfeld, Mathias

    2017-06-30

    HIV-1 causes a chronic, incurable disease due to its persistence in CD4+ T cells that contain replication-competent provirus, but exhibit little or no active viral gene expression and effectively resist combination antiretroviral therapy (cART). These latently infected T cells represent an extremely small proportion of all circulating CD4+ T cells but possess a remarkable long-term stability and typically persist throughout life, for reasons that are not fully understood. Here we performed massive single-genome, near-full-length next-generation sequencing of HIV-1 DNA derived from unfractionated peripheral blood mononuclear cells, ex vivo-isolated CD4+ T cells, and subsets of functionally polarized memory CD4+ T cells. This approach identified multiple sets of independent, near-full-length proviral sequences from cART-treated individuals that were completely identical, consistent with clonal expansion of CD4+ T cells harboring intact HIV-1. Intact, near-full-genome HIV-1 DNA sequences that were derived from such clonally expanded CD4+ T cells constituted 62% of all analyzed genome-intact sequences in memory CD4 T cells, were preferentially observed in Th1-polarized cells, were longitudinally detected over a duration of up to 5 years, and were fully replication- and infection-competent. Together, these data suggest that clonal proliferation of Th1-polarized CD4+ T cells encoding for intact HIV-1 represents a driving force for stabilizing the pool of latently infected CD4+ T cells.

  13. Cloaked similarity between HIV-1 and SARS-CoV suggests an anti-SARS strategy

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    Kliger Yossef

    2003-09-01

    Full Text Available Abstract Background Severe acute respiratory syndrome (SARS is a febrile respiratory illness. The disease has been etiologically linked to a novel coronavirus that has been named the SARS-associated coronavirus (SARS-CoV, whose genome was recently sequenced. Since it is a member of the Coronaviridae, its spike protein (S2 is believed to play a central role in viral entry by facilitating fusion between the viral and host cell membranes. The protein responsible for viral-induced membrane fusion of HIV-1 (gp41 differs in length, and has no sequence homology with S2. Results Sequence analysis reveals that the two viral proteins share the sequence motifs that construct their active conformation. These include (1 an N-terminal leucine/isoleucine zipper-like sequence, and (2 a C-terminal heptad repeat located upstream of (3 an aromatic residue-rich region juxtaposed to the (4 transmembrane segment. Conclusions This study points to a similar mode of action for the two viral proteins, suggesting that anti-viral strategy that targets the viral-induced membrane fusion step can be adopted from HIV-1 to SARS-CoV. Recently the FDA approved Enfuvirtide, a synthetic peptide corresponding to the C-terminal heptad repeat of HIV-1 gp41, as an anti-AIDS agent. Enfuvirtide and C34, another anti HIV-1 peptide, exert their inhibitory activity by binding to a leucine/isoleucine zipper-like sequence in gp41, thus inhibiting a conformational change of gp41 required for its activation. We suggest that peptides corresponding to the C-terminal heptad repeat of the S2 protein may serve as inhibitors for SARS-CoV entry.

  14. Novel HIV-1 recombinants spreading across multiple risk groups in the United Kingdom: the identification and phylogeography of Circulating Recombinant Form (CRF 50_A1D.

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    Geraldine M Foster

    Full Text Available An increase in non-B HIV-1 infections among men who have sex with men (MSM in the United Kingdom (UK has created opportunities for novel recombinants to arise and become established. We used molecular mapping to characterize the importance of such recombinants to the UK HIV epidemic, in order to gain insights into transmission dynamics that can inform control strategies.A total of 55,556 pol (reverse transcriptase and protease sequences in the UK HIV Drug Resistance Database were analyzed using Subtype Classification Using Evolutionary Algorithms (SCUEAL. Overall 72 patients shared the same A1/D recombination breakpoint in pol, comprising predominantly MSM but also heterosexuals and injecting drug users (IDUs. In six MSM, full-length single genome amplification of plasma HIV-1 RNA was performed in order to characterize the A1/D recombinant. Subtypes and recombination breakpoints were identified using sliding window and jumping profile hidden markov model approaches. Global maximum likelihood trees of gag, pol and env genes were drawn using FastTree version 2.1. Five of the six strains showed the same novel A1/D recombinant (8 breakpoints, which has been classified as CRF50_A1D. The sixth strain showed a complex CRF50_A1D/B/U structure. Divergence dates and phylogeographic inferences were determined using Bayesian Evolutionary Analysis using Sampling Trees (BEAST. This estimated that CRF50_A1D emerged in the UK around 1992 in MSM, with subsequent transmissions to heterosexuals and IDUs. Analysis of CRF50_A1D/B/U demonstrated that around the year 2000 CRF50_A1D underwent recombination with a subtype B strain.We report the identification of CRF50_A1D, a novel circulating recombinant that emerged in UK MSM around 1992, with subsequent onward transmission to heterosexuals and IDUs, and more recent recombination with subtype B. These findings highlight the changing dynamics of HIV transmission in the UK and the converging of the two previously

  15. Novel tetra-peptide insertion in Gag-p6 ALIX-binding motif in HIV-1 subtype C associated with protease inhibitor failure

    Science.gov (United States)

    Neogi, Ujjwal; RAO, Shwetha D; BONTELL, Irene; VERHEYEN, Jens; RAO, Vasudev R; GORE, Sagar C; SONI, Neelesh; SHET, Anita; SCHÜLTER, Eugen; EKSTRAND, Maria L.; WONDWOSSEN, Amogne; KAISER, Rolf; MADHUSUDHAN, Mallur S.; PRASAD, Vinayaka R; SONNERBORG, Anders

    2014-01-01

    A novel tetra-peptide insertion was identified in Gag-p6 ALIX-binding region which is appears in protease inhibitor (PI) failure Indian HIV-1C sequences (Odds Ratio 17.1, p<0.001) but naturally present in half of untreated Ethiopian sequences. The insertion will probably restore the ALIX mediated virus release pathway, which is lacking in HIV-1C. The clinical importance of such insertion need to be evaluated in HIV-1C dominating regions were PI-drugs are being scaled up as second line treatment options. PMID:25102091

  16. BI-2 destabilizes HIV-1 cores during infection and Prevents Binding of CPSF6 to the HIV-1 Capsid.

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    Fricke, Thomas; Buffone, Cindy; Opp, Silvana; Valle-Casuso, Jose; Diaz-Griffero, Felipe

    2014-12-11

    The recently discovered small-molecule BI-2 potently blocks HIV-1 infection. BI-2 binds to the N-terminal domain of HIV-1 capsid. BI-2 utilizes the same capsid pocket used by the small molecule PF74. Although both drugs bind to the same pocket, it has been proposed that BI-2 uses a different mechanism to block HIV-1 infection when compared to PF74. This work demonstrates that BI-2 destabilizes the HIV-1 core during infection, and prevents the binding of the cellular factor CPSF6 to the HIV-1 core. Overall this short-form paper suggests that BI-2 is using a similar mechanism to the one used by PF74 to block HIV-1 infection.

  17. The Origin and Evolutionary History of HIV-1 Subtype C in Senegal

    Science.gov (United States)

    Jung, Matthieu; Leye, Nafissatou; Vidal, Nicole; Fargette, Denis; Diop, Halimatou; Toure Kane, Coumba; Gascuel, Olivier; Peeters, Martine

    2012-01-01

    Background The classification of HIV-1 strains in subtypes and Circulating Recombinant Forms (CRFs) has helped in tracking the course of the HIV pandemic. In Senegal, which is located at the tip of West Africa, CRF02_AG predominates in the general population and Female Sex Workers (FSWs). In contrast, 40% of Men having Sex with Men (MSM) in Senegal are infected with subtype C. In this study we analyzed the geographical origins and introduction dates of HIV-1 C in Senegal in order to better understand the evolutionary history of this subtype, which predominates today in the MSM population Methodology/Principal Findings We used a combination of phylogenetic analyses and a Bayesian coalescent-based approach, to study the phylogenetic relationships in pol of 56 subtype C isolates from Senegal with 3,025 subtype C strains that were sampled worldwide. Our analysis shows a significantly well supported cluster which contains all subtype C strains that circulate among MSM in Senegal. The MSM cluster and other strains from Senegal are widely dispersed among the different subclusters of African HIV-1 C strains, suggesting multiple introductions of subtype C in Senegal from many different southern and east African countries. More detailed analyses show that HIV-1 C strains from MSM are more closely related to those from southern Africa. The estimated date of the MRCA of subtype C in the MSM population in Senegal is estimated to be in the early 80's. Conclusions/Significance Our evolutionary reconstructions suggest that multiple subtype C viruses with a common ancestor originating in the early 1970s entered Senegal. There was only one efficient spread in the MSM population, which most likely resulted from a single introduction, underlining the importance of high-risk behavior in spread of viruses. PMID:22470456

  18. Evaluation of a cost effective in-house method for HIV-1 drug resistance genotyping using plasma samples.

    Directory of Open Access Journals (Sweden)

    Devidas N Chaturbhuj

    Full Text Available OBJECTIVES: Validation of a cost effective in-house method for HIV-1 drug resistance genotyping using plasma samples. DESIGN: The validation includes the establishment of analytical performance characteristics such as accuracy, reproducibility, precision and sensitivity. METHODS: The accuracy was assessed by comparing 26 paired Virological Quality Assessment (VQA proficiency testing panel sequences generated by in-house and ViroSeq Genotyping System 2.0 (Celera Diagnostics, US as a gold standard. The reproducibility and precision were carried out on five samples with five replicates representing multiple HIV-1 subtypes (A, B, C and resistance patterns. The amplification sensitivity was evaluated on HIV-1 positive plasma samples (n = 88 with known viral loads ranges from 1000-1.8 million RNA copies/ml. RESULTS: Comparison of the nucleotide sequences generated by ViroSeq and in-house method showed 99.41±0.46 and 99.68±0.35% mean nucleotide and amino acid identity respectively. Out of 135 Stanford HIVdb listed HIV-1 drug resistance mutations, partial discordance was observed at 15 positions and complete discordance was absent. The reproducibility and precision study showed high nucleotide sequence identities i.e. 99.88±0.10 and 99.82±0.20 respectively. The in-house method showed 100% analytical sensitivity on the samples with HIV-1 viral load >1000 RNA copies/ml. The cost of running the in-house method is only 50% of that for ViroSeq method (112$ vs 300$, thus making it cost effective. CONCLUSIONS: The validated cost effective in-house method may be used to collect surveillance data on the emergence and transmission of HIV-1 drug resistance in resource limited countries. Moreover, the wide applications of a cost effective and validated in-house method for HIV-1 drug resistance testing will facilitate the decision making for the appropriate management of HIV infected patients.

  19. HIV-1, HTLV-I and the interleukin-2 receptor: insights into transcriptional control.

    Science.gov (United States)

    Böhnlein, E; Lowenthal, J W; Wano, Y; Franza, B R; Ballard, D W; Greene, W C

    1989-01-01

    In this study, we present direct evidence for the binding of the inducible cellular protein, HIVEN86A, to a 12-bp element present in the IL-2R alpha promoter. This element shares significant sequence similarity with the NF-kappa B binding sites present in the HIV-1 and kappa immunoglobulin enhancers. Transient transfection studies indicate that this kappa B element is both necessary and sufficient to confer tax or mitogen inducibility to a heterologous promoter. As summarized schematically in Fig. 5, the findings suggest that the HIVEN86A protein may play a central role in the activation of cellular genes required for T-cell growth, specifically the IL-2R alpha gene. In addition, the induced HIVEN86A protein also binds to a similar sequence present in the HIV-1 LTR leading to enhanced viral gene expression and ultimately T-cell death. Thus, mitogen activation of the HIV-1 LTR appears to involve the same inducible transcription factor(s) that normally regulates IL-2R alpha gene expression and T-cell growth. These findings further underscore the importance of the state of T-cell activation in the regulation of HIV-1 replication. Our results also demonstrate that HIVEN86A is induced by the tax protein of HTLV-I. Thus, in HTLV-I infected cells, normally the tight control of the transient expression of the IL-2R alpha gene is lost. The constitutive high-level display of IL-2 receptors may play a role in leukemic transformation mediated by HTLV-I (ATL). Apparently by the same mechanism, the tax protein also activates the HIV-1 LTR through the induction of HIVEN86A.(ABSTRACT TRUNCATED AT 250 WORDS)

  20. Short-Term Dynamic and Local Epidemiological Trends in the South American HIV-1B Epidemic.

    Directory of Open Access Journals (Sweden)

    Dennis Maletich Junqueira

    Full Text Available The human displacement and sexual behavior are the main factors driving the HIV-1 pandemic to the current profile. The intrinsic structure of the HIV transmission among different individuals has valuable importance for the understanding of the epidemic and for the public health response. The aim of this study was to characterize the HIV-1 subtype B (HIV-1B epidemic in South America through the identification of transmission links and infer trends about geographical patterns and median time of transmission between individuals. Sequences of the protease and reverse transcriptase coding regions from 4,810 individuals were selected from GenBank. Maximum likelihood phylogenies were inferred and submitted to ClusterPicker to identify transmission links. Bayesian analyses were applied only for clusters including ≥5 dated samples in order to estimate the median maximum inter-transmission interval. This study analyzed sequences sampled from 12 South American countries, from individuals of different exposure categories, under different antiretroviral profiles, and from a wide period of time (1989-2013. Continentally, Brazil, Argentina and Venezuela were revealed important sites for the spread of HIV-1B among countries inside South America. Of note, from all the clusters identified about 70% of the HIV-1B infections are primarily occurring among individuals living in the same geographic region. In addition, these transmissions seem to occur early after the infection of an individual, taking in average 2.39 years (95% CI 1.48-3.30 to succeed. Homosexual/Bisexual individuals transmit the virus as quickly as almost half time of that estimated for the general population sampled here. Public health services can be broadly benefitted from this kind of information whether to focus on specific programs of response to the epidemic whether as guiding of prevention campaigns to specific risk groups.

  1. Short-Term Dynamic and Local Epidemiological Trends in the South American HIV-1B Epidemic.

    Science.gov (United States)

    Junqueira, Dennis Maletich; de Medeiros, Rubia Marília; Gräf, Tiago; Almeida, Sabrina Esteves de Matos

    2016-01-01

    The human displacement and sexual behavior are the main factors driving the HIV-1 pandemic to the current profile. The intrinsic structure of the HIV transmission among different individuals has valuable importance for the understanding of the epidemic and for the public health response. The aim of this study was to characterize the HIV-1 subtype B (HIV-1B) epidemic in South America through the identification of transmission links and infer trends about geographical patterns and median time of transmission between individuals. Sequences of the protease and reverse transcriptase coding regions from 4,810 individuals were selected from GenBank. Maximum likelihood phylogenies were inferred and submitted to ClusterPicker to identify transmission links. Bayesian analyses were applied only for clusters including ≥5 dated samples in order to estimate the median maximum inter-transmission interval. This study analyzed sequences sampled from 12 South American countries, from individuals of different exposure categories, under different antiretroviral profiles, and from a wide period of time (1989-2013). Continentally, Brazil, Argentina and Venezuela were revealed important sites for the spread of HIV-1B among countries inside South America. Of note, from all the clusters identified about 70% of the HIV-1B infections are primarily occurring among individuals living in the same geographic region. In addition, these transmissions seem to occur early after the infection of an individual, taking in average 2.39 years (95% CI 1.48-3.30) to succeed. Homosexual/Bisexual individuals transmit the virus as quickly as almost half time of that estimated for the general population sampled here. Public health services can be broadly benefitted from this kind of information whether to focus on specific programs of response to the epidemic whether as guiding of prevention campaigns to specific risk groups.

  2. Enhanced anti-HIV-1 activity of G-quadruplexes comprising locked nucleic acids and intercalating nucleic acids

    DEFF Research Database (Denmark)

    Pedersen, Erik Bjerregaard; Nielsen, Jakob Toudahl; Nielsen, Claus

    2011-01-01

    Two G-quadruplex forming sequences, 50-TGGGAG and the 17-mer sequence T30177, which exhibit anti-HIV-1 activity on cell lines, were modified using either locked nucleic acids (LNA) or via insertions of (R)-1-O-(pyren-1-ylmethyl)glycerol (intercalating nucleic acid, INA) or (R)-1-O-[4-(1......-pyrenylethynyl)phenylmethyl]glycerol (twisted intercalating nucleic acid, TINA). Incorporation of LNA or INA/TINA monomers provide as much as 8-fold improvement of anti-HIV-1 activity. We demonstrate for the first time a detailed analysis of the effect the incorporation of INA/TINA monomers in quadruplex forming...

  3. The anti-HIV-1 effect of scutellarin

    International Nuclear Information System (INIS)

    Zhang Gaohong; Wang Qian; Chen Jijun; Zhang Xuemei; Tam, S.-C.; Zheng Yongtang

    2005-01-01

    Scutellarin was purified from the plant Erigeron breviscapus (Vant.) Hand.-Mazz. The activity against 3 strains of human immunodeficiency virus (HIV) was determined in vitro in this study. These were laboratory-derived virus (HIV-1 IIIB ), drug-resistant virus (HIV-1 74V ), and low-passage clinical isolated virus (HIV-1 KM018 ). From syncytia inhibition study, the EC 50 of scutellarin against HIV-1 IIIB direct infection in C8166 cells was 26 μM with a therapeutic index of 36. When the mode of infection changed from acute infection to cell-to-cell infection, this compound became even more potent and the EC 50 reduced to 15 μM. This suggested that cell fusion might be affected by this compound. By comparing the inhibitory effects on p24 antigen, scutellarin was also found to be active against HIV-1 74V (EC 50 253 μM) and HIV-1 KM018 (EC 50 136 μM) infection with significant difference in potency. The mechanism of its action was also explored in this study. At a concentration of 433 μM, scutellarin inhibited 48% of the cell free recombinant HIV-1 RT activity. It also caused 82% inhibition of HIV-1 particle attachment and 45% inhibition of fusion at the concentrations of 54 μM. In summary, scutellarin was found to inhibit several strains of HIV-1 replication with different potencies. It appeared to inhibit HIV-1 RT activity, HIV-1 particle attachment and cell fusion. These are essential activities for viral transmission and replication

  4. Preferential susceptibility of Th9 and Th2 CD4+ T cells to X4-tropic HIV-1 infection.

    Science.gov (United States)

    Orlova-Fink, Nina; Chowdhury, Fatema Z; Sun, Xiaoming; Harrington, Sean; Rosenberg, Eric S; Yu, Xu G; Lichterfeld, Mathias

    2017-10-23

    The functional polarization of CD4 T cells determines their antimicrobial effector profile, but may also impact the susceptibility to infection with HIV-1. Here, we analyzed the susceptibility of CD4 T cells with different functional polarization to infection with X4 and R5-tropic HIV-1. CD4 T cells with a Th1, Th2, Th17, and Th9 polarization were subjected to in-vitro infection assays with X4, R5, or vesicular stomatitis virus-G protein-pseudotyped HIV-1. In addition, we sorted differentially polarized CD4 T-cell subsets from individuals treated with antiretroviral therapy and analyzed the tropism of viral env sequences. Th9-polarized CD4 T cells and, to a lesser extent, Th2-polarized CD4 T cells expressed higher surface levels of CXCR4, and are more permissive to X4-tropic infection in vitro. In contrast, Th1 and Th17 CD4 T cells exhibited stronger surface expression of CCR5, and were more susceptible to infection with R5-tropic viruses. Correspondingly, the distribution of X4-tropic viral sequences in antiretroviral therapy-treated HIV-1-infected patients was biased toward Th9/Th2 cells, whereas R5-tropic sequences were more frequently observed in Th17 cells. CD4 T-cell polarization is associated with a distinct susceptibility to X4 and R5-tropic HIV-1 infection.

  5. Rapid selection of escape mutants by the first CD8 T cell responses in acute HIV-1 infection

    Energy Technology Data Exchange (ETDEWEB)

    Korber, Bette Tina Marie [Los Alamos National Laboratory

    2008-01-01

    The recent failure of a vaccine that primes T cell responses to control primary HIV-1 infection has raised doubts about the role of CD8+ T cells in early HIV-1 infection. We studied four patients who were identified shortly after HIV-1 infection and before seroconversion. In each patient there was very rapid selection of multiple HIV-1 escape mutants in the transmitted virus by CD8 T cells, including examples of complete fixation of non-synonymous substitutions within 2 weeks. Sequencing by single genome amplification suggested that the high rate of virus replication in acute infection gave a selective advantage to virus molecules that contained simultaneous and gained sequential T cell escape mutations. These observations show that whilst early HIV-1 specific CD8 T cells can act against virus, rapid escape means that these T cell responses are unlikely to benefit the patient and may in part explain why current HIV-1 T cell vaccines may not be protective.

  6. Editing of HIV-1 RNA by the double-stranded RNA deaminase ADAR1 stimulates viral infection

    Science.gov (United States)

    Doria, Margherita; Neri, Francesca; Gallo, Angela; Farace, Maria Giulia; Michienzi, Alessandro

    2009-01-01

    Adenosine deaminases that act on dsRNA (ADARs) are enzymes that target double-stranded regions of RNA converting adenosines into inosines (A-to-I editing) thus contributing to genome complexity and fine regulation of gene expression. It has been described that a member of the ADAR family, ADAR1, can target viruses and affect their replication process. Here we report evidence showing that ADAR1 stimulates human immuno deficiency virus type 1 (HIV-1) replication by using both editing-dependent and editing-independent mechanisms. We show that over-expression of ADAR1 in HIV-1 producer cells increases viral protein accumulation in an editing-independent manner. Moreover, HIV-1 virions generated in the presence of over-expressed ADAR1 but not an editing-inactive ADAR1 mutant are released more efficiently and display enhanced infectivity, as demonstrated by challenge assays performed with T cell lines and primary CD4+ T lymphocytes. Finally, we report that ADAR1 associates with HIV-1 RNAs and edits adenosines in the 5′ untranslated region (UTR) and the Rev and Tat coding sequence. Overall these results suggest that HIV-1 has evolved mechanisms to take advantage of specific RNA editing activity of the host cell and disclose a stimulatory function of ADAR1 in the spread of HIV-1. PMID:19651874

  7. HIV-1 evolution, drug resistance, and host genetics: The Indian scenario

    Directory of Open Access Journals (Sweden)

    U Shankarkumar

    2009-03-01

    Full Text Available U Shankarkumar, A Pawar, K GhoshNational Institute of Immunohaematology (ICMR, KEM Hospital, Parel, Mumbai, Maharashtra, IndiaAbstract: A regimen with varied side effects and compliance is of paramount importance to prevent viral drug resistance. Most of the drug-resistance studies, as well as interpretation algorithms, are based on sequence data from HIV-1 subtype B viruses. Increased resistance to antiretroviral drugs leads to poor prognosis by restricting treatment options. Due to suboptimal adherence to antiretroviral therapy there is an emergence of drug-resistant HIV-1 strains. The other factors responsible for this viral evolution are antiretroviral drug types and host genetics, especially major histocompatibility complex (MHC. Both primary and secondary drug resistances occur due to mutations in specific epitopes of viral protein regions which may influence the T cell recognition by immune system through MHC Class I and class II alleles. Mutations in viral epitopes enable the virus to escape the immune system. New drugs under clinical trials are being added but their exorbitant costs limit their access in developing countries. Thus the environmental consequences and, the impact of both viral and host genetic variations on the therapy in persons infected with HIV-1 clade C from India need to be determined.Keywords: HIV-1 C drug resistance, virus adaptation, HARRT, India

  8. LEDGF/p75 Deficiency Increases Deletions at the HIV-1 cDNA Ends.

    Science.gov (United States)

    Bueno, Murilo T D; Reyes, Daniel; Llano, Manuel

    2017-09-15

    Processing of unintegrated linear HIV-1 cDNA by the host DNA repair system results in its degradation and/or circularization. As a consequence, deficient viral cDNA integration generally leads to an increase in the levels of HIV-1 cDNA circles containing one or two long terminal repeats (LTRs). Intriguingly, impaired HIV-1 integration in LEDGF/p75-deficient cells does not result in a correspondent increase in viral cDNA circles. We postulate that increased degradation of unintegrated linear viral cDNA in cells lacking the lens epithelium-derived growth factor (LEDGF/p75) account for this inconsistency. To evaluate this hypothesis, we characterized the nucleotide sequence spanning 2-LTR junctions isolated from LEDGF/p75-deficient and control cells. LEDGF/p75 deficiency resulted in a significant increase in the frequency of 2-LTRs harboring large deletions. Of note, these deletions were dependent on the 3' processing activity of integrase and were not originated by aberrant reverse transcription. Our findings suggest a novel role of LEDGF/p75 in protecting the unintegrated 3' processed linear HIV-1 cDNA from exonucleolytic degradation.

  9. Interplay of HIV-1 phenotype and neutralizing antibody response in pathogenesis of AIDS.

    Science.gov (United States)

    Scarlatti, G; Leitner, T; Hodara, V; Jansson, M; Karlsson, A; Wahlberg, J; Rossi, P; Uhlén, M; Fenyö, E M; Albert, J

    1996-06-01

    A majority of human immunodeficiency virus type 1 (HIV-1) infected individuals display a rapid loss of CD4+ lymphocytes with fast progression towards overt acquired immunodeficiency syndrome (AIDS). However, a small proportion of individuals infected by HIV-1 remain immunologically intact for many years. In order to identify factors that might influence the pathogenesis of HIV-1 infection, 21 Italian mothers and 11 Swedish homosexual men were studied for the presence of autologous neutralizing antibodies in serum, biological phenotype of virus isolates and envelope variable region 3 (V3) sequences. The results were compared to the risk of mother-to-child transmission and progression of the disease. The presence of a neutralizing antibody response to the autologous virus as well as a virus with slow replicative capacity were linked both to low risk of mother-to-child transmission and non-progression of the disease. Patients whose peripheral blood mononuclear cells contained a mutation in the tip of the V3 loop (Arg318 to serine, lysine or leucine) significantly more often had neutralizing antibodies to autologous virus isolates containing arginine at this position. Thus, it appears that the interplay and balance between neutralizing antibody response of the host and the biological phenotype of HIV-1 strongly influence pathogenesis.

  10. Selection of unadapted, pathogenic SHIVs encoding newly transmitted HIV-1 envelope proteins.

    Science.gov (United States)

    Del Prete, Gregory Q; Ailers, Braiden; Moldt, Brian; Keele, Brandon F; Estes, Jacob D; Rodriguez, Anthony; Sampias, Marissa; Oswald, Kelli; Fast, Randy; Trubey, Charles M; Chertova, Elena; Smedley, Jeremy; LaBranche, Celia C; Montefiori, David C; Burton, Dennis R; Shaw, George M; Markowitz, Marty; Piatak, Michael; KewalRamani, Vineet N; Bieniasz, Paul D; Lifson, Jeffrey D; Hatziioannou, Theodora

    2014-09-10

    Infection of macaques with chimeric viruses based on SIVMAC but expressing the HIV-1 envelope (Env) glycoproteins (SHIVs) remains the most powerful model for evaluating prevention and therapeutic strategies against AIDS. Unfortunately, only a few SHIVs are currently available. Furthermore, their generation has required extensive adaptation of the HIV-1 Env sequences in macaques so they may not accurately represent HIV-1 Env proteins circulating in humans, potentially limiting their translational utility. We developed a strategy for generating large numbers of SHIV constructs expressing Env proteins from newly transmitted HIV-1 strains. By inoculating macaques with cocktails of multiple SHIV variants, we selected SHIVs that can replicate and cause AIDS-like disease in immunologically intact rhesus macaques without requiring animal-to-animal passage. One of these SHIVs could be transmitted mucosally. We demonstrate the utility of the SHIVs generated by this method for evaluating neutralizing antibody administration as a protection against mucosal SHIV challenge. Copyright © 2014 Elsevier Inc. All rights reserved.

  11. Partial protective effect of CCR5-Delta 32 heterozygosity in a cohort of heterosexual Italian HIV-1 exposed uninfected individuals

    Directory of Open Access Journals (Sweden)

    Cauda Roberto

    2006-09-01

    Full Text Available Abstract Despite multiple sexual exposure to HIV-1 virus, some individuals remain HIV-1 seronegative (exposed seronegative, ESN. The mechanisms underlying this resistance remain still unclear, although a multifactorial pathogenesis can be hypothesised. Although several genetic factors have been related to HIV-1 resistance, the homozigosity for a mutation in CCR5 gene (the 32 bp deletion, i.e. CCR5-Delta32 allele is presently considered the most relevant one. In the present study we analysed the genotype at CCR5 locus of 30 Italian ESN individuals (case group who referred multiple unprotected heterosexual intercourse with HIV-1 seropositive partner(s, for at least two years. One hundred and twenty HIV-1 infected patients and 120 individuals representative of the general population were included as control groups. Twenty percent of ESN individuals had heterozygous CCR5-Delta 32 genotype, compared to 7.5% of HIV-1 seropositive and 10% of individuals from the general population, respectively. None of the analysed individuals had CCR5-Delta 32 homozygous genotype. Sequence analysis of the entire open reading frame of CCR5 was performed in all ESN subjects and no polymorphisms or mutations were identified. Moreover, we determined the distribution of C77G variant in CD45 gene, which has been previously related to HIV-1 infection susceptibility. The frequency of the C77G variant showed no significant difference between ESN subjects and the two control groups. In conclusion, our data show a significantly higher frequency of CCR5-Delta 32 heterozygous genotype (p = 0.04 among the Italian heterosexual ESN individuals compared to HIV-1 seropositive patients, suggesting a partial protective role of CCR5-Delta 32 heterozygosity in this cohort.

  12. HIV-1 Genetic Diversity and Transmitted Drug Resistance Mutations among Patients from the North, Central and South Regions of Angola

    Science.gov (United States)

    Afonso, Joana Morais; Bello, Gonzalo; Guimarães, Monick L.; Sojka, Marta; Morgado, Mariza G.

    2012-01-01

    Background Angola presents a very complex HIV-1 epidemic characterized by the co-circulation of several HIV-1 group M subtypes, intersubtype recombinants and unclassified (U) variants. The viral diversity outside the major metropolitan regions (Luanda and Cabinda) and the prevalence of transmitted drug resistance mutations (DRM) since the introduction of HAART in 2004, however, has been barely studied. Methods One hundred and one individuals from the Central (n = 44), North (n = 35), and South (n = 22) regions of Angola were diagnosed as HIV-1 positive and had their blood collected between 2008 and 2010, at one of the National Referral Centers for HIV diagnosis, the Kifangondo Medical Center, located in the border between the Luanda and Bengo provinces. Angolan samples were genotyped based on phylogenetic and bootscanning analyses of the pol (PR/RT) gene and their drug resistance profile was analyzed. Results Among the 101 samples analyzed, 51% clustered within a pure group M subtype, 42% were classified as intersubtype recombinants, and 7% were denoted as U. We observed an important variation in the prevalence of different HIV-1 genetic variants among country regions, with high frequency of subtype F1 in the North (20%), intersubtype recombinants in the Central (42%), and subtype C in the South (45%). Statistically significant difference in HIV-1 clade distribution was only observed in subtype C prevalence between North vs South (p = 0.0005) and Central vs South (p = 0.0012) regions. DRM to NRTI and/or NNRTI were detected in 16.3% of patients analyzed. Conclusions These results demonstrate a heterogeneous distribution of HIV-1 genetic variants across different regions in Angola and also revealed an unexpected high frequency of DRM to RT inhibitors in patients that have reported no antiretroviral usage, which may decrease the efficiency of the standard first-line antiretroviral regimens currently used in the country. PMID:22952625

  13. The origin and emergence of an HIV-1 epidemic:

    DEFF Research Database (Denmark)

    Bruhn, Christian Anders Wathne; Audelin, Anne M.; Helleberg, Marie

    2014-01-01

    To describe, at patient-level detail, the determining events and factors involved in the development of a country's HIV-1 epidemic.......To describe, at patient-level detail, the determining events and factors involved in the development of a country's HIV-1 epidemic....

  14. Schistosomiasis and HIV-1 infection in rural Zimbabwe

    DEFF Research Database (Denmark)

    Kallestrup, Per; Zinyama, Rutendo; Gomo, Exnevia

    2005-01-01

    Stunted development and reduced fecundity of Schistosoma parasites in immunodeficient mice and the impaired ability of human immunodeficiency virus 1 (HIV-1)-infected humans to excrete schistosome eggs have been described. This study explores the effect that HIV-1-associated immunodeficiency has...

  15. HIV-1 Nef control of cell signalling molecules: multiple strategies

    Indian Academy of Sciences (India)

    HIV-1 has at its disposal numerous proteins encoded by its genome which provide the required arsenal to establish and maintain infection in its host for a considerable number of years. One of the most important and enigmatic of these proteins is Nef. The Nef protein of HIV-1 plays a fundamental role in the virus life cycle.

  16. Iron chelators ICL670 and 311 inhibit HIV-1 transcription

    International Nuclear Information System (INIS)

    Debebe, Zufan; Ammosova, Tatyana; Jerebtsova, Marina; Kurantsin-Mills, Joseph; Niu, Xiaomei; Charles, Sharroya; Richardson, Des R.; Ray, Patricio E.; Gordeuk, Victor R.; Nekhai, Sergei

    2007-01-01

    HIV-1 replication is induced by an excess of iron and iron chelation by desferrioxamine (DFO) inhibits viral replication by reducing proliferation of infected cells. Treatment of cells with DFO and 2-hydroxy-1-naphthylaldehyde isonicotinoyl hydrazone (311) inhibit expression of proteins that regulate cell-cycle progression, including cycle-dependent kinase 2 (CDK2). Our recent studies showed that CDK2 participates in HIV-1 transcription and viral replication suggesting that inhibition of CDK2 by iron chelators might also affect HIV-1 transcription. Here we evaluated the effect of a clinically approved orally effective iron chelator, 4-[3,5-bis-(hydroxyphenyl)-1,2,4-triazol-1-yl]-benzoic acid (ICL670) and 311 on HIV-1 transcription. Both ICL670 and 311 inhibited Tat-induced HIV-1 transcription in CEM-T cells, 293T and HeLa cells. Neither ICL670 nor 311 induced cytotoxicity at concentrations that inhibited HIV-1 transcription. The chelators decreased cellular activity of CDK2 and reduced HIV-1 Tat phosphorylation by CDK2. Neither ICL670A or 311 decreased CDK9 protein level but significantly reduced association of CDK9 with cyclin T1 and reduced phosphorylation of Ser-2 residues of RNA polymerase II C-terminal domain. In conclusion, our findings add to the evidence that iron chelators can inhibit HIV-1 transcription by deregulating CDK2 and CDK9. Further consideration should be given to the development of iron chelators for future anti-retroviral therapeutics

  17. Molecular Mechanisms in Activation of Latent HIV-1

    NARCIS (Netherlands)

    H. Rafati (Haleh)

    2014-01-01

    markdownabstract__Abstract__ Finding a cure for the human immunodeficiency virus type 1 (HIV-1) is extremely challenging. Development of highly active anti-retroviral therapy (HAART), transformed HIV-1 infection from an acute syndrome into chronic disease. Although using HAART results in

  18. Vaginalmycosis and HIV-1 infection in Kaduna, Nigeria. | Eni ...

    African Journals Online (AJOL)

    ... mycosis in HIV-1positive women and managed accordingly. Proper management of these two conditions will improve reproductive health of women in Nigeria. Keywords: Vaginal mycosis, Genital candidiasis, Reproductive health: Candida albicans: HIV-1 infection. Journal of Biomedical Investigation Vol. 3 (1) 2005: pp.

  19. Antiviral Therapy by HIV-1 Broadly Neutralizing and Inhibitory Antibodies

    Directory of Open Access Journals (Sweden)

    Zhiqing Zhang

    2016-11-01

    Full Text Available Human immunodeficiency virus type 1 (HIV-1 infection causes acquired immune deficiency syndrome (AIDS, a global epidemic for more than three decades. HIV-1 replication is primarily controlled through antiretroviral therapy (ART but this treatment does not cure HIV-1 infection. Furthermore, there is increasing viral resistance to ART, and side effects associated with long-term therapy. Consequently, there is a need of alternative candidates for HIV-1 prevention and therapy. Recent advances have discovered multiple broadly neutralizing antibodies against HIV-1. In this review, we describe the key epitopes on the HIV-1 Env protein and the reciprocal broadly neutralizing antibodies, and discuss the ongoing clinical trials of broadly neutralizing and inhibitory antibody therapy as well as antibody combinations, bispecific antibodies, and methods that improve therapeutic efficacy by combining broadly neutralizing antibodies (bNAbs with latency reversing agents. Compared with ART, HIV-1 therapeutics that incorporate these broadly neutralizing and inhibitory antibodies offer the advantage of decreasing virus load and clearing infected cells, which is a promising prospect in HIV-1 prevention and treatment.

  20. Cold denaturation of the HIV-1 protease monomer

    DEFF Research Database (Denmark)

    Rösner, Heike Ilona; Caldarini, Martina; Prestel, Andreas

    2017-01-01

    The HIV-1-protease is a complex protein which in its active form adopts a homodimer dominated by -sheet structures. We have discovered a cold-denatured state of the monomeric subunit of HIV-1-protease which is populated above 0ºC and therefore directly accessible to various spectroscopic approac...

  1. Raltegravir with optimized background therapy for resistant HIV-1 infection

    DEFF Research Database (Denmark)

    Steigbigel, Roy T; Cooper, David A; Kumar, Princy N

    2008-01-01

    BACKGROUND: Raltegravir (MK-0518) is an inhibitor of human immunodeficiency virus type 1 (HIV-1) integrase active against HIV-1 susceptible or resistant to older antiretroviral drugs. METHODS: We conducted two identical trials in different geographic regions to evaluate the safety and efficacy of...

  2. HIV-1 evolution, drug resistance, and host genetics: The Indian scenario

    OpenAIRE

    Shankarkumar, U.; Pawar,Aruna; Ghosh,Kanjaksha

    2009-01-01

    U Shankarkumar, A Pawar, K GhoshNational Institute of Immunohaematology (ICMR), KEM Hospital, Parel, Mumbai, Maharashtra, IndiaAbstract: A regimen with varied side effects and compliance is of paramount importance to prevent viral drug resistance. Most of the drug-resistance studies, as well as interpretation algorithms, are based on sequence data from HIV-1 subtype B viruses. Increased resistance to antiretroviral drugs leads to poor prognosis by restricting treatment optio...

  3. Mutation of HIV-1 Genomes in a Clinical Population Treated with the Mutagenic Nucleoside KP1461

    OpenAIRE

    Mullins, James I.; Heath, Laura; Hughes, James P.; Kicha, Jessica; Styrchak, Sheila; Wong, Kim G.; Rao, Ushnal; Hansen, Alexis; Harris, Kevin S.; Laurent, Jean-Pierre; Li, Deyu; Simpson, Jeffrey H.; Essigmann, John M.; Loeb, Lawrence A.; Parkins, Jeffrey

    2011-01-01

    The deoxycytidine analog KP1212, and its prodrug KP1461, are prototypes of a new class of antiretroviral drugs designed to increase viral mutation rates, with the goal of eventually causing the collapse of the viral population. Here we present an extensive analysis of viral sequences from HIV-1 infected volunteers from the first "mechanism validation" phase II clinical trial of a mutagenic base analog in which individuals previously treated with antiviral drugs received 1600 mg of KP1461 twic...

  4. Alterations in HIV-1 LTR promoter activity during AIDS progression

    International Nuclear Information System (INIS)

    Hiebenthal-Millow, Kirsten; Greenough, Thomas C.; Bretttler, Doreen B.; Schindler, Michael; Wildum, Steffen; Sullivan, John L.; Kirchhoff, Frank

    2003-01-01

    HIV-1 variants evolving in AIDS patients frequently show increased replicative capacity compared to those present during early asymptomatic infection. It is known that late stage HIV-1 variants often show an expanded coreceptor tropism and altered Nef function. In the present study we investigated whether enhanced HIV-1 LTR promoter activity might also evolve during disease progression. Our results demonstrate increased LTR promoter activity after AIDS progression in 3 of 12 HIV-1-infected individuals studied. Further analysis revealed that multiple alterations in the U3 core-enhancer and in the transactivation-response (TAR) region seem to be responsible for the enhanced functional activity. Our findings show that in a subset of HIV-1-infected individuals enhanced LTR transcription contributes to the increased replicative potential of late stage virus isolates and might accelerate disease progression

  5. HIV-1 protease inhibitory substances from Cassia garrettiana

    Directory of Open Access Journals (Sweden)

    Jindaporn Puripattanvong

    2007-01-01

    Full Text Available Cassia garrettiana Craib, a Thai medicinal plant locally known as Samae-sarn, was investigated for its active constituents against HIV-1 protease (HIV-1 PR. Bioassay-guided fractionation of the heart woodof this plant led to the isolation of a stilbene derivative (1, piceatannol and an anthraquinone derivative (2, chrysophanol. Piceatannol exhibited appreciable inhibitory effect against HIV-1 PR with an IC50 value of25.4 μg/ml, whereas that of chrysophanol was 73.5 μg/ml. In addition, other two stilbenoids together with three anthraquinone derivatives were also investigated for their anti-HIV-1 PR activities. The resultindicated that resveratrol possessed anti-HIV-1 PR activity with an IC50 value of 85.0 μg/ml, whereas other stilbenoid (oxyresveratrol and anthraquinone derivatives (emodin, aloe-emodin, rhein were inactive (IC50 > 100 μg/ml.

  6. Comparison of glycerolisation with cryopreservation methods on HIV-1 inactivation

    International Nuclear Information System (INIS)

    Van Baare, J.; Pagnon, J.; Cameron, P.; Vardaxis, N.; Middlekoop, E.; Crowe, S.

    1999-01-01

    Cryopreservation and glycerolisation are two successful long-term preservation methods for human cadaveric donor skin, which is used in the treatment of bum patients. High concentrations of glycerol has been shown to be antibacterial and virucidal. Because fear of possible transmission of HIV-1 following allograft transplantation, this study was undertaken to investigate whether HIV can be effectively eliminated from skin explants. HIV-1 Ba-L, which has been shown to infect monocytes in skin explants and also dendritic cells, was. For the experiments we used cell-free virus, exogenously HIV infected peripheral blood mononuclear cells (PBMCs) and exogenously HIV infected cadaver split skin. Different concentrations of glycerol at various temperatures and the glycerolisation procedure as used by the Euro Skin Bank were used to determine the effects on HIV-1 Ba-L infectivity. For the cryopreservation technique we used 10% DMSO and a controlled rate freezer. HIV-1 Ba-L transfer was determined by adding uninfected PBMCs to the infected material and reverse transcriptase was measured. Cell-free HIV-1 Ba-L was not inactivated by 50% glycerol but was effectively inactivated within 30 minutes by 70% and 85% glycerol at 4 degree C, room temperature and 37 degree C. In contrast, cell-free HIV-1 Ba-L was not inactivated by cryopreservation. Most importantly, we have shown that HIV-1 Ba-L present in split skin is inactivated by incubating skin in 70% glycerol for three hours at 37-C. HIV in exogenously infected skin was not inactivated by cryopreservation. High concentrations of glycerol effectively inactivates free HIV-1 Ba-L and intracellular HIV-1 Ba-L. Also the current glycerolisation procedure carried out by the Euro Skin Bank effectively inactivates infectious virus. However, the cryopreservation technique did not show any reduction in HIV-1 Ba-L infectivity

  7. Sexually transmitted infections among HIV-1-discordant couples.

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    Brandon L Guthrie

    2009-12-01

    Full Text Available More new HIV-1 infections occur within stable HIV-1-discordant couples than in any other group in Africa, and sexually transmitted infections (STIs may increase transmission risk among discordant couples, accounting for a large proportion of new HIV-1 infections. Understanding correlates of STIs among discordant couples will aid in optimizing interventions to prevent HIV-1 transmission in these couples.HIV-1-discordant couples in which HIV-1-infected partners were HSV-2-seropositive were tested for syphilis, chlamydia, gonorrhea, and trichomoniasis, and HIV-1-uninfected partners were tested for HSV-2. We assessed sociodemographic, behavioral, and biological correlates of a current STI.Of 416 couples enrolled, 16% were affected by a treatable STI, and among these both partners were infected in 17% of couples. A treatable STI was found in 46 (11% females and 30 (7% males. The most prevalent infections were trichomoniasis (5.9% and syphilis (2.6%. Participants were 5.9-fold more likely to have an STI if their partner had an STI (P<0.01, and STIs were more common among those reporting any unprotected sex (OR = 2.43; P<0.01 and those with low education (OR = 3.00; P<0.01. Among HIV-1-uninfected participants with an HSV-2-seropositive partner, females were significantly more likely to be HSV-2-seropositive than males (78% versus 50%, P<0.01.Treatable STIs were common among HIV-1-discordant couples and the majority of couples affected by an STI were discordant for the STI, with relatively high HSV-2 discordance. Awareness of STI correlates and treatment of both partners may reduce HIV-1 transmission.ClinicalTrials.gov NCT00194519.

  8. The Depsipeptide Romidepsin Reverses HIV-1 Latency In Vivo.

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    Ole S Søgaard

    2015-09-01

    Full Text Available Pharmacologically-induced activation of replication competent proviruses from latency in the presence of antiretroviral treatment (ART has been proposed as a step towards curing HIV-1 infection. However, until now, approaches to reverse HIV-1 latency in humans have yielded mixed results. Here, we report a proof-of-concept phase Ib/IIa trial where 6 aviremic HIV-1 infected adults received intravenous 5 mg/m2 romidepsin (Celgene once weekly for 3 weeks while maintaining ART. Lymphocyte histone H3 acetylation, a cellular measure of the pharmacodynamic response to romidepsin, increased rapidly (maximum fold range: 3.7–7.7 relative to baseline within the first hours following each romidepsin administration. Concurrently, HIV-1 transcription quantified as copies of cell-associated un-spliced HIV-1 RNA increased significantly from baseline during treatment (range of fold-increase: 2.4–5.0; p = 0.03. Plasma HIV-1 RNA increased from <20 copies/mL at baseline to readily quantifiable levels at multiple post-infusion time-points in 5 of 6 patients (range 46–103 copies/mL following the second infusion, p = 0.04. Importantly, romidepsin did not decrease the number of HIV-specific T cells or inhibit T cell cytokine production. Adverse events (all grade 1–2 were consistent with the known side effects of romidepsin. In conclusion, romidepsin safely induced HIV-1 transcription resulting in plasma HIV-1 RNA that was readily detected with standard commercial assays demonstrating that significant reversal of HIV-1 latency in vivo is possible without blunting T cell-mediated immune responses. These finding have major implications for future trials aiming to eradicate the HIV-1 reservoir.clinicaltrials.gov NTC02092116.

  9. Cis elements and trans-acting factors involved in the RNA dimerization of the human immunodeficiency virus HIV-1.

    Science.gov (United States)

    Darlix, J L; Gabus, C; Nugeyre, M T; Clavel, F; Barré-Sinoussi, F

    1990-12-05

    The retroviral genome consists of two identical RNA molecules joined at their 5' ends by the Dimer Linkage Structure (DLS). To study the mechanism of dimerization and the DLS of HIV-1 RNA, large amounts of bona fide HIV-1 RNA and of mutants have been synthesized in vitro. We report that HIV-1 RNA forms dimeric molecules and that viral nucleocapsid (NC) protein NCp15 greatly activates dimerization. Deletion mutagenesis in the RNA 5' 1333 nucleotides indicated that a small domain of 100 nucleotides, located between positions 311 to 415 from the 5' end, is necessary and sufficient to promote HIV-1 RNA dimerization. This dimerization domain encompasses an encapsidation element located between the 5' splice donor site and initiator AUG of gag and shows little sequence variations in different strains of HIV-1. Furthermore, cross-linking analysis of the interactions between NC and HIV-1 RNA (311 to 415) locates a major contact site in the encapsidation element of HIV-1 RNA. The genomic RNA dimer is tightly associated with nucleocapsid protein molecules in avian and murine retroviruses, and this ribonucleoprotein structure is believed to be the template for reverse transcription. Genomic RNA-protein interactions have been analyzed in human immunodeficiency virus (HIV) virions and results showed that NC protein molecules are tightly bound to the genomic RNA dimer. Since retroviral RNA dimerization and packaging appear to be under the control of the same cis element, the encapsidation sequences, and trans-acting factor, the NC protein, they are probably related events in the course of virion assembly.

  10. The Number and Complexity of Pure and Recombinant HIV-1 Strains Observed within Incident Infections during the HIV and Malaria Cohort Study Conducted in Kericho, Kenya, from 2003 to 2006.

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    Erik Billings

    Full Text Available Characterization of HIV-1 subtype diversity in regions where vaccine trials are conducted is critical for vaccine development and testing. This study describes the molecular epidemiology of HIV-1 within a tea-plantation community cohort in Kericho, Kenya. Sixty-three incident infections were ascertained in the HIV and Malaria Cohort Study conducted in Kericho from 2003 to 2006. HIV-1 strains from 58 of those individuals were full genome characterized and compared to two previous Kenyan studies describing 41 prevalent infections from a blood bank survey (1999-2000 and 21 infections from a higher-risk cohort containing a mix of incident and prevalent infections (2006. Among the 58 strains from the community cohort, 43.1% were pure subtypes (36.2% A1, 5.2% C, and 1.7% G and 56.9% were inter-subtype recombinants (29.3% A1D, 8.6% A1CD, 6.9% A1A2D, 5.2% A1C, 3.4% A1A2CD, and 3.4% A2D. This diversity and the resulting genetic distance between the observed strains will need to be addressed when vaccine immunogens are chosen. In consideration of current vaccine development efforts, the strains from these three studies were compared to five candidate vaccines (each of which are viral vectored, carrying inserts corresponding to parts of gag, pol, and envelope, which have been developed for possible use in sub-Saharan Africa. The sequence comparison between the observed strains and the candidate vaccines indicates that in the presence of diverse recombinants, a bivalent vaccine is more likely to provide T-cell epitope coverage than monovalent vaccines even when the inserts of the bivalent vaccine are not subtype-matched to the local epidemic.

  11. The Number and Complexity of Pure and Recombinant HIV-1 Strains Observed within Incident Infections during the HIV and Malaria Cohort Study Conducted in Kericho, Kenya, from 2003 to 2006

    Science.gov (United States)

    Billings, Erik; Sanders-Buell, Eric; Bose, Meera; Bradfield, Andrea; Lei, Esther; Kijak, Gustavo H.; Arroyo, Miguel A.; Kibaya, Rukia M.; Scott, Paul T.; Wasunna, Monique K.; Sawe, Frederick K.; Shaffer, Douglas N.; Birx, Deborah L.; McCutchan, Francine E.; Michael, Nelson L.; Robb, Merlin L.; Kim, Jerome H.; Tovanabutra, Sodsai

    2015-01-01

    Characterization of HIV-1 subtype diversity in regions where vaccine trials are conducted is critical for vaccine development and testing. This study describes the molecular epidemiology of HIV-1 within a tea-plantation community cohort in Kericho, Kenya. Sixty-three incident infections were ascertained in the HIV and Malaria Cohort Study conducted in Kericho from 2003 to 2006. HIV-1 strains from 58 of those individuals were full genome characterized and compared to two previous Kenyan studies describing 41 prevalent infections from a blood bank survey (1999–2000) and 21 infections from a higher-risk cohort containing a mix of incident and prevalent infections (2006). Among the 58 strains from the community cohort, 43.1% were pure subtypes (36.2% A1, 5.2% C, and 1.7% G) and 56.9% were inter-subtype recombinants (29.3% A1D, 8.6% A1CD, 6.9% A1A2D, 5.2% A1C, 3.4% A1A2CD, and 3.4% A2D). This diversity and the resulting genetic distance between the observed strains will need to be addressed when vaccine immunogens are chosen. In consideration of current vaccine development efforts, the strains from these three studies were compared to five candidate vaccines (each of which are viral vectored, carrying inserts corresponding to parts of gag, pol, and envelope), which have been developed for possible use in sub-Saharan Africa. The sequence comparison between the observed strains and the candidate vaccines indicates that in the presence of diverse recombinants, a bivalent vaccine is more likely to provide T-cell epitope coverage than monovalent vaccines even when the inserts of the bivalent vaccine are not subtype-matched to the local epidemic. PMID:26287814

  12. HIV-1 persistence following extremely early initiation of antiretroviral therapy (ART during acute HIV-1 infection: An observational study.

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    Timothy J Henrich

    2017-11-01

    cells/mouse experienced very low level viremia (201 copies/mL; sequence confirmation was unsuccessful. PrEP Participant A stopped ART and remained aviremic for 7.4 months, rebounding with HIV RNA of 36 copies/mL that rose to 59,805 copies/mL 6 days later. ART was restarted promptly. Rebound plasma HIV sequences were identical to those obtained during acute infection by single-genome sequencing. Mathematical modeling predicted that the latent reservoir size was approximately 200 cells prior to ATI and that only around 1% of individuals with a similar HIV burden may achieve lifelong ART-free remission. Furthermore, we observed that lymphocytes expressing the tumor marker CD30 increased in frequency weeks to months prior to detectable HIV-1 RNA in plasma. This study was limited by the small sample size, which was a result of the rarity of individuals presenting during hyperacute infection.We report HIV relapse despite initiation of ART at one of the earliest stages of acute HIV infection possible. Near complete or complete loss of detectable HIV in blood and tissues did not lead to indefinite ART-free HIV remission. However, the small numbers of latently infected cells in individuals treated during hyperacute infection may be associated with prolonged ART-free remission.

  13. HIV-1-Specific IgA Monoclonal Antibodies from an HIV-1 Vaccinee Mediate Galactosylceramide Blocking and Phagocytosis

    Science.gov (United States)

    2018-01-01

    ABSTRACT Vaccine-elicited humoral immune responses comprise an array of antibody forms and specificities, with only a fraction contributing to protective host immunity. Elucidation of antibody effector functions responsible for protective immunity against human immunodeficiency virus type 1 (HIV-1) acquisition is a major goal for the HIV-1 vaccine field. Immunoglobulin A (IgA) is an important part of the host defense against pathogens; however, little is known about the role of vaccine-elicited IgA and its capacity to mediate antiviral functions. To identify the antiviral functions of HIV-1-specific IgA elicited by vaccination, we cloned HIV-1 envelope-specific IgA monoclonal antibodies (MAbs) by memory B cell cultures from peripheral blood mononuclear cells from an RV144 vaccinee and produced two IgA clonal cell lines (HG129 and HG130) producing native, nonrecombinant IgA MAbs. The HG129 and HG130 MAbs mediated phagocytosis by monocytes, and HG129 blocked HIV-1 Env glycoprotein binding to galactosylceramide, an alternative HIV-1 receptor. These findings elucidate potential antiviral functions of vaccine-elicited HIV-1 envelope-specific IgA that may act to block HIV-1 acquisition at the portal of entry by preventing HIV-1 binding to galactosylceramide and mediating antibody Fc receptor-mediated virion phagocytosis. Furthermore, these findings highlight the complex and diverse interactions of vaccine-elicited IgA with pathogens that depend on IgA fine specificity and form (e.g., multimeric or monomeric) in the systemic circulation and mucosal compartments. IMPORTANCE Host-pathogen interactions in vivo involve numerous immune mechanisms that can lead to pathogen clearance. Understanding the nature of antiviral immune mechanisms can inform the design of efficacious HIV-1 vaccine strategies. Evidence suggests that both neutralizing and nonneutralizing antibodies can mediate some protection against HIV in animal models. Although numerous studies have characterized the

  14. High Multiplicity Infection by HIV-1 in Men Who Have Sex with Men.

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    Hui Li

    2010-05-01

    Full Text Available Elucidating virus-host interactions responsible for HIV-1 transmission is important for advancing HIV-1 prevention strategies. To this end, single genome amplification (SGA and sequencing of HIV-1 within the context of a model of random virus evolution has made possible for the first time an unambiguous identification of transmitted/founder viruses and a precise estimation of their numbers. Here, we applied this approach to HIV-1 env analyses in a cohort of acutely infected men who have sex with men (MSM and found that a high proportion (10 of 28; 36% had been productively infected by more than one virus. In subjects with multivariant transmission, the minimum number of transmitted viruses ranged from 2 to 10 with viral recombination leading to rapid and extensive genetic shuffling among virus lineages. A combined analysis of these results, together with recently published findings based on identical SGA methods in largely heterosexual (HSX cohorts, revealed a significantly higher frequency of multivariant transmission in MSM than in HSX [19 of 50 subjects (38% versus 34 of 175 subjects (19%; Fisher's exact p = 0.008]. To further evaluate the SGA strategy for identifying transmitted/founder viruses, we analyzed 239 overlapping 5' and 3' half genome or env-only sequences from plasma viral RNA (vRNA and blood mononuclear cell DNA in an MSM subject who had a particularly well-documented virus exposure history 3-6 days before symptom onset and 14-17 days before peak plasma viremia (47,600,000 vRNA molecules/ml. All 239 sequences coalesced to a single transmitted/founder virus genome in a time frame consistent with the clinical history, and a molecular clone of this genome encoded replication competent virus in accord with model predictions. Higher multiplicity of HIV-1 infection in MSM compared with HSX is consistent with the demonstrably higher epidemiological risk of virus acquisition in MSM and could indicate a greater challenge for HIV-1

  15. Trans-dissemination of exosomes from HIV-1-infected cells fosters both HIV-1 trans-infection in resting CD4+ T lymphocytes and reactivation of the HIV-1 reservoir.

    Science.gov (United States)

    Chiozzini, Chiara; Arenaccio, Claudia; Olivetta, Eleonora; Anticoli, Simona; Manfredi, Francesco; Ferrantelli, Flavia; d'Ettorre, Gabriella; Schietroma, Ivan; Andreotti, Mauro; Federico, Maurizio

    2017-09-01

    Intact HIV-1 and exosomes can be internalized by dendritic cells (DCs) through a common pathway leading to their transmission to CD4 + T lymphocytes by means of mechanisms defined as trans-infection and trans-dissemination, respectively. We previously reported that exosomes from HIV-1-infected cells activate both uninfected quiescent CD4 + T lymphocytes, which become permissive to HIV-1, and latently infected cells, with release of HIV-1 particles. However, nothing is known about the effects of trans-dissemination of exosomes produced by HIV-1-infected cells on uninfected or latently HIV-1-infected CD4 + T lymphocytes. Here, we report that trans-dissemination of exosomes from HIV-1-infected cells induces cell activation in resting CD4 + T lymphocytes, which appears stronger with mature than immature DCs. Using purified preparations of both HIV-1 and exosomes, we observed that mDC-mediated trans-dissemination of exosomes from HIV-1-infected cells to resting CD4 + T lymphocytes induces efficient trans-infection and HIV-1 expression in target cells. Most relevant, when both mDCs and CD4 + T lymphocytes were isolated from combination anti-retroviral therapy (ART)-treated HIV-1-infected patients, trans-dissemination of exosomes from HIV-1-infected cells led to HIV-1 reactivation from the viral reservoir. In sum, our data suggest a role of exosome trans-dissemination in both HIV-1 spread in the infected host and reactivation of the HIV-1 reservoir.

  16. Interaction between HIV-1 Tat and DNA-PKcs modulates HIV transcription and class switch recombination.

    Science.gov (United States)

    Zhang, Shi-Meng; Zhang, He; Yang, Tian-Yi; Ying, Tian-Yi; Yang, Pei-Xiang; Liu, Xiao-Dan; Tang, Sheng-Jian; Zhou, Ping-Kun

    2014-01-01

    HIV-1 tat targets a variety of host cell proteins to facilitate viral transcription and disrupts host cellular immunity by inducing lymphocyte apoptosis, but whether it influences humoral immunity remains unclear. Previously, our group demonstrated that tat depresses expression of DNA-PKcs, a critical component of the non-homologous end joining pathway (NHEJ) of DNA double-strand breaks repair, immunoglobulin class switch recombination (CSR) and V(D)J recombination, and sensitizes cells to ionizing radiation. In this study, we demonstrated that HIV-1 Tat down-regulates DNA-PKcs expression by directly binding to the core promoter sequence. In addition, Tat interacts with and activates the kinase activity of DNA-PKcs in a dose-dependent and DNA independent manner. Furthermore, Tat inhibits class switch recombination (CSR) at low concentrations (≤ 4 µg/ml) and stimulates CSR at high concentrations (≥ 8 µg/ml). On the other hand, low protein level and high kinase activity of DNA-PKcs promotes HIV-1 transcription, while high protein level and low kinase activity inhibit HIV-1 transcription. Co-immunoprecipitation results revealed that DNA-PKcs forms a large complex comprised of Cyclin T1, CDK9 and Tat via direct interacting with CDK9 and Tat but not Cyclin T1. Taken together, our results provide new clues that Tat regulates host humoral immunity via both transcriptional depression and kinase activation of DNA-PKcs. We also raise the possibility that inhibitors and interventions directed towards DNA-PKcs may inhibit HIV-1 transcription in AIDS patients.

  17. Broad and potent immune responses to a low dose intradermal HIV-1 DNA boosted with HIV-1 recombinant MVA among healthy adults in Tanzania☆,☆☆

    Science.gov (United States)

    Bakari, Muhammad; Aboud, Said; Nilsson, Charlotta; Francis, Joel; Buma, Deus; Moshiro, Candida; Aris, Eric A.; Lyamuya, Eligius F.; Janabi, Mohamed; Godoy-Ramirez, Karina; Joachim, Agricola; Polonis, Victoria R.; Bråve, Andreas; Earl, Patricia; Robb, Merlin; Marovich, Mary; Wahren, Britta; Pallangyo, Kisali; Biberfeld, Gunnel; Mhalu, Fred; Sandström, Eric

    2016-01-01

    Background We conducted a phase I/II randomized placebo-controlled trial with the aim of exploring whether priming with a low intradermal dose of a multiclade, multigene HIV-1 DNA vaccine could improve the immunogenicity of the same vaccine given intramuscularly prior to boosting with a heterologous HIV-1 MVA among healthy adults in Dar es Salaam, Tanzania. Methods Sixty HIV-uninfected volunteers were randomized to receive DNA plasmid vaccine 1 mg intradermally (id), n = 20, or 3.8 mg intramuscularly (im), n = 20, or placebo, n = 20, using a needle-free injection device. DNA plasmids encoding HIV-1 genes gp160 subtype A, B, C; rev B; p17/p24 gag A, B and Rtmut B were given at weeks 0, 4 and 12. Recombinant MVA (108 pfu) expressing HIV-1 Env, Gag, Pol of CRF01_AE or placebo was administered im at month 9 and 21. Results The vaccines were well tolerated. Two weeks after the third HIV-DNA injection, 22/38 (58%) vaccinees had IFN-γ ELISpot responses to Gag. Two weeks after the first HIV-MVA boost all 35 (100%) vaccinees responded to Gag and 31 (89%) to Env. Two to four weeks after the second HIV-MVA boost, 28/29 (97%) vaccinees had IFN-γ ELISpot responses, 27 (93%) to Gag and 23 (79%) to Env. The id-primed recipients had significantly higher responses to Env than im recipients. Intracellular cytokine staining for Gag-specific IFN-γ/IL-2 production showed both CD8+ and CD4+ T cell responses. All vaccinees had HIV-specific lymphoproliferative responses. All vaccinees reacted in diagnostic HIV serological tests and 26/29 (90%) had antibodies against gp160 after the second HIV-MVA boost. Furthermore, while all of 29 vaccinee sera were negative for neutralizing antibodies against clade B, C and CRF01 AE pseudoviruses in the TZM-bl neutralization assay, in a PBMC assay, the response rate ranged from 31% to 83% positives, depending upon the clade B or CRF01_AE virus tested. This vaccine approach is safe and highly immunogenic. Low dose, id HIV-DNA priming elicited higher

  18. Controlling cellular P-TEFb activity by the HIV-1 transcriptional transactivator Tat.

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    Lisa Muniz

    Full Text Available The human immunodeficiency virus 1 (HIV-1 transcriptional transactivator (Tat is essential for synthesis of full-length transcripts from the integrated viral genome by RNA polymerase II (Pol II. Tat recruits the host positive transcription elongation factor b (P-TEFb to the HIV-1 promoter through binding to the transactivator RNA (TAR at the 5'-end of the nascent HIV transcript. P-TEFb is a general Pol II transcription factor; its cellular activity is controlled by the 7SK small nuclear RNA (snRNA and the HEXIM1 protein, which sequester P-TEFb into transcriptionally inactive 7SK/HEXIM/P-TEFb snRNP. Besides targeting P-TEFb to HIV transcription, Tat also increases the nuclear level of active P-TEFb through promoting its dissociation from the 7SK/HEXIM/P-TEFb RNP by an unclear mechanism. In this study, by using in vitro and in vivo RNA-protein binding assays, we demonstrate that HIV-1 Tat binds with high specificity and efficiency to an evolutionarily highly conserved stem-bulge-stem motif of the 5'-hairpin of human 7SK snRNA. The newly discovered Tat-binding motif of 7SK is structurally and functionally indistinguishable from the extensively characterized Tat-binding site of HIV TAR and importantly, it is imbedded in the HEXIM-binding elements of 7SK snRNA. We show that Tat efficiently replaces HEXIM1 on the 7SK snRNA in vivo and therefore, it promotes the disassembly of the 7SK/HEXIM/P-TEFb negative transcriptional regulatory snRNP to augment the nuclear level of active P-TEFb. This is the first demonstration that HIV-1 specifically targets an important cellular regulatory RNA, most probably to promote viral transcription and replication. Demonstration that the human 7SK snRNA carries a TAR RNA-like Tat-binding element that is essential for the normal transcriptional regulatory function of 7SK questions the viability of HIV therapeutic approaches based on small drugs blocking the Tat-binding site of HIV TAR.

  19. Anti-HIV-1 activity of flavonoid myricetin on HIV-1 infection in a dual-chamber in vitro model.

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    Silvana Pasetto

    Full Text Available HIV infection by sexual transmission remains an enormous global health concern. More than 1 million new infections among women occur annually. Microbicides represent a promising prevention strategy that women can easily control. Among emerging therapies, natural small molecules such as flavonoids are an important source of new active substances. In this study we report the in vitro cytotoxicity and anti-HIV-1 and microbicide activity of the following flavonoids: Myricetin, Quercetin and Pinocembrin. Cytotoxicity tests were conducted on TZM-bl, HeLa, PBMC, and H9 cell cultures using 0.01-100 µM concentrations. Myricetin presented the lowest toxic effect, with Quercetin and Pinocembrin relatively more toxic. The anti-HIV-1 activity was tested with TZM-bl cell plus HIV-1 BaL (R5 tropic, H9 and PBMC cells plus HIV-1 MN (X4 tropic, and the dual tropic (X4R5 HIV-1 89.6. All flavonoids showed anti-HIV activity, although Myricetin was more effective than Quercetin or Pinocembrin. In TZM-bl cells, Myricetin inhibited ≥90% of HIV-1 BaL infection. The results were confirmed by quantification of HIV-1 p24 antigen in supernatant from H9 and PBMC cells following flavonoid treatment. In H9 and PBMC cells infected by HIV-1 MN and HIV-1 89.6, Myricetin showed more than 80% anti-HIV activity. Quercetin and Pinocembrin presented modest anti-HIV activity in all experiments. Myricetin activity was tested against HIV-RT and inhibited the enzyme by 49%. Microbicide activities were evaluated using a dual-chamber female genital tract model. In the in vitro microbicide activity model, Myricetin showed promising results against different strains of HIV-1 while also showing insignificant cytotoxic effects. Further studies of Myricetin should be performed to identify its molecular targets in order to provide a solid biological foundation for translational research.

  20. Transmission of single and multiple viral variants in primary HIV-1 subtype C infection.

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    Vladimir Novitsky

    2011-02-01

    Full Text Available To address whether sequences of viral gag and env quasispecies collected during the early post-acute period can be utilized to determine multiplicity of transmitted HIV's, recently developed approaches for analysis of viral evolution in acute HIV-1 infection [1,2] were applied. Specifically, phylogenetic reconstruction, inter- and intra-patient distribution of maximum and mean genetic distances, analysis of Poisson fitness, shape of highlighter plots, recombination analysis, and estimation of time to the most recent common ancestor (tMRCA were utilized for resolving multiplicity of HIV-1 transmission in a set of viral quasispecies collected within 50 days post-seroconversion (p/s in 25 HIV-infected individuals with estimated time of seroconversion. The decision on multiplicity of HIV infection was made based on the model's fit with, or failure to explain, the observed extent of viral sequence heterogeneity. The initial analysis was based on phylogeny, inter-patient distribution of maximum and mean distances, and Poisson fitness, and was able to resolve multiplicity of HIV transmission in 20 of 25 (80% cases. Additional analysis involved distribution of individual viral distances, highlighter plots, recombination analysis, and estimation of tMRCA, and resolved 4 of the 5 remaining cases. Overall, transmission of a single viral variant was identified in 16 of 25 (64% cases, and transmission of multiple variants was evident in 8 of 25 (32% cases. In one case multiplicity of HIV-1 transmission could not be determined. In primary HIV-1 subtype C infection, samples collected within 50 days p/s and analyzed by a single-genome amplification/sequencing technique can provide reliable identification of transmission multiplicity in 24 of 25 (96% cases. Observed transmission frequency of a single viral variant and multiple viral variants were within the ranges of 64% to 68%, and 32% to 36%, respectively.

  1. Correlates of HIV-1 genital shedding in Tanzanian women.

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    Clare Tanton

    2011-03-01

    Full Text Available Understanding the correlates of HIV shedding is important to inform strategies to reduce HIV infectiousness. We examined correlates of genital HIV-1 RNA in women who were seropositive for both herpes simplex virus (HSV-2 and HIV-1 and who were enrolled in a randomised controlled trial of HSV suppressive therapy (aciclovir 400 mg b.i.d vs. placebo in Tanzania.Samples, including a cervico-vaginal lavage, were collected and tested for genital HIV-1 and HSV and reproductive tract infections (RTIs at randomisation and 6, 12 and 24 months follow-up. Data from all women at randomisation and women in the placebo arm during follow-up were analysed using generalised estimating equations to determine the correlates of cervico-vaginal HIV-1 RNA detection and load.Cervico-vaginal HIV-1 RNA was detected at 52.0% of 971 visits among 482 women, and was independently associated with plasma viral load, presence of genital ulcers, pregnancy, bloody cervical or vaginal discharge, abnormal vaginal discharge, cervical ectopy, Neisseria gonorrhoeae, Chlamydia trachomatis, Trichomonas vaginalis, an intermediate bacterial vaginosis score and HSV DNA detection. Similar factors were associated with genital HIV-1 RNA load.RTIs were associated with increased presence and quantity of genital HIV-1 RNA in this population. These results highlight the importance of integrating effective RTI treatment into HIV care services.

  2. Novel host restriction factors implicated in HIV-1 replication.

    Science.gov (United States)

    Ghimire, Dibya; Rai, Madhu; Gaur, Ritu

    2018-04-01

    Human immunodeficiency virus-1 (HIV-1) is known to interact with multiple host cellular proteins during its replication in the target cell. While many of these host cellular proteins facilitate viral replication, a number of them are reported to inhibit HIV-1 replication at various stages of its life cycle. These host cellular proteins, which are known as restriction factors, constitute an integral part of the host's first line of defence against the viral pathogen. Since the discovery of apolipoprotein B mRNA-editing enzyme 3G (APOBEC3G) as an HIV-1 restriction factor, several human proteins have been identified that exhibit anti-HIV-1 restriction. While each restriction factor employs a distinct mechanism of inhibition, the HIV-1 virus has equally evolved complex counter strategies to neutralize their inhibitory effect. APOBEC3G, tetherin, sterile alpha motif and histidine-aspartate domain 1 (SAMHD1), and trim-5α are some of the best known HIV-1 restriction factors that have been studied in great detail. Recently, six novel restriction factors were discovered that exhibit significant antiviral activity: endoplasmic reticulum α1,2-mannosidase I (ERManI), translocator protein (TSPO), guanylate-binding protein 5 (GBP5), serine incorporator (SERINC3/5) and zinc-finger antiviral protein (ZAP). The focus of this review is to discuss the antiviral mechanism of action of these six restriction factors and provide insights into the probable counter-evasion strategies employed by the HIV-1 virus. The recent discovery of new restriction factors substantiates the complex host-pathogen interactions occurring during HIV-1 pathogenesis and makes it imperative that further investigations are conducted to elucidate the molecular basis of HIV-1 replication.

  3. Modeling HIV-1 drug resistance as episodic directional selection.

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    Murrell, Ben; de Oliveira, Tulio; Seebregts, Chris; Kosakovsky Pond, Sergei L; Scheffler, Konrad

    2012-01-01

    The evolution of substitutions conferring drug resistance to HIV-1 is both episodic, occurring when patients are on antiretroviral therapy, and strongly directional, with site-specific resistant residues increasing in frequency over time. While methods exist to detect episodic diversifying selection and continuous directional selection, no evolutionary model combining these two properties has been proposed. We present two models of episodic directional selection (MEDS and EDEPS) which allow the a priori specification of lineages expected to have undergone directional selection. The models infer the sites and target residues that were likely subject to directional selection, using either codon or protein sequences. Compared to its null model of episodic diversifying selection, MEDS provides a superior fit to most sites known to be involved in drug resistance, and neither one test for episodic diversifying selection nor another for constant directional selection are able to detect as many true positives as MEDS and EDEPS while maintaining acceptable levels of false positives. This suggests that episodic directional selection is a better description of the process driving the evolution of drug resistance.

  4. Modeling HIV-1 drug resistance as episodic directional selection.

    Directory of Open Access Journals (Sweden)

    Ben Murrell

    Full Text Available The evolution of substitutions conferring drug resistance to HIV-1 is both episodic, occurring when patients are on antiretroviral therapy, and strongly directional, with site-specific resistant residues increasing in frequency over time. While methods exist to detect episodic diversifying selection and continuous directional selection, no evolutionary model combining these two properties has been proposed. We present two models of episodic directional selection (MEDS and EDEPS which allow the a priori specification of lineages expected to have undergone directional selection. The models infer the sites and target residues that were likely subject to directional selection, using either codon or protein sequences. Compared to its null model of episodic diversifying selection, MEDS provides a superior fit to most sites known to be involved in drug resistance, and neither one test for episodic diversifying selection nor another for constant directional selection are able to detect as many true positives as MEDS and EDEPS while maintaining acceptable levels of false positives. This suggests that episodic directional selection is a better description of the process driving the evolution of drug resistance.

  5. Phase I safety and immunogenicity evaluation of MVA-CMDR, a multigenic, recombinant modified vaccinia Ankara-HIV-1 vaccine candidate.

    Science.gov (United States)

    Currier, Jeffrey R; Ngauy, Viseth; de Souza, Mark S; Ratto-Kim, Silvia; Cox, Josephine H; Polonis, Victoria R; Earl, Patricia; Moss, Bernard; Peel, Sheila; Slike, Bonnie; Sriplienchan, Somchai; Thongcharoen, Prasert; Paris, Robert M; Robb, Merlin L; Kim, Jerome; Michael, Nelson L; Marovich, Mary A

    2010-11-15

    We conducted a Phase I randomized, dose-escalation, route-comparison trial of MVA-CMDR, a candidate HIV-1 vaccine based on a recombinant modified vaccinia Ankara viral vector expressing HIV-1 genes env/gag/pol. The HIV sequences were derived from circulating recombinant form CRF01_AE, which predominates in Thailand. The objective was to evaluate safety and immunogenicity of MVA-CMDR in human volunteers in the US and Thailand. MVA-CMDR or placebo was administered intra-muscularly (IM; 10(7) or 10(8) pfu) or intradermally (ID; 10(6) or 10(7) pfu) at months 0, 1 and 3, to 48 healthy volunteers at low risk for HIV-1 infection. Twelve volunteers in each dosage group were randomized to receive MVA-CMDR or placebo (10∶2). Volunteers were actively monitored for local and systemic reactogenicity and adverse events post vaccination. Cellular immunogenicity was assessed by a validated IFNγ Elispot assay, an intracellular cytokine staining assay, lymphocyte proliferation and a (51)Cr-release assay. Humoral immunogenicity was assessed by ADCC for gp120 and binding antibody ELISAs for gp120 and p24. MVA-CMDR was safe and well tolerated with no vaccine related serious adverse events. Cell-mediated immune responses were: (i) moderate in magnitude (median IFNγ Elispot of 78 SFC/10(6) PBMC at 10(8) pfu IM), but high in response rate (70% (51)Cr-release positive; 90% Elispot positive; 100% ICS positive, at 10(8) pfu IM); (ii) predominantly HIV Env-specific CD4(+) T cells, with a high proliferative capacity and durable for at least 6 months (100% LPA response rate by the IM route); (iv) dose- and route-dependent with 10(8) pfu IM being the most immunogenic treatment. Binding antibodies against gp120 and p24 were detectable in all vaccination groups with ADCC capacity detectable at the highest dose (40% positive at 10(8) pfu IM). MVA-CMDR delivered both intramuscularly and intradermally was safe, well-tolerated and elicited durable cell-mediated and humoral immune responses

  6. Phase I safety and immunogenicity evaluation of MVA-CMDR, a multigenic, recombinant modified vaccinia Ankara-HIV-1 vaccine candidate.

    Directory of Open Access Journals (Sweden)

    Jeffrey R Currier

    2010-11-01

    Full Text Available We conducted a Phase I randomized, dose-escalation, route-comparison trial of MVA-CMDR, a candidate HIV-1 vaccine based on a recombinant modified vaccinia Ankara viral vector expressing HIV-1 genes env/gag/pol. The HIV sequences were derived from circulating recombinant form CRF01_AE, which predominates in Thailand. The objective was to evaluate safety and immunogenicity of MVA-CMDR in human volunteers in the US and Thailand.MVA-CMDR or placebo was administered intra-muscularly (IM; 10(7 or 10(8 pfu or intradermally (ID; 10(6 or 10(7 pfu at months 0, 1 and 3, to 48 healthy volunteers at low risk for HIV-1 infection. Twelve volunteers in each dosage group were randomized to receive MVA-CMDR or placebo (10∶2. Volunteers were actively monitored for local and systemic reactogenicity and adverse events post vaccination. Cellular immunogenicity was assessed by a validated IFNγ Elispot assay, an intracellular cytokine staining assay, lymphocyte proliferation and a (51Cr-release assay. Humoral immunogenicity was assessed by ADCC for gp120 and binding antibody ELISAs for gp120 and p24. MVA-CMDR was safe and well tolerated with no vaccine related serious adverse events. Cell-mediated immune responses were: (i moderate in magnitude (median IFNγ Elispot of 78 SFC/10(6 PBMC at 10(8 pfu IM, but high in response rate (70% (51Cr-release positive; 90% Elispot positive; 100% ICS positive, at 10(8 pfu IM; (ii predominantly HIV Env-specific CD4(+ T cells, with a high proliferative capacity and durable for at least 6 months (100% LPA response rate by the IM route; (iv dose- and route-dependent with 10(8 pfu IM being the most immunogenic treatment. Binding antibodies against gp120 and p24 were detectable in all vaccination groups with ADCC capacity detectable at the highest dose (40% positive at 10(8 pfu IM.MVA-CMDR delivered both intramuscularly and intradermally was safe, well-tolerated and elicited durable cell-mediated and humoral immune responses

  7. HIV-1 Replication and the Cellular Eukaryotic Translation Apparatus

    Directory of Open Access Journals (Sweden)

    Santiago Guerrero

    2015-01-01

    Full Text Available Eukaryotic translation is a complex process composed of three main steps: initiation, elongation, and termination. During infections by RNA- and DNA-viruses, the eukaryotic translation machinery is used to assure optimal viral protein synthesis. Human immunodeficiency virus type I (HIV-1 uses several non-canonical pathways to translate its own proteins, such as leaky scanning, frameshifting, shunt, and cap-independent mechanisms. Moreover, HIV-1 modulates the host translation machinery by targeting key translation factors and overcomes different cellular obstacles that affect protein translation. In this review, we describe how HIV-1 proteins target several components of the eukaryotic translation machinery, which consequently improves viral translation and replication.

  8. HIV-1 Non-Nucleoside Reverse Transcriptase Inhibitors

    DEFF Research Database (Denmark)

    Vanangamudi, Murugesan; Poongavanam, Vasanthanathan; Namasivayam, Vigneshwaran

    2017-01-01

    BACKGROUND: Design of inhibitors for HIV-1 reverse transcriptase inhibition (HIV-1 RT) is one of the successful chemotherapies for the treatment of HIV infection. Among the inhibitors available for HIV-1 RT, non-nucleoside reverse transcriptase inhibitors (NNRTIs) have shown to be very promising......: The conformation dependent-alignment based (CoMFA and CoMSIA) methods have been proven very successful ligand based strategy in the drug design. Here, CoMFA and CoMSIA studies reported for structurally distinct NNRTIs including thiazolobenzimidazole, dipyridodiazepinone, 1,1,3-trioxo [1,2,4]-thiadiazine...

  9. Novel Latency Reversal Agents for HIV-1 Cure.

    Science.gov (United States)

    Spivak, Adam M; Planelles, Vicente

    2018-01-29

    Antiretroviral therapy (ART) has rendered HIV-1 infection a treatable illness; however, ART is not curative owing to the persistence of replication-competent, latent proviruses in long-lived resting T cells. Strategies that target these latently infected cells and allow immune recognition and clearance of this reservoir will be necessary to eradicate HIV-1 in infected individuals. This review describes current pharmacologic approaches to reactivate the latent reservoir so that infected cells can be recognized and targeted, with the ultimate goal of achieving an HIV-1 cure.

  10. Towards an HIV-1 cure: measuring the latent reservoir.

    Science.gov (United States)

    Bruner, Katherine M; Hosmane, Nina N; Siliciano, Robert F

    2015-04-01

    The latent reservoir (LR) of HIV-1 in resting memory CD4(+) T cells serves as a major barrier to curing HIV-1 infection. While many PCR- and culture-based assays have been used to measure the size of the LR, correlation between results of different assays is poor and recent studies indicate that no available assay provides an accurate measurement of reservoir size. The discrepancies between assays are a hurdle to clinical trials that aim to measure the efficacy of HIV-1 eradication strategies. Here we describe the advantages and disadvantages of various approaches to measuring the LR. Copyright © 2015 Elsevier Ltd. All rights reserved.

  11. Towards an HIV-1 cure: measuring the latent reservoir

    Science.gov (United States)

    Bruner, Katherine M.; Hosmane, Nina N.; Siliciano, Robert F.

    2015-01-01

    The latent reservoir of HIV-1 in resting memory CD4+ T cells serves as a major barrier to curing HIV-1 infection. While many PCR- and culture-based assays have been used to measure the size of the latent reservoir, correlation between results of different assays is poor and recent studies indicate that no available assay provides an accurate measurement of reservoir size. The discrepancies between assays are a hurdle to clinical trials that aim to measure the efficacy of HIV-1 eradication strategies. Here we describe the advantages and disadvantages of various approaches to measure the latent reservoir. PMID:25747663

  12. Stable 293 T and CHO cell lines expressing cleaved, stable HIV-1 envelope glycoprotein trimers for structural and vaccine studies

    NARCIS (Netherlands)

    Chung, Nancy P. Y.; Matthews, Katie; Kim, Helen J.; Ketas, Thomas J.; Golabek, Michael; de Los Reyes, Kevin; Korzun, Jacob; Yasmeen, Anila; Sanders, Rogier W.; Klasse, Per Johan; Wilson, Ian A.; Ward, Andrew B.; Marozsan, Andre J.; Moore, John P.; Cupo, Albert

    2014-01-01

    Recombinant soluble, cleaved HIV-1 envelope glycoprotein SOSIP.664 gp140 trimers based on the subtype A BG505 sequence are being studied structurally and tested as immunogens in animals. For these trimers to become a vaccine candidate for human trials, they would need to be made in appropriate

  13. A Combinatorial CRISPR-Cas9 Attack on HIV-1 DNA Extinguishes All Infectious Provirus in Infected T Cell Cultures

    NARCIS (Netherlands)

    Wang, Gang; Zhao, Na; Berkhout, Ben; Das, Atze T.

    2016-01-01

    Current drug therapies effectively suppress HIV-1 replication but do not inactivate the provirus that persists in latent reservoirs. Recent studies have found that the guide RNA (gRNA)-directed CRISPR/Cas9 system can be used for sequence-specific attack on this proviral DNA. Although potent

  14. Human antibody response to a strain-specific HIV-1 gp120 epitope associated with cell fusion inhibition

    NARCIS (Netherlands)

    Goudsmit, J.; Boucher, C. A.; Meloen, R. H.; Epstein, L. G.; Smit, L.; van der Hoek, L.; Bakker, M.

    1988-01-01

    PEPSCAN analysis, performed using 536 overlapping nonapeptides derived from the HTLV-III B nucleotide sequence of the region encoding the external envelope protein of 120 kDa (gp120), identified in the V3 region of gp120 a major binding site for antibodies of HIV-1-infected humans. The minimal amino

  15. Tracing the HIV-1 subtype B mobility in Europe: a phylogeographic approach

    Energy Technology Data Exchange (ETDEWEB)

    Leitner, Thomas [Los Alamos National Laboratory; Paraskevis, D [KATHOLIEKE UNIV; Pybus, O [UNIV OF OXFORD; Magiorkinis, G [KATHOLIEKE UNIV; Hatzakis, A [KATHOLIEKE UNIV

    2008-01-01

    The prevalence and the origin of HIV-1 subtype B, the most prevalent circulating clade among the long-term residents in Europe, have been studied extensively. However the spatial diffusion of the epidemic from the perspective of the virus has not previously been traced. In the current study we inferred the migration history of HIV-1 subtype B by way of a phylogeography of viral sequences sampled from 16 European countries and Israel. Migration events were inferred from viral phylogenies by character reconstruction using parsimony. With regard to the spatial dispersal of the HIV subtype B sequences across viral phylogenies, in most of the countries in Europe the epidemic was introduced by multiple sources and subsequently spread within local networks. Poland provides an exception where most of the infections were the result of a single point introduction. According to the significant migratory pathways, we show that there are considerable differences across Europe. Specifically, Greece, Portugal, Serbia and Spain, provide sources shedding HIV-1; Austria, Belgium and Luxembourg, on the other hand, are migratory targets, while for Denmark, Germany, Italy, Israel, Norway, the Netherlands, Sweden, Switzerland and the UK we inferred significant bidirectional migration. For Poland no significant migratory pathways were inferred. Subtype B phylogeographies provide a new insight about the geographical distribution of viral lineages, as well as the significant pathways of virus dispersal across Europe, suggesting that intervention strategies should also address tourists, travellers and migrants.

  16. A Rough Set-Based Model of HIV-1 Reverse Transcriptase Resistome

    Directory of Open Access Journals (Sweden)

    Marcin Kierczak

    2009-10-01

    Full Text Available Reverse transcriptase (RT is a viral enzyme crucial for HIV-1 replication. Currently, 12 drugs are targeted against the RT. The low fidelity of the RT-mediated transcription leads to the quick accumulation of drug-resistance mutations. The sequence-resistance relationship remains only partially understood. Using publicly available data collected from over 15 years of HIV proteome research, we have created a general and predictive rule-based model of HIV-1 resistance to eight RT inhibitors. Our rough set-based model considers changes in the physicochemical properties of a mutated sequence as compared to the wild-type strain. Thanks to the application of the Monte Carlo feature selection method, the model takes into account only the properties that significantly contribute to the resistance phenomenon. The obtained results show that drug-resistance is determined in more complex way than believed. We confirmed the importance of many resistance-associated sites, found some sites to be less relevant than formerly postulated and— more importantly—identified several previously neglected sites as potentially relevant. By mapping some of the newly discovered sites on the 3D structure of the RT, we were able to suggest possible molecular-mechanisms of drug-resistance. Importantly, our model has the ability to generalize predictions to the previously unseen cases. The study is an example of how computational biology methods can increase our understanding of the HIV-1 resistome.

  17. Tracing the HIV-1 subtype B mobility in Europe: a phylogeographic approach

    Directory of Open Access Journals (Sweden)

    Perrin Luc

    2009-05-01

    Full Text Available Abstract Background The prevalence and the origin of HIV-1 subtype B, the most prevalent circulating clade among the long-term residents in Europe, have been studied extensively. However the spatial diffusion of the epidemic from the perspective of the virus has not previously been traced. Results In the current study we inferred the migration history of HIV-1 subtype B by way of a phylogeography of viral sequences sampled from 16 European countries and Israel. Migration events were inferred from viral phylogenies by character reconstruction using parsimony. With regard to the spatial dispersal of the HIV subtype B sequences across viral phylogenies, in most of the countries in Europe the epidemic was introduced by multiple sources and subsequently spread within local networks. Poland provides an exception where most of the infections were the result of a single point introduction. According to the significant migratory pathways, we show that there are considerable differences across Europe. Specifically, Greece, Portugal, Serbia and Spain, provide sources shedding HIV-1; Austria, Belgium and Luxembourg, on the other hand, are migratory targets, while for Denmark, Germany, Italy, Israel, Norway, the Netherlands, Sweden, Switzerland and the UK we inferred significant bidirectional migration. For Poland no significant migratory pathways were inferred. Conclusion Subtype B phylogeographies provide a new insight about the geographical distribution of viral lineages, as well as the significant pathways of virus dispersal across Europe, suggesting that intervention strategies should also address tourists, travellers and migrants.

  18. The molecular epidemiology of HIV-1 in the Comunidad Valenciana (Spain): analysis of transmission clusters.

    Science.gov (United States)

    Patiño-Galindo, Juan Ángel; Torres-Puente, Manoli; Bracho, María Alma; Alastrué, Ignacio; Juan, Amparo; Navarro, David; Galindo, María José; Ocete, Dolores; Ortega, Enrique; Gimeno, Concepción; Belda, Josefina; Domínguez, Victoria; Moreno, Rosario; González-Candelas, Fernando

    2017-09-14

    HIV infections are still a very serious concern for public heath worldwide. We have applied molecular evolution methods to study the HIV-1 epidemics in the Comunidad Valenciana (CV, Spain) from a public health surveillance perspective. For this, we analysed 1804 HIV-1 sequences comprising protease and reverse transcriptase (PR/RT) coding regions, sampled between 2004 and 2014. These sequences were subtyped and subjected to phylogenetic analyses in order to detect transmission clusters. In addition, univariate and multinomial comparisons were performed to detect epidemiological differences between HIV-1 subtypes, and risk groups. The HIV epidemic in the CV is dominated by subtype B infections among local men who have sex with men (MSM). 270 transmission clusters were identified (>57% of the dataset), 12 of which included ≥10 patients; 11 of subtype B (9 affecting MSMs) and one (n = 21) of CRF14, affecting predominately intravenous drug users (IDUs). Dated phylogenies revealed these large clusters to have originated from the mid-80s to the early 00 s. Subtype B is more likely to form transmission clusters than non-B variants and MSMs to cluster than other risk groups. Multinomial analyses revealed an association between non-B variants, which are not established in the local population yet, and different foreign groups.

  19. Structural Study of a New HIV-1 Entry Inhibitor and Interaction with the HIV-1 Fusion Peptide in Dodecylphosphocholine Micelles.

    Science.gov (United States)

    Pérez, Yolanda; Gómara, Maria José; Yuste, Eloísa; Gómez-Gutierrez, Patricia; Pérez, Juan Jesús; Haro, Isabel

    2017-08-25

    Previous studies support the hypothesis that the envelope GB virus C (GBV-C) E1 protein interferes the HIV-1 entry and that a peptide, derived from the region 139-156 of this protein, has been defined as a novel HIV-1 entry inhibitor. In this work, we firstly focus on the characterization of the structural features of this peptide, which are determinant for its anti-HIV-1 activity and secondly, on the study of its interaction with the proposed viral target (i.e., the HIV-1 fusion peptide). We report the structure of the peptide determined by NMR spectroscopy in dodecylphosphocholine (DPC) micelles solved by using restrained molecular dynamics calculations. The acquisition of different NMR experiments in DPC micelles (i.e., peptide-peptide titration, diffusion NMR spectroscopy, and addition of paramagnetic relaxation agents) allows a proposal of an inhibition mechanism. We conclude that a 18-mer peptide from the non-pathogenic E1 GBV-C protein, with a helix-turn-helix structure inhibits HIV-1 by binding to the HIV-1 fusion peptide at the membrane level, thereby interfering with those domains in the HIV-1, which are critical for stabilizing the six-helix bundle formation in a membranous environment. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Short Communication: Evolution of HIV-1 Coreceptor Usage and Coreceptor Switching During Pregnancy

    OpenAIRE

    Ransy, Doris G.; Motorina, Alena; Merindol, Natacha; Akouamba, Bertine S.; Samson, Johanne; Lie, Yolanda; Napolitano, Laura A.; Lapointe, Normand; Boucher, Marc; Soudeyns, Hugo

    2014-01-01

    Coreceptor switch from CCR5 to CXCR4 is associated with HIV disease progression. To document the evolution of coreceptor tropism during pregnancy, a longitudinal study of envelope gene sequences was performed in a group of pregnant women infected with HIV-1 of clade B (n=10) or non-B (n=9). Polymerase chain reaction (PCR) amplification of the V1-V3 region was performed on plasma viral RNA, followed by cloning and sequencing. Using geno2pheno and PSSMX4R5, the presence of X4 variants was predi...

  1. Compartmentalization of HIV-1 within the female genital tract is due to monotypic and low-diversity variants not distinct viral populations.

    Science.gov (United States)

    Bull, Marta; Learn, Gerald; Genowati, Indira; McKernan, Jennifer; Hitti, Jane; Lockhart, David; Tapia, Kenneth; Holte, Sarah; Dragavon, Joan; Coombs, Robert; Mullins, James; Frenkel, Lisa

    2009-09-22

    Compartmentalization of HIV-1 between the genital tract and blood was noted in half of 57 women included in 12 studies primarily using cell-free virus. To further understand differences between genital tract and blood viruses of women with chronic HIV-1 infection cell-free and cell-associated virus populations were sequenced from these tissues, reasoning that integrated viral DNA includes variants archived from earlier in infection, and provides a greater array of genotypes for comparisons. Multiple sequences from single-genome-amplification of HIV-1 RNA and DNA from the genital tract and blood of each woman were compared in a cross-sectional study. Maximum likelihood phylogenies were evaluated for evidence of compartmentalization using four statistical tests. Genital tract and blood HIV-1 appears compartmentalized in 7/13 women by >/=2 statistical analyses. These subjects' phylograms were characterized by low diversity genital-specific viral clades interspersed between clades containing both genital and blood sequences. Many of the genital-specific clades contained monotypic HIV-1 sequences. In 2/7 women, HIV-1 populations were significantly compartmentalized across all four statistical tests; both had low diversity genital tract-only clades. Collapsing monotypic variants into a single sequence diminished the prevalence and extent of compartmentalization. Viral sequences did not demonstrate tissue-specific signature amino acid residues, differential immune selection, or co-receptor usage. In women with chronic HIV-1 infection multiple identical sequences suggest proliferation of HIV-1-infected cells, and low diversity tissue-specific phylogenetic clades are consistent with bursts of viral replication. These monotypic and tissue-specific viruses provide statistical support for compartmentalization of HIV-1 between the female genital tract and blood. However, the intermingling of these clades with clades comprised of both genital and blood sequences and the absence

  2. Molecular Epidemiology of HIV-1 Infection among Men who Have Sex with Men in Taiwan in 2012

    Science.gov (United States)

    Huang, Szu-Wei; Wang, Sheng-Fan; Cowó, Ángel E.; Chen, Marcelo; Lin, Yu-Ting; Hung, Chun-Po; Chen, Yi-Hsien; Yang, Jyh-Yuan; Tang, Hung-Jen; Chen, Yi-Ming Arthur

    2015-01-01

    The number of men who have sex with men (MSM) infected with HIV-1 in Taiwan has increased rapidly in the past few years. The goal of this study was to conduct a molecular epidemiological study of HIV-1 infection among MSM in Taiwan to identify risk factors for intervention. Voluntary counseling program and anonymous testing were provided to patrons at 1 gay bar, 7 night clubs and 3 gay saunas in Taipei and New Taipei Cities in 2012. HIV-1 subtypes were determined using gag subtype-specific PCR and phylogenetic analysis by env sequences. Recent HIV-1 infection was determined using LAg-Avidity EIA. In-depth interviews and questionnaires were used to identify risk factors. The prevalence and incidence of HIV-1 among MSM in Taiwan were 4.38% (53/1,208) and 3.29 per 100 person-years, respectively. Of 49 cases genotyped, 48 (97.9%) were infected with subtype B and 1 with CRF01_AE (2%). Phylogenetic analysis of 46 HIV-1 strains showed that 25 (54.4%) subtype B strains formed 9 clusters with each other or with other local strains. The CRF01_AE case clustered with a reference strain from a Thai blood donor with bootstrap value of 99. Multivariate logistic regression analysis showed that risk factors associated with HIV-1 infection included use of oil-based solution as lubricant (vs. saliva or water-based lubricants, OR= 4.23; p Taiwan in 2012. Misuse of oil-based solution as lubricant is a new risk factor identified among MSM in Taiwan. The Taiwan’s Centers for Disease Control has created a video (www.youtube.com/watch?v=BinExvvOTMM&feature=iv&src_vid=BW81-PfmY3E&annotation_id=annotation_2436493705) to correct such misconception in its AIDS prevention campaign. PMID:26039757

  3. Varied sensitivity to therapy of HIV-1 strains in CD4+ lymphocyte sub-populations upon ART initiation

    Directory of Open Access Journals (Sweden)

    Paxton William A

    2010-12-01

    Full Text Available Abstract Background Although antiretroviral therapy (ART has proven its success against HIV-1, the long lifespan of infected cells and viral latency prevent eradication. In this study we analyzed the sensitivity to ART of HIV-1 strains in naïve, central memory and effector memory CD4+ lymphocyte subsets. Methods From five patients cellular HIV-1 infection levels were quantified before and after initiation of therapy (2-5 weeks. Through sequencing the C2V3 region of the HIV-1 gp120 envelope, we studied the effect of short-term therapy on virus variants derived from naïve, central memory and effector memory CD4+ lymphocyte subsets. Results During short-term ART, HIV-1 infection levels declined in all lymphocyte subsets but not as much as RNA levels in serum. Virus diversity in the naïve and central memory lymphocyte populations remained unchanged, whilst diversity decreased in serum and the effector memory lymphocytes. ART differentially affected the virus populations co-circulating in one individual harboring a dual HIV-1 infection. Changes in V3 charge were found in all individuals after ART initiation with increases within the effector memory subset and decreases found in the naïve cell population. Conclusions During early ART virus diversity is affected mainly in the serum and effector memory cell compartments. Differential alterations in V3 charge were observed between effector memory and naïve populations. While certain cell populations can be targeted preferentially during early ART, some virus strains demonstrate varied sensitivity to therapy, as shown from studying two strains within a dual HIV-1 infected individual.

  4. Molecular Epidemiology of HIV-1 Infection among Men who Have Sex with Men in Taiwan in 2012.

    Science.gov (United States)

    Huang, Szu-Wei; Wang, Sheng-Fan; Cowó, Ángel E; Chen, Marcelo; Lin, Yu-Ting; Hung, Chun-Po; Chen, Yi-Hsien; Yang, Jyh-Yuan; Tang, Hung-Jen; Chen, Yi-Ming Arthur

    2015-01-01

    The number of men who have sex with men (MSM) infected with HIV-1 in Taiwan has increased rapidly in the past few years. The goal of this study was to conduct a molecular epidemiological study of HIV-1 infection among MSM in Taiwan to identify risk factors for intervention. Voluntary counseling program and anonymous testing were provided to patrons at 1 gay bar, 7 night clubs and 3 gay saunas in Taipei and New Taipei Cities in 2012. HIV-1 subtypes were determined using gag subtype-specific PCR and phylogenetic analysis by env sequences. Recent HIV-1 infection was determined using LAg-Avidity EIA. In-depth interviews and questionnaires were used to identify risk factors. The prevalence and incidence of HIV-1 among MSM in Taiwan were 4.38% (53/1,208) and 3.29 per 100 person-years, respectively. Of 49 cases genotyped, 48 (97.9%) were infected with subtype B and 1 with CRF01_AE (2%). Phylogenetic analysis of 46 HIV-1 strains showed that 25 (54.4%) subtype B strains formed 9 clusters with each other or with other local strains. The CRF01_AE case clustered with a reference strain from a Thai blood donor with bootstrap value of 99. Multivariate logistic regression analysis showed that risk factors associated with HIV-1 infection included use of oil-based solution as lubricant (vs. saliva or water-based lubricants, OR= 4.23; p watch?v=BinExvvOTMM&feature=iv&src_vid=BW81-PfmY3E&annotation_id=annotation_2436493705) to correct such misconception in its AIDS prevention campaign.

  5. Molecular and phylogenetic analysis of HIV-1 variants circulating among injecting drug users in Mashhad-Iran

    Directory of Open Access Journals (Sweden)

    Buonaguro FM

    2006-09-01

    Full Text Available Abstract Genetic and phylogenetic information on the HIV-1 epidemic in Middle-East Countries, and in particular in Iran, are extremely limited. By March 2004, the Iranian Ministry of Health officially reported a cumulative number of 6'532 HIV positive individuals and 214 AIDS cases in the Iranian HIV-1 epidemic. The intra-venous drug users (IDUs represent the group at highest risk for HIV-1 infection in Iran, accounting for almost 63% of all HIV-infected population. In this regards, a molecular phylogenetic study has been performed on a sentinel cohort of HIV-1 seropositive IDUs enrolled at the end of 2005 at the University of Mashhad, the largest city North East of Tehran. The study has been performed on both gag and env subgenomic regions amplified by Polymerase Chain Reaction (PCR from peripheral blood mononuclear cells (PBMCs and characterized by direct DNA sequence analysis. The results reported here show that the HIV-1 subtype A is circulating in this IDUs sentinel cohort. Moreover, the single phylogenetic cluster as well as the intra-group low nucleotide divergence is indicative of a recent outbreak. Unexpectedly, the Iranian samples appear to be phylogenetically derived from African Sub-Saharan subtype A viruses, raising stirring speculations on HIV-1 introduction into the IDUs epidemic in Mashhad. This sentinel study could represent the starting point for a wider molecular survey of the HIV-1 epidemics in Iran to evaluate in detail the distribution of genetic subtypes and possible natural drug-resistant variants, which are extremely helpful information to design diagnostic and therapeutic strategies.

  6. HIV-1 subtype F1 epidemiological networks among Italian heterosexual males are associated with introduction events from South America.

    Science.gov (United States)

    Lai, Alessia; Simonetti, Francesco R; Zehender, Gianguglielmo; De Luca, Andrea; Micheli, Valeria; Meraviglia, Paola; Corsi, Paola; Bagnarelli, Patrizia; Almi, Paolo; Zoncada, Alessia; Paolucci, Stefania; Gonnelli, Angela; Colao, Grazia; Tacconi, Danilo; Franzetti, Marco; Ciccozzi, Massimo; Zazzi, Maurizio; Balotta, Claudia

    2012-01-01

    About 40% of the Italian HIV-1 epidemic due to non-B variants is sustained by F1 clade, which circulates at high prevalence in South America and Eastern Europe. Aim of this study was to define clade F1 origin, population dynamics and epidemiological networks through phylogenetic approaches. We analyzed pol sequences of 343 patients carrying F1 subtype stored in the ARCA database from 1998 to 2009. Citizenship of patients was as follows: 72.6% Italians, 9.3% South Americans and 7.3% Rumanians. Heterosexuals, Homo-bisexuals, Intravenous Drug Users accounted for 58.1%, 24.0% and 8.8% of patients, respectively. Phylogenetic analysis indicated that 70% of sequences clustered in 27 transmission networks. Two distinct groups were identified; the first clade, encompassing 56 sequences, included all Rumanian patients. The second group involved the remaining clusters and included 10 South American Homo-bisexuals in 9 distinct clusters. Heterosexual modality of infection was significantly associated with the probability to be detected in transmission networks. Heterosexuals were prevalent either among Italians (67.2%) or Rumanians (50%); by contrast, Homo-bisexuals accounted for 71.4% of South Americans. Among patients with resistant strains the proportion of clustering sequences was 57.1%, involving 14 clusters (51.8%). Resistance in clusters tended to be higher in South Americans (28.6%) compared to Italian (17.7%) and Rumanian patients (14.3%). A striking proportion of epidemiological networks could be identified in heterosexuals carrying F1 subtype residing in Italy. Italian Heterosexual males predominated within epidemiological clusters while foreign patients were mainly Heterosexual Rumanians, both males and females, and South American Homo-bisexuals. Tree topology suggested that F1 variant from South America gave rise to the Italian F1 epidemic through multiple introduction events. The contact tracing also revealed an unexpected burden of resistance in epidemiological

  7. Multiple barriers to recombination between divergent HIV-1 variants revealed by a dual-marker recombination assay

    DEFF Research Database (Denmark)

    Nikolaitchik, Olga A; Galli, Andrea; Moore, Michael D

    2011-01-01

    Recombination is a major force for generating human immunodeficiency virus type 1 (HIV-1) diversity and produces numerous recombinants circulating in the human population. We previously established a cell-based system using green fluorescent protein gene (gfp) as a reporter to study the mechanisms...... of HIV-1 recombination. We now report an improved system capable of detecting recombination using authentic viral sequences. Frameshift mutations were introduced into the gag gene so that parental viruses do not express full-length Gag; however, recombination can generate a progeny virus that expresses...

  8. Induction of novel CD8+ T-cell responses during chronic untreated HIV-1 infection by immunization with subdominant cytotoxic T-lymphocyte epitopes

    DEFF Research Database (Denmark)

    Kloverpris, Henrik; Karlsson, Ingrid; Bonde, Jesper

    2009-01-01

    OBJECTIVE:: To investigate the potential to induce additional cytotoxic T-lymphocyte (CTL) immunity during chronic HIV-1 infection. DESIGN:: We selected infrequently targeted or subdominant but conserved HLA-A*0201-binding epitopes in Gag, Pol, Env, Vpu and Vif. These relatively immune silent...... epitopes were modified as anchor-optimized peptides to improve immunogenicity and delivered on autologous monocyte-derived dendritic cells (MDDCs). METHODS:: Twelve treatment-naïve HLA-A*0201 HIV-1-infected Danish individuals received 1 x 10 MDDCs subcutaneously (s.c.) (weeks 0, 2, 4 and 8), pulsed......-cell counts was observed. CONCLUSION:: These data show that it is possible to generate new T-cell responses in treatment-naive HIV-1-infected individuals despite high viral loads, and thereby redirect immunity to target new multiple and rationally selected subdominant CTL epitopes. Further optimization could...

  9. Hepatitis B Virus Vaccination in HIV-1-Infected Young Adults: A Tool to Reduce the Size of HIV-1 Reservoirs?

    Science.gov (United States)

    Bekele, Yonas; Graham, Rebecka Lantto; Soeria-Atmadja, Sandra; Nasi, Aikaterini; Zazzi, Maurizio; Vicenti, Ilaria; Naver, Lars; Nilsson, Anna; Chiodi, Francesca

    2017-01-01

    During anti-retroviral therapy (ART) HIV-1 persists in cellular reservoirs, mostly represented by CD4+ memory T cells. Several approaches are currently being undertaken to develop a cure for HIV-1 infection through elimination (or reduction) of these reservoirs. Few studies have so far been conducted to assess the possibility of reducing the size of HIV-1 reservoirs through vaccination in virologically controlled HIV-1-infected children. We recently conducted a vaccination study with a combined hepatitis A virus (HAV) and hepatitis B virus (HBV) vaccine in 22 HIV-1-infected children. We assessed the size of the virus reservoir, measured as total HIV-1 DNA copies in blood cells, pre- and postvaccination. In addition, we investigated by immunostaining whether the frequencies of CD4+ and CD8+ T cells and parameters of immune activation and proliferation on these cells were modulated by vaccination. At 1 month from the last vaccination dose, we found that 20 out of 22 children mounted a serological response to HBV; a majority of children had antibodies against HAV at baseline. The number of HIV-1 DNA copies in blood at 1 month postvaccination was reduced in comparison to baseline although this reduction was not statistically significant. A significant reduction of HIV-1 DNA copies in blood following vaccination was found in 12 children. The frequencies of CD4+ (naïve, effector memory) and CD8+ (central memory) T-cell subpopulations changed following vaccinations and a reduction in the activation and proliferation pattern of these cells was also noticed. Multivariate linear regression analysis revealed that the frequency of CD8+ effector memory T cells prior to vaccination was strongly predictive of the reduction of HIV-1 DNA copies in blood following vaccination of the 22 HIV-1-infected children. The results of this study suggest a beneficial effect of vaccination to reduce the size of virus reservoir in HIV-1-infected children receiving ART. A reduced frequency of

  10. Hepatitis B Virus Vaccination in HIV-1-Infected Young Adults: A Tool to Reduce the Size of HIV-1 Reservoirs?

    Directory of Open Access Journals (Sweden)

    Yonas Bekele

    2018-01-01

    Full Text Available During anti-retroviral therapy (ART HIV-1 persists in cellular reservoirs, mostly represented by CD4+ memory T cells. Several approaches are currently being undertaken to develop a cure for HIV-1 infection through elimination (or reduction of these reservoirs. Few studies have so far been conducted to assess the possibility of reducing the size of HIV-1 reservoirs through vaccination in virologically controlled HIV-1-infected children. We recently conducted a vaccination study with a combined hepatitis A virus (HAV and hepatitis B virus (HBV vaccine in 22 HIV-1-infected children. We assessed the size of the virus reservoir, measured as total HIV-1 DNA copies in blood cells, pre- and postvaccination. In addition, we investigated by immunostaining whether the frequencies of CD4+ and CD8+ T cells and parameters of immune activation and proliferation on these cells were modulated by vaccination. At 1 month from the last vaccination dose, we found that 20 out of 22 children mounted a serological response to HBV; a majority of children had antibodies against HAV at baseline. The number of HIV-1 DNA copies in blood at 1 month postvaccination was reduced in comparison to baseline although this reduction was not statistically significant. A significant reduction of HIV-1 DNA copies in blood following vaccination was found in 12 children. The frequencies of CD4+ (naïve, effector memory and CD8+ (central memory T-cell subpopulations changed following vaccinations and a reduction in the activation and proliferation pattern of these cells was also noticed. Multivariate linear regression analysis revealed that the frequency of CD8+ effector memory T cells prior to vaccination was strongly predictive of the reduction of HIV-1 DNA copies in blood following vaccination of the 22 HIV-1-infected children. The results of this study suggest a beneficial effect of vaccination to reduce the size of virus reservoir in HIV-1-infected children receiving ART. A reduced

  11. Hepatitis B Virus Vaccination in HIV-1-Infected Young Adults: A Tool to Reduce the Size of HIV-1 Reservoirs?

    Science.gov (United States)

    Bekele, Yonas; Graham, Rebecka Lantto; Soeria-Atmadja, Sandra; Nasi, Aikaterini; Zazzi, Maurizio; Vicenti, Ilaria; Naver, Lars; Nilsson, Anna; Chiodi, Francesca

    2018-01-01

    During anti-retroviral therapy (ART) HIV-1 persists in cellular reservoirs, mostly represented by CD4+ memory T cells. Several approaches are currently being undertaken to develop a cure for HIV-1 infection through elimination (or reduction) of these reservoirs. Few studies have so far been conducted to assess the possibility of reducing the size of HIV-1 reservoirs through vaccination in virologically controlled HIV-1-infected children. We recently conducted a vaccination study with a combined hepatitis A virus (HAV) and hepatitis B virus (HBV) vaccine in 22 HIV-1-infected children. We assessed the size of the virus reservoir, measured as total HIV-1 DNA copies in blood cells, pre- and postvaccination. In addition, we investigated by immunostaining whether the frequencies of CD4+ and CD8+ T cells and parameters of immune activation and proliferation on these cells were modulated by vaccination. At 1 month from the last vaccination dose, we found that 20 out of 22 children mounted a serological response to HBV; a majority of children had antibodies against HAV at baseline. The number of HIV-1 DNA copies in blood at 1 month postvaccination was reduced in comparison to baseline although this reduction was not statistically significant. A significant reduction of HIV-1 DNA copies in blood following vaccination was found in 12 children. The frequencies of CD4+ (naïve, effector memory) and CD8+ (central memory) T-cell subpopulations changed following vaccinations and a reduction in the activation and proliferation pattern of these cells was also noticed. Multivariate linear regression analysis revealed that the frequency of CD8+ effector memory T cells prior to vaccination was strongly predictive of the reduction of HIV-1 DNA copies in blood following vaccination of the 22 HIV-1-infected children. The results of this study suggest a beneficial effect of vaccination to reduce the size of virus reservoir in HIV-1-infected children receiving ART. A reduced frequency of

  12. Therapeutic strategies to fight HIV-1 latency: progress and challenges

    CSIR Research Space (South Africa)

    Manoto, Sello L

    2017-10-01

    Full Text Available —1112, 2017 Therapeutic strategies to fight HIV-1 latency: progress and challenges Sello Lebohang Manoto, Lebogang Thobakgale, Rudzani Malabi, Charles Maphanga, Saturnin Ombinda-Lemboumba, Patience Mthunzi-Kufa Abstract: The life...

  13. Distribution of HIV-1 resistance-conferring polymorphic alleles SDF ...

    Indian Academy of Sciences (India)

    Unknown

    involved with delay in disease progression. ... proteins in 525 healthy individuals without any history of HIV-1 infection from 11 diverse populations of ... in three populations (Yamani, Pathan and Kamma), all in low frequencies (i.e. 1% to 3%).

  14. The latest evidence for possible HIV-1 curative strategies.

    Science.gov (United States)

    Pham, Hanh Thi; Mesplède, Thibault

    2018-01-01

    Human immunodeficiency virus type 1 (HIV-1) infection remains a major health issue worldwide. In developed countries, antiretroviral therapy has extended its reach from treatment of people living with HIV-1 to post-exposure prophylaxis, treatment as prevention, and, more recently, pre-exposure prophylaxis. These healthcare strategies offer the epidemiological tools to curve the epidemic in rich settings and will be concomitantly implemented in developing countries. One of the remaining challenges is to identify an efficacious curative strategy. This review manuscript will focus on some of the current curative strategies aiming at providing a sterilizing or functional cure to HIV-1-positive individuals. These include the following: early treatment initiation in post-treatment controllers as a long-term HIV-1 remission strategy, latency reversal, gene editing with or without stem cell transplantation, and antibodies against either the viral envelope protein or the host integrin α4β7.

  15. Towards HIV-1 remission: potential roles for broadly neutralizing antibodies.

    Science.gov (United States)

    Halper-Stromberg, Ariel; Nussenzweig, Michel C

    2016-02-01

    Current antiretroviral drug therapies do not cure HIV-1 because they do not eliminate a pool of long-lived cells harboring immunologically silent but replication-competent proviruses - termed the latent reservoir. Eliminating this reservoir and stimulating the immune response to control infection in the absence of therapy remain important but unsolved goals of HIV-1 cure research. Recently discovered broadly neutralizing antibodies (bNAbs) exhibit remarkable breadth and potency in their ability to neutralize HIV-1 in vitro, and recent studies have demonstrated new therapeutic applications for passively administered bNAbs in vivo. This Review discusses the roles bNAbs might play in HIV-1 treatment regimens, including prevention, therapy, and cure.

  16. Human immunodeficiency virus type-1 (HIV-1) genetic diversity and ...

    African Journals Online (AJOL)

    PROGMANAGER

    2013-04-24

    Apr 24, 2013 ... objective of this study was to determine the genetic diversity of HIV-1 and the prevalence of antiretroviral (ARV) ... individuals in resource limited settings. Key words: ... management of HIV infection even as antiretroviral (ARV).

  17. Purinergic Receptors: Key Mediators of HIV-1 infection and inflammation

    Directory of Open Access Journals (Sweden)

    Talia H Swartz

    2015-11-01

    Full Text Available Human immunodeficiency virus (HIV-1 causes a chronic infection that afflicts more than 38 million individuals worldwide. While the infection can be suppressed with potent anti-retroviral therapies, individuals infected with HIV have elevated levels of inflammation as indicated by increased T cell activation, soluble biomarkers, and associated morbidity and mortality. A single mechanism linking HIV pathogenesis to this inflammation has yet to be identified. Purinergic receptors are known to mediate inflammation and have been shown to be required for HIV-1 infection at the level of HIV-1 membrane fusion. Here we review the literature on the role of purinergic receptors in HIV-1 infection and associated inflammation and describe a role for these receptors as potential therapeutic targets.

  18. Profile of the HIV epidemic in Cape Verde: molecular epidemiology and drug resistance mutations among HIV-1 and HIV-2 infected patients from distinct islands of the archipelago.

    Science.gov (United States)

    de Pina-Araujo, Isabel Inês M; Guimarães, Monick L; Bello, Gonzalo; Vicente, Ana Carolina P; Morgado, Mariza G

    2014-01-01

    HIV-1 and HIV-2 have been detected in Cape Verde since 1987, but little is known regarding the genetic diversity of these viruses in this archipelago, located near the West African coast. In this study, we characterized the molecular epidemiology of HIV-1 and HIV-2 and described the occurrence of drug resistance mutations (DRM) among antiretroviral therapy naïve (ARTn) patients and patients under treatment (ARTexp) from different Cape Verde islands. Blood samples, socio-demographic and clinical-laboratory data were obtained from 221 HIV-positive individuals during 2010-2011. Phylogenetic and bootscan analyses of the pol region (1300 bp) were performed for viral subtyping. HIV-1 and HIV-2 DRM were evaluated for ARTn and ARTexp patients using the Stanford HIV Database and HIV-GRADE e.V. Algorithm Homepage, respectively. Among the 221 patients (169 [76.5%] HIV-1, 43 [19.5%] HIV-2 and 9 [4.1%] HIV-1/HIV-2 co-infections), 67% were female. The median ages were 34 (IQR = 1-75) and 47 (IQR = 12-84) for HIV-1 and HIV-2, respectively. HIV-1 infections were due to subtypes G (36.6%), CRF02_AG (30.6%), F1 (9.7%), URFs (10.4%), B (5.2%), CRF05_DF (3.0%), C (2.2%), CRF06_cpx (0.7%), CRF25_cpx (0.7%) and CRF49_cpx (0.7%), whereas all HIV-2 infections belonged to group A. Transmitted DRM (TDRM) was observed in 3.4% (2/58) of ARTn HIV-1-infected patients (1.7% NRTI, 1.7% NNRTI), but not among those with HIV-2. Among ARTexp patients, DRM was observed in 47.8% (33/69) of HIV-1 (37.7% NRTI, 37.7% NNRTI, 7.4% PI, 33.3% for two classes) and 17.6% (3/17) of HIV-2-infections (17.6% NRTI, 11.8% PI, 11.8% both). This study indicates that Cape Verde has a complex and unique HIV-1 molecular epidemiological scenario dominated by HIV-1 subtypes G, CRF02_AG and F1 and HIV-2 subtype A. The occurrence of TDRM and the relatively high level of DRM among treated patients are of concern. Continuous monitoring of patients on ART, including genotyping, are public policies to be implemented.

  19. Determination of HIV-1 co-receptor usage.

    Science.gov (United States)

    Cavarelli, Mariangela; Scarlatti, Gabriella

    2014-01-01

    Human immunodeficiency virus type I (HIV-1) infects target cells through interaction with the CD4 molecule and chemokine receptors, mainly the β-chemokine receptor 5 (CCR5) and the α-chemokine receptor 4 (CXCR4). Viral isolates can be phenotypically classified based on the co-receptor they utilize to infect target cells. In this chapter, methods to determine the co-receptor usage of HIV-1 variants are described.

  20. Increased T cell trafficking as adjunct therapy for HIV-1

    OpenAIRE

    Fryer, HR; Wolinsky, SM; McLean, AR

    2018-01-01

    Although antiretroviral drug therapy suppresses human immunodeficiency virus-type 1 (HIV-1) to undetectable levels in the blood of treated individuals, reservoirs of replication competent HIV-1 endure. Upon cessation of antiretroviral therapy, the reservoir usually allows outgrowth of virus and approaches to targeting the reservoir have had limited success. Ongoing cycles of viral replication in regions with low drug penetration contribute to this persistence. Here, we use a mathematical mode...