Improving HIV-1 tropism determination by combining geno2pheno and V3 net charge calculation

Objective: The aim of this study was to evaluate tropism prediction performances of three algorithms [geno2pheno false-positive rate 10% (G2P10), position-specific scoring matrix (PSSM) and a combination of the 11/25 and net charge rules] and to investigate the viral and host factors potentially involved in the X4 or R5 prediction in human immunodeficiency virus-1 (HIV-1) patients. Methods: Viral tropism was determined in 179 HIV-1-infected patients eligible for CCR5 antagonist therapy. HIV-1 RNA or DNA was extracted and amplified for env gp120 sequencing. In parallel, demographic, viral, immunological and clinical determinants were analyzed. Results: According to the G2P10 algorithm, 48 patients harbored X4 or X4R5 virus. The tropism prediction was concordant for 87.7 and 88.2% of samples when comparing G2P10 with PSSM or with a combination of the 11/25 and net charge rules, respectively. X4 prediction was significantly associated with more than 35 amino acids in the V3 domain (p < 0.0001) and loss of an N-linked glycosylation site (p < 0.0001). Of the factors studied, only the nadir CD4 T-cell count was significantly associated with X4 tropism (p = 0.01). Conclusion: We determined that the X4 virus detection is closely linked to the nadir CD4 T-cell count below 100 cells/mm3 that must be taken into account when considering a CCR5 antagonist therapy switch.

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Primary HIV-1 R5 isolates from end-stage disease display enhanced viral fitness in parallel with increased gp120 net charge

Virology ◽

10.1016/j.virol.2008.06.014 ◽

2008 ◽

Vol 379 (1) ◽

pp. 125-134 ◽

Cited By ~ 33

Author(s):

Johanna Repits ◽

Jasminka Sterjovski ◽

Daniel Badia-Martinez ◽

Mattias Mild ◽

Lachlan Gray ◽

...

Keyword(s):

Viral Fitness ◽

Net Charge ◽

Stage Disease ◽

End Stage ◽

Hiv 1

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Mechanisms and evolutionary implications of integration of different functions within HIV-1 Tat

10.1101/2021.04.20.440437 ◽

2021 ◽

Author(s):

Margarita A. Kurnaeva ◽

Arthur O. Zalevsky ◽

Eugene A. Arifulin ◽

Olga M. Lisitsyna ◽

Anna V. Tvorogova ◽

...

Keyword(s):

Amino Acids ◽

Viral Proteins ◽

Net Charge ◽

Origin And Evolution ◽

Nuclear Localization Signals ◽

Protein Size ◽

Charged Amino Acids ◽

A Charge ◽

Hiv 1 ◽

Positively Charged

During evolution, viruses had to adapt to an increasingly complex environment of eukaryotic cells. Viral proteins that need to enter the cell nucleus or associate with nucleoli possess nuclear localization signals (NLSs) and nucleolar localization signals (NoLSs) for nuclear and nucleolar accumulation, respectively. As viral proteins are relatively small, acquisition of novel sequences seems to be a more complicated task for viruses than for eukaryotes. Here, we carried out a comprehensive analysis of the basic domain (BD) of HIV-1 Tat to show how viral proteins might evolve with NLSs and NoLSs without an increase in protein size. The HIV-1 Tat BD is involved in several functions, the most important being the transactivation of viral transcription. The BD also functions as an NLS, although it is substantially longer than a typical NLS. It seems that different regions in the BD could function as NLSs due to its enrichment with positively charged amino acids. Additionally, the high positive net charge inevitably causes the BD to function as an NoLS through a charge-specific mechanism. The integration of NLSs and NoLSs into functional domains enriched with positively charged amino acids might be a mechanism that allows the condensation of different functional sequences in small protein regions and, as a result, to reduce protein size, influencing the origin and evolution of NLSs and NoLSs in viruses.

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Impact of Mutations Outside the V3 Region on Coreceptor Tropism Phenotypically Assessed in Patients Infected with HIV-1 Subtype B

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00743-11 ◽

2011 ◽

Vol 55 (11) ◽

pp. 5078-5084 ◽

Cited By ~ 20

Author(s):

Laura Monno ◽

Annalisa Saracino ◽

Luigia Scudeller ◽

Grazia Punzi ◽

Gaetano Brindicci ◽

...

Keyword(s):

Amino Acid ◽

Statistical Significance ◽

Net Charge ◽

Variable Regions ◽

Coreceptor Tropism ◽

Enhanced Sensitivity ◽

Subtype B ◽

End Stage ◽

The Impact ◽

Hiv 1

ABSTRACTHIV coreceptor tropism (CTR) testing is a prerequisite for prescribing a coreceptor antagonist. CTR is increasingly deduced by analyzing the V3 loop sequence of gp120. We investigated the impact of mutations outside V3 on CTR as determined by the enhanced-sensitivity Trofile assay (ESTA). Paired ESTA and gp120 sequencing (population sequencing; from codon 32 of the conserved C1 to the variable V5 domains) were obtained from 60 antiretroviral treatment (ART)-naïve patients (15 with AIDS) infected with subtype B HIV-1. For gp120 sequence analysis, nucleotide mixtures were considered when the second highest electropherogram peak was >25%; sequences were translated into all possible permutations and classified as X4, dual/mixed (DM), and R5 based on coincident ESTA results. ESTA identified R5 and DM viruses in 72 and 28% of patients, respectively; no pure X4 was labeled. Forty percent of AIDS patients had R5 strains. Thirty-two positions, mostly outside V3, were significantly (P< 0.05) different between R5 and DM sequences. According to multivariate analysis, amino acid changes at 9 and 7 positions within the C1 to C4 and V1 to V5 regions, respectively, maintained a statistical significance, as did the net charge of V3 and C4. When analyzing only R5 sequences, 6 positions in the variable regions were found which, along with the V4 net charge, were significantly different for sequences from early- and end-stage disease patients. This study identifies specific amino acid changes outside V3 which contribute to CTR. Extending the analysis to include pure X4 and increasing the sample size would be desirable to define gp120 variables/changes which should be included in predictive algorithms.

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Phenotypic and Genotypic Co-receptor Tropism Testing in HIV-1 Epidemic Region of Tanzania Where Multiple Non-B Subtypes Co-circulate

Frontiers in Microbiology ◽

10.3389/fmicb.2021.703041 ◽

2021 ◽

Vol 12 ◽

Author(s):

George P. Judicate ◽

Godfrey Barabona ◽

Doreen Kamori ◽

Macdonald Mahiti ◽

Toong Seng Tan ◽

...

Keyword(s):

Human Immunodeficiency Virus ◽

Entry Inhibitor ◽

Type I ◽

Cd4 Cells ◽

Net Charge ◽

Prediction Algorithms ◽

Immunodeficiency Virus ◽

Treatment Naïve ◽

Almost All ◽

Hiv 1

HIV human immunodeficiency virus type I (HIV-1) entry inhibitor potency is dependent on viral co-receptor tropisms and thereby tropism determination is clinically important. However, phenotypic tropisms of HIV-1 non-B subtypes have been poorly investigated and the genotypic prediction algorithms remain insufficiently validated. To clarify this issue, we recruited 52 treatment-naïve, HIV-1-infected patients in Tanzania, where multiple HIV-1 non-B subtypes co-circulate. Sequence analysis of 93 infectious envelope clones isolated from their plasma viral RNA revealed the co-circulation of subtypes A1, C, D, and inter-subtype recombinant forms (isRFs). Phenotypic tropism assays revealed that lentivirus reporters pseudotyped with 75 (80.6%) and 5 (5.4%) envelope clones could establish infection toward U87.CD4 cells expressing CCR5 (R5) and CXCR4 (X4), respectively; whereas the remaining 13 (14%) clones could infect both cells. Genotypic analyses by widely used algorithms including V3 net charge, Geno2pheno, WebPSSM, and PhenoSeq showed that almost all phenotypic X4-tropic clones and only 15 of 75 phenotypic R5-tropic clones were concordantly predicted. However, the remaining 60 phenotypic R5-tropic clones were discordantly predicted by at least one algorithm. In particular, 2 phenotypic R5-tropic clones were discordantly predicted by all algorithms tested. Taken together, the results demonstrate the limitation of currently available genotypic algorithms for predicting co-receptor inference among co-circulating multiple non-B subtypes and emerging isRFs. Also, the phenotypic tropism dataset presented here could be valuable for retraining of the widely used genotypic prediction algorithms to enhance their performance.

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Ultrastructural observations of human monocytes infected with HIV-1

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100084843 ◽

1991 ◽

Vol 49 ◽

pp. 108-109

Author(s):

James K. Koehler ◽

Steven G. Reed ◽

Joao S. Silva

Keyword(s):

Gradient Centrifugation ◽

Virus Production ◽

Human Monocyte ◽

Red Cross ◽

Human Monocytes ◽

Blood Monocytes ◽

Hiv Replication ◽

Electron Microscopic ◽

Ultrastructural Studies ◽

Hiv 1

As part of a larger study involving the co-infection of human monocyte cultures with HIV and protozoan parasites, electron microscopic observations were made on the course of HIV replication and infection in these cells. Although several ultrastructural studies of the cytopathology associated with HIV infection have appeared, few studies have shown the details of virus production in “normal,” human monocytes/macrophages, one of the natural targets of the virus, and suspected of being a locus of quiescent virus during its long latent period. In this report, we detail some of the interactions of developing virons with the membranes and organelles of the monocyte host.Peripheral blood monocytes were prepared from buffy coats (Portland Red Cross) by Percoll gradient centrifugation, followed by adherence to cover slips. 90-95% pure monocytes were cultured in RPMI with 5% non-activated human AB serum for four days and infected with 100 TCID50/ml of HIV-1 for four hours, washed and incubated in fresh medium for 14 days.

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Characteristics of an Electrostatic Phase Plate

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100096345 ◽

1972 ◽

Vol 30 ◽

pp. 618-619

Author(s):

William Krakow ◽

Benjamin Siegel

Keyword(s):

Phase Contrast ◽

Objective Lens ◽

Phase Plate ◽

Net Charge ◽

Phase Plates ◽

Beam Currents ◽

Contrast Image ◽

Bright Phase ◽

Additional Phase Shift ◽

Additional Phase

Unwin has used a metallized non-conducting thread in the back focal plane of the objective lens that stops out a portion of the unscattered beam, takes on a localized positive charge and thus produces an additional phase shift to give a different transfer function of the lens. Under the particular conditions Unwin used, the phase contrast image was shifted to bright phase contrast for optimum focus.We have investigated the characteristics of this type of electrostatic phase plate, both analytically and experimentally, as functions of the magnitude of charge and defocus. Phase plates have been constructed by using Wollaston wire to mount 0.25μ diameter platinum wires across apertures ranging from 50 to 200μ diameter and vapor depositing SiO and gold on the mounted wires to give them the desired charging characteristics. The net charge was varied by adjusting only the bias on the Wehnelt shield of the gun, and hence the beam currents and effective size of the source.

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Direct imaging of charge transfer

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100165860 ◽

1996 ◽

Vol 54 ◽

pp. 680-681

Author(s):

Yimei Zhu ◽

J. Tafto

Keyword(s):

Charge Transfer ◽

Electron Diffraction ◽

Scattering Amplitude ◽

High Temperature Superconductors ◽

Charge Carriers ◽

Image Charge ◽

Net Charge ◽

Long Time ◽

Electron Holes ◽

First Time

The electron holes confined to the CuO2-plane are the charge carriers in high-temperature superconductors, and thus, the distribution of charge plays a key role in determining their superconducting properties. While it has been known for a long time that in principle, electron diffraction at low angles is very sensitive to charge transfer, we, for the first time, show that under a proper TEM imaging condition, it is possible to directly image charge in crystals with a large unit cell. We apply this new way of studying charge distribution to the technologically important Bi2Sr2Ca1Cu2O8+δ superconductors.Charged particles interact with the electrostatic potential, and thus, for small scattering angles, the incident particle sees a nuclei that is screened by the electron cloud. Hence, the scattering amplitude mainly is determined by the net charge of the ion. Comparing with the high Z neutral Bi atom, we note that the scattering amplitude of the hole or an electron is larger at small scattering angles. This is in stark contrast to the displacements which contribute negligibly to the electron diffraction pattern at small angles because of the short g-vectors.

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