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Understanding the drivers and markers of clonally expanding HIV-1-infected CD4+ T cells is essential for HIV-1 eradication. We used single-cell ECCITE-seq, which captures surface protein expression, cellular transcriptome, HIV-1 RNA, and TCR sequences within the same single cell to track clonal expansion dynamics in longitudinally archived samples from six HIV-1-infected individuals (during viremia and after suppressive antiretroviral therapy) and two uninfected individuals, in unstimulated conditions and after CMV and HIV-1 antigen stimulation. Despite antiretroviral therapy, persistent antigen and TNF responses shaped T cell clonal expansion. HIV-1 resided in Th1-polarized, antigen-responding T cells expressing BCL2 and SERPINB9 that may resist cell death. HIV-1 RNA+ T cell clones were larger in clone size, established during viremia, persistent after viral suppression, and enriched in GZMB+ cytotoxic effector memory Th1 cells. Targeting HIV-1-infected cytotoxic CD4+ T cells and drivers of clonal expansion provides another direction for HIV-1 eradication.
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Infecções por HIV , HIV-1 , Linfócitos T CD4-Positivos , Células Clonais , Humanos , RNA , ViremiaRESUMO
BACKGROUND: Synaptic injury is a pathological hallmark of neurological impairment in people living with human immunodeficiency virus (HIV, PLWH), a common complication despite viral suppression with antiretroviral therapy (ART). Measurement of synaptic density in living humans may allow better understanding of HIV neuropathogenesis and provide a dynamic biomarker for therapeutic studies. We applied novel synaptic vesical protein 2A (SV2A) positron emission tomographic (PET) imaging to investigate synaptic density in the frontostriatalthalamic region in PLWH and HIV-uninfected participants. METHODS: In this cross-sectional pilot study,13 older male PLWH on ART underwent magnetic resonance imaging (MRI) and PET scanning with the SV2A ligand [11C]UCB-J with partial volume correction and had neurocognitive assessments. SV2A binding potential (BPND) in the frontostriatalthalamic circuit was compared to 13 age-matched HIV-uninfected participants and assessed with respect to neurocognitive performance in PLWH. RESULTS: PLWH had 14% lower frontostriatalthalamic SV2A synaptic density compared to HIV-uninfected (PLWH: mean [SD], 3.93 [0.80]; HIV-uninfected: 4.59 [0.43]; Pâ =â .02, effect size 1.02). Differences were observed in widespread additional regions in exploratory analyses. Higher frontostriatalthalamic SV2A BPND associated with better grooved pegboard performance, a measure of motor coordination, in PLWH (râ =â 0.61, Pâ =â .03). CONCLUSIONS: In a pilot study, SV2A PET imaging reveals reduced synaptic density in older male PLWH on ART compared to HIV-uninfected in the frontostriatalthalamic circuit and other cortical areas. Larger studies controlling for factors in addition to age are needed to determine whether differences are attributable to HIV or comorbidities in PLWH. SV2A imaging is a promising biomarker for studies of neuropathogenesis and therapeutic interventions in HIV.
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Infecções por HIV , Tomografia por Emissão de Pósitrons , Idoso , Estudos Transversais , HIV , Infecções por HIV/complicações , Infecções por HIV/tratamento farmacológico , Humanos , Masculino , Projetos PilotoRESUMO
BACKGROUND: COVID-19 is caused by the severe acute respiratory syndrome virus SARS-CoV-2. It is widely recognized as a respiratory pathogen, but neurologic complications can be the presenting manifestation in a subset of infected patients. CASE PRESENTATION: We describe a 78-year old immunocompromised woman who presented with altered mental status after witnessed seizure-like activity at home. She was found to have SARS-CoV-2 infection and associated neuroinflammation. In this case, we undertake the first detailed analysis of cerebrospinal fluid (CSF) cytokines during COVID-19 infection and find a unique pattern of inflammation in CSF, but no evidence of viral neuroinvasion. CONCLUSION: Our findings suggest that neurologic symptoms such as encephalopathy and seizures may be the initial presentation of COVID-19. Central nervous system inflammation may associate with neurologic manifestations of disease.
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Betacoronavirus , Infecções por Coronavirus , Citocinas/líquido cefalorraquidiano , Encefalite Viral , Pandemias , Pneumonia Viral , Doença Aguda , Idoso , Biomarcadores/líquido cefalorraquidiano , COVID-19 , Feminino , Humanos , SARS-CoV-2 , ConvulsõesRESUMO
OBJECTIVE: Approximately 40% of adults living with HIV experience cognitive deficits. Little is known about the risk factors for cognitive impairment and its association with myelin content in young adults living with perinatally acquired HIV (YApHIV), which is assessed in our cross-sectional study. DESIGN: A prospective, observational cohort study. METHODS: All participants underwent an 11-test cognitive battery and completed medical and social history surveys. Cognitive impairment was defined as Z scores falling at least 1.5 SD below the mean in at least two domains. Twelve participants underwent myelin water imaging. Neuroimaging data were compared to age and sex-matched HIV-uninfected controls. Regression analyses were used to evaluate for risk factors of lower cognitive domain scores and association between myelin content and cognition in YApHIV. RESULTS: We enrolled 21 virally suppressed YApHIV across two sites in the United States. Ten participants (48%) met criteria for cognitive impairment. Participants with any non-HIV related medical comorbidity scored lower across multiple cognitive domains compared to participants without comorbidities. Myelin content did not differ between YApHIV and controls after adjusting for years of education. Lower cognitive scores were associated with lower myelin content in the cingulum and corticospinal tract in YApHIV participants after correcting for multiple comparisons. CONCLUSION: Poor cognition in YApHIV may be exacerbated by non-HIV related comorbidities as noted in older adults with horizontally acquired HIV. The corticospinal tract and cingulum may be vulnerable to the legacy effect of untreated HIV in infancy. Myelin content may be a marker of cognitive reserve in YApHIV.
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The central nervous system HIV reservoir is incompletely understood and is a major barrier to HIV cure. We profiled people with HIV (PWH) and uninfected controls through single-cell transcriptomic and T cell receptor (TCR) sequencing to understand the dynamics of HIV persistence in the CNS. In PWH on ART, we found that most participants had single cells containing HIV-1 RNA, which was found predominantly in CD4 central memory T cells, in both cerebrospinal fluid (CSF) and blood. HIV-1 RNA-containing cells were found more frequently in CSF than blood, indicating a higher burden of reservoir cells in the CNS than blood for some PWH. Most CD4 T cell clones containing infected cells were compartment specific, while some (22%) - including rare clones with members of the clone containing detectable HIV RNA in both blood and CSF - were found in both CSF and blood. These results suggest that infected T cells trafficked between tissue compartments and that maintenance and expansion of infected T cell clones contributed to the CNS reservoir in PWH on ART.
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Infecções por HIV , HIV-1 , Humanos , HIV-1/genética , Sistema Nervoso Central , RNA , Células ClonaisRESUMO
People with HIV (PWH) on antiretroviral therapy (ART) experience elevated rates of neurological impairment, despite controlling for demographic factors and comorbidities, suggesting viral or neuroimmune etiologies for these deficits. Here, we apply multimodal and cross-compartmental single-cell analyses of paired cerebrospinal fluid (CSF) and peripheral blood in PWH and uninfected controls. We demonstrate that a subset of central memory CD4+ T cells in the CSF produced HIV-1 RNA, despite apparent systemic viral suppression, and that HIV-1-infected cells were more frequently found in the CSF than in the blood. Using cellular indexing of transcriptomes and epitopes by sequencing (CITE-seq), we show that the cell surface marker CD204 is a reliable marker for rare microglia-like cells in the CSF, which have been implicated in HIV neuropathogenesis, but which we did not find to contain HIV transcripts. Through a feature selection method for supervised deep learning of single-cell transcriptomes, we find that abnormal CD8+ T cell activation, rather than CD4+ T cell abnormalities, predominated in the CSF of PWH compared with controls. Overall, these findings suggest ongoing CNS viral persistence and compartmentalized CNS neuroimmune effects of HIV infection during ART and demonstrate the power of single-cell studies of CSF to better understand the CNS reservoir during HIV infection.
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Infecções por HIV , HIV-1 , Infecções por HIV/tratamento farmacológico , Infecções por HIV/patologia , HIV-1/genética , Humanos , Estudos Longitudinais , Microglia/patologia , Transcrição ViralRESUMO
Individuals with coronavirus disease 2019 (COVID-19) frequently develop neurological symptoms, but the biological underpinnings of these phenomena are unknown. Through single-cell RNA sequencing (scRNA-seq) and cytokine analyses of cerebrospinal fluid (CSF) and blood from individuals with COVID-19 with neurological symptoms, we find compartmentalized, CNS-specific T cell activation and B cell responses. All affected individuals had CSF anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibodies whose target epitopes diverged from serum antibodies. In an animal model, we find that intrathecal SARS-CoV-2 antibodies are present only during brain infection and not elicited by pulmonary infection. We produced CSF-derived monoclonal antibodies from an individual with COVID-19 and found that these monoclonal antibodies (mAbs) target antiviral and antineural antigens, including one mAb that reacted to spike protein and neural tissue. CSF immunoglobulin G (IgG) from 5 of 7 patients showed antineural reactivity. This immune survey reveals evidence of a compartmentalized immune response in the CNS of individuals with COVID-19 and suggests a role of autoimmunity in neurologic sequelae of COVID-19.
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Despite effective antiretroviral therapy, HIV-1-infected cells continue to produce viral antigens and induce chronic immune exhaustion. We propose to identify HIV-1-suppressing agents that can inhibit HIV-1 reactivation and reduce HIV-1-induced immune activation. Using a newly developed dual-reporter system and a high-throughput drug screen, we identified FDA-approved drugs that can suppress HIV-1 reactivation in both cell line models and CD4+ T cells from virally suppressed HIV-1-infected individuals. We identified 11 cellular pathways required for HIV-1 reactivation as druggable targets. Using differential expression analysis, gene set enrichment analysis, and exon-intron landscape analysis, we examined the impact of drug treatment on the cellular environment at a genome-wide level. We identified what we believe to be a new function of a JAK inhibitor, filgotinib, that suppresses HIV-1 splicing. First, filgotinib preferentially suppresses spliced HIV-1 RNA transcription. Second, filgotinib suppresses HIV-1-driven aberrant cancer-related gene expression at the integration site. Third, we found that filgotinib suppresses HIV-1 transcription by inhibiting T cell activation and by modulating RNA splicing. Finally, we found that filgotinib treatment reduces the proliferation of HIV-1-infected cells. Overall, the combination of a drug screen and transcriptome analysis provides systematic understanding of cellular targets required for HIV-1 reactivation and drug candidates that may reduce HIV-1-related immune activation.
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Linfócitos T CD4-Positivos/imunologia , Infecções por HIV/tratamento farmacológico , HIV-1/imunologia , Ativação Linfocitária/efeitos dos fármacos , Piridinas/farmacologia , Splicing de RNA/efeitos dos fármacos , Transcrição Gênica/efeitos dos fármacos , Triazóis/farmacologia , Linfócitos T CD4-Positivos/patologia , Infecções por HIV/imunologia , Infecções por HIV/patologia , Humanos , Células Jurkat , Splicing de RNA/imunologia , Transcrição Gênica/imunologiaRESUMO
INTRODUCTION: Up to 30% of individuals treated with antiretroviral therapy (ART) during chronic HIV fail to recover CD4 counts to >500 cells/mm3 despite plasma viral suppression. We investigated the frequency and associations of suboptimal CD4 recovery after ART started during acute HIV infection (AHI). METHODS: Participants who started ART in Fiebig I to V AHI with ≥48 weeks of continuous documented HIV-RNA < 50 copies/mL were stratified by CD4 count at latest study visit to suboptimal immune recovery (SIR; CD4 < 350 cells/mm3 ), intermediate immune recovery (IIR; 350 ≤ CD4 < 500) and complete immune recovery (CIR; CD4 ≥ 500). Clinical and laboratory parameters were assessed at pre-ART baseline and latest study visit. Additional inflammatory and neurobehavioral endpoints were examined at baseline and 96 weeks. RESULTS: Of 304 participants (96% male, median 26 years old) evaluated after median 144 (range 60 to 420) weeks of ART initiated at median 19 days (range 1 to 62) post-exposure, 3.6% (n = 11) had SIR and 14.5% (n = 44) had IIR. Pre-ART CD4 count in SIR compared to CIR participants was 265 versus 411 cells/mm3 (p = 0.002). Individuals with SIR or IIR had a slower CD4 rate of recovery compared to those with CIR. Timing of ART initiation by Fiebig stage did not affect CD4 count during treatment. Following ART, the CD8+ T cell count (p = 0.001) and CD4/CD8 ratio (p = 0.047) were lower in SIR compared to CIR participants. Compared to the CIR group at week 96, the combined SIR and IIR groups had higher sCD14 (p = 0.008) and lower IL-6 (p = 0.04) in plasma, without differences in neuropsychological or psychiatric indices. CONCLUSIONS: Despite immediate and sustained treatment in AHI, suboptimal CD4 recovery occurs uncommonly and is associated with low pre-ART CD4 count as well as persistent low CD8 count and CD4/CD8 ratio during treatment.
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Fármacos Anti-HIV/uso terapêutico , Linfócitos T CD4-Positivos/efeitos dos fármacos , Infecções por HIV/tratamento farmacológico , HIV-1/imunologia , Adulto , Terapia Antirretroviral de Alta Atividade , Contagem de Linfócito CD4 , Relação CD4-CD8 , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Estudos de Coortes , Infecções por HIV/sangue , Infecções por HIV/imunologia , HIV-1/efeitos dos fármacos , HIV-1/genética , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Adulto JovemRESUMO
Understanding HIV-1-host interactions can identify the cellular environment supporting HIV-1 reactivation and mechanisms of clonal expansion. We developed HIV-1 SortSeq to isolate rare HIV-1-infected cells from virally suppressed, HIV-1-infected individuals upon early latency reversal. Single-cell transcriptome analysis of HIV-1 SortSeq+ cells revealed enrichment of nonsense-mediated RNA decay and viral transcription pathways. HIV-1 SortSeq+ cells up-regulated cellular factors that can support HIV-1 transcription (IMPDH1 and JAK1) or promote cellular survival (IL2 and IKBKB). HIV-1-host RNA landscape analysis at the integration site revealed that HIV-1 drives high aberrant host gene transcription downstream, but not upstream, of the integration site through HIV-1-to-host aberrant splicing, in which HIV-1 RNA splices into the host RNA and aberrantly drives host RNA transcription. HIV-1-induced aberrant transcription was driven by the HIV-1 promoter as shown by CRISPR-dCas9-mediated HIV-1-specific activation and could be suppressed by CRISPR-dCas9-mediated inhibition of HIV-1 5' long terminal repeat. Overall, we identified cellular factors supporting HIV-1 reactivation and HIV-1-driven aberrant host gene transcription as potential therapeutic targets to disrupt HIV-1 persistence.
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Infecções por HIV , HIV-1 , Regulação Viral da Expressão Gênica , Infecções por HIV/tratamento farmacológico , Infecções por HIV/genética , HIV-1/genética , Humanos , Transcrição Gênica , Ativação Viral , Latência ViralRESUMO
BACKGROUND: COVID-19 is caused by the severe acute respiratory syndrome virus SARS-CoV-2. It is widely recognized as a respiratory pathogen, but neurologic complications can be the presenting manifestation in a subset of infected patients. CASE PRESENTATION: We describe a 78-year old immunocompromised woman who presented with altered mental status after witnessed seizure-like activity at home. She was found to have SARS-CoV-2 infection and associated neuroinflammation. In this case, we undertake the first detailed analysis of cerebrospinal fluid (CSF) cytokines during COVID-19 infection and find a unique pattern of inflammation in CSF, but no evidence of viral neuroinvasion. CONCLUSION: Our findings suggest that neurologic symptoms such as encephalopathy and seizures may be the initial presentation of COVID-19. Central nervous system inflammation may associate with neurologic manifestations of disease.
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One third of COVID-19 patients develop significant neurological symptoms, yet SARS-CoV-2 is rarely detected in central nervous system (CNS) tissue, suggesting a potential role for parainfectious processes, including neuroimmune responses. We therefore examined immune parameters in cerebrospinal fluid (CSF) and blood samples from a cohort of patients with COVID-19 and significant neurological complications. We found divergent immunological responses in the CNS compartment, including increased levels of IL-12 and IL-12-associated innate and adaptive immune cell activation. Moreover, we found increased proportions of B cells in the CSF relative to the periphery and evidence of clonal expansion of CSF B cells, suggesting a divergent intrathecal humoral response to SARS-CoV-2. Indeed, all COVID-19 cases examined had anti-SARS-CoV-2 IgG antibodies in the CSF whose target epitopes diverged from serum antibodies. We directly examined whether CSF resident antibodies target self-antigens and found a significant burden of CNS autoimmunity, with the CSF from most patients recognizing neural self-antigens. Finally, we produced a panel of monoclonal antibodies from patients' CSF and show that these target both anti-viral and anti-neural antigens-including one mAb specific for the spike protein that also recognizes neural tissue. This exploratory immune survey reveals evidence of a compartmentalized and self-reactive immune response in the CNS meriting a more systematic evaluation of neurologically impaired COVID-19 patients.
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BACKGROUND: Neurocognitive dysfunction remains prevalent among people living with HIV (PLWH), even after viral suppression on combination antiretroviral therapy (cART). We investigated associations between neuropsychological performance (NP) and patterns of circulating exosomal microRNA (exo-miRNA) expression in PLWH on cART. SETTING: A cross-sectional examination of plasma exo-miRNA among PLWH on cART with systemic viral suppression and volunteers without HIV infection. METHODS: Thirty-one PLWH who started cART during early infection (n = 19) or chronic infection (n = 12) participated in phlebotomy and an 11-test neuropsychological battery after >1 year on treatment. NP higher- or lower-performing participants were categorized based on normalized neuropsychological scores. Total RNA was extracted from purified exosomes of 31 PLWH and 5 volunteers without HIV and subject to small RNA sequencing. Differential expression of exo-miRNAs was examined and biological functions were predicted. RESULTS: Eleven exo-miRNAs were up-regulated in NP lower-performing (n = 18) relative to higher-performing PLWH (n = 13). A high proportion of the differentiating exo-miRNA target the axon guidance KEGG pathway and neurotrophin tyrosine receptor kinase signaling Gene Ontology pathway. Differential expression analysis of exo-miRNAs between NP lower- (n = 7) and higher-performing (n = 12) PLWH within the early infection group alone confirmed largely consistent findings. CONCLUSIONS: Plasma exo-miRNA content differed between NP higher- and lower-performing PLWH. Several differentially expressed exo-miRNAs were predicted to be involved in inflammation and neurodegeneration pathways. Exo-miRNA in plasma may indicate cross-talk between the circulation and central nervous system and thus may be clinically relevant for neurocognitive dysfunction in PLWH.
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Exossomos/metabolismo , Infecções por HIV/sangue , Infecções por HIV/psicologia , MicroRNAs/sangue , Transtornos Neurocognitivos/sangue , Adulto , Fármacos Anti-HIV/uso terapêutico , Estudos Transversais , Quimioterapia Combinada , Exossomos/genética , Feminino , Infecções por HIV/tratamento farmacológico , Humanos , Masculino , MicroRNAs/genética , Pessoa de Meia-Idade , Transtornos Neurocognitivos/virologia , Testes Neuropsicológicos , Transdução de Sinais/genéticaRESUMO
The presence of quantifiable HIV RNA in cerebrospinal fluid (CSF) during antiretroviral therapy (ART) can associate with central nervous system (CNS) pathology, but the significance of RNA detected below the limit of quantification (LOQ) on a standard assay during ART remains unknown. We compared CNS parameters between individuals with CSF RNA detected below the LOQ (20 copies/mL) with those with HIV RNA not detected. Detection of CSF HIV RNA associated with decreased blood-brain barrier integrity and with decreased executive function, but not with CNS immune activation or poorer performance in overall neuropsychological testing.
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This case-control study assesses cerebrospinal fluid markers of neuroinflammation and blood-brain barrier disruption in individuals with postCOVID-19 condition who reported neuropsychiatric symptoms.
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COVID-19 , Doenças Neuroinflamatórias , Humanos , Autorrelato , EncéfaloRESUMO
Central nervous system (CNS) immune activation is an important driver of neuronal injury during several neurodegenerative and neuroinflammatory diseases. During HIV infection, CNS immune activation is associated with high rates of neurocognitive impairment, even during sustained long-term suppressive antiretroviral therapy (ART). However, the cellular subsets that drive immune activation and neuronal damage in the CNS during HIV infection and other neurological conditions remain unknown, in part because CNS cells are difficult to access in living humans. Using single-cell RNA sequencing (scRNA-seq) on cerebrospinal fluid (CSF) and blood from adults with and without HIV, we identified a rare (<5% of cells) subset of myeloid cells that are found only in CSF and that present a gene expression signature that overlaps significantly with neurodegenerative disease-associated microglia. This highlights the power of scRNA-seq of CSF to identify rare CNS immune cell subsets that may perpetuate neuronal injury during HIV infection and other conditions.
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Líquido Cefalorraquidiano/imunologia , Infecções por HIV/imunologia , Microglia/imunologia , Análise de Sequência de RNA , Adulto , Fármacos Anti-HIV/uso terapêutico , Terapia Antirretroviral de Alta Atividade , Sangue , Sistema Nervoso Central/virologia , Expressão Gênica , Humanos , Inflamação , Células Mieloides , Doenças Neurodegenerativas/genética , Doenças Neurodegenerativas/imunologiaRESUMO
BACKGROUND: Two inhibitors of the HIV-1 integrase enzyme (INIs) are in late stage clinical development. To date, approximately 42 mutations within the HIV-1 integrase (IN) gene have been associated with INI drug resistance. Naturally occurring IN gene polymorphisms may have important implications for INI development. In this study, we evaluated clinical HIV-1 strains from INI-naive patients to determine the prevalence of IN gene polymorphisms, and the frequency of naturally occurring amino acid (aa) substitutions at positions associated with INI resistance and at sites crucial for LEDGF/p75 binding and HIV-1 integration. METHODS: The IN gene from 67 INI-naive, HIV-1 clade B-infected patients were sequenced using standard population-based DNA sequencing methods. In addition, 176 unique full-length HIV-1 clade B IN gene sequences from INI-naive patients obtained from the HIV Los Alamos database were analysed. RESULTS: Analysis of 243 IN genes from HIV-1 clade B, INI-naive clinical strains revealed that 64% of the aa positions were polymorphic. Of the 42 aa substitutions currently associated with INI resistance, 21 occurred as natural polymorphisms: V72I, L74I, T97A, T112I, A128T, E138K, Q148H, V151I, S153Y/A, M154I, N155H, K156N, E157Q, G163R, V165I, V201I, I203M, T206S, S230N and R263K. IN aa positions crucial to LEDGF/P75 binding and HIV-1 integration were well conserved. CONCLUSION: Major INI mutations within the catalytic domain and extended active sites associated with high level resistance to the compounds in late stage development, especially strand transfer inhibitors (STIs), were infrequent in our study, which may help explain the excellent virological responses demonstrated in clinical trials.
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Farmacorresistência Viral/genética , Inibidores de Integrase de HIV/farmacologia , Integrase de HIV/genética , HIV-1/efeitos dos fármacos , Mutação , Polimorfismo Genético , Sequência de Aminoácidos , Sequência Consenso , Infecções por HIV/virologia , Integrase de HIV/química , Integrase de HIV/efeitos dos fármacos , HIV-1/classificação , HIV-1/enzimologia , HIV-1/genética , Humanos , Dados de Sequência Molecular , RNA Viral/sangue , RNA Viral/genética , Análise de Sequência de DNARESUMO
BACKGROUND: Population-based and longitudinal information regarding sexual risk behavior among patients with multidrug resistant (MDR) HIV and their sexual partners is of great public health and clinical importance. OBJECTIVE: To characterize the HIV sexual risk behaviors of patients with and without drug-resistant HIV in the clinical care setting over time. MEASUREMENTS: 393 HIV-positive patients completed questionnaires of self-reported sexual risk behaviors at approximate 6-month intervals extending over 24 months. HIV viral load and genotypic drug resistance obtained during the same time points were matched to the behavioral data. Multidrug resistance was defined as having resistance to 2 or 3 antiretroviral (ARV) drug classes. RESULTS: In serial cross-sectional analyses, 393 patients (44% female and 79% heterosexual) contributed 919 matched behavioral and virologic results over the 24 months of data collection. Of these, 250 patients (64%) reported having sex during at least 1 survey period resulting in greater than 10,000 sexual events with more than 1000 partners. Unprotected sexual behavior was reported by 45% of sexually active patients, resulting in 34% of all sex events that exposed 29% of all partners. Of these patients with unprotected sexual events, 31% had HIV drug resistance--11.6% with resistance to 2 classes of ARVs (2-class), and 1.8% with 3-class ARV resistance at the time of a sexual risk event. Close to 1000 or 28% of all unprotected sexual events involved resistant strains (11% of these with resistance to 2 classes and 0.2% with 3-class resistance, exposing 20% of unprotected sexual partners to resistant HIV (8% to 2-class and 0.6% to 3-class resistance). In longitudinal analysis among the 78 patients who reported a cumulative total of 12 months of sexual history and had resistance testing, 38% reported engaging in unprotected sexual behavior. There was substantial and complex variation in the distribution of unprotected sexual events and in the detection of resistance over time. CONCLUSION: In this study of HIV sexual risk and resistance over time among HIV-infected patients in clinical care, a substantial proportion engaged in unprotected sex and had drug-resistant HIV, frequently exposing partners to 1- or 2-class resistant HIV strains. However, relatively few exposures involved 3-class resistance. The dynamics of sexual risk behavior and HIV drug resistance are complex and vary over time and urgently require both general and targeted interventions to reduce transmission of resistant HIV.
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Farmacorresistência Viral Múltipla , Infecções por HIV/tratamento farmacológico , Comportamento Sexual , Adulto , Estudos Transversais , Feminino , Humanos , Estudos Longitudinais , Masculino , Assunção de RiscosRESUMO
OBJECTIVE: Determine HIV drug resistance mutations (DRMs) prevalence at low and high levels in ART-experienced patients experiencing virologic failure (VF). METHODS: 29 subjects from 18 counties in Hunan Province that experienced VF were evaluated for the prevalence of DRMs (Stanford DRMs with an algorithm value ≥15, include low-, intermediate and high-level resistance) by both Sanger sequencing (SS) and deep sequencing (DS) to 1% frequency levels. RESULTS: DS was performed on samples from 29 ART-experienced subjects; the median viral load 4.95×10(4) c/ml; 82.76% subtype CRF01_AE. 58 DRMs were detected by DS. 18 DRMs were detected by SS. Of the 58 mutations detected by DS, 40 were at levels <20% frequency (26 NNRTI, 12 NRTI and 2 PI) and the majority of these 95.00% (38/40) were not detected by standard genotyping. Of these 40 low-level DRMs, 16 (40%) were detected at frequency levels of 1-4% and 24 (60%) at levels of 5-19%. SS detected 15 of 17 (88.24%) DRMs at levels ≥ 20% that were detected by DS. The only variable associated with the detection of DRMs by DS was ART adherence (missed doses in the prior 7 days); all patients that reported missing a dose in the last 7 days had DRMs detected by DS. CONCLUSIONS: DS of VF samples from treatment experienced subjects infected with primarily AE subtype frequently identified Stanford HIVdb NRTI and NNRTI resistance mutations with an algorithm value 15. Low frequency level resistant variants detected by DS were frequently missed by standard genotyping in VF specimens from antiretroviral-experienced subjects.
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Farmacorresistência Viral , Infecções por HIV/virologia , HIV-1/efeitos dos fármacos , HIV-1/genética , Mutação , Adulto , Idoso , Terapia Antirretroviral de Alta Atividade , China , Feminino , Infecções por HIV/diagnóstico , Infecções por HIV/tratamento farmacológico , Infecções por HIV/epidemiologia , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Incidência , Masculino , Adesão à Medicação , Pessoa de Meia-Idade , Prevalência , Fatores de Risco , Falha de Tratamento , Carga Viral , Adulto JovemRESUMO
BACKGROUND: Limited data exist comparing viral quasispecies between cerebrospinal fluid (CSF) and plasma compartments during primary HIV infection. Deep sequencing is a new method to examine the HIV plasma and CSF quasispecies. METHODS: In this pilot study, deep sequencing of protease (PR) and reverse transcriptase (RT) was performed in plasma and CSF from participants during primary HIV infection. Estimated mutational load was calculated by mutant variant frequency multiplied by HIV-RNA level. RESULTS: Paired plasma and CSF samples were studied from five antiretroviral therapy-naïve male participants with median 109 days post estimated transmission, age 32 years, CD4 cell count 580 cells/µL, HIV-RNA 5.18 log10 copies/mL in plasma and 3.67 log10 copies/mL in CSF. Plasma samples averaged 7,124 reads of PR and 2,448 reads of RT, whereas CSF samples averaged 7,082 and 2,792 reads, respectively. A distinct drug-resistance pattern with linked mutations present at significant levels (5-10%) was detected in one participant in CSF. Other low abundance variants (>0.2%) were detected in plasma and CSF of four out of five participants. CONCLUSIONS: Deep sequencing of CSF HIV is technically possible with sufficient HIV-RNA levels. Differences between the quasispecies in the two compartments detected in one participant, which were present with a high mutational load in CSF at an estimated 3.6 months after HIV infection, suggest that early CNS compartmentalisation may be revealed by sensitive deep-sequencing methods. The presence of distinct low abundance (<1%) resistance variants in plasma and CSF of three other subjects may be significant, but further investigation is needed.