Human Immunodeficiency Virus DNA Research Articles

Label-free and portable detection of HIV-DNA by a handheld luminometer

BackgroundThe human immunodeficiency virus (HIV) remains a major worldwide health problem. Nowadays, many methods have been developed for quantitative detecting human immunodeficiency virus DNA (HIV-DNA), such as fluorescence and colorimetry. However, these methods still have the disadvantages of being expensive and requiring professional technicians. Early diagnosis of pathogens is increasingly dependent on portable instruments and simple point-of-care testing (POCT). Therefore, it is meaningful and necessary to develop portable and cheap methods for detecting disease markers. ResultsIn this work, a label-free chemiluminescence (CL) method was developed for detecting HIV-DNA via a handheld luminometer. To achieve label-free target detection, the CL catalyst, G-triplex-hemin DNAzyme (G3-hemin DNAzyme), was in-situ assembled in the presence of HIV-DNA. For improving sensitivity, HIV-DNA induced the cyclic strand displacement reaction (SDR), which can form three G3-hemin DNAzymes in one cycle. So, the chemiluminescence reaction between luminol and H2O2 was highly effectively catalyzed, and the CL intensity was linearly related with the concentration of HIV-DNA in the range of 0.05–10 nM with a detection limit of 29.0 pM. Due to the high specificity of hairpin DNA, single-base mismatch can be discriminated, which ensured the specific detection of HIV-DNA. SignificanceIn-situ formation of G3-hemin DNAzyme led to label-free and selective detection without complex synthesis and functionalization. Therefore, it offers a cheap, selective, sensitive and portable method for detecting disease-related genes, which is promising for POCT of clinical diagnosis in resource-limited settings.

Development of a droplet digital polymerase chain reaction assay for the sensitive detection of total and integrated HIV-1 DNA.

Total human immunodeficiency virus (HIV) DNA and integrated HIV DNA are widely used markers of HIV persistence. Droplet digital polymerase chain reaction (ddPCR) can be used for absolute quantification without needing a standard curve. Here, we developed duplex ddPCR assays to detect and quantify total HIV DNA and integrated HIV DNA. The limit of detection, dynamic ranges, sensitivity, and reproducibility were evaluated by plasmid constructs containing both the HIV long terminal repeat (LTR) and human CD3 gene (for total HIV DNA) and ACH-2 cells (for integrated HIV DNA). Forty-two cases on stable suppressive antiretroviral therapy (ART) were assayed in total HIV DNA and integrated HIV DNA. Correlation coefficient analysis was performed on the data related to DNA copies and cluster of differentiation 4 positive (CD4 + ) T-cell counts, CD8 + T-cell counts and CD4/CD8 T-cell ratio, respectively. The assay linear dynamic range and lower limit of detection (LLOD) were also assessed. The assay could detect the presence of HIV-1 copies 100% at concentrations of 6.3 copies/reaction, and the estimated LLOD of the ddPCR assay was 4.4 HIV DNA copies/reaction (95% confidence intervals [CI]: 3.6-6.5 copies/reaction) with linearity over a 5-log 10 -unit range in total HIV DNA assay. For the integrated HIV DNA assay, the LLOD was 8.0 copies/reaction (95% CI: 5.8-16.6 copies/reaction) with linearity over a 3-log 10 -unit range. Total HIV DNA in CD4 + T cells was positively associated with integrated HIV DNA ( r = 0.76, P <0.0001). Meanwhile, both total HIV DNA and integrated HIV DNA in CD4 + T cells were inversely correlated with the ratio of CD4/CD8 but positively correlated with the CD8 + T-cell counts. This ddPCR assay can quantify total HIV DNA and integrated HIV DNA efficiently with robustness and sensitivity. It can be readily adapted for measuring HIV DNA with non-B clades, and it could be beneficial for testing in clinical trials.

Cannabis Use Associates With Reduced Proviral Burden and Inflammatory Cytokine in Tissues From Men With Clade C HIV-1 on Suppressive Antiretroviral Therapy.

Human immunodeficiency virus 1 (HIV-1) tissue reservoirs remain the main obstacle against an HIV cure. Limited information exists regarding cannabis's effects on HIV-1 infections in vivo, and the impact of cannabis use on HIV-1 parenchymal tissue reservoirs is unexplored. To investigate whether cannabis use alters HIV-1 tissue reservoirs, we systematically collected 21 postmortem brain and peripheral tissues from 20 men with subtype C HIV-1 and with suppressed viral load enrolled in Zambia, 10 of whom tested positive for cannabis use. The tissue distribution and copies of subtype C HIV-1 LTR, gag, env DNA and RNA, and the relative mRNA levels of cytokines IL-1β, IL-6, IL-10, and TGF-β1 were quantified using PCR-based approaches. Utilizing generalized linear mixed models we compared persons with HIV-1 and suppressed viral load, with and without cannabis use. The odds of tissues harboring HIV-1 DNA and the viral DNA copies in those tissues were significantly lower in persons using cannabis. Moreover, the transcription levels of proinflammatory cytokines IL-1β and IL-6 in lymphoid tissues of persons using cannabis were also significantly lower. Our findings suggested that cannabis use is associated with reduced sizes and inflammatory cytokine expression of subtype C HIV-1 reservoirs in men with suppressed viral load.

DAA-mediated HCV cure reduces HIV DNA levels in HCV/HIV coinfected people.

Antiretroviral therapy (ART) for human immunodeficiency virus (HIV) can control virus replication and prolong the life of people living with HIV (PLWH). However, the virus remains dormant within immune cells in what is called the HIV reservoir. Furthermore, 2.3 million PLWH are also coinfected with hepatitis C virus (HCV) and are at risk of developing chronic liver disease and cancer. HCV treatment with direct acting antivirals (DAA) can completely cure the infection in more than 95% of treated individuals and improve their long-term health outcomes. In this study, we investigated how HCV treatment and cure affect the HIV reservoir. We demonstrate the beneficial impact of DAA treatment as it reduces the HIV reservoirs in particular in people infected with HCV before HIV. These results support the need for early ART and DAA treatment in HIV/HCV coinfections.

Severe intestinal barrier damage in HIV-infected immunological non-responders

The intestinal epithelial barrier plays an important role during human immunodeficiency virus (HIV) disease progression. However, the extent to which the intestinal epithelial barrier is damaged in immunological non-responders (INRs) and immunological responders (IRs) is largely unknown. In this study, we investigated and compared the levels of intestinal gland damage and related molecules, including the tight junction protein claudin-1, apoptosis marker caspase-3, HIV DNA, CD4+ T cell count, and inflammation marker tumor necrosis factor-α (TNF-α) among the IRs (n = 10), INRs (n = 8), and healthy controls (HCs, n = 7). Intestinal damage was not completely restored in both INRs and IRs and was more serious in INRs than that in IRs. Moreover, intestinal damage was positively correlated with HIV DNA levels and negatively correlated with CD4+ T cell counts. These results provide insight into understanding the characteristics of intestinal epithelial barrier damage between IRs and INRs.

Cell membranes cloaked magnetic nanoparticles for target recycling detection of nucleic acid

Accurate analysis of human immunodeficiency virus (HIV) related molecules are significant for early diagnosis and therapy of acquired immune deficiency syndrome (AIDS). In this work, a novel electrochemical biosensor for HIV DNA detection is developed. Target induced recycling amplification is designed to enhance the sensitivity. Magnetic nanoparticles are used for facile separation, which are first cloaked by red blood cell membranes and then modified with DNA strands. Target initiated formation of multiple DNA three-way junctions occurs at the interface and the nanoconjugates can be conveniently immobilized on the surface of magnetic electrode. The measured responses from electrochemical species carried on the nanoconjugates can be used to reflect the level of target DNA. This proposed electrochemical method shows excellent sensing performances with a linear range from 0.1 fM to 1 pM. The limit of detection is as low as 35 aM. It is also successfully applied in clinical serum samples with satisfactory results. Therefore, this approach has great potential use for point-of-care testing of HIV DNA and provides a general strategy for various virus analysis.

Ultrasensitive detection of HIV DNA using an electrochemical biosensor with branched DNA amplification

This paper presents a novel strategy for ultrasensitive detection of human immunodeficiency virus (HIV) DNA by an electrochemical biosensor. By the shearing cycle of the target DNA (T-DNA), a large amount of complimentary sequence probes (CS probe DNA) are obtained. Then, the Zr4+ functional probe DNA containing many branched DNAs is captured. Next, the initiator α-bromophenylacetic acid is connected to the branched chains of probe DNA by esterification reaction. Numerous electroactive monomers, namely, ferrocenylmethyl methacrylate (FMMA), are produced at the end of the branched chains by electrochemically mediated atomic transfer radical polymerization reaction. The signal of FMMA is greatly enhanced due to a large number of branched chain-loaded probe clusters containing numerous FMMA. T-DNA can be detected with a linear range of 0.1 fM–10 nM with a detection limit of 0.037 fM. The sensor is highly sensitive and selective. It has been applied to the detection of HIV DNA in human serum samples, and it has potential application prospects in the early clinical diagnosis of AIDS.

HIV Tissue Reservoirs: Current Advances in Research.

Acquired immunodeficiency syndrome (AIDS), caused by the human immunodeficiency virus (HIV), has become a heavy burden of disease and an important public health problem in the world. Although current antiretroviral therapy (ART) is effective at suppressing the virus in the blood, HIV still remains in two different types of reservoirs-the latently infected cells (represented by CD4+ T cells) and the tissues containing those cells, which may block access to ART, HIV-neutralizing antibodies and latency-reversing agents. The latter is the focus of our review, as blood viral load drops below detectable levels after ART, a deeper and more systematic understanding of the HIV tissue reservoirs is imperative. In this review, we take the lymphoid system (including lymph nodes, gut-associated lymphoid tissue, spleen and bone marrow), nervous system, respiratory system, reproductive system (divided into male and female), urinary system as the order, focusing on the particularity and importance of each tissue in HIV infection, the infection target cell types of each tissue, the specific infection situation of each tissue quantified by HIV DNA or HIV RNA and the evidence of compartmentalization and pharmacokinetics. In summary, we found that the present state of HIV in different tissues has both similarities and differences. In the future, the therapeutic principle we need to follow is to respect the discrepancy on the basis of grasping the commonality. The measures taken to completely eliminate the virus in the whole body cannot be generalized. It is necessary to formulate personalized treatment strategies according to the different characteristics of the HIV in the various tissues, so as to realize the prospect of curing AIDS as soon as possible.

Monocyte-derived macrophages contain persistent latent HIV reservoirs

The development of persistent cellular reservoirs of latent human immunodeficiency virus (HIV) is a critical obstacle to viral eradication since viral rebound takes place once anti-retroviral therapy (ART) is interrupted. Previous studies show that HIV persists in myeloid cells (monocytes and macrophages) in blood and tissues in virologically suppressed people with HIV (vsPWH). However, how myeloid cells contribute to the size of the HIV reservoir and what impact they have on rebound after treatment interruption remain unclear. Here we report the development of a human monocyte-derived macrophage quantitative viral outgrowth assay (MDM-QVOA) and highly sensitive T cell detection assays to confirm purity. We assess the frequency of latent HIV in monocytes using this assay in a longitudinal cohort of vsPWH (n = 10, 100% male, ART duration 5–14 yr) and find half of the participants showed latent HIV in monocytes. In some participants, these reservoirs could be detected over several years. Additionally, we assessed HIV genomes in monocytes from 30 vsPWH (27% male, ART duration 5–22 yr) utilizing a myeloid-adapted intact proviral DNA assay (IPDA) and demonstrate that intact genomes were present in 40% of the participants and higher total HIV DNA correlated with reactivatable latent reservoirs. The virus produced in the MDM-QVOA was capable of infecting bystander cells resulting in viral spread. These findings provide further evidence that myeloid cells meet the definition of a clinically relevant HIV reservoir and emphasize that myeloid reservoirs should be included in efforts towards an HIV cure.

3-D DNA walker-mediated ultrasensitive dual-signal biosensor for HIV DNA detection

Here, we successfully designed a dual-signal sensing system based on 3-D DNA walker signal amplification and target recycling amplification strategies for ultrasensitive determination of the human immunodeficiency virus（HIV）DNA. The DNA walker is constructed by locked swing arm and track strand onto the surface of the magnetic beads (MBs). After adding HIV DNA, the swing arm was activated to hybridize with the track strand to form ds-DNA, which could be cleaved by Exonuclease III (Exo III) to release G-quadruplex. Subsequently, hemin combined with G-quadruplex to construct DNAzymes, which catalyzed the oxidation of o-phenylenediamine (OPD) into the yellow product 2,3 diaminophenazine (DAP), resulting in the fluorescence quenching of MoS2 QDs at 420 nm, and the fluorescence enhancement of DAP at 560 nm. Therefore, the amounts of HIV DNA can be detected by Uv-vis absorbance and fluorescence dual-signal changes. Besides, each swing arm cleaved multiple track strands to amplify the signal, while the target recycling amplification was also in progress. Such double-amplified signal strategy not only provided excellent sensitivity and selectivity, but also realized magnetic separation, eliminating interferences in complex samples. In addition, our designed sensing platform can detect different DNAs by adjusting the recognition sequence, providing a general strategy for various virus analysis.

A Rapid and Sensitive Detection of HIV-1 with a One-Pot Two-Stage Reverse Transcription Recombinase Aided Real-Time PCR Assay.

Human immunodeficiency virus 1 (HIV-1) attacks the immune system, making people susceptible to various diseases, thus increasing their risk of death. Comprehensive detection of major HIV-1 strains circulating in China is vital for effective HIV-1 infection prevention and treatment. HIV-1 nucleic acid detection is considered effective for HIV-1 diagnosis since traditional immunological testing may fail to detect HIV-1 infection during the window period. This work demonstrates a one-pot two-stage amplification assay (RT-RAP), a combination of reverse transcription recombinase (RT- RAA), and quantitative real-time polymerase chain reaction (qRT-PCR). The turn-around time of the assay is only 50 min and can be performed with commonly available laboratory equipment, the qPCR devices. The RT-RAP assay could detect approximately 5 and 14 copies/reaction of HIV-1 DNA and RNA using recombinant plasmids and standard reference strains, respectively. Additionally, we found that the clinical performance of RT-RAP (detected 169 samples out of 170 specimens) was consistent with that of qRT-PCR. The sensitivity and specificity of RT-RAP were 100.00% (99/99) and 98.59% (70/71), respectively, while its positive and negative predictive values were 99.00% (99/100) and 100.00% (70/70), respectively. The total coincidence rate of the RT-RAP was 99.41% (169/170), with a kappa value of 0.988 (p < 0.05). We demonstrated that RT-RAP could rapidly detect the common HIV-1 subtypes commonly circulating in China with comparable sensitivity and specificity to qRT-PCR.

Pt/Ti3C2Tx nanozyme-amplified colorimetric lateral flow biosensor for dual-readout detection of HIV-DNA

Rapid and sensitive virus genes detection with point-of-care testing is the key to improving the probability of survival and lowering infectious rate. Here, we proposed a sensitive and quantitative colorimetric lateral flow biosensor (CLFB) for human immunodeficiency virus-DNA (HIV-DNA) detection with a novel nanomaterial (Pt nanoparticles decorated Ti3C2Tx MXene, Pt/Ti3C2Tx) as signal reporters. Pt/Ti3C2Tx was synthesized by a self-reduction method with Ti3C2Tx as both reducing agent and supporter. Each Pt/Ti3C2Tx contained hundreds of Pt nanoparticles to achieve enhanced colorimetric dual-readout, which can not only show gray color itself, but also catalyze its substrate to produce more obvious red color within 3 min. The integration of mimic enzyme property of Pt/Ti3C2Tx improved the colorimetric signal brightness and decreased naked-eye detection limit of HIV-DNA to 0.1 nM relative to original 0.8 nM. Capture DNA was further modified onto the Pt/Ti3C2Tx to realize specific binding with HIV-DNA, enabling the CLFB good reproducibility with an average intra-assay deviation of 4.3% and an inter-assay deviation of 4.2% respectively. Furthermore, the successful detection of HIV-DNA in serum samples demonstrated its great application potential in real samples and timely diagnosis.

Eradication of human immunodeficiency virus-1 reservoir in the brain microglia.

Eradication of human immunodeficiency virus-1 reservoir in the brain microglia.

Fluorine-nitrogen co-doped carbon dots with stable and strong electrochemiluminescence as an emitter for ultrasensitive detection of HIV-DNA fragment

Herein, the fluorine-nitrogen co-doped carbon dots (FNCDs) with stable and strong electrochemiluminescence (ECL) was used as a novel ECL emitter for constructing an ultrasensitive biosensor to detect human immunodeficiency virus (HIV) DNA fragment. The doping of fluorine and nitrogen in the carbon dots (CDs) not only effectively formed hydrogen bonds to hinder the free rotation of CDs for increasing the radiation transition probability, but also changed the charge distribution of CDs leading to a narrower energy gap, which improved the ECL intensity and stability of FNCDs. Moreover, exonuclease III (Exo III) -assisted multiple recycling amplification could convert the trace HIV-DNA fragment into a large amount of output DNA to trigger horizontal hybridization chain reaction (H-HCR) to form Y-supported three-dimensional (3D) network DNA nanostructure for immobilizing a lot of doxorubicin-ferrocenecarboxylic acid (Dox-FcCOOH) as ECL quencher, which significantly improved the sensitivity of the proposed biosensor. Thus, the fabricated ECL biosensor realized ultrasensitive detection of HIV-DNA fragment in the concentration range of 10 aM to 10 pM with a detection limit as low as 6.34 aM. This strategy provided a novel ECL emitter to construct biosensors for detection of biomolecule and early clinical disease diagnosis.

Association Between the Development of Subclinical Cardiovascular Disease and Human Immunodeficiency Virus (HIV) Reservoir Markers in People With HIV on Suppressive Antiretroviral Therapy.

We report that people with human immunodeficiency virus (HIV) diagnosed with coronary artery atherosclerotic plaques display higher levels of HIV DNA compared with those without atherosclerotic plaques. In a multivariable prediction model that included 27 traditional and HIV-related risk factors, measures of HIV DNA were among the most important predictors of atherosclerotic plaque formation.

Systems biology analyses reveal enhanced chronic morphine distortion of gut-brain interrelationships in simian human immunodeficiency virus infected rhesus macaques.

BackgroundCommonly used opioids, such as morphine have been implicated in augmented SIV/HIV persistence within the central nervous system (CNS). However, the extent of myeloid cell polarization and viral persistence in different brain regions remains unclear. Additionally, the additive effects of morphine on SIV/HIV dysregulation of gut-brain crosstalk remain underexplored. Therefore, studies focused on understanding how drugs of abuse such as morphine affect immune dynamics, viral persistence and gut-brain interrelationships are warranted.Materials and methodsFor a total of 9 weeks, rhesus macaques were ramped-up, and twice daily injections of either morphine (n = 4) or saline (n = 4) administered. This was later followed with infection with SHIVAD8EO variants. At necropsy, mononuclear cells were isolated from diverse brain [frontal lobe, cerebellum, medulla, putamen, hippocampus (HIP) and subventricular zone (SVZ)] and gut [lamina propria (LP) and muscularis (MUSC) of ascending colon, duodenum, and ileum] regions. Multiparametric flow cytometry was used to were profile for myeloid cell polarity/activation and results corroborated with indirect immunofluorescence assays. Simian human immunodeficiency virus (SHIV) DNA levels were measured with aid of the digital droplet polymerase chain reaction (PCR) assay. Luminex assays were then used to evaluate soluble plasma/CSF biomarker levels. Finally, changes in the fecal microbiome were evaluated using 16S rRNA on the Illumina NovaSeq platform.ResultsFlow Cytometry-based semi-supervised analysis revealed that morphine exposure led to exacerbated M1 (CD14/CD16)/M2 (CD163/CD206) polarization in activated microglia that spanned across diverse brain regions. This was accompanied by elevated SHIV DNA within the sites of neurogenesis–HIP and SVZ. HIP/SVZ CD16+ activated microglia positively correlated with SHIV DNA levels in the brain (r = 0.548, p = 0.042). Simultaneously, morphine dependence depleted butyrate-producing bacteria, including Ruminococcus (p = 0.05), Lachnospira (p = 0.068) genera and Roseburia_sp_831b (p = 0.068). Finally, morphine also altered the regulation of CNS inflammation by reducing the levels of IL1 Receptor antagonist (IL1Ra).ConclusionThese findings are suggestive that morphine promotes CNS inflammation by altering receptor modulation, increasing myeloid brain activation, distorting gut-brain crosstalk, and causing selective enhancement of SHIV persistence in sites of neurogenesis.

Synthetic gRNA/Cas9 Ribonucleoprotein Inhibits HIV Reactivation and Replication.

The current antiretroviral therapy (ART) for human immunodeficiency virus (HIV) can halt viral replication but cannot eradicate HIV infection because proviral DNA integrated into the host genome remains genetically silent in reservoir cells and is replication-competent upon interruption or cessation of ART. CRISPR/Cas9-based technology is widely used to edit target genes via mutagenesis (i.e., nucleotide insertion/deletion and/or substitution) and thus can inactivate integrated proviral DNA. However, CRISPR/Cas9 delivery systems often require viral vectors, which pose safety concerns for therapeutic applications in humans. In this study, we used synthetic guide RNA (gRNA)/Cas9-ribonucleoprotein (RNP) as a non-viral formulation to develop a novel HIV gene therapy. We designed a series of gRNAs targeting different HIV genes crucial for HIV replication and tested their antiviral efficacy and cellular cytotoxicity in lymphoid and monocytic latent HIV cell lines. Compared with the scramble gRNA control, HIV-gRNA/Cas9 RNP-treated cells exhibited efficient viral suppression with no apparent cytotoxicity, as evidenced by the significant inhibition of latent HIV DNA reactivation and RNA replication. Moreover, HIV-gRNA/Cas9 RNP inhibited p24 antigen expression, suppressed infectious viral particle production, and generated specific DNA cleavages in the targeted HIV genes that are confirmed by DNA sequencing. Because of its rapid DNA cleavage, low off-target effects, low risk of insertional mutagenesis, easy production, and readiness for use in clinical application, this study provides a proof-of-concept that synthetic gRNA/Cas9 RNP drugs can be utilized as a novel therapeutic approach for HIV eradication.

Intact HIV Proviruses Persist in the Brain Despite Viral Suppression with ART.

Human immunodeficiency virus (HIV) persistence in blood and tissue reservoirs, including the brain, is a major barrier to HIV cure and possible cause of comorbid disease. However, the size and replication competent nature of the central nervous system (CNS) reservoir is unclear. Here, we used the intact proviral DNA assay (IPDA) to provide the first quantitative assessment of the intact and defective HIV reservoir in the brain of people with HIV (PWH). Total, intact, and defective HIV proviruses were measured in autopsy frontal lobe tissue from viremic (n=18) or virologically suppressed (n=12) PWH. Total or intact/defective proviruses were measured by detection of HIV pol or the IPDA, respectively, through use of droplet digital polymerase chain reaction (ddPCR). HIV-seronegative individuals were included as controls (n=6). Total HIV DNA was present at similar levels in brain tissues from untreated viremic and antiretroviral (ART)-suppressed individuals (median=22.3 vs 26.2 HIV pol copies/106 cells), reflecting a stable CNS reservoir of HIV that persists despite therapy. Furthermore, 8 of 10 viremic and 6 of 9 virally suppressed PWH also harbored intact proviruses in the CNS (4.63 vs 12.7 intact copies/106 cells). Viral reservoirs in CNS and matched lymphoid tissue were similar in the composition of intact and/or defective proviruses, albeit at lower levels in the brain. Importantly, CNS resident CD68+ myeloid cells in virally suppressed individuals harbored HIV DNA, directly showing the presence of a CNS resident HIV reservoir. Our results demonstrate the first evidence for an intact, potentially replication competent HIV reservoir in the CNS of virally suppressed PWH. ANN NEUROL 2022;92:532-544.

Mother-to-Child Transmission of HIV Within 24 Months After Delivery in Women Initiating Lifelong Antiretroviral Therapy Pre/Post-Conception or Postnatally; Effects of Adolescent Girl and Young Woman Status and Plasma Viremia Late in Pregnancy

IntroductionMother-to-child-transmission (MTCT) of human immunodeficiency virus (HIV) can occur in pregnancy/in utero (IU), during childbirth/intrapartum (IP), or postpartum (PP) through breastfeeding from an infected mother to her infant. Burden of PP-MTCT and associated risk factors remain poorly described, especially in adolescent girls and young women (AGYW) aged 15–24 years. Furthermore, despite concerns on high postnatal seroconversions, there is paucity of data on the burden of subsequent MTCT rates.MethodsPregnant women ≥20 weeks of gestation were enrolled into the University of Zimbabwe Birth Cohort from four primary health centers in Harare, Zimbabwe. Mother–infant dyads were followed up from delivery, week(s) 1, 6, 10, 14, 24, 36, 48, 72, and 96 after birth. Women who were uninfected at baseline were re-tested for HIV on subsequent visits. Plasma HIV RNA was quantified using reverse transcriptase polymerase chain reaction. Exposed babies were tested for HIV using qualitative/quantitative proviral DNA PCR on dried blood spots. Maternal–infant factors were tested in univariable/multivariable regression analyses for HIV-MTCT predictors.ResultsA total of 600 HIV-uninfected and 608 HIV-infected pregnant women on Tenofovir/Lamivudine/Efavirenz regimen were enrolled from 2016 to 2019. Postnatal HIV incidence was 0.42 cases/100 women-years [95% confidence interval (CI): 0.12–1.1]. Postnatal seroconverters were less likely to have children/pregnancies sharing same father and unaware of their spouses/intimate partner’s HIV status: p = 0.008 and p = 0.02, respectively, compared with non-seroconverters.Overall HIV-MTCT rate was (15/549): 2.7% (CI: 1.3–4.1%); (7/93) 7.5% observed in AGYW against 1.7%; in women aged &amp;gt;24, p = 0.008. PP-MTCT was the predominant 9/15 (60%) route, followed by IP-MTCT 4/15 (26.6%), whereas IU and postnatal MTCT rates each contributed 6.7% of all infant infections. Postnatal MTCT incidence was 12.8 (CI: 0.3–71.4) infant HIV infections/100 child-years of breastfeeding; a rate 14 times higher than PP-MTCT rate in babies born to women HIV-infected pre/post-conception whose babies were HIV DNA PCR–negative at six weeks.Antenatal HIV RNA &amp;gt;1,000 copies/ml was independently associated with MTCT; odds ratio [CI: 9.3 (2.6–43.1)]. Infected infants’ pre–HIV treatment HIV RNA levels correlated positively with maternal viral load; Spearman’s rank correlation. r = 0.6; p = 0.03.DiscussionMothers were 9.3 times more likely to transmit if HIV RNA was &amp;gt;1,000 copies/ml, disproportionately occurring in vulnerable AGYW. Breastfeeding-associated PP-MTCT remains high; therefore, it is imperative that HIV-infected women commence antiretroviral therapy early in pregnancy to suppress HIV RNA until weaning to decrease the risk of MTCT and possibly reduce the severity of disease in infected infants. HIV-uninfected lactating mothers should be continuously counseled on the risks of postnatal seroconversion.www.clinicaltrials.gov, trial registration number: NCT04087239.

Recognition of HIV-1 capsid by PQBP1 licenses an innate immune sensing of nascent HIV-1 DNA

We have previously described polyglutamine-binding protein 1 (PQBP1) as an adapter required for the cyclic GMP-AMP synthase (cGAS)-mediated innate response to the human immunodeficiency virus 1 (HIV-1) and other lentiviruses. Cytoplasmic HIV-1 DNA is a transient and low-abundance pathogen-associated molecular pattern (PAMP), and the mechanism for its detection and verification is not fully understood. Here, we show a two-factor authentication strategy by the innate surveillance machinery to selectively respond to the low concentration of HIV-1 DNA, while distinguishing these species from extranuclear DNA molecules. We find that, upon HIV-1 infection, PQBP1 decorates the intact viral capsid, and this serves as a primary verification step for the viral nucleic acid cargo. As reverse transcription and capsid disassembly initiate, cGAS is recruited to the capsid in a PQBP1-dependent manner. This positions cGAS at the site of PAMP generation and sanctions its response to a low-abundance DNA PAMP.