Determinants Of Hiv-1 Transmission Fitness

DETERMINANTS OF HIV-1 TRANSMISSION FITNESS Shilpa S. Iyer Beatrice H. Hahn HIV-1 is predominantly transmitted by mucosal routes and almost 80 percent of new infections are initiated by a single variant. The elucidation of the biological properties of transmitted viruses which distinguish them from non-transmitted variants are critical for the development of therapeutic interventions. To identify such properties, we characterized the biology of 300 limiting dilution-derived virus isolates from the plasma and genital secretions of eight HIV-1 donor and recipient transmission pairs representing the most prevalent subtypes (B and C). Recipient viruses were more infectious per viral particle as determined on a reporter cell line, replicated to higher titers and were released more efficiently from infected primary CD4+ T cells than the corresponding donor isolates. Recipient viruses were more resistant to the inhibitory effects of IFN-α2 and IFN-β evidenced as higher half-maximal inhibitory concentrations and higher replication at the maximal doses of IFN-α2 and IFN-β than corresponding donor isolates. Interestingly, pretreatment of CD4+ T cells with IFN-β, but not IFN-α2 selected donor plasma isolates that exhibited phenotypes similar to transmitted viruses. This suggests that transmitted variants are distinct and that the selective pressure imposed by type I interferons may in part be responsible for the bottleneck associated with mucosal transmission. We next wanted to assess the role of the interferon stimulated gene, tetherin in the antiviral state established by type I IFNs. Thus, we introduced mutations into the vpu gene of various HIV-1 constructs to specifically disrupt their Vpu-mediated tetherin antagonism, and determined the effect on replication and release from infected cells in the presence and absence of IFN-α2. Mutations at key residues in Vpu reduced the viral particle production and release from infected primary CD4+ T cells and this was particularly evident in IFN-α2-treated cells. Interestingly, transmitted HIV-1 variants were released to higher levels from infected cells than chronic control viruses, even in the absence of Vpu. Thus, the counteraction of tetherin resulting in efficient particle release is an important determinant of the interferon resistance of mucosally transmitted HIV-1

Recombinant Mitochondrial Transcription Factor A with N-terminal Mitochondrial Transduction Domain Increases Respiration and Mitochondrial Gene Expression in G11778A Leber's Hereditary Optic Neuropathy Cybrid Cells

Diseases involving mitochondrial defects usually manifest themselves in high-energy, post-mitotic tissues such as brain, retina, skeletal and cardiac muscle and frequently cause deficiencies in mitochondrial bioenergetics. We have developed a scalable procedure to produce recombinant human mitochondrial transcription factor A (TFAM) modified with an N-terminal protein transduction domain (PTD) and mitochondrial localization signal (MLS) that allow it to cross membranes and enter mitochondria through its "mitochondrial transduction domain" (MTD,=PTD+MLS). _In vitro_ studies in a classic mitochondrial disease cell model demonstrated that Alexa488-labeled MTD-TFAM rapidly entered the mitochondrial compartment. MTD-TFAM treatment of these cell lines reversibly increased oxygen consumption (respiration) rates 3-fold, levels of respiratory proteins and mitochondrial gene expression. _In vivo_ results demonstrated that respiration increased to lesser degrees in mitochondria from tissues of mice injected with MTD-TFAM. MTD-TFAM can alter mitochondrial bioenergetics and holds promise for treatment of mitochondrial diseases involving deficiencies of energy production

Efficient Web Navigator for Multi-Constrained Spatial Keyword Queries

The mobile technology revolutionizes the world of communications opening up new possibilities for applications such as location based web search. It involves retrieving the user point of interest (POI) from the web documents based on the query relative to a particular place or region. Existing Location based Applications on mobiles finds the nearest neighbors from the POI database not from the web documents. The existing query searches are limited to POI and do not include data objects brands like Model name, color and price etc. This paper introduces a Spatial Web Crawler (SWC) for multi-constrained keyword queries. New algorithms were developed which provides desired data objects from the web pages basing on the working hours of data objects, keywords and priority such as cost, quality and popularity of data objects etc. The SWC also provides the shortest path to reach point of interest based on travel time

Can patients independently identify their urinary incontinence symptoms?

INTRODUCTION AND HYPOTHESIS: The objective of our study is to compare patient self-reported urinary incontinence symptoms based on the International Consultation on Incontinence Questionnaire- Short Form (ICIQ-SF) question number 6 (When does urine leak?) with physician-assessed interpretation of the patient\u27s urinary incontinence symptoms. METHODS: This trial is a cross-sectional study of patients who presented to a tertiary urogynecology center with symptoms of urinary incontinence between January 2014 and August 2016. We compared patient-reported symptoms on the ICIQ-SF with physician interpretation of urinary complaints during their initial visit. The urinary incontinence symptoms included stress urinary incontinence (SUI), urgency urinary incontinence (UUI), insensible urine loss, nocturnal enuresis, and post-micturition dribbling. RESULTS: A total of 432 patients with a mean age of 61 were included in this evaluation. The most common urinary incontinence symptoms according to the physician were UUI (n = 357, 83%), followed by SUI (n = 308, 71%). Of the patients who were diagnosed by a physician with the symptom of UUI, only 61% self-identified as having this symptom based on the ICIQ-SF, and for SUI, only 66% self-identified as having SUI symptoms based on the ICIQ-SF. Overall UUI (κ = 0.30) appears to have poor agreement, as does nocturnal enuresis (κ = 0.39), when compared with physician historical assessment. CONCLUSION: There is a discrepancy between patient-reported urinary incontinence symptoms on the ICIQ-SF and physician-assessed symptoms. Symptomatology entered into electronic medical records by patients is often inaccurate. Physician validation is essential in understanding the underlying the precise symptomatology

Telomeric circles are abundant in the stn1-M1 mutant that maintains its telomeres through recombination

Some human cancers maintain their telomeres using the alternative lengthening of telomeres (ALT) mechanism; a process thought to involve recombination. Different types of recombinational telomere elongation pathways have been identified in yeasts. In senescing yeast telomerase deletion (ter1-Δ) mutants with very short telomeres, it has been hypothesized that copying a tiny telomeric circle (t-circle) by a rolling circle mechanism is the key event in telomere elongation. In other cases more closely resembling ALT cells, such as the stn1-M1 mutant of Kluyveromyces lactis, the telomeres appear to be continuously unstable and routinely reach very large sizes. By employing two-dimensional gel electrophoresis and electron microscopy, we show that stn1-M1 cells contain abundant double stranded t-circles ranging from ∼100 to 30 000 bp in size. We also observed small single-stranded t-circles, specifically composed of the G-rich telomeric strand and tailed circles resembling rolling circle replication intermediates. The t-circles most likely arose from recombination events that also resulted in telomere truncations. The findings strengthen the possibility that t-circles contribute to telomere maintenance in stn1-M1 and ALT cells

Mitigating the effects of COVID-19 on HIV treatment and care in Lusaka, Zambia: A before-after cohort study using mixed effects regression

INTRODUCTION: The Zambian Ministry of Health (MoH) issued COVID-19 mitigation guidance for HIV care immediately after the first COVID-19 case was confirmed in Zambia on 18 March 2020. The Centre for Infectious Disease Research in Zambia implemented MoH guidance by: 1) extending antiretroviral therapy (ART) refill duration to 6 multi-month dispensation (6MMD) and 2) task-shifting communication and mobilisation of those in HIV care to collect their next ART refill early. We assessed the impact of COVID-19 mitigation guidance on HIV care 3 months before and after guidance implementation. METHODS: We reviewed all ART pharmacy visit data in the national HIV medical record for PLHIV in care having ≥1 visit between 1 January-30 June 2020 at 59 HIV care facilities in Lusaka Province, Zambia. We undertook a before-after evaluation using mixed-effects Poisson regression to examine predictors and marginal probability of early clinic return (pharmacy visit \u3e7 days before next appointment), proportion of late visit (\u3e7 days late for next appointment) and probability of receiving a 6MMD ART refill. RESULTS: A total of 101 371 individuals (64% female, median age 39) with 130 486 pharmacy visits were included in the analysis. We observed a significant increase in the adjusted prevalence ratio (4.63; 95% CI 4.45 to 4.82) of early return before compared with after guidance implementation. Receipt of 6MMD increased from a weekly mean of 47.9% (95% CI 46.6% to 49.2%) before to 73.4% (95% CI 72.0% to 74.9%) after guidance implementation. The proportion of late visits (8-89 days late) was significantly higher before (18.8%, 95% CI17.2%to20.2%) compared with after (15.1%, 95% CI13.8%to16.4%) guidance implementation . CONCLUSIONS: Timely issuance and implementation of COVID-19 mitigation guidance involving task-shifted patient communication and mobilisation alongside 6MMD significantly increased early return to ART clinic, potentially reducing interruptions in HIV care during a global public health emergency

The effect of HIV-1 Vif polymorphisms on A3G anti-viral activity in an in vivo mouse model

Abstract Humans encode seven APOBEC3 proteins (A–H), with A3G, 3F and 3H as the major factors restricting HIV-1 replication. HIV-1, however, encodes Vif, which counteracts A3 proteins by chaperoning them to the proteasome where they are degraded. Vif polymorphisms found in HIV-1s isolated from infected patients have varying anti-A3G potency when assayed in vitro, but there are few studies demonstrating this in in vivo models. Here, we created Friend murine leukemia viruses encoding vif alleles that were previously shown to differentially neutralize A3G in cell culture or that were originally found in primary HIV-1 isolates. Infection of transgenic mice expressing different levels of human A3G showed that these naturally occurring Vif variants differed in their ability to counteract A3G during in vivo infection, although the effects on viral replication were not identical to those seen in cultured cells. We also found that the polymorphic Vifs that attenuated viral replication reverted to wild type only in A3G transgenic mice. Finally, we found that the level of A3G-mediated deamination was inversely correlated with the level of viral replication. This animal model should be useful for studying the functional significance of naturally occurring vif polymorphisms, as well as viral evolution in the presence of A3G

Comparable exposure to SARS-CoV-2 in young children and healthcare workers in Zambia

Background: Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is an ongoing global health crisis that has caused large scale morbidity and mortality. We aimed to determine the exposure to SARS-CoV-2 among young children and healthcare workers by measurement of anti-S1 antigen (spike protein) specific immunoglobulin G (IgG) using an in-house optimized indirect enzyme-linked immunosorbent assay (ELISA) method. Methods: Plasma samples were collected from cohorts of healthcare workers (n = 287) and young children aged from 6 weeks to 2 years old (n = 150) pre-COVID-19 pandemic between September 2018 and November 2019 and post-COVID-19 pandemic between August and December 2020 were simultaneously tested for anti-SARS-CoV-2 S1 specific IgG. The arithmetic mean of natural logarithm-transformed ELISA relative absorbance reading + (3 x standard deviation) of pre-pandemic plasma was used as the cut-off to determine SARS-CoV-2 IgG seropositivity of post-pandemic plasma, Results: There was no reactivity to SARS-CoV-2 S1 antigen detected in pre-pandemic plasma but in post pandemic plasma an 8.0% (23/287) IgG seropositivity in healthcare workers’ and 6.0% (9/150) seropositivity in children aged 2 years old was detected. Conclusions: Comparable levels of SARS-CoV-2 IgG seropositivity in healthcare workers and children suggest widespread exposure to SARS-CoV-2 in Zambia during the first wave of the pandemic. This finding has implications for continued acquisition and transmission of infection in the healthcare setting, household, and wider community

Transmitted/Founder and Chronic Subtype C HIV-1 Use CD4 and CCR5 Receptors with Equal Efficiency and Are Not Inhibited by Blocking the Integrin α4β7

Sexual transmission of human immunodeficiency virus type 1 (HIV-1) most often results from productive infection by a single transmitted/founder (T/F) virus, indicating a stringent mucosal bottleneck. Understanding the viral traits that overcome this bottleneck could have important implications for HIV-1 vaccine design and other prevention strategies. Most T/F viruses use CCR5 to infect target cells and some encode envelope glycoproteins (Envs) that contain fewer potential N-linked glycosylation sites and shorter V1/V2 variable loops than Envs from chronic viruses. Moreover, it has been reported that the gp120 subunits of certain transmitted Envs bind to the gut-homing integrin α4β7, possibly enhancing virus entry and cell-to-cell spread. Here we sought to determine whether subtype C T/F viruses, which are responsible for the majority of new HIV-1 infections worldwide, share biological properties that increase their transmission fitness, including preferential α4β7 engagement. Using single genome amplification, we generated panels of both T/F (n = 20) and chronic (n = 20) Env constructs as well as full-length T/F (n = 6) and chronic (n = 4) infectious molecular clones (IMCs). We found that T/F and chronic control Envs were indistinguishable in the efficiency with which they used CD4 and CCR5. Both groups of Envs also exhibited the same CD4+ T cell subset tropism and showed similar sensitivity to neutralization by CD4 binding site (CD4bs) antibodies. Finally, saturating concentrations of anti-α4β7 antibodies failed to inhibit infection and replication of T/F as well as chronic control viruses, although the growth of the tissue culture-adapted strain SF162 was modestly impaired. These results indicate that the population bottleneck associated with mucosal HIV-1 acquisition is not due to the selection of T/F viruses that use α4β7, CD4 or CCR5 more efficiently

Resistance to Type 1 Interferons is a Major Determinant of HIV-1 Transmission Fitness

Sexual transmission of HIV-1 is an inefficient process, with only one or few variants of the donor quasispecies establishing the new infection. A critical, and as yet unresolved, question is whether the mucosal bottleneck selects for viruses with increased transmission fitness. Here, we characterized 300 limiting dilution-derived virus isolates from the plasma, and in some instances genital secretions, of eight HIV-1 donor and recipient pairs. Although there were no differences in the amount of virion-associated envelope glycoprotein, recipient isolates were on average threefold more infectious (P = 0.0001), replicated to 1.4-fold higher titers (P = 0.004), were released from infected cells 4.2-fold more efficiently (P < 0.00001), and were significantly more resistant to type I IFNs than the corresponding donor isolates. Remarkably, transmitted viruses exhibited 7.8-fold higher IFNα2 (P < 0.00001) and 39-fold higher IFNβ (P < 0.00001) half-maximal inhibitory concentrations (IC50) than did donor isolates, and their odds of replicating in CD4+ T cells at the highest IFNα2 and IFNβ doses were 35-fold (P < 0.00001) and 250-fold (P < 0.00001) greater, respectively. Interestingly, pretreatment of CD4+ T cells with IFNβ, but not IFNα2, selected donor plasma isolates that exhibited a transmitted virus-like phenotype, and such viruses were also detected in the donor genital tract. These data indicate that transmitted viruses are phenotypically distinct, and that increased IFN resistance represents their most distinguishing property. Thus, the mucosal bottleneck selects for viruses that are able to replicate and spread efficiently in the face of a potent innate immune response