The mechanism of resistance to favipiravir in influenza.

Favipiravir is a broad-spectrum antiviral that has shown promise in treatment of influenza virus infections. While emergence of resistance has been observed for many antiinfluenza drugs, to date, clinical trials and laboratory studies of favipiravir have not yielded resistant viruses. Here we show evolution of resistance to favipiravir in the pandemic H1N1 influenza A virus in a laboratory setting. We found that two mutations were required for robust resistance to favipiravir. We demonstrate that a K229R mutation in motif F of the PB1 subunit of the influenza virus RNA-dependent RNA polymerase (RdRP) confers resistance to favipiravir in vitro and in cell culture. This mutation has a cost to viral fitness, but fitness can be restored by a P653L mutation in the PA subunit of the polymerase. K229R also conferred favipiravir resistance to RNA polymerases of other influenza A virus strains, and its location within a highly conserved structural feature of the RdRP suggests that other RNA viruses might also acquire resistance through mutations in motif F. The mutations identified here could be used to screen influenza virus-infected patients treated with favipiravir for the emergence of resistance

HIV Prevention in a Time of COVID-19: A Report from the HIVR4P // Virtual Conference 2021.

The HIV Research for Prevention (HIVR4P) conference catalyzes knowledge sharing on biomedical HIV prevention interventions such as HIV vaccines, antibody infusions, pre-exposure prophylaxis, and microbicides in totality-from the molecular details and delivery formulations to the behavioral, social, and structural underpinnings. HIVR4P // Virtual was held over the course of 2 weeks on January 27-28 and February 3-4, 2021 as the coronavirus disease 2019 (COVID-19) pandemic continued to inflict unprecedented harm globally. The HIVR4P community came together with 1,802 researchers, care providers, policymakers, implementers, and advocates from 92 countries whose expertise spanned the breadth of the HIV prevention pipeline from preclinical to implementation. The program included 113 oral and 266 poster presentations. This article presents a brief summary of the conference highlights. Complete abstracts, webcasts, and daily rapporteur summaries may be found on the conference website (https://www.hivr4p.org/)

The evolving SARS-CoV-2 epidemic in Africa: Insights from rapidly expanding genomic surveillance

INTRODUCTION Investment in Africa over the past year with regard to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequencing has led to a massive increase in the number of sequences, which, to date, exceeds 100,000 sequences generated to track the pandemic on the continent. These sequences have profoundly affected how public health officials in Africa have navigated the COVID-19 pandemic. RATIONALE We demonstrate how the first 100,000 SARS-CoV-2 sequences from Africa have helped monitor the epidemic on the continent, how genomic surveillance expanded over the course of the pandemic, and how we adapted our sequencing methods to deal with an evolving virus. Finally, we also examine how viral lineages have spread across the continent in a phylogeographic framework to gain insights into the underlying temporal and spatial transmission dynamics for several variants of concern (VOCs). RESULTS Our results indicate that the number of countries in Africa that can sequence the virus within their own borders is growing and that this is coupled with a shorter turnaround time from the time of sampling to sequence submission. Ongoing evolution necessitated the continual updating of primer sets, and, as a result, eight primer sets were designed in tandem with viral evolution and used to ensure effective sequencing of the virus. The pandemic unfolded through multiple waves of infection that were each driven by distinct genetic lineages, with B.1-like ancestral strains associated with the first pandemic wave of infections in 2020. Successive waves on the continent were fueled by different VOCs, with Alpha and Beta cocirculating in distinct spatial patterns during the second wave and Delta and Omicron affecting the whole continent during the third and fourth waves, respectively. Phylogeographic reconstruction points toward distinct differences in viral importation and exportation patterns associated with the Alpha, Beta, Delta, and Omicron variants and subvariants, when considering both Africa versus the rest of the world and viral dissemination within the continent. Our epidemiological and phylogenetic inferences therefore underscore the heterogeneous nature of the pandemic on the continent and highlight key insights and challenges, for instance, recognizing the limitations of low testing proportions. We also highlight the early warning capacity that genomic surveillance in Africa has had for the rest of the world with the detection of new lineages and variants, the most recent being the characterization of various Omicron subvariants. CONCLUSION Sustained investment for diagnostics and genomic surveillance in Africa is needed as the virus continues to evolve. This is important not only to help combat SARS-CoV-2 on the continent but also because it can be used as a platform to help address the many emerging and reemerging infectious disease threats in Africa. In particular, capacity building for local sequencing within countries or within the continent should be prioritized because this is generally associated with shorter turnaround times, providing the most benefit to local public health authorities tasked with pandemic response and mitigation and allowing for the fastest reaction to localized outbreaks. These investments are crucial for pandemic preparedness and response and will serve the health of the continent well into the 21st century

The impact of human immunodeficiency virus-1 viral replicative capacity on infection of human cervical and rectal tissue explants ex vivo

Characterization of the targets and kinetics of HIV infection within rectal and genital tissues is central to the assessment of novel prevention agents. The ex vivo models available are helpful for studying the initial stages of HIV‐1 infection, such as defining the primary HIV target cells and the factors that influence viral replication kinetics. Five HIV-1 clade C, 1 clade A and 1 clade A/D HIV-1 TF variants were isolated from IAVI Protocol C; a prospective acute infection cohort, and their infection of PBMCs (n=7), rectal (n=59) and endo- and ecto-cervical (n=49) tissue explants were assessed relative to HIV1BaL over 15 days in culture. There was no statistical difference in the rates of replication of the HIV-1 variants relative to HIV-1BaL in endo- and ectocervical tissues with the exception of HIV-1K4791 which replicated significantly higher in ectocervix than endocervix. Interestingly, HIV-1 variants defined as having “high replicative capacity” (RC) replicated to similar rates as those defined as low RC in cervical tissues in these experiments. We observed high levels of inter-donor variation in all tissues tested, however we detected HIV-1 replication in PBMC, rectal and cervical tissues with varied kinetics. HIV-1 variants infected both CD3+Th17 cells as well as non-T cells within rectal and cervical tissues 2 days after infection. The infected non-T cells were large and irregularly shaped; a phenotype consistent with myeloid dendritic or macrophages, and we aim to confirm the identity of these cells using tailored combinations of antibodies in future using existing stored samples. We have successfully transferred and established an explant culture system at KAVI-ICR for studying early events of HIV‐1 transmission in cervical and rectal tissues ex vivo. They are valuable tools to test the efficacy of microbicides in the prevention of HIV‐1 transmission study HIV-1, as well to evaluate HIV-1 vaccine candidatesOpen Acces

Metabolic and mitochondrial dysregulation in CD4+ T cells from HIV-positive women on combination anti-retroviral therapy.

BackgroundFor optimal functionality, immune cells require a robust and adaptable metabolic program that is fueled by dynamic mitochondrial activity. In this study, we investigate the metabolic alterations occurring in immune cells during HIV infection and antiretroviral therapy by analyzing the uptake of metabolic substrates and mitochondrial phenotypes. By delineating changes in immune cell metabolic programming during HIV, we may identify novel potential therapeutic targets to improve anti-viral immune responses.MethodsAfter consent and voluntary participation was confirmed, whole blood was drawn from HIV uninfected women and women with chronic HIV infection on long-term combination antiretroviral therapy (HIV/cART). Peripheral blood mononuclear cells-derived immune cells were directly incubated with different fluorescently tagged metabolites and markers of mitochondrial activity: FITC-2-NBDG (2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) amino]-2-deoxy-D-glucose), FITC-BODIPY (4,4-Difluoro-5,7-Dimethyl-4-Bora-3a,4a-Diaza-s-Indacene-3-Hexadecanoic Acid), FITC-MitoTracker Green and APC-MitoTracker Deep Red. The uptake of glucose and fats and the mitochondrial mass and potential were measured using flow cytometry. All values are reported quantitatively as geometric means of fluorescence intensity.ResultsDuring chronic HIV infection, cellular uptake of glucose increases in HIV+ dendritic cells in particular. CD4+ T cells had the lowest uptake of glucose and fats compared to all other cells regardless of HIV status, while CD8+ T cells took up more fatty acids. Interestingly, despite the lower utilization of glucose and fats in CD4+ T cells, mitochondrial mass increased in HIV+ CD4+ T cells compared to HIV negative CD4+ T-cells. HIV+ CD4+ T cells also had the highest mitochondrial potential.ConclusionsSignificant disparities in the utilization of substrates by leukocytes during chronic HIV/cART exist. Innate immune cells increased utilization of sugars and fats while adaptive immune cells displayed lower glucose and fat utilization despite having a higher mitochondrial activity. Our findings suggest that cART treated HIV-infected CD4+ T cells be dysfunctional or may prefer alternative fuel sources not included in these studies. This underscores the importance of understanding the metabolic effects of HIV treatment on immune function

Women in Health and their Economic, Equity and Livelihood statuses during Emergency Preparedness and Response (WHEELER) protocol: a mixed methods study in Kenya

Introduction Kenya reported its first COVID-19 case on 13 March 2020. Pandemic-driven health system changes followed and unforeseen societal, economic and health effects reported. This protocol aims to describe the methods used to identify the gender equality and health equity gaps and possible disproportional health and socioeconomic impacts experienced by paid and unpaid (community health volunteer) female healthcare providers in Kilifi and Mombasa Counties, Kenya during the COVID-19 pandemic.Methods and analysis Participatory mixed methods framed by gender analysis and human-centred design will be used. Research implementation will follow four of the five phases of the human-centred design approach. Community research advisory groups and local advisory boards will be established to ensure integration and the sustainability of participatory research design.Ethics and dissemination Ethical approval was obtained from the Institutional Scientific and Ethics Review Committee at the Aga Khan University and the University of Manitoba.This study will generate evidence on root cultural, structural, socioeconomic and political factors that perpetuate gender inequities and female disadvantage in the paid and unpaid health sectors. It will also identify evidence-based policy options for future safeguarding of the unpaid and paid female health workforce during emergency preparedness, response and recovery periods

Correlations of substrate uptake with CD4 counts and duration of treatment in HIV⁺ participants.

(A) Correlation of substrate uptake with CD4 counts in chronically infected HIV/cART participants and (B) Correlation of substrate uptake with time on ARVs. Statistical significance shown in charts.</p

Substrate uptake and assessment of mitochondrial phenotype and activity in HIV/cART leukocytes.

(A) Assessment of mitochondrial mass (via MTG staining) in uninfected and chronically infected HIV adaptive and innate immune cells. (B) Glucose uptake (via 2NBDG staining) in uninfected and chronically infected HIV+ adaptive and innate immune cells. (C) Fat uptake (via BODIPY staining) in uninfected and chronically infected HIV adaptive and innate immune cells. (D) Assessment of mitochondrial potential (via MTDR staining) in uninfected and chronically infected HIV adaptive and innate immune cells (* p<0.05, ** p<0.01, *** p<0.001, **** p<0.0001).</p