Ongoing Replication Research Articles

Reported impact of creativity in the Wakakosha ('You're Worth It') internal stigma intervention for young people living with HIV in Harare, Zimbabwe.

Internal stigma (negative judgements towards oneself) continues to be a barrier to HIV treatment, management and care, and has global public health consequences. People living with HIV (PLHIV) who report internal stigma are less likely to seek care, adhere to treatment and can experience increased depression and lower quality of life. The Wakakosha ('You're Worth It') programme sought to reduce internal stigma among young PLHIV using inquiry-based stress reduction (IBSR), a cognitive and awareness-based methodology. This sub-study focused on creativity, exploring how it promotes young PLHIV's well-being. We analysed individual interviews (n = 14), focus groups (n = 3), poems (n = 5), songs (n = 2) and self-compassion letters (n = 38) and 23 activity journals from November 2021 to March 2022, utilising inductive thematic analysis to identify themes across the data. Creativity saturated the Wakakosha intervention through modalities such as drawing, colouring, body mapping, music and letter writing. These engaged participants and gave them a space in which to shift their internal stigmatising beliefs, and helped them in multiple ways including: facilitating emotional regulation, self-acceptance, self-compassion, self-worth and body positivity. Creativity also enhanced participants' self-image and allowed them to see their own abilities. Music improved the therapeutic environment by helping participants focus, emote, retain messages and connect within the group setting. Letters to self/others allowed participants to let go of stigma towards themselves and the circumstances of their infection. Tools such as drawing and body mapping assisted participants with emotional recognition and expression. This sub-study provided insight into the potential of creativity, when integrated into an IBSR intervention, in counteracting internal stigma. Ongoing replication is needed to continue to evolve best practices for internal stigma interventions. Future work should use more structured and specific interviews with participants regarding their creative processes.

HIV replication and tuberculosis risk among people living with HIV in Europe: A multicohort analysis, 1983-2015.

HIV replication leads to a change in lymphocyte phenotypes that impairs immune protection against opportunistic infections. We examined current HIV replication as an independent risk factor for tuberculosis (TB). We included people living with HIV from 25 European cohorts 1983-2015. Individuals <16 years or with previous TB were excluded. Person-time was calculated from enrolment (baseline) to the date of TB diagnosis or last follow-up information. We used adjusted Poisson regression and general additive regression models. We included 272,548 people with a median follow-up of 5.9 years (interquartile range [IQR] 2.3-10.9). At baseline, the median CD4 cell count was 355 cells/μL (IQR 193-540) and the median HIV-RNA level 22,000 copies/mL (IQR 1,300-103,000). During 1,923,441 person-years of follow-up, 5,956 (2.2%) people developed TB. Overall, TB incidence was 3.1 per 1,000 person-years (95% confidence interval [CI] 3.02-3.18) and was four times higher in patients with HIV-RNA levels of 10,000 compared with levels <400 copies/mL in any CD4 stratum. CD4 and HIV-RNA time-updated analyses showed that the association between HIV-RNA and TB incidence was independent of CD4. The TB incidence rate ratio for people born in TB-endemic countries compared with those born in Europe was 1.8 (95% CI 1.5-2.2). Our results indicate that ongoing HIV replication (suboptimal HIV control) is an important risk factor for TB, independent of CD4 count. Those at highest risk of TB are people from TB-endemic countries. Close monitoring and TB preventive therapy for people with suboptimal HIV control is important.

PCNA-binding activity separates RNF168 functions in DNA replication and DNA double-stranded break signaling.

RNF168 orchestrates a ubiquitin-dependent DNA damage response to regulate the recruitment of repair factors, such as 53BP1 to DNA double-strand breaks (DSBs). In addition to its canonical functions in DSB signaling, RNF168 may facilitate DNA replication fork progression. However, the precise role of RNF168 in DNA replication remains unclear. Here, we demonstrate that RNF168 is recruited to DNA replication factories in a manner that is independent of the canonical DSB response pathway regulated by Ataxia-Telangiectasia Mutated (ATM) and RNF8. We identify a degenerate Proliferating Cell Nuclear Antigen (PCNA)-interacting peptide (DPIP) motif in the C-terminus of RNF168, which together with its Motif Interacting with Ubiquitin (MIU) domain mediates binding to mono-ubiquitylatedPCNA at replication factories. An RNF168 mutant harboring inactivating substitutions in its DPIP box and MIU1 domain (termed RNF168 ΔDPIP/ΔMIU1) is not recruited to sites of DNA synthesis and fails to support ongoing DNA replication. Notably, the PCNA interaction-deficient RNF168 ΔDPIP/ΔMIU1 mutant fully rescues the ability of RNF168-/- cells to form 53BP1 foci in response to DNA DSBs. Therefore, RNF168 functions in DNA replication and DSB signaling are fully separable. Our results define a new mechanism by which RNF168 promotes DNA replication independently of its canonical functions in DSB signaling.

Molecular and microbiological methods for the identification of nonreplicating Mycobacterium tuberculosis.

Chronic tuberculosis (TB) disease, which requires months-long chemotherapy with multiple antibiotics, is defined by diverse pathological manifestations and bacterial phenotypes. Targeting drug-tolerant bacteria in the host is critical to achieving a faster and durable cure for TB. In order to facilitate this field of research, we need to consider the physiology of persistent MTB during infection, which is often associated with the nonreplicating (NR) state. However, the traditional approach to quantifying bacterial burden through colony enumeration alone only informs on the abundance of live bacilli at the time of sampling, and provides an incomplete picture of the replicative state of the pathogen and the extent to which bacterial replication is balanced by ongoing cell death. Modern approaches to profiling bacterial replication status provide a better understanding of inter- and intra-population dynamics under different culture conditions and in distinct host microenvironments. While some methods use molecular markers of DNA replication and cell division, other approaches take advantage of advances in the field of microfluidics and live-cell microscopy. Considerable effort has been made over the past few decades to develop preclinical in vivo models of TB infection and some are recognized for more closely recapitulating clinical disease pathology than others. Unique lesion compartments presenting different environmental conditions produce significant heterogeneity between Mycobacterium tuberculosis populations within the host. While cellular lesion compartments appear to be more permissive of ongoing bacterial replication, caseous foci are associated with the maintenance of M. tuberculosis in a state of static equilibrium. The accurate identification of nonreplicators and where they hide within the host have significant implications for the way novel chemotherapeutic agents and regimens are designed for persistent infections.

USP50 suppresses alternative RecQ helicase use and deleterious DNA2 activity during replication

Mammalian DNA replication relies on various DNA helicase and nuclease activities to ensure accurate genetic duplication, but how different helicase and nuclease activities are properly directed remains unclear. Here, we identify the ubiquitin-specific protease, USP50, as a chromatin-associated protein required to promote ongoing replication, fork restart, telomere maintenance, cellular survival following hydroxyurea or pyridostatin treatment, and suppression of DNA breaks near GC-rich sequences. We find that USP50 supports proper WRN-FEN1 localisation at or near stalled replication forks. Nascent DNA in cells lacking USP50 shows increased association of the DNA2 nuclease and RECQL4 and RECQL5 helicases and replication defects in cells lacking USP50, or FEN1 are driven by these proteins. Consequently, suppression of DNA2 or RECQL4/5 improves USP50-depleted cell resistance to agents inducing replicative stress and restores telomere stability. These data define an unexpected regulatory protein that promotes the balance of helicase and nuclease use at ongoing and stalled replication forks.

Liquid-liquid phase inclusion bodies in acute and persistent parainfluenaza virus type 5 infections.

Cytoplasmic inclusion bodies (IBs) are a common feature of single-stranded, non-segmented, negative-strand RNA virus (Mononegavirales) infections and are thought to be regions of active virus transcription and replication. Here we followed the dynamics of IB formation and maintenance in cells infected with persistent and lytic/acute variants of the paramyxovirus, parainfluenza virus type 5 (PIV5). We show that there is a rapid increase in the number of small inclusions bodies up until approximately 12 h post-infection. Thereafter the number of inclusion bodies decreases but they increase in size, presumably due to the fusion of these liquid organelles that can be disrupted by osmotically shocking cells. No obvious differences were observed at these times between inclusion body formation in cells infected with lytic/acute and persistent viruses. IBs are also readily detected in cells persistently infected with PIV5, including in cells in which there is little or no ongoing virus transcription or replication. In situ hybridization shows that genomic RNA is primarily located in IBs, whilst viral mRNA is more diffusely distributed throughout the cytoplasm. Some, but not all, IBs show incorporation of 5-ethynyl-uridine (5EU), which is integrated into newly synthesized RNA, at early times post-infection. These results strongly suggest that, although genomic RNA is present in all IBs, IBs are not continuously active sites of virus transcription and replication. Disruption of IBs by osmotically shocking persistently infected cells does not increase virus protein synthesis, suggesting that in persistently infected cells most of the virus genomes are in a repressed state. The role of IBs in PIV5 replication and the establishment and maintenance of persistence is discussed.

No evidence for ongoing replication on ART in SIV-infected macaques

The capacity of HIV-1 to replicate during optimal antiretroviral therapy (ART) is challenging to assess directly. To gain greater sensitivity to detect evolution on ART, we used a nonhuman primate (NHP) model providing precise control over the level of pre-ART evolution and more comprehensive analyses than are possible with clinical samples. We infected 21 rhesus macaques (RMs) with the barcoded virus SIVmac239M and initiated ART early to minimize baseline genetic diversity. RMs were treated for 285–1200 days. We used several tests of molecular evolution to compare 1352 near-full-length (nFL) SIV DNA single genome sequences from PBMCs, lymph nodes, and spleen obtained near the time of ART initiation and those present after long-term ART, none of which showed significant changes to the SIV DNA population during ART in any animal. To investigate the possibility of ongoing replication in unsampled putative tissue sanctuaries during ART, we discontinued treatment in four animals and confirmed that none of the 336 nFL SIV RNA sequences obtained from rebound plasma viremia showed evidence of evolution. The rigorous nature of our analyses reinforced the emerging consensus of a lack of appreciable ongoing replication on effective ART and validates the relevance of this NHP model for cure studies.

The TIMELESS and PARP1 interaction suppresses replication-associated DNA gap accumulation.

TIMELESS (TIM) in the fork protection complex acts as a scaffold of the replisome to prevent its uncoupling and ensure efficient DNA replication fork progression. Nevertheless, its underlying basis for coordinating leading and lagging strand synthesis to limit single-stranded DNA (ssDNA) exposure remains elusive. Here, we demonstrate that acute degradation of TIM at ongoing DNA replication forks induces the accumulation of ssDNA gaps stemming from defective Okazaki fragment (OF) processing. Cells devoid of TIM fail to support the poly(ADP-ribosyl)ation necessary for backing up the canonical OF processing mechanism mediated by LIG1 and FEN1. Consequently, recruitment of XRCC1, a known effector of PARP1-dependent single-strand break repair, to post-replicative ssDNA gaps behind replication forks is impaired. Physical disruption of the TIM-PARP1 complex phenocopies the rapid loss of TIM, indicating that the TIM-PARP1 interaction is critical for the activation of this compensatory pathway. Accordingly, combined deficiency of FEN1 and the TIM-PARP1 interaction leads to synergistic DNA damage and cytotoxicity. We propose that TIM is essential for the engagement of PARP1 to the replisome to coordinate lagging strand synthesis with replication fork progression. Our study identifies TIM as a synthetic lethal target of OF processing enzymes that can be exploited for cancer therapy.

Adaptive use of error-prone DNA polymerases provides flexibility in genome replication during tumorigenesis.

Human cells possess many different polymerase enzymes, which collaborate in conducting DNA replication and genome maintenance to ensure faithful duplication of genetic material. Each polymerase performs a specialized role, together providing a balance of accuracy and flexibility to the replication process. Perturbed replication increases the requirement for flexibility to ensure duplication of the entire genome. Flexibility is provided via the use of error-prone polymerases, which maintain the progression of challenged DNA replication at the expense of mutagenesis, an enabling characteristic of cancer. This review describes our recent understanding of mechanisms that alter the usage of polymerases during tumorigenesis and examines the implications of this for cell survival and tumor progression. Although expression levels of polymerases are often misregulated in cancers, this does not necessarily alter polymerase usage since an additional regulatory step may govern the use of these enzymes. We therefore also examine how the regulatory mechanisms of DNA polymerases, such as Rad18-mediated PCNA ubiquitylation, may impact the functionalization of error-prone polymerases to tolerate oncogene-induced replication stress. Crucially, it is becoming increasingly evident that cancer cells utilize error-prone polymerases to sustain ongoing replication in response to oncogenic mutations which inactivate key DNA replication and repair pathways, such as BRCA deficiency. This accelerates mutagenesis and confers chemoresistance, but also presents a dependency that can potentially be exploited by therapeutics.

Nonnegligible Contribution of Nonlymphoid Tissue to Viral Reservoir During the Short-Term Early cART in SIVmac239-Infected Chinese Rhesus Macaques.

HIV/AIDS cannot be cured because of the persistence of the viral reservoir. Because of the complexity of the cellular composition and structure of the human organs, HIV reservoirs of anatomical site are also complex. Recently, although a variety of molecules have been reported to be involved in the establishment and maintenance of the viral reservoirs, or as marker of latent cells, the research mainly focuses on blood and lymph nodes. Now, the characteristics of the viral reservoir in tissue are not yet fully understood. In this study, various tissues were collected from SIVmac239-infected monkeys, and the level of total SIV DNA, SIV 2-LTR DNA, and cell-associated virus RNA in them were compared with character of the anatomical viral reservoir under early treatment. The results showed that short-term combination antiretroviral therapy (cART) starting from 3 days after infection could significantly inhibit viremia and reduce the size of the anatomical viral reservoir, but it could not eradicate de novo infections and ongoing replication of virus. Moreover, the effects of early cART on the level of total SIV DNA, SIV 2-LTR DNA, and cell-associated virus RNA in different tissues were different, which changed the size distribution of viral reservoir in anatomical site. Finally, the contribution of nonlymphoid tissues, especially liver and lung, to the viral reservoir increased after treatment, while the contribution of intestinal lymphoid to the viral reservoir significantly reduced. These results suggested that early treatment effectively decreased the size of viral reservoir, and that the effects of cART on the tissue viral reservoir varied greatly by tissue type. The results implied that persistent existence of virus in nonlymphoid tissues after short-term treatment suggested that the role of nonlymphoid tissues cannot be ignored in development strategies for AIDS therapy.

Timeless-Tipin interactions with MCM and RPA mediate DNA replication stress response.

The accuracy of replication is one of the most important mechanisms ensuring the stability of the genome. The fork protection complex prevents premature replisome stalling and/or premature disassembly upon stress. Here, we characterize the Timeless-Tipin complex, a component of the fork protection complex. We used microscopy approaches, including colocalization analysis and proximity ligation assay, to investigate the spatial localization of the complex during ongoing replication in human cells. Taking advantage of the replication stress induction and the ensuing polymerase-helicase uncoupling, we characterized the Timeless-Tipin localization within the replisome. Replication stress was induced using hydroxyurea (HU) and aphidicolin (APH). While HU depletes the substrate for DNA synthesis, APH binds directly inside the catalytic pocket of DNA polymerase and inhibits its activity. Our data revealed that the Timeless-Tipin complex, independent of the stress, remains bound on chromatin upon stress induction and progresses together with the replicative helicase. This is accompanied by the spatial dissociation of the complex from the blocked replication machinery. Additionally, after stress induction, Timeless interaction with RPA, which continuously accumulates on ssDNA, was increased. Taken together, the Timeless-Tipin complex acts as a universal guardian of the mammalian replisome in an unperturbed S-phase progression as well as during replication stress.

Treating Target Behaviors of Autistic Individuals With Applied Behavior Analysis: An Ongoing Replication Study.

Background Ongoing outcome data replication on target behaviors with autistic individuals using applied behavior analysis (ABA) confirms its effectiveness and remains an essential evidenced-based standard of care. This replication study aims tofurther confirm the impact of discrete trial training and mass trials on general target behaviors within a naturalistic environment. Methods Data was gathered from 92 children and four adult autistic individuals over one month from 7/7/23to 8/8/23using a repeated measures design. This study used a retrospective chart review with general target behaviors to determine the effectiveness of ABA treatments using discrete trial training and mass trials across time and age categories in a naturalistic environment. Results A mixed analysis of variance (ANOVA) indicated statistical significance (sphericity assumed), F(2,168) = 31.663, p < 0.001 (time). Multiple comparisons using bootstrapped paired t-tests indicated p < 0.001 on the three comparisons. There was a significant interaction effect (sphericity assumed) with timex age category, F(8,168) = 2.918, p = 0.004. Interaction contrasts indicated statistically significant differences over time within the 1-4 years, 5-8 years, and a portion of 9-12 years, and not within the 13-16 years and 17-73 years age groups. Conclusions Autistic individuals receiving ABA demonstrated statistically significant improvement in target behaviors over one month. There was a significant interaction between timeand ageon target behaviors, suggesting a significant association between time and age categories.The reporting of ongoing intervention outcomes provides further justification for continued treatmentsrelative to target behavior mastery with autistic individuals.

Hepatic macrophage niche: a bridge between HBV-mediated metabolic changes with intrahepatic inflammation.

Hepatitis B Virus (HBV) is a stealthy and insidious pathogen capable of inducing chronic necro-inflammatory liver disease and hepatocellular carcinoma (HCC), resulting in over one million deaths worldwide per year. The traditional understanding of Chronic Hepatitis B (CHB) progression has focused on the complex interplay among ongoing virus replication, aberrant immune responses, and liver pathogenesis. However, the dynamic progression and crucial factors involved in the transition from HBV infection to immune activation and intrahepatic inflammation remain elusive. Recent insights have illuminated HBV's exploitation of the sodium taurocholate co-transporting polypeptide (NTCP) and manipulation of the cholesterol transport system shared between macrophages and hepatocytes for viral entry. These discoveries deepen our understanding of HBV as a virus that hijacks hepatocyte metabolism. Moreover, hepatic niche macrophages exhibit significant phenotypic and functional diversity, zonal characteristics, and play essential roles, either in maintaining liver homeostasis or contributing to the pathogenesis of chronic liver diseases. Therefore, we underscore recent revelations concerning the importance of hepatic niche macrophages in the context of viral hepatitis. This review particularly emphasizes the significant role of HBV-induced metabolic changes in hepatic macrophages as a key factor in the transition from viral infection to immune activation, ultimately culminating in liver inflammation. These metabolic alterations in hepatic macrophages offer promising targets for therapeutic interventions and serve as valuable early warning indicators, shedding light on the disease progression.

Prolonged Mpox Disease in People With Advanced HIV: Characterization of Mpox Skin Lesions.

We report 3 complicated and prolonged cases of mpox in people with advanced human immunodeficiency virus (HIV) not on antiretroviral therapy (ART) at mpox diagnosis. Multiple medical countermeasures were used, including prolonged tecovirimat treatment and immune optimization with ART initiation. Immunofluorescence of skin biopsies demonstrated a dense immune infiltrate of predominantly myeloid and CD8+ T cells, with a strong type I interferon local response. RNAscope detected abundant replication of monkeypox virus (MPXV) in epithelial cells and dendritic cells. These data suggest that prolonged mpox in people with advanced HIV may be due to ongoing MPXV replication, warranting aggressive medical countermeasures and immune optimization.

Growth From Uncertainty: Understanding the Replication ‘Crisis’ in Infant Cognition

AbstractPsychology is a discipline that has a high number of failed replications, which has been characterized as a “crisis” on the assumption that failed replications are indicative of untrustworthy research. This article uses Chang’s concept of epistemic iteration to show how a research program can advance epistemic goals despite many failed replications. It illustrates this by analyzing an ongoing large-scale replication attempt of Southgate et al.’s work exploring infants’ understanding of false beliefs. It concludes that epistemic iteration offers a way of understanding the value of replications—both failed and successful—that contradicts the narrative centered around distrust.

Ongoing HIV replication in lymph node sanctuary sites in treated individuals contributes to the total latent HIV at a very slow rate

Lymph nodes (LNs) serve as a sanctuary site for HIV viruses due to the heterogeneous distribution of the antiretrovirals (ARVs) inside the LNs. There is an ongoing debate whether this represents ongoing cycles of viral replication in the LNs or merely residual virus production by latently infected cells. Previous work has claimed that the measured levels of genetic variation in proviruses sampled from the blood were inconsistent with ongoing replication. However, it is not clear what rate of variation is consistent with ongoing replication in small sanctuary sites.In this study, we used a spherically symmetric compartmental ODE model to track the HIV viral dynamics in the LN and predict the contribution of ongoing replication within the LN to the whole-body proviral pool in an ARV-suppressed person living with HIV. This model tracks the reaction–diffusion dynamics of uninfected, actively infected, and latently infected T-cells as well as free virus within the LN parenchyma and the blood, and distinguishes between latently infected cells created before ARV therapy and during ARV therapy.We simulated suppressive therapy beginning in year 5 post-infection. Each LN sanctuary site had a volume of 1 ml, and we considered cases of 1 ml, 30 ml, and 250 ml total volume, which represent a single active sanctuary site, moderate systemic involvement, and involvement of the total lymphoid tissue. Viral load in the blood rapidly dropped and remained below the limit of detection in all cases but remained high in the LN sanctuary sites. Novel latent cells increased systemically over time but very slowly, taking between 25 and 50 years to reach 5 % of the total latent pool, depending on the volume of lymphoid tissue involvement.Putative sanctuary sites in LNs are limited in volume and produce novel latent cells slowly. Assays to detect genetic drift due to such sites would require very deep sequencing if sampling only from the blood. Previous studies showing a lack of genetic drift are consistent with the expected contribution of ongoing replication in lymph node sanctuary sites.

Power priors for replication studies

The ongoing replication crisis in science has increased interest in the methodology of replication studies. We propose a novel Bayesian analysis approach using power priors: The likelihood of the original study’s data is raised to the power of α\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\\alpha $$\\end{document}, and then used as the prior distribution in the analysis of the replication data. Posterior distribution and Bayes factor hypothesis tests related to the power parameter α\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\\alpha $$\\end{document} quantify the degree of compatibility between the original and replication study. Inferences for other parameters, such as effect sizes, dynamically borrow information from the original study. The degree of borrowing depends on the conflict between the two studies. The practical value of the approach is illustrated on data from three replication studies, and the connection to hierarchical modeling approaches explored. We generalize the known connection between normal power priors and normal hierarchical models for fixed parameters and show that normal power prior inferences with a beta prior on the power parameter α\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\\alpha $$\\end{document} align with normal hierarchical model inferences using a generalized beta prior on the relative heterogeneity variance I2\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$I^2$$\\end{document}. The connection illustrates that power prior modeling is unnatural from the perspective of hierarchical modeling since it corresponds to specifying priors on a relative rather than an absolute heterogeneity scale.

Time of day dependent longitudinal changes in resting-state fMRI.

Longitudinal studies have become more common in the past years due to their superiority over cross-sectional samples. In light of the ongoing replication crisis, the factors that may introduce variability in resting-state networks have been widely debated. This publication aimed to address the potential sources of variability, namely, time of day, sex, and age, in longitudinal studies within individual resting-state fMRI data. DCM was used to analyze the fMRI time series, extracting EC connectivity measures and parameters that define the BOLD signal. In addition, a two-way ANOVA was used to assess the change in EC and parameters that define the BOLD signal between data collection waves. The results indicate that time of day and gender have significant model evidence for the parameters that define the BOLD signal but not EC. From the ANOVA analysis, findings indicate that there was a significant change in the two nodes of the DMN and their connections with the fronto-parietal network. Overall, these findings suggest that in addition to age and gender, which are commonly accounted for in the fMRI data collection, studies should note the time of day, possibly treating it as a covariate in longitudinal samples.

Cytolytic CD8+ T cells infiltrate germinal centers to limit ongoing HIV replication in spontaneous controller lymph nodes.

Follicular CD8+ T cells (fCD8) mediate surveillance in lymph node (LN) germinal centers against lymphotropic infections and cancers, but the precise mechanisms by which these cells mediate immune control remain incompletely resolved. To address this, we investigated functionality, clonotypic compartmentalization, spatial localization, phenotypic characteristics, and transcriptional profiles of LN-resident virus-specific CD8+ T cells in persons who control HIV without medications. Antigen-induced proliferative and cytolytic potential consistently distinguished spontaneous controllers from noncontrollers. T cell receptor analysis revealed complete clonotypic overlap between peripheral and LN-resident HIV-specific CD8+ T cells. Transcriptional analysis of LN CD8+ T cells revealed gene signatures of inflammatory chemotaxis and antigen-induced effector function. In HIV controllers, the cytotoxic effectors perforin and granzyme B were elevated among virus-specific CXCR5+ fCD8s proximate to foci of HIV RNA within germinal centers. These results provide evidence consistent with cytolytic control of lymphotropic infection supported by inflammatory recruitment, antigen-specific proliferation, and cytotoxicity of fCD8s.

Successful Combination of Remdesivir and Convalescent Plasma to Treat a Patient with Rituximab-Related B-Cell Deficiency and Prolonged COVID-19: A Case Report

Background: Treatment of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in immuno-compromised patients with complete B cell depletion can be really challenging due to the lack of seroconversion and long-lasting disease. Case Report: We describe a case of long-lasting coronavirus disease (COVID-19) in a female patient with rheumatoid arthritis who was treated with rituximab and continued to show B-cell depletion. An ongoing replication of SARS-CoV-2 was demonstrated for a period of 8 months when nasopharyngeal swabs were tested. She was treated once with remdesivir but without lasting resolution, and she was then treated with convalescent plasma but with a similar effect. Only with a combination of both treatments was clinical resolution achieved. The patient's lack of seroconversion and the prolonged course of the disease illustrate the importance of humoral immunity in resolving SARS-CoV-2 infection. This case report highlights challenges in managing immunocompromised hosts, who may act as persistent shedders and sources of transmission. Conclusions: The combination of remdesivir and convalescent plasma resulted in successfully achieving clinical resolution of SARS-CoV-2 infection in our patient.