High-throughput drug screen identifies calcium and calmodulin inhibitors that reduce JCPyV infection.

JC polyomavirus (JCPyV) is a nonenveloped, double-stranded DNA virus that infects the majority of the population. Immunocompetent individuals harbor infection in their kidneys, while severe immunosuppression can result in JCPyV spread to the brain, causing the neurodegenerative disease progressive multifocal leukoencephalopathy (PML). Due to a lack of approved therapies to treat JCPyV and PML, the disease results in rapid deterioration, and is often fatal. In order to identify potential antiviral treatments for JCPyV, a high-throughput, large-scale drug screen was performed using the National Institutes of Health Clinical Collection (NCC). Drugs from the NCC were tested for inhibitory effects on JCPyV infection, and drugs from various classes that reduced JCPyV infection were identified, including receptor agonists and antagonists, calcium signaling modulators, and enzyme inhibitors. Given the role of calcium signaling in viral infection including Merkel cell polyomavirus and simian virus 40 polyomavirus (SV40), calcium signaling inhibitors were further explored for the capacity to impact JCPyV infection. Calcium and calmodulin inhibitors trifluoperazine (TFP), W-7, tetrandrine, and nifedipine reduced JCPyV infection, and TFP specifically reduced viral internalization. Additionally, TFP and W-7 reduced infection by BK polyomavirus, SV40, and SARS-CoV-2. These results highlight specific inhibitors, some FDA-approved, for the possible treatment and prevention of JCPyV and several other viruses, and further illuminate the calcium and calmodulin pathway as a potential target for antiviral drug development.

Quantitative Prediction of Sling Events in Turbulence at High Reynolds Numbers.

Collisional growth of droplets, such as occurring in warm clouds, is known to be significantly enhanced by turbulence. Whether particles collide depends on their flow history, in particular on their encounters with highly intermittent small-scale turbulent structures, which despite their rarity can dominate the overall collision rate. Here, we develop a quantitative criterion for sling events based on the velocity gradient history along particle paths. We show by a combination of theory and simulations that the problem reduces to a one-dimensional localization problem as encountered in condensed matter physics. The reduction demonstrates that the creation of slings is controlled by the minimal real eigenvalue of the velocity gradient tensor. We use fully resolved turbulence simulations to confirm our predictions and study their Stokes and Reynolds number dependence. We also discuss extrapolations to the parameter range relevant for typical cloud droplets, showing that sling events at high Reynolds numbers are enhanced by an order of magnitude for small Stokes numbers. Thus, intermittency could be a significant ingredient in the collisional growth of rain droplets.

Frailty Among Sexual and Gender Minority Older Adults: The All of Us Database.

BACKGROUND: Despite known disparities in health status among older sexual and gender minority adults (OSGM), the prevalence of frailty is unknown. The aim of this study was to develop and validate a deficit-accumulation frailty index (AoU-FI) for the All of Us database to describe and compare frailty between OSGM and non-OSGM participants. METHODS: Developed using a standardized approach, the AoU-FI consists of 33 deficits from baseline survey responses of adults aged 50+. OSGM were self-reported as "not straight" or as having discordant gender and sex assigned at birth. Descriptive statistics characterized the AoU-FI. Regression was used to assess the association between frailty, age, and gender. Validation of the AoU-FI used Cox proportional hazard models to test the association between frailty categories (robust <0.15, 0.15 ≤ pre-frail ≤ 0.25, frail >0.25) and mortality. RESULTS: There were 9 110 OSGM and 67 420 non-OSGM with sufficient data to calculate AoU-FI; 41% OSGM versus 50% non-OSGM were robust, whereas 34% versus 32% were pre-frail, and 26% versus 19% were frail. Mean AoU-FI was 0.19 (95% confidence interval [CI]: 0.187, 0.191) for OSGM and 0.168 (95% CI: 0.167, 0.169) for non-OSGM. Compared to robust, odds of mortality were higher among frail OSGM (odds ratio [OR] 6.40; 95% CI: 1.84, 22.23) and non-OSGM (OR 3.96; 95% CI: 2.96, 5.29). CONCLUSIONS: The AoU-FI identified a higher burden of frailty, increased risk of mortality, and an attenuated impact of age on frailty among OSGM compared to non-OSGM. Future work is needed to understand how frailty affects the OSGM population.

Dynamics and Patterning of 5-Hydroxytryptamine 2 Subtype Receptors in JC Polyomavirus Entry.

The organization and dynamics of plasma membrane receptors are a critical link in virus-receptor interactions, which finetune signaling efficiency and determine cellular responses during infection. Characterizing the mechanisms responsible for the active rearrangement and clustering of receptors may aid in developing novel strategies for the therapeutic treatment of viruses. Virus-receptor interactions are poorly understood at the nanoscale, yet they present an attractive target for the design of drugs and for the illumination of viral infection and pathogenesis. This study utilizes super-resolution microscopy and related techniques, which surpass traditional microscopy resolution limitations, to provide both a spatial and temporal assessment of the interactions of human JC polyomavirus (JCPyV) with 5-hydroxytrypamine 2 receptors (5-HT2Rs) subtypes during viral entry. JCPyV causes asymptomatic kidney infection in the majority of the population and can cause fatal brain disease, and progressive multifocal leukoencephalopathy (PML), in immunocompromised individuals. Using Fluorescence Photoactivation Localization Microscopy (FPALM), the colocalization of JCPyV with 5-HT2 receptor subtypes (5-HT2A, 5-HT2B, and 5-HT2C) during viral attachment and viral entry was analyzed. JCPyV was found to significantly enhance the clustering of 5-HT2 receptors during entry. Cluster analysis of infected cells reveals changes in 5-HT2 receptor cluster attributes, and radial distribution function (RDF) analyses suggest a significant increase in the aggregation of JCPyV particles colocalized with 5-HT2 receptor clusters in JCPyV-infected samples. These findings provide novel insights into receptor patterning during viral entry and highlight improved technologies for the future development of therapies for JCPyV infection as well as therapies for diseases involving 5-HT2 receptors.

Elongation enhances encounter rates between phytoplankton in turbulence.

Phytoplankton come in a stunning variety of shapes but elongated morphologies dominate-typically 50% of species have aspect ratio above 5, and bloom-forming species often form chains whose aspect ratios can exceed 100. How elongation affects encounter rates between phytoplankton in turbulence has remained unknown, yet encounters control the formation of marine snow in the ocean. Here, we present simulations of encounters among elongated phytoplankton in turbulence, showing that encounter rates between neutrally buoyant elongated cells are up to 10-fold higher than for spherical cells and even higher when cells sink. Consequently, we predict that elongation can significantly speed up the formation of marine snow compared to spherical cells. This unexpectedly large effect of morphology in driving encounter rates among plankton provides a potential mechanistic explanation for the rapid clearance of many phytoplankton blooms.

Rearrangement in the Hypervariable Region of JC Polyomavirus Genomes Isolated from Patient Samples and Impact on Transcription Factor-Binding Sites and Disease Outcomes.

JC polyomavirus (JCPyV) is the causative agent of the fatal, incurable, neurological disease, progressive multifocal leukoencephalopathy (PML). The virus is present in most of the adult population as a persistent, asymptotic infection in the kidneys. During immunosuppression, JCPyV reactivates and invades the central nervous system. A main predictor of disease outcome is determined by mutations within the hypervariable region of the viral genome. In patients with PML, JCPyV undergoes genetic rearrangements in the noncoding control region (NCCR). The outcome of these rearrangements influences transcription factor binding to the NCCR, orchestrating viral gene transcription. This study examines 989 NCCR sequences from patient isolates deposited in GenBank to determine the frequency of mutations based on patient isolation site and disease status. The transcription factor binding sites (TFBS) were also analyzed to understand how these rearrangements could influence viral transcription. It was determined that the number of TFBS was significantly higher in PML samples compared to non-PML samples. Additionally, TFBS that could promote JCPyV infection were more prevalent in samples isolated from the cerebrospinal fluid compared to other locations. Collectively, this research describes the extent of mutations in the NCCR that alter TFBS and how they correlate with disease outcome.

The statistical geometry of material loops in turbulence.

Material elements - which are lines, surfaces, or volumes behaving as passive, non-diffusive markers - provide an inherently geometric window into the intricate dynamics of chaotic flows. Their stretching and folding dynamics has immediate implications for mixing in the oceans or the atmosphere, as well as the emergence of self-sustained dynamos in astrophysical settings. Here, we uncover robust statistical properties of an ensemble of material loops in a turbulent environment. Our approach combines high-resolution direct numerical simulations of Navier-Stokes turbulence, stochastic models, and dynamical systems techniques to reveal predictable, universal features of these complex objects. We show that the loop curvature statistics become stationary through a dynamical formation process of high-curvature folds, leading to distributions with power-law tails whose exponents are determined by the large-deviations statistics of finite-time Lyapunov exponents of the flow. This prediction applies to advected material lines in a broad range of chaotic flows. To complement this dynamical picture, we confirm our theory in the analytically tractable Kraichnan model with an exact Fokker-Planck approach.

Emergence of behaviour in a self-organized living matter network.

What is the origin of behaviour? Although typically associated with a nervous system, simple organisms also show complex behaviours. Among them, the slime mold Physarum polycephalum, a giant single cell, is ideally suited to study emergence of behaviour. Here, we show how locomotion and morphological adaptation behaviour emerge from self-organized patterns of rhythmic contractions of the actomyosin lining of the tubes making up the network-shaped organism. We quantify the spatio-temporal contraction dynamics by decomposing experimentally recorded contraction patterns into spatial contraction modes. Notably, we find a continuous spectrum of modes, as opposed to a few dominant modes. Our data suggests that the continuous spectrum of modes allows for dynamic transitions between a plethora of specific behaviours with transitions marked by highly irregular contraction states. By mapping specific behaviours to states of active contractions, we provide the basis to understand behaviour's complexity as a function of biomechanical dynamics.

Self-generated oxygen gradients control collective aggregation of photosynthetic microbes.

For billions of years, photosynthetic microbes have evolved under the variable exposure to sunlight in diverse ecosystems and microhabitats all over our planet. Their abilities to dynamically respond to alterations of the luminous intensity, including phototaxis, surface association and diurnal cell cycles, are pivotal for their survival. If these strategies fail in the absence of light, the microbes can still sustain essential metabolic functionalities and motility by switching their energy production from photosynthesis to oxygen respiration. For suspensions of motile C. reinhardtii cells above a critical density, we demonstrate that this switch reversibly controls collective microbial aggregation. Aerobic respiration dominates over photosynthesis in conditions of low light, which causes the microbial motility to sensitively depend on the local availability of oxygen. For dense microbial populations in self-generated oxygen gradients, microfluidic experiments and continuum theory based on a reaction-diffusion mechanism show that oxygen-regulated motility enables the collective emergence of highly localized regions of high and low cell densities.

PI3K/AKT/mTOR Signaling Pathway Is Required for JCPyV Infection in Primary Astrocytes.

Astrocytes are a main target of JC polyomavirus (JCPyV) in the central nervous system (CNS), where the destruction of these cells, along with oligodendrocytes, leads to the fatal disease progressive multifocal leukoencephalopathy (PML). There is no cure currently available for PML, so it is essential to discover antivirals for this aggressive disease. Additionally, the lack of a tractable in vivo models for studying JCPyV infection makes primary cells an accurate alternative for elucidating mechanisms of viral infection in the CNS. This research to better understand the signaling pathways activated in response to JCPyV infection reveals and establishes the importance of the PI3K/AKT/mTOR signaling pathway in JCPyV infection in primary human astrocytes compared to transformed cell lines. Using RNA sequencing and chemical inhibitors to target PI3K, AKT, and mTOR, we have demonstrated the importance of this signaling pathway in JCPyV infection of primary astrocytes not observed in transformed cells. Collectively, these findings illuminate the potential for repurposing drugs that are involved with inhibition of the PI3K/AKT/mTOR signaling pathway and cancer treatment as potential therapeutics for PML, caused by this neuroinvasive virus.

Metacarpal bone diameter increases constantly in adult females, but escalated resorption of the inner surface at menopause explains the observed decreased bone mineral density at digital X-ray radiogrammetry.

OBJECTIVE: To assess how bone morphology and mineralization changes with age, in women by digital X-ray radiogrammetry (DXR). DXR has potential as a screening tool for osteoporosis, as it can evaluate bone mineralization similarly to dual-energy X-ray absorptiometry. METHODS: The nondominant hand was analyzed with DXR in 13,285 women ages 40-74 years undergoing mammography. 1,556 women attended two consecutive examinations with 18 to 24 months interval. Changes in bone parameters were calculated and compared in 5-year age groups. Regression analysis and ANOVA tests were performed. RESULTS: Univariate linear regression showed no significant difference in age or bone size between the groups with single or consecutive measurements. In the group with consecutive measurements, the average inner diameter (DXR-ID) of the metacarpals significantly increased with age from 0.38 cm in the 40-45 years age span to 0.47 cm in the 65+ age group (P < 0.001), whereas DXR bone mineral density (DXR-BMD) decreased from 0.59 g/cm2 to 0.50 g/cm2 in the same age groups (P < 0.001). Intraindividual measurements showed a fourfold increase in yearly DXR-ID increase and concurrent DXR-BMD loss between 50 and 59 years of age (P < 0.001). The outer diameter only increased significantly between the youngest and oldest age group (P < 0.001). CONCLUSIONS: The faster decrease in DXR-BMD observed during and after menopause is caused by resorption of the inner cortical surface, not matched by outer diameter increase. We speculate that most bones in the human body grow in the same pattern as observed in the metacarpals, partly explaining decreasing BMD at DXR and dual-energy X-ray absorptiometry.

Video Summary:http://links.lww.com/MENO/A832 .

The MAPK/ERK Pathway and the Role of DUSP1 in JCPyV Infection of Primary Astrocytes.

JC polyomavirus (JCPyV) is a neuroinvasive pathogen causing a fatal, demyelinating disease of the central nervous system (CNS) known as progressive multifocal leukoencephalopathy (PML). Within the CNS, JCPyV predominately targets two cell types: oligodendrocytes and astrocytes. The underlying mechanisms of astrocytic infection are poorly understood, yet recent findings suggest critical differences in JCPyV infection of primary astrocytes compared to a widely studied immortalized cell model. RNA sequencing was performed in primary normal human astrocytes (NHAs) to analyze the transcriptomic profile that emerges during JCPyV infection. Through a comparative analysis, it was validated that JCPyV requires the mitogen-activated protein kinase, extracellular signal-regulated kinase (MAPK/ERK) pathway, and additionally requires the expression of dual-specificity phosphatases (DUSPs). Specifically, the expression of DUSP1 is needed to establish a successful infection in NHAs, yet this was not observed in an immortalized cell model of JCPyV infection. Additional analyses demonstrated immune activation uniquely observed in NHAs. These results support the hypothesis that DUSPs within the MAPK/ERK pathway impact viral infection and influence potential downstream targets and cellular pathways. Collectively, this research implicates DUSP1 in JCPyV infection of primary human astrocytes, and most importantly, further resolves the signaling events that lead to successful JCPyV infection in the CNS.

Emergence and melting of active vortex crystals.

Melting of two-dimensional (2D) equilibrium crystals is a complex phenomenon characterized by the sequential loss of positional and orientational order. In contrast to passive systems, active crystals can self-assemble and melt into an active fluid by virtue of their intrinsic motility and inherent non-equilibrium stresses. Currently, the non-equilibrium physics of active crystallization and melting processes is not well understood. Here, we establish the emergence and investigate the melting of self-organized vortex crystals in 2D active fluids using a generalized Toner-Tu theory. Performing extensive hydrodynamic simulations, we find rich transition scenarios. On small domains, we identify a hysteretic transition as well as a transition featuring temporal coexistence of active vortex lattices and active turbulence, both of which can be controlled by self-propulsion and active stresses. On large domains, an active vortex crystal with solid order forms within the parameter range corresponding to active vortex lattices. The melting of this crystal proceeds through an intermediate hexatic phase. Generally, these results highlight the differences and similarities between crystalline phases in active fluids and their equilibrium counterparts.

Sending mixed signals: polyomavirus entry and trafficking.

Polyomaviruses are mostly non-pathogenic, yet some can cause human disease especially under conditions of immunosuppression, including JC, BK, and Merkel cell polyomaviruses. Direct interactions between viruses and the host early during infection dictate the outcome of disease, many of which remain enigmatic. However, significant work in recent years has contributed to our understanding of how this virus family establishes an infection, largely due to advances made for animal polyomaviruses murine and SV40. Here we summarize the major findings that have contributed to our understanding of polyomavirus entry, trafficking, disassembly, signaling, and immune evasion during the infectious process and highlight major unknowns in these processes that are open areas of study.

The challenges of containing SARS-CoV-2 via test-trace-and-isolate.

Without a cure, vaccine, or proven long-term immunity against SARS-CoV-2, test-trace-and-isolate (TTI) strategies present a promising tool to contain its spread. For any TTI strategy, however, mitigation is challenged by pre- and asymptomatic transmission, TTI-avoiders, and undetected spreaders, which strongly contribute to "hidden" infection chains. Here, we study a semi-analytical model and identify two tipping points between controlled and uncontrolled spread: (1) the behavior-driven reproduction number [Formula: see text] of the hidden chains becomes too large to be compensated by the TTI capabilities, and (2) the number of new infections exceeds the tracing capacity. Both trigger a self-accelerating spread. We investigate how these tipping points depend on challenges like limited cooperation, missing contacts, and imperfect isolation. Our results suggest that TTI alone is insufficient to contain an otherwise unhindered spread of SARS-CoV-2, implying that complementary measures like social distancing and improved hygiene remain necessary.

GRK2 mediates ß-arrestin interactions with 5-HT2 receptors for JC polyomavirus endocytosis.

JC polyomavirus (JCPyV) infects the majority of the population, establishing a lifelong, asymptomatic infection in the kidney of healthy individuals. People that become severely immunocompromised may experience JCPyV reactivation, which can cause progressive multifocal leukoencephalopathy (PML), a neurodegenerative disease. Due to a lack of therapeutic options, PML results in fatality or significant debilitation among affected individuals. Cellular internalization of JCPyV is mediated by serotonin 5-hydroxytryptamine subfamily 2 receptors (5-HT2Rs) via clathrin-mediated endocytosis. The JCPyV entry process requires the clathrin-scaffolding proteins ß-arrestin, adaptor protein 2 (AP2), and dynamin. Further, a ß-arrestin interacting domain, the Ala-Ser-Lys (ASK) motif, within the C-terminus of 5-HT2AR is important for JCPyV internalization and infection. Interestingly, 5-HT2R subtypes A, B, and C equally support JCPyV entry and infection, and all subtypes contain an ASK motif, suggesting a conserved mechanism for viral entry. However, the role of the 5-HT2R ASK motifs and the activation of ß-arrestin-associated proteins during internalization has not been fully elucidated. Through mutagenesis, the ASK motifs within 5-HT2BR and 5-HT2CR were identified as critical for JCPyV internalization and infectivity. Further, utilizing biochemical pulldown techniques, mutagenesis of the ASK motifs in 5-HT2BR and 5-HT2CR resulted in reduced ß-arrestin binding. Utilizing small-molecule chemical inhibitors and RNA interference, G-protein receptor kinase 2 (GRK2) was determined to be required for JCPyV internalization and infection by mediating interactions between ß-arrestin and the ASK motif of 5-HT2Rs. These findings demonstrate that GRK2 and ß-arrestin interactions with 5-HT2Rs are critical for JCPyV entry by clathrin-mediated endocytosis and resultant infection.IMPORTANCE As intracellular parasites, viruses require a host cell to replicate and cause disease. Therefore, virus-host interactions contribute to viral pathogenesis. JC polyomavirus (JCPyV) infects most of the population, establishing a lifelong asymptomatic infection within the kidney. Under conditions of severe immunosuppression JCPyV may spread to the central nervous system, causing the fatal demyelinating disease progressive multifocal leukoencephalopathy (PML). Individuals living with HIV or undergoing immunomodulatory therapies are at risk for developing PML. The mechanisms of how JCPyV uses specific receptors on the surface of host cells to initiate internalization and infection is a poorly understood process. We have further identified cellular proteins involved in JCPyV internalization and infection and elucidated their specific interactions that are responsible for activation of receptors. Collectively, these findings illuminate how viruses usurp cellular receptors during infection, contributing to current development efforts for therapeutic options for the treatment or prevention of PML.

Role of Advective Inertia in Active Nematic Turbulence.

Suspensions of active agents with nematic interactions exhibit complex spatiotemporal dynamics such as mesoscale turbulence. Since the Reynolds number of microscopic flows is very small on the scale of individual agents, inertial effects are typically excluded in continuum theories of active nematic turbulence. Whether active stresses can collectively excite inertial flows is currently unclear. To address this question, we investigate a two-dimensional continuum theory for active nematic turbulence. In particular, we compare mesoscale turbulence with and without the effects of advective inertia. We find that inertial effects can influence the flow already close to the onset of the turbulent state and, moreover, give rise to large-scale fluid motion for strong active driving. A detailed analysis of the kinetic energy budget reveals an energy transfer to large scales mediated by inertial advection. While this transfer is small in comparison to energy injection and dissipation, its effects accumulate over time. The inclusion of friction, which is typically present in experiments, can compensate for this effect. The findings suggest that the inclusion of inertia and friction may be necessary for dynamically consistent theories of active nematic turbulence.

Capturing Velocity Gradients and Particle Rotation Rates in Turbulence.

Turbulent fluid flows exhibit a complex small-scale structure with frequently occurring extreme velocity gradients. Particles probing such swirling and straining regions respond with an intricate shape-dependent orientational dynamics, which sensitively depends on the particle history. Here, we systematically develop a reduced-order model for the small-scale dynamics of turbulence, which captures the velocity gradient statistics along particle paths. An analysis of the resulting stochastic dynamical system allows pinpointing the emergence of non-Gaussian statistics and nontrivial temporal correlations of vorticity and strain, as previously reported from experiments and simulations. Based on these insights, we use our model to predict the orientational statistics of anisotropic particles in turbulence, enabling a host of modeling applications for complex particulate flows.

Inferring change points in the spread of COVID-19 reveals the effectiveness of interventions.

As coronavirus disease 2019 (COVID-19) is rapidly spreading across the globe, short-term modeling forecasts provide time-critical information for decisions on containment and mitigation strategies. A major challenge for short-term forecasts is the assessment of key epidemiological parameters and how they change when first interventions show an effect. By combining an established epidemiological model with Bayesian inference, we analyzed the time dependence of the effective growth rate of new infections. Focusing on COVID-19 spread in Germany, we detected change points in the effective growth rate that correlate well with the times of publicly announced interventions. Thereby, we could quantify the effect of interventions and incorporate the corresponding change points into forecasts of future scenarios and case numbers. Our code is freely available and can be readily adapted to any country or region.