Precision medicine for pandemics: stratification of COVID-19 molecular phenotypes defined by topological analysis of global blood gene expression. (preprint)

Precision medicine offers a promising avenue for better therapeutic responses to pandemics such as COVID-19. This study leverages independent patient cohorts in Florence and Liege gathered under the umbrella of the DRAGON consortium for the stratification of molecular phenotypes associated with COVID-19 using topological analysis of global blood gene expression. Whole blood from 173 patients was collected and RNA was sequenced on the Novaseq platform. Molecular phenotypes were defined through topological analysis of gene expression relative to the biological network using the TopMD algorithm. The two cohorts from Florence and Liege allowed for independent validation of the findings in this study. Clustering of the topological maps of differential pathway activation revealed three distinct molecular phenotypes of COVID-19 in the Florence patient cohort, which were also observed in the Liege cohort. Cluster 1, was characterised by high activation of pathways associated with ESC pluripotency, NRF2, and TGF-B; receptor signalling. Cluster 2 displayed high activation of pathways including focal adhesion-PI3K-Akt-mTOR signalling and type I interferon induction and signalling, while Cluster 3 exhibited low IRF7-related pathway activation. TopMD was also used with the Drug-Gene Interaction Database (DGIdb), revealing pharmaceutical interventions targeting mechanisms across multiple phenotypes and individuals. The data illustrates the utility of molecular phenotyping from topological analysis of blood gene expression, and holds promise for informing personalised therapeutic strategies not only for COVID-19 but also for Disease X. Its potential transferability across multiple diseases highlights the value in pandemic response efforts, offering insights before large-scale clinical studies are initiated.

Distinct Type 1 Immune Networks Underlie the Severity of Restrictive Lung Disease after COVID-19 (preprint)

The variable etiology of persistent breathlessness after COVID-19 have confounded efforts to decipher the immunopathology of lung sequelae. Here, we analyzed hundreds of cellular and molecular features in the context of discrete pulmonary phenotypes to define the systemic immune landscape of post-COVID lung disease. Cluster analysis of lung physiology measures highlighted two phenotypes of restrictive lung disease that differed by their impaired diffusion and severity of fibrosis. Machine learning revealed marked CCR5+CD95+ CD8+ T-cell perturbations in mild-to-moderate lung disease, but attenuated T-cell responses hallmarked by elevated CXCL13 in more severe disease. Distinct sets of cells, mediators, and autoantibodies distinguished each restrictive phenotype, and differed from those of patients without significant lung involvement. These differences were reflected in divergent T-cell-based type 1 networks according to severity of lung disease. Our findings, which provide an immunological basis for active lung injury versus advanced disease after COVID-19, might offer new targets for treatment.

The inflammatory microenvironment of the lung at the time of infection governs innate control of SARS-CoV-2 replication (preprint)

SARS-CoV-2 infection leads to vastly divergent clinical outcomes ranging from asymptomatic infection to fatal disease. Co-morbidities, sex, age, host genetics and vaccine status are known to affect disease severity. Yet, how the inflammatory milieu of the lung at the time of SARS-CoV-2 exposure impacts the control of viral replication remains poorly understood. We demonstrate here that immune events in the mouse lung closely preceding SARS-CoV-2 infection significantly impact viral control and we identify key innate immune pathways required to limit viral replication. A diverse set of pulmonary inflammatory stimuli, including resolved antecedent respiratory infections with S. aureus or influenza, ongoing pulmonary M. tuberculosis infection, ovalbumin/alum-induced asthma or airway administration of defined TLR ligands and recombinant cytokines, all establish an antiviral state in the lung that restricts SARS-CoV-2 replication upon infection. In addition to antiviral type I interferons, the broadly inducible inflammatory cytokines TNF and IL-1 precondition the lung for enhanced viral control. Collectively, our work shows that SARS-CoV-2 may benefit from an immunologically quiescent lung microenvironment and suggests that heterogeneity in pulmonary inflammation that precedes or accompanies SARS-CoV-2 exposure may be a significant factor contributing to the population-wide variability in COVID-19 disease outcomes.

Pre-existing interferon gamma conditions the lung to mediate early control of SARS-CoV-2 (preprint)

Interferons (IFNs) are critical for anti-viral host defence. Type-1 and type-3 IFNs are typically associated with early control of viral replication and promotion of inflammatory immune responses; however, less is known about the role of IFN{gamma} in anti-viral immunity, particularly in the context of SARS-CoV-2. We have previously observed that lung infection with attenuated bacteria Mycobacterium bovis BCG achieved though intravenous (iv) administration provides strong protection against SARS-CoV-2 (SCV2) infection and disease in two mouse models. Assessment of the pulmonary cytokine milieu revealed that iv BCG induces a robust IFN{gamma} response and low levels of IFN{beta}. Here we examined the role of ongoing IFN{gamma} responses due to pre-established bacterial infection on SCV2 disease outcomes in two murine models. We report that IFN{gamma} is required for iv BCG induced reduction in pulmonary viral loads and that this outcome is dependent on IFN{gamma} receptor expression by non-hematopoietic cells. Further analysis revealed that BCG infection promotes the upregulation of interferon-stimulated genes (ISGs) with reported anti-viral activity by pneumocytes and bronchial epithelial cells in an IFN{gamma}-dependent manner, suggesting a possible mechanism for the observed protection. Finally, we confirmed the importance of IFN{gamma} in these anti-viral effects by demonstrating that the recombinant cytokine itself provides strong protection against SCV2 challenge when administered intranasally. Together, our data show that a pre-established IFN{gamma} response within the lung is protective against SCV2 infection, suggesting that concurrent or recent infections that drive IFN{gamma} may limit the pathogenesis of SCV2 and supporting possible prophylactic uses of IFN{gamma} in COVID-19 management.

Establishing SARS-CoV-2 Membrane protein-specific antibodies as a valuable serological target via high-content microscopy

The prevalence and strength of serological responses mounted towards SARS-CoV-2 proteins other than nucleocapsid (N) and spike (S), which may be of use as additional serological markers, remains underexplored. Using high content microscopy to assess antibody responses against full length StrepTagged SARS-CoV-2 proteins, we found that 85% (166/196) of unvaccinated individuals with RT-PCR confirmed SARS-CoV-2 infections and 74% (31/42) of individuals infected after being vaccinated developed detectable IgG against the structural protein M, which is higher than previous estimates. Compared with N antibodies, M IgG displayed a shallower time-dependent decay and greater specificity. Sensitivity for SARS-CoV-2 seroprevalence was enhanced when N and M IgG detection was combined. These findings indicate that screening for M seroconversion may be a good approach for detecting additional vaccine breakthrough infections and highlight the potential to use HCM as a rapidly deployable method to identify the most immunogenic targets of newly emergent pathogens. Graphical

Simultaneous induced membrane technique to reconstruct an obliterated floating knee.

The induced membrane technique, first described by Masquelet, is a powerful surgical approach that can be used to address segmental bone loss of various aetiologies. Despite ongoing debate regarding optimal delivery, the indications and limits of its application have been tested in increasingly complex situations, highlighting its considerable potential. We present a case of a devastating open lower limb injury with simultaneous femoral and ipsilateral tibial bone loss including articular injury on both sides of the joint. The Masquelet technique was used to successfully address both segments of bone loss within the same limb.

Changes in anticoagulant and antiplatelet drug supply at US hospitals before and during the COVID-19 pandemic (2018-2021).

PURPOSE: Antithrombotic agents have a role in coronavirus disease 2019 (COVID-19) treatment, but the pandemic disrupted medication supply. This study examined changes in the volume of oral and parenteral anticoagulant and antiplatelet medications at US hospitals during the pandemic. METHODS: IQVIA National Sales Perspective (NSP) data was used to determine the monthly volume of anticoagulants and antiplatelets purchased at US hospitals between January 2018 and February 2021. Mean monthly medication volumes, reported as extended units (EUs), and year-over-year changes in medication volume were determined. A single-group interrupted time series analysis was used to evaluate changes in the rate of growth of monthly medication volumes before (January 2019-February 2020) and during (March 2020-February 2021) the COVID-19 pandemic. RESULTS: Overall, there was a 43.4% decline in the total volume of anticoagulants and antiplatelets at US hospitals in March 2020, driven by a decrease in heparin volume. Mean monthly volumes decreased significantly (P < 0.05) for parenteral anticoagulants (-106,691,340 EU [95% CI, -200,033,910 to -13,348,780]), oral anticoagulants (-354,800 EU [95% CI, -612,180 to -97,420]), and parenteral antiplatelets (-391,880 EU [95% CI, -535,420 to -248,330]). During the pandemic, the monthly volume of oral anticoagulants, parenteral anticoagulants, and parenteral antiplatelets grew significantly more than in the prepandemic period. This growth was primarily seen in volumes of apixaban, argatroban, enoxaparin, heparin, eptifibatide, and tirofiban. Apixaban and heparin volumes continued a prepandemic uptrend, while argatroban and eptifibatide volumes reversed trend. CONCLUSION: Rapid changes in anticoagulant and antiplatelet volume at US hospitals during the COVID-19 pandemic highlight the need for institutional protocols to manage fluctuating medication volume demands.

The impact of the COVID-19 pandemic on inequalities in lifestyle behaviours and mental health and wellbeing of elementary school children in northern Canada.

Objectives: The COVID-19 pandemic has negatively affected children's lifestyle behaviours and mental health and wellbeing, and concerns have been raised that COVID-19 has also increased health inequalities. No study to date has quantified the impact of COVID-19 on health inequalities among children. We compared pre-pandemic vs. post-lockdown inequalities in lifestyle behaviours and mental health and wellbeing among children living in rural and remote northern communities. Methods: We surveyed 473 grade 4-6 students (9-12 years of age) from 11 schools in rural and remote communities in northern Canada in 2018 (pre-pandemic), and 443 grade 4-6 students from the same schools in 2020 (post-lockdown). The surveys included questions on sedentary behaviours, physical activity, dietary intake, and mental health and wellbeing. We measured inequality in these behaviors using the Gini coefficient, a unitless measure ranging from 0 to 1 with a higher value indicating greater inequality. We used temporal changes (2020 vs. 2018) in Gini coefficients to assess the impact of COVID-19 on inequalities in lifestyle behaviours and mental health and wellbeing separately among girls and boys. Results: Inequalities in all examined lifestyle behaviours increased between 2018 and 2020. Inequalities in watching TV, playing video games, and using a cell phone increased among girls, while inequalities in playing video games, using computers and tablets, and consumption of sugar, salt, saturated fat and total fat increased among boys. Changes in inequalities in mental health and wellbeing were small and not statistically significant. Conclusion: The findings suggest that the COVID-19 pandemic has exacerbated inequalities in lifestyle behaviours among children living in rural and remote northern communities. If not addressed, these differences may translate into exacerbated inequalities in future health. The findings further suggest that school health programs can help mitigate the negative impact of the pandemic on lifestyle behaviours and mental health and wellbeing.

Pornography and Sociosexual Attitudes and Behaviors in a Nationally Representative Survey: Potential Pandemic and Method Effects.

Due to the COVID-19 pandemic, The General Social Survey (GSS) switched from an in-person interview to a self-administered online survey for its most recent data collection. This modality switch makes it possible to compare sociosexual data gathered using the GSS's last in-person survey (2018) to its first-ever self-administered online survey (2021), an oft suggested format for reducing social desirability bias. This study compared sociosexual data collected in the 2018 GSS to the 2021 GSS, with a primary focus on pornography use. Results suggested that among men neither the direction nor the magnitude of the association between pornography use and more nontraditional sociosexual attitudes and behaviors are affected by whether surveys are conducted in-person or online; that among women the magnitude of the positive association between pornography use and certain nontraditional sexual behaviors could be attenuated by in-person interviews; a pandemic period increase in pornography use among both men and women; a pandemic period decrease in men's nonrelational sexual behavior; and that men's and women's reporting of certain nontraditional sexual attitudes may be reduced by in-person interviews. It is important to emphasize that alternative explanations for 2018-2021 change are possible. The goal of the present study was to promote interpretive dialogue rather than definitive answers.

Proteomic analysis of circulating immune cells identifies cellular phenotypes associated with COVID-19 severity.

Certain serum proteins, including C-reactive protein (CRP) and D-dimer, have prognostic value in patients with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Nonetheless, these factors are non-specific, providing limited mechanistic insight into the peripheral blood mononuclear cell (PBMC) populations that drive the pathogenesis of severe COVID-19. To identify cellular phenotypes associated with disease, we performed a comprehensive, unbiased analysis of total and plasma-membrane PBMC proteomes from 40 unvaccinated individuals with SARS-CoV-2, spanning the whole disease spectrum. Combined with RNA sequencing (RNA-seq) and flow cytometry from the same donors, we define a comprehensive multi-omic profile for each severity level, revealing that immune-cell dysregulation progresses with increasing disease. The cell-surface proteins CEACAMs1, 6, and 8, CD177, CD63, and CD89 are strongly associated with severe COVID-19, corresponding to the emergence of atypical CD3+CD4+CEACAM1/6/8+CD177+CD63+CD89+ and CD16+CEACAM1/6/8+ mononuclear cells. Utilization of these markers may facilitate real-time patient assessment by flow cytometry and identify immune populations that could be targeted to ameliorate immunopathology.

Risks of neurological and psychiatric sequelae 2 years after hospitalisation or intensive care admission with COVID-19 compared to admissions for other causes.

The association between COVID-19 and subsequent neurological and psychiatric disorders is well established. However, two important questions remain unanswered. First, what are the risks in those admitted to intensive care unit (ICU) with COVID-19? Admission to ICU is itself associated with neurological and psychiatric sequelae and it is not clear whether COVID-19 further increases those risks or changes their profile. Second, what are the trajectories of neurological and psychiatric risks in patients admitted to hospital or ICU with COVID-19, and when do the risks subside? We sought to answer these two questions using a retrospective cohort study based on electronic health records (EHR) data from the TriNetX Analytics Network (covering 89 million patients, mostly in the USA). Cohorts of patients admitted to hospital or ICU with COVID-19 were propensity score-matched (for 82 covariates capturing risk factors for COVID-19 and more severe COVID-19 illness) to patients admitted to hospital or ICU (respectively) for any other reason. Matched cohorts were followed for up to two years and the risk of 14 neurological and psychiatric outcomes were compared. A total of 280,173 patients admitted to hospital and 46,573 patients admitted to ICU with COVID-19 were successfully matched to an equal number of patients admitted to hospital or ICU for any other reason. Those hospitalised with COVID-19 were found to be at a greater risk of a range of neurological and psychiatric outcomes including seizure/epilepsy, encephalitis, myoneural junction/muscle disease, Guillain-Barré syndrome (GBS), dementia, cognitive deficits, psychotic disorder, mood and anxiety disorders, but not ischaemic stroke or intracranial haemorrhage. When risks were elevated after COVID-19, most remained so for the whole two years of follow-up (except for mood and anxiety disorders). Risk profiles and trajectories were substantially different among those admitted to ICU: compared to those admitted for any other reasons, those admitted with COVID-19 were at a greater risk of myoneural junction/muscle disease, GBS, cognitive deficits and anxiety disorder, but at a significantly lower risk of ischaemic stroke, intracranial haemorrhage, encephalitis, and mood disorder. When elevated, the risks in those admitted to ICU with COVID-19 were mostly short-lived. In summary, risks of neurological and psychiatric sequelae in patients hospitalised with COVID-19 are wide ranging and long standing whereas those in patients admitted to ICU with COVID-19 are similar to, or lower than, the risks observed post-ICU admission for any other cause. These contrasting risk trajectories are relevant for researchers, clinicians, patients, and policymakers.

An approximate diffusion process for environmental stochasticity in infectious disease transmission modelling.

Modelling the transmission dynamics of an infectious disease is a complex task. Not only it is difficult to accurately model the inherent non-stationarity and heterogeneity of transmission, but it is nearly impossible to describe, mechanistically, changes in extrinsic environmental factors including public behaviour and seasonal fluctuations. An elegant approach to capturing environmental stochasticity is to model the force of infection as a stochastic process. However, inference in this context requires solving a computationally expensive "missing data" problem, using data-augmentation techniques. We propose to model the time-varying transmission-potential as an approximate diffusion process using a path-wise series expansion of Brownian motion. This approximation replaces the "missing data" imputation step with the inference of the expansion coefficients: a simpler and computationally cheaper task. We illustrate the merit of this approach through three examples: modelling influenza using a canonical SIR model, capturing seasonality using a SIRS model, and the modelling of COVID-19 pandemic using a multi-type SEIR model.

The Impact of COVID-19 Redeployment on an Orthopedic Surgery Department at a Tertiary Care Medical Center in New York City.

BACKGROUND: The SARS-CoV-2 (COVID-19) pandemic resulted in new, non-orthopedic roles for many members of our New York City based orthopedic department, including redeployment to medicine wards, emergency departments, and intensive care units. The purpose of this study was to determine if certain areas of redeployment predisposed individuals to higher likelihood of positive diagnostic or serologic testing for COVID-19. METHODS: In this study, attendings, residents, and phy-sician assistants within our orthopedic department were surveyed to determine their roles during the COVID-19 pandemic and whether they were tested via diagnostic or serologic methods for detecting COVID-19. Additionally, symptoms and missed days of work were reported. RESULTS: No significant association between redeployment site and rate of positive COVID-19 diagnostic (p = 0.91) or serologic (p = 0.38) testing was detected. Sixty individuals responded to the survey, with 88.3% of respondents rede-ployed during the pandemic. Nearly half (n = 28) of those redeployed experienced at least one COVID-19 related symptom. Two respondents had a positive diagnostic test, and 10 had a positive serologic test. CONCLUSIONS: Area of redeployment during the COVID-19 pandemic is not associated with an increased risk of subse-quently having a positive diagnostic or serologic COVID-19 test.

Blood gene expression predicts intensive care unit admission in hospitalised patients with COVID-19.

Background: The COVID-19 pandemic has created pressure on healthcare systems worldwide. Tools that can stratify individuals according to prognosis could allow for more efficient allocation of healthcare resources and thus improved patient outcomes. It is currently unclear if blood gene expression signatures derived from patients at the point of admission to hospital could provide useful prognostic information. Methods: Gene expression of whole blood obtained at the point of admission from a cohort of 78 patients hospitalised with COVID-19 during the first wave was measured by high resolution RNA sequencing. Gene signatures predictive of admission to Intensive Care Unit were identified and tested using machine learning and topological data analysis, TopMD. Results: The best gene expression signature predictive of ICU admission was defined using topological data analysis with an accuracy: 0.72 and ROC AUC: 0.76. The gene signature was primarily based on differentially activated pathways controlling epidermal growth factor receptor (EGFR) presentation, Peroxisome proliferator-activated receptor alpha (PPAR-α) signalling and Transforming growth factor beta (TGF-ß) signalling. Conclusions: Gene expression signatures from blood taken at the point of admission to hospital predicted ICU admission of treatment naïve patients with COVID-19.

Coordination in a not-for-profit organisation during the COVID-19 pandemic: organisational sensemaking during planning meetings

PurposeThe authors examine how a not-for-profit organisation (NPO) coordinates NPO's actions during the coronavirus disease 2019 (COVID-19) global pandemic to remain focussed on strategic and operational goals.Design/methodology/approachThe authors conducted a live case study of an NPO as the crises caused by the COVID-19 pandemic unfolded. Drawing on a sensemaking perspective that incorporates sensegiving, the authors develop a framework of five types of organisational sensemaking. The authors analyse weekly planning meetings during which managers discussed past performance, forecast performance and the forecast duration of current cash reserves.FindingsThe authors show how three of the five types of organisational sensemaking helped to coordinate actions. The authors highlight how accounting information triggers organisational sensemaking processes;but depending on the type of organisational sensemaking, accounting information has little further role. The authors also show that the stability of decisions depends on the types of organisational sensemaking.Practical implicationsThe authors show how coordination as a management control practice is enabled by organisational sensemaking within an NPO during a crisis. Organisational sensemaking enabled the agreement of actions, which enabled coordination. Accounting practices provided trigger mechanisms to facilitate organisational sensemaking.Originality/valueSince this study is the first to examine sensemaking processes and accounting practices in coordination in an NPO in a pandemic, the authors contribute to the limited research on NPOs during crises and on the management control practice of coordination. The authors extend the accounting literature on sensemaking by showing that, whilst accounting triggers organisational sensemaking, accounting is only implicated in one type of organisational sensemaking and by revealing the different outcomes of the different types of organisational sensemaking.

Risk factors for SARS-CoV-2 seroprevalence following the first pandemic wave in UK healthcare workers in a large NHS Foundation Trust.

Background: We aimed to measure SARS-CoV-2 seroprevalence in a cohort of healthcare workers (HCWs) during the first UK wave of the COVID-19 pandemic, explore risk factors associated with infection, and investigate the impact of antibody titres on assay sensitivity. Methods: HCWs at Sheffield Teaching Hospitals NHS Foundation Trust were prospectively enrolled and sampled at two time points. We developed an in-house ELISA for testing participant serum for SARS-CoV-2 IgG and IgA reactivity against Spike and Nucleoprotein. Data were analysed using three statistical models: a seroprevalence model, an antibody kinetics model, and a heterogeneous sensitivity model. Results: Our in-house assay had a sensitivity of 99·47% and specificity of 99·56%. We found that 24·4% (n=311/1275) of HCWs were seropositive as of 12th June 2020. Of these, 39·2% (n=122/311) were asymptomatic. The highest adjusted seroprevalence was measured in HCWs on the Acute Medical Unit (41·1%, 95% CrI 30·0-52·9) and in Physiotherapists and Occupational Therapists (39·2%, 95% CrI 24·4-56·5). Older age groups showed overall higher median antibody titres. Further modelling suggests that, for a serological assay with an overall sensitivity of 80%, antibody titres may be markedly affected by differences in age, with sensitivity estimates of 89% in those over 60 years but 61% in those ≤30 years. Conclusions:  HCWs in acute medical units and those working closely with COVID-19 patients were at highest risk of infection, though whether these are infections acquired from patients or other staff is unknown. Current serological assays may underestimate seroprevalence in younger age groups if validated using sera from older and/or more severe COVID-19 cases.

A Fluorescence Polarization Assay for Macrodomains Facilitates the Identification of Potent Inhibitors of the SARS-CoV-2 Macrodomain.

Viral macrodomains, which can bind to and/or hydrolyze adenine diphosphate ribose (ADP-ribose or ADPr) from proteins, have been suggested to counteract host immune response and be viable targets for the development of antiviral drugs. Therefore, developing high-throughput screening (HTS) techniques for macrodomain inhibitors is of great interest. Herein, using a novel tracer TAMRA-ADPr, an ADP-ribose compound conjugated with tetramethylrhodamine, we developed a robust fluorescence polarization assay for various viral and human macrodomains including SARS-CoV-2 Macro1, VEEV Macro, CHIKV Macro, human MacroD1, MacroD2, and PARP9 Macro2. Using this assay, we validated Z8539 (IC50 6.4 µM) and GS441524 (IC50 15.2 µM), two literature-reported small-molecule inhibitors of SARS-CoV-2 Macro1. Our data suggest that GS441524 is highly selective for SARS-CoV-2 Macro1 over other human and viral macrodomains. Furthermore, using this assay, we identified pNP-ADPr (ADP-ribosylated p-nitrophenol, IC50 370 nM) and TFMU-ADPr (ADP-ribosylated trifluoromethyl umbelliferone, IC50 590 nM) as the most potent SARS-CoV-2 Macro1 binders reported to date. An X-ray crystal structure of SARS-CoV-2 Macro1 in complex with TFMU-ADPr revealed how the TFMU moiety contributes to the binding affinity. Our data demonstrate that this fluorescence polarization assay is a useful addition to the HTS methods for the identification of macrodomain inhibitors.

CD4+ T cell calibration of antigen-presenting cells optimizes antiviral CD8+ T cell immunity.

Antiviral CD8+ T cell immunity depends on the integration of various contextual cues, but how antigen-presenting cells (APCs) consolidate these signals for decoding by T cells remains unclear. Here, we describe gradual interferon-α/interferon-ß (IFNα/ß)-induced transcriptional adaptations that endow APCs with the capacity to rapidly activate the transcriptional regulators p65, IRF1 and FOS after CD4+ T cell-mediated CD40 stimulation. While these responses operate through broadly used signaling components, they induce a unique set of co-stimulatory molecules and soluble mediators that cannot be elicited by IFNα/ß or CD40 alone. These responses are critical for the acquisition of antiviral CD8+ T cell effector function, and their activity in APCs from individuals infected with severe acute respiratory syndrome coronavirus 2 correlates with milder disease. These observations uncover a sequential integration process whereby APCs rely on CD4+ T cells to select the innate circuits that guide antiviral CD8+ T cell responses.