Mid-density Gene Expression profiling of SARSCOV-2 infected samples using Applied BiosystemsTM TaqManTM Flexible Array Panels

Background/Objectives: The broad spectrum of clinical manifestations from SARS-COV-2 infection and observed risk factors for severe disease highlight the importance of understanding molecular mechanisms underlying SARS-CoV-2 associated disease pathogenesis. Research studies have identified a large number of host proteins that play roles in viral entry, innate immune response, or immune signalling during infection. The ability to interrogate subsets of these genes simultaneously within SARSCOV-2 infected samples is critical to understanding how their expression contribute to phenotypic variability of the disease caused by the pathogen. Method(s): 30 Nasopharyngeal swab were obtained and included SARS-CoV-2 infected and control samples. RNA was extracted, reverse transcribed and loaded onto flexible TaqMan array panels designed specifically for targeting the most cited genes related to SARS-COV-2 entry and restriction factors as well as cytokines, chemokines, and growth factors involved in the pathogenesis. Result(s): Our data indicated that not only were the levels of several of these host factors differentially modulated between the two study groups, but also that SARS-CoV-2 infected subjects presented with greater frequency of several important inflammatory cytokines and chemokines such as CCL2, CCL3, IFNG, entry receptors such as ACE2, TMRPS11A, and host restriction factors including LY6E and ZBP1. Conclusion(s): TaqMan array plates provide a fast, midthroughput solution to determine the levels of several virus and host-associated factors in various cell types and add to our understanding of how the pathogenesis may vary depending on gender, age, infection site etc.

Optimizing mRNA vaccines and virusmediated CAR T-cell therapy for immuno-oncology: Synthetic tools for gene modification

With the success of mRNA vaccines during the COVID-19 pandemic and CAR T-cell therapies in clinical trials, there is growing opportunity for immunotherapies in the treatment of many types of cancers. Lentiviral vectors have proven effective at delivery of genetic material or gene editing technology for ex vivo processing, but the benefits and promise of Adeno-associated virus (AAV) and mRNA tools for in vivo immunotherapy have garnered recent interest. Here we describe complete synthetic solutions for immuno-oncology research programs using either mRNA-vaccines or virus-mediated cell and gene engineering. These solutions optimize workflows to minimize screening time while maximizing successful research results through: (1) Efficiency in lentiviral packaging with versatility in titer options for high-quality particles. (2) A highthroughput viral packaging process to enable rapid downstream screening. (3) Proprietary plasmid synthesis and preparation techniques to maintain ITR integrity through AAV packaging and improve gene delivery. (4) Rapid synthesis, in vitro transcription, and novel sequencing of mRNA constructs for complete characterization of critical components such as the polyA tail. The reported research demonstrates a streamlined approach that improves data quality through innovative synthesis and sequencing methodologies as compared to current standard practices.

Virtual learning before and during the COVID-19 pandemic: A mixed methods study of Pharmacy students

Background It is a challenge for pharmacy courses worldwide to combine theoretical knowledge with practical skills to equip students for their future practice. Computer-based simulation offers a way of building a bridge between theory and practice. In recent years, digital simulation has expanded rapidly as a new technique of virtual learning. The digital platform ''Pharmacy Simulator'' proposes computer-based encounters with virtual patients to train clinical and communication skills in a community pharmacy setting. However, during the COVID-19 pandemic, while students were digitally resilient and endured the endless challenges of online lectures, many were dealing with Zoom and screen fatigue. Purpose To investigate pharmacy students' acceptance of Pharmacy Simulator before and during a pandemic situation. This focuses on students' self-assessment and confidence in counselling after playing the scenarios on Pharmacy Simulator. Method Two cohorts of Master of Pharmacy students at The University of Western Australia played two scenarios on Pharmacy Simulator in 2019 (anaphylaxis and salbutamol) and 2021 (anaphylaxis and vaccination). A mixed-method analysis was performed with data from (i) qualitative semi-structured interviews carried out in 2019 pertaining to participants' acceptance of Pharmacy Simulator and in 2021 (ii) a questionnaire with 25 items derived from the interviews. The interviews were transcribed verbatim into electronic format with the data management assistance MAXQDA and analyzed inductively using the Framework Method. Questionnaire responses were analyzed in Microsoft Excel using descriptive statistics. Openended questions were evaluated inductively. Findings Data were collected from 20 interviews and 31 answered questionnaires. In 2019, participants reported that Pharmacy Simulator was a fun, engaging, and straightforward learning tool and, therefore, user-friendly. They reported the feedback at the end of the session to be most valuable. The platform was perceived to fill the gap between the theory from lectures and community pharmacy practice. In 2021, participants ''agreed'' (median: 4, on a 5-point Likert scale) with seven statements about Pharmacy Simulator's usability, such as it being a helpful tool for acquiring new knowledge. Participants' confidence in counselling regarding the scenario topics improved. One participant stated, ''It taught me more through trial and error''. Conclusion Pharmacy students reported similar acceptance levels of Pharmacy Simulator before and during the COVID-19 pandemic. The use of simulation during virtual patient encounters seems to facilitate the transfer from theory to practice, independently of learning conditions that were predominantly screen-based.

A Qualitative Analysis of Physician, Staff, and Patient Perspectives During the Discharge Process for Vascular Patients

Objectives: Evidence demonstrates that when hospitals focus on the discharge process patient safety improves and overall costs decline. Hospital discharge requires care coordination of multiple disciplines, often leading to fragmented care, and adverse outcomes after discharge include emergency department visits and hospital readmissions. The Re-Engineered Discharge (RED) process was developed as an evidence-based strategy to improve the hospital discharge. We evaluated perspectives and priorities of physicians, health care workers, and patients involved in the vascular discharge process using RED as a framework. Method(s): A single-center qualitative analysis using a semi-structured focus groups and an interview guide based on the RED process. Focus groups were Zoom platform recorded, transcribed into text files, independently coded, and analyzed with Dedoose qualitative software using a directed content analysis approach. Thematic concepts were created, and comparisons between groups were analyzed by coding frequency. Researchers independently thematically coded each transcript;prior to analysis, all redundancy of codes was resolved;and all team members agreed on text categorization and coding frequency. Result(s): Eight focus groups with 38 participants were performed. Participants included: physicians (n = 13), nursing and ancillary staff (n = 19), and patients/caregivers (n = 6). Transcript analyses revealed facilitators and barriers to discharge. Overarching themes identified from the qualitative analysis frequencies are displayed by stakeholder role (Fig 1). Themes identified with the greatest coding frequencies included helpfulness of discharge instructions, patient health literacy, patient medical complexity, poor interdisciplinary team communication, time constraints during discharge, technology literacy of patients, barriers to obtaining medications for patients, barriers to organizing outpatient services for health care workers, barriers for patients to obtain help after discharge, and the impact of COVID-19. Conclusion(s): These findings identify the need to strengthen efforts to overcome stakeholder barriers to improve patient safety at the interface of the hospital to create a well-organized discharge. Physicians were most concerned with low patient health literacy, patient understanding of discharge instructions, organizing outpatient services, and overall patient medical complexity hindering a smooth discharge. Health care staff identified time constraints, obtaining medications and, and inter-team communication as their greatest obstacles to an organized discharge. Patients found the complexity and amount of discharge instructions, the impact of COVID-19 on support systems, and technology utilization after discharge most challenging. Modifications to address individual stakeholder barriers within the discharge process are needed to develop a national standardized discharge specific for vascular surgery patients to improve patient safety and satisfaction. [Formula presented]Copyright © 2023

A qualitative study exploring how youth use TikTok to learn about mental health and mental health services in British Columbia, Canada

Aims: Youth are increasingly seeking health information through online platforms, such as websites, social media, and online forums. TikTok emerged as a popular platform for disseminating and consuming health information during the COVID-19 pandemic. As such, this study aimed to explore how youth used TikTok to access information about mental health and mental health services during the pandemic. Method(s): Twenty-one interviews were conducted over Zoom with youth (ages 12-24) who lived in British Columbia, Canada and had accessed TikTok for mental health information during the pandemic. Interviews were audio-recorded, transcribed verbatim, and analysed thematically using an inductive approach. Result(s): Youth described TikTok as a safe place to talk about mental health and share personal experiences. This helped youth feel less alone with their struggles and facilitated conversations about mental health with friends, family, and service providers. Participants also described how mental health content on TikTok helped them be more mindful of their own mental health and the different resources and coping strategies available and encouraged them to seek services. For those hesitant or unable to access services, TikTok provided immediate support. Youth appreciated the ease of accessing this information, given the platform's engaging and digestible format (i.e., short videos) and predictive nature of its algorithm. However, participants expressed concerns with the spread of misinformation and the lack of verifiable information on the platform. Conclusion(s): TikTok is as a practical platform to disseminate mental health information to youth. However, efforts to establish strategies for preventing and reporting misinformation are warranted.

Increases in expression of carbohydrate sulfotransferases CHST11 and CHST15 and decline in N-acetylgalactosamine-4-sulfatase (Arylsulfatase B, ARSB) require phospho-p38-MAPK following exposure to SARS-CoV-2 spike protein receptor binding domain in human a

Objective: Immunohistochemistry of post-mortem lung tissue from Covid-19 patients with diffuse alveolar damage demonstrated marked increases in chondroitin sulfate and CHST15 and decline in N-acetylgalactosamine-4-sulfatase. Studies were undertaken to identify the mechanisms involved in these effects. Method(s): Human primary small airway epithelial cells (PCS 301-010;ATCC) were cultured and exposed to the SARSCoV- 2 spike protein receptor binding domain (SPRBD;AA: Lys310-Leu560;Amsbio). Expression of the spike protein receptor, angiotensin converting enzyme 2 (ACE2), was enhanced by treatment with Interferon-beta. Promoter activation, DNA-binding, RNA silencing, QPCR, Western blots, ELISAs, and specific enzyme inhibitors were used to elucidate the underlying molecular mechanisms. Result(s): Treatment of the cultured cells by the SPRBD led to increased CHST15 and CHST11 expression and decline in ARSB expression. Sulfotransferase activity, total chondroitin sulfate, and sulfated glycosaminoglycan (GAG) content were increased. Phospho-T180/T182-p38-MAPK and phospho- S423/S425-Smad3 were required for the activation of the CHST15 and CHST11 promoters. Inhibition by SB203580, a phospho-p38 MAPK inhibitor, and by SIS3, a Smad3 inhibitor, blocked the CHST15 and CHST11 promoter activation. SB203580 reversed the SPRBD-induced decline in ARSB expression, but SIS3 had no effect on ARSB expression or promoter activation. Phospho-p38 MAPK was shown to reduce retinoblastoma protein (RB) S807/S811 phosphorylation and increase RB S249/T252 phosphorylation. E2F-DNA binding declined following exposure to SPRBD, and SB203580 reversed this effect. This indicates a mechanism by which SPRBD, phospho-p38 MAPK, E2F, and RB can regulate ARSB expression and thereby impact on chondroitin 4-sulfate and dermatan sulfate and molecules that bind to these sulfated GAGs, including Interleukin-8, bone morphogenetic protein-4, galectin-3 and SHP-2 (Src homology region 2-containing protein tyrosine phosphatase 2). Conclusion(s): The enzyme ARSB is required for the degradation of chondroitin 4-sulfate and dermatan sulfate, and accumulation of these sulfated GAGs can contribute to lung pathophysiology, as evident in Covid-19. Some effects of the SPRBD may be attributable to unopposed Angiotensin II, when Ang1-7 counter effects are diminished due to binding of ACE2 with the SARS-CoV-2 spike protein and reduced production of Ang1-7. Aberrant cell signaling and activation of the phospho-p38 MAPK and Smad3 pathways increase CHST15 and CHST11 production, which can contribute to increased chondroitin sulfate in infected cells. Decline in ARSB may occur as a consequence of effects of phospho-p38 MAPK on RB phosphorylation and E2F1 availability. Decline in ARSB and the resulting impaired degradation of sulfated GAGs have profound consequences on cellular metabolic, signaling, and transcriptional events. Funding is VA Merit Award.Copyright © 2023 The American Society for Biochemistry and Molecular Biology, Inc.

Qualitative methods in a learning healthcare system: Balancing rigour and the need for rapid analysis and dissemination

Qualitative methods are used to capture stakeholder perspectives within learning healthcare systems (LHS), but there is a need to specify methods that balance rigour and pragmatic approaches to inform quality improvement (QI). Utilizing examples from two QI projects within the OTNY LHS, we illustrate methods and strategies that build team capacity and flexibility to respond to an evolving LHS. Method(s): Qualitative methods were tailored to fit each project's timelines and goals, to inform both practice and research. Tools to facilitate rapid cycle feedback included interview/focus group summary templates, aggregate summaries that synthesize findings by stakeholder group, case matrix templates for rapid extraction and systematic categorization of data along topic areas, and dissemination materials adapted for stakeholder audience and project phases. Strategies to maintain rigour included processes for data reduction and interpretation, a multi-disciplinary approach for analysis, frequent consensus-based meetings, data triangulation, and member checks. Result(s): Rapid cycle approaches yielded interim results that reshaped research questions or identified critical gaps. Case summary analysis exploring the impact of COVID-19 revealed limited information on telehealth challenges amongst OTNY participants, necessitating a shift in recruitment and interview focus. For another project, analytic methods were sequenced to rapidly inventory suggestions from interview summaries on how to enhance OTNY practice to better address racism, while subsequent thematic analysis of transcripts captured participants' experiences of racism for context. Challenges included concurrent alignment of data collection and analysis, tailoring summary templates to maximize utility for rapid analysis, and maintaining flexibility to respond to evolving findings and LHS stakeholder input. Conclusion(s): The diverse methods and strategies illustrated by these projects offer guidance for balancing.

Determination of the Effects of Prenatal Remdesivir Exposure on Placenta Morphology, Placental IgG Transfer, and Fetal Growth in a Humanized Mouse Model

Remdesivir (RDV) is an antiviral medication used most recently for the treatment of COVID-19. Although no adverse effects were observed on perinatal parameters in reproductive and development toxicology studies at doses up to four-fold clinical area under the curve (AUC) exposures, some researchers have reported that therapeutic levels of RDV may impair early embryogenesis, as observed by in vitro studies. In addition, the influence of prenatal RDV exposure on maternal IgG transfer in the placenta is still unknown. Administration of RDV in pregnant humanized mouse model (Tg32), which expresses the human Fc gamma receptor and transporter (FCGRT) gene, was used to further evaluate potential effects on IgG transfer and concurrent perinatal endpoints. Animals were dosed daily from gestational days (GDs) 10- 14 with 25 mg/kg RDV (GS-5734) via intravenous injection (n=3-5 per group). Concurrent vehicle control animals were dosed intravenously with 12% sulfobutyl ether- beta-cyclodextrin in water (pH3.5;NaOH/HCl). All animals were administered 2 mg/kg human IgG via intravenous injection on GD 14. Placentae and fetuses were collected from dams on GD 14, 15, 16, and 18 and evaluated using histopathology and qPCR for inflammation markers. No abnormal morphologies (necrosis/apoptosis) of placentae were observed between the concurrent control and RDVdosed groups. Additionally, no differences in maternal body weights were observed. There were no statistically significant differences in placenta weights. There were no statistically significant changes in pregnancy parameters (implantation sites and dead fetuses/litter) and fetal weights between the RDV-dosed group and concurrent controls at GD 14, 15, 16, and 18. No changes were observed in transcript levels of inflammation markers in the RDV-dosed group when compared to the concurrent control group. There was a slightly lower ratio of fetal IgG level to maternal IgG levels in the RDV-dosed group;however, no statistically significant differences were observed between the RDV-dosed group and concurrent controls on GD 14, 15, 16, and 18. Our results suggest that a daily dose of 25 mg/kg RDV on GDs 10-14 in humanized mice did not cause adverse effects on placenta and fetal development. (Funded by the Perinatal Health Center of Excellence: E0300201.).

A Qualitative Investigation of the Impact of the Sars-Cov-2 Pandemic on Goal Attainment Scaling in a Clinical Research Setting

Objectives: Goal Attainment Scaling (GAS) quantifies the effect of interventions on individuals' personal goals. Goals that are meaningful to individual patients are set by GAS interviewers (physicians/academics, experienced in clinical research/trials) in collaboration with patients and/or their caregivers. The SARS-CoV-2 pandemic presented a major challenge to health care worldwide. We investigated how the SARS-CoV-2 pandemic affected the use and implementation of GAS in clinical/research settings. Method(s): Eleven GAS interviewers in Canada, the US, UK and Australia with current/past experience working with patient-caregivers to identify goals and build scales to measure goal attainment as an outcome were interviewed using a semi-structured approach. They described how the pandemic affected GAS use in clinical/research settings. Interviews were recorded and transcribed, then themes were identified and coded in Nvivo 12. Result(s): Most GAS interviews were moved to the virtual environment during the pandemic (9/11). GAS interviewers identified few negative impacts. Some goals required modification (e.g. going to the gym or shops). Pandemic stress impacted both interviewers and interviewees. Even so, most GAS interviewers (8/11) emphasized positive impacts of the pandemic. These were: 1) the virtual environment meant patients were more readily available as they did not need to travel;2) because interviewers did not have to travel, patient recruitment could be expanded into more remote geographical areas;3) researchers could recruit and retain more participants and collect data more quickly;and 4) interviewers commented they became more open to technology because of the pandemic (9/11). Conclusion(s): Despite the challenges of the SARS-CoV-2 pandemic for health care, most GAS interviewers found that the impacts were largely positive for using GAS in clinical/research settings. The ability to deploy GAS through a virtual platform could facilitate the uptake of GAS as a patient-centric outcome measure.Copyright © 2023

Using the framework for reporting adaptations and modifications for evidence-based interventions (FRAME) for reporting adaptations to coordinated specialty care during the COVID-19 pandemic

Throughout the COVID-19 pandemic, OnTrackNY teams provided coordinated specialty care (CSC) in a radically shifted environment. This presentation describes adaptations to OnTrackNY's model implemented during the pandemic. Method(s): OnTrackNY providers were recruited to participate in indepth, qualitative interviews conducted using phone and video platforms. The project team co-developed the qualitative interview guides with OnTrack Central trainers for each of the six team member roles. A coding team used the FRAME to identify the top three role-based adaptations to the OnTrackNY model from transcripts of qualitative interviews. Result(s): Twenty-three providers (n = 3-4 providers per role) discussed challenges and adaptations of providing CSC services during the pandemic. Use of telehealth was a major adaptation applied across all roles. Adaptations to outreach included narrowing community outreach to inpatient and emergency settings, increasing communication with referral sources, increasing contact with newly referred participants and families. Peer specialist adaptations include conducting physically-distanced groups, discussing current events and expanding online resources for engagement. SEES adaptations included monitoring evolving employment opportunities, conducting mock job interviews remotely, and supporting online learning. Adaptations to PCP/RN roles included sending equipment home for monitoring heath, changing methods and frequency of administering medication, and providing education regarding COVID-19 and vaccination. Adaptations to the Primary Clinician role included increasing informal 'check-ins', using screen sharing to complete assessments and safety plans, and addressing increased stress due to the pandemic. Conclusion(s): Adaptations to CSC were common with providers most frequently making changes to format and setting of care delivery and content modifications. Future work will examine implications of adaptations and OnTrackNY fidelity indicators.

Transcriptomic analysis in patients with SARS-CoV-2 virus infection. Identification of the association of risk/ protective profiles against the disease

Background/Objectives: SARS-CoV2 causes the COVID-19 disease, capable of producing a severe acute respiratory syndrome. Several clinical variables and genetic variants have been related to a worse prognosis. The aim of this study is to measure if difference in the gene expression are associated with COVID-19 severity. Method(s): We performed RNA-seq Transcriptome in RNA extracted from lymphoblastoid cell line in 20 patients who require hospitalization (10 from the intensive care unit) in a GeneStudio S5 Plus Sequencer (Ion Torrent Technology). FASTQ files were obtained and trimmed using BBtools, BBduk for cutting, filtering and masking the data, and Dedupe for the elimination of duplicates. Mapping and counting matrix was done in bash using the Salmon program. Differential expression analysis and subsequent functional enrichment was performed using Rstudio (DESeq2, ClusterProfiler, GO and KEGG). Result(s): We observed that 2042 differentially expressed genes (1996 overexpressed, LFC>0 and 406 underexpressed, LFC<0) were obtained between patients who require hospitalization versus those in the intensive care unit. We found some genes previously SARS-CoV-2 associated (PGLYRP1, HDAC9 and FUT4). Furthermore, genes involved in the activity of the immune system and in inflammatory processes showed significant differences between cohorts (ABCF1 (LFC = -25.14, padj = 1.05e-13), ABHD16A (LFC = 25.00, padj = 1.05e-13) and IER3 (LFC = -24.45, padj = 2.43e-13)). Conclusion(s): We described differential expression in genes of the immune system and inflammatory processes that might be have a role in the risk of develop severe symptoms of COVID-19, including admission in the intensive care unit. This results should be validated by additional functional studies.

Remote care for chronic pain: A qualitative evaluation with managers, practitioners, and patients

The Covid-19 pandemic obliged many healthcare providers to transition rapidly to a remote-only model of care. Concerns have been expressed about patient access to remote services, their appropriateness for sensitive consultations and physical examinations. Pre-pandemic research into telemedicine showed evidence of its effectiveness, but patient, staff and service user perspectives on remote care approaches remain unclear. This study explored the experiences and perceptions of care among patients, practitioners and managers in a single United Kingdom chronic pain organisation whose services were delivered remotely (telephone and online) during the pandemic. Nineteen participants (seven patients, seven practitioners, five managers) took part in in-depth qualitative interviews, conducted via telephone or online. Transcripts were analysed thematically. Five service provider and four service user themes were generated. Service provider themes comprised "The change process," "Accessibility and efficiency," "Effective when remote: Contact, support and education," "Concerns about communication, connection and disembodied work," and "Supporting and sustaining the team." Patients' themes comprised "Preferences, expectations and acceptance of remote care," "Convenience and accessibility," "Sense of support" and "Delivery modality matters." The study provides evidence from the qualitative evaluation of a single remote only service of its benefits and limitations as perceived by stakeholders. Findings suggest that service providers could address limitations, and progress to a blended care package, based on for patient need and choice. Further attention could be paid to services delivered by telephone, and to staff communication skills, resources, time management and wellbeing needs.Copyright © Nova Science Publishers, Inc.

Keeping occupational possibilities in pandemic times: Perspectives of young adults experiencing a first episode and their mental health practitioners

Aims: (1) To describe how occupational possibilities of young adults experiencing a first episode of psychosis changed during the pandemic;(2) to describe how mental health practitioners, working in two first episode clinics, strived to assist their clients in maintaining a meaningful daily life during the COVID-19 pandemic. Method(s): This exploratory study used an interpretative descriptive design. Young adults (n = 6), attending a specialized clinic, participated in two semi-directed interviews: they explored how their daily lives changed during the pandemic, their (re)actions in light of the changes and which occupations were (im)possible to pursue. A focus group discussion with the practitioners will allow to explore the adjustments made to clinical practices to enrich young adults' daily lives. The interviews and focus groups were transcribed and analysed independently by two members of the research team, using a thematic analysis approach. Result(s): Preliminary results suggest that if some occupational possibilities were lost, many were transformed or newly envisioned by young adults. They engaged in occupations within existing niches. The presentation will also include the perspective of mental health practitioners, as focus groups will be concluded in early Winter 2023. Conclusion(s): This presentation offers a novel perspective on the daily lives of young adults during the pandemic, focusing on the nature of their occupational engagement, whilst considering how the context shaped their occupational possibilities. Likewise, it will document the innovative practices and challenges experienced by practitioners, as they strived to expand possibilities for their clients and to support their transition into adulthood.

Outcomes of a Pilot Contribution Database for the ICU Pharmacy Team at the University Hospitals of Leicester NHS Trust

Introduction It has long been felt that many contributions made by the ICU Pharmacy team, are not well showcased by the yearly regional network multi-speciality contributions audit. Themes specific to ICU are diluted amongst Trust and region wide data, and valuable learning for the multi-disciplinary team (MDT) is subsequently overlooked. Objective(s): The aims of this project were to: * Develop and pilot a MicrosoftTM Access © database for the ICU pharmacy team to record significant contributions. * Enable the production of reports to the ICU Quality & Safety board, to raise awareness, disseminate concerns, and influence future quality improvement projects. * Provide examples to contribute to the training of the whole MDT. * Generate evidence of team effectiveness and encourage further investment. * Provide team members with a means to recall contributions, for revalidation, appraisal, prescribing re-affirmation and framework mapping. Method(s): * A database was built with a user-friendly data-entry form to prevent overwriting. Fields were agreed with peers who would be using the database. * The team were invited to voluntarily enter their contributions which they thought added value and provided useful learning. * The pilot phase ceased with the emergence of the Omicron SARS-CoV-2 variant, due to staffing pressures and surge planning. Result(s): * Between 12/07/2021 and 25/11/2021, a total of 211 contributions were recorded. * Pharmacists entered 88.6% and a single technician entered 11.4% of these. * Independent Prescribing was utilised in 52.13% of contributions, and deprescribing in 25.12%. * Figure 1 demonstrates the contributions by drug group * The top 5 drugs associated with contributions were: ? Dalteparin ? Vancomycin ? Voriconazole ? Meropenem ? Co-trimoxazole * Treatment optimisation was an outcome for 76.3% of all contributions. Figure 2 stratifies these by type. Contributions. * Drug suitability was a cause for intervention in 12.8% of all contributions, encompassing allergies, contraindications, cautions and interactions and routes. * Medicines reconciliation accounted for 17.54% of all contributions, which almost half were Technician led. Admission was the most common stage to intervene (81.08%), followed by transcription. * Of all contributions, 37.91% were classified as patient safety incidents. Reassuringly 76.25% of these were prevented by the Pharmacy team. Themes have been extracted from these incidents and are presented in Table 1. Conclusion(s): PROTECTED-UK1 demonstrated the value pharmacists contribute to the quality and safety of patient care on ICU. Studies of similar quality and scale including Pharmacy Technicians are lacking, but even in this pilot study, it is evident how important their input is. Independent prescribing is a fundamental and well utilised part of our ICU Pharmacist skillset, supporting the GPICS2 recommendation that ICU pharmacists should be encouraged to become prescribers. Compiling a team interventions database is a useful tool to highlight local priority areas for guideline development;training;and ensuring that appropriate decision support is built into electronic prescribing systems. To improve the usefulness of the data, further stratification of contributions according to the Eadon Criteria3 may be worthwhile, to expand its use as a medication safety thermometer for ICU.

"working in the dark but expected to be an expert": Reflecting on critical care preparations for COVID-19

Introduction: Before spring 2020, many healthcare organisations did not possess detailed plans for the expansion and delivery of critical care during a pandemic. Furthermore, there was little directly-relevant individual or institutional experience to draw upon. Local, national and international guidance was drawn up rapidly and subject to frequent revision.1 Reflecting on these challenges, we designed a study to explore critical care and anaesthetic doctors' experiences of preparation for the provision of critical care services in the first wave of COVID-19. Objective(s): 1. To establish what factors facilitated and hindered the expansion and delivery of critical care services. 2. To identify important learning points for the provision of critical care during future pandemics. Method(s): We conducted semi-structured interviews with medical staff from the anaesthesia and critical care departments of our hospital, a tertiary centre with general and cardiothoracic intensive care units, including an ECMO service. We classified participants into two groups;1. Decision makers - individuals instrumental in shaping the critical care response, e.g., clinical directors and college tutors. 2. Staff members - clinicians working within the departments, including consultants and trainees. Thirteen interviews were conducted with 15 participants: eight decision makers and seven staff members. The interviews were recorded, transcribed and anonymised. We manually coded transcripts, and carried out an inductive thematic analysis.2 Results: Eight themes were generated from our analysis: * Problem solving with simulation: simulation exercises allowed experienced clinicians to troubleshoot practical issues and helped staff to prepare for unfamiliar tasks. * A sense of togetherness: staff reported that the "all hands-on deck" ethos was protective against fatigue, although this was short-lived. * Delayed and changing guidance: frequent guideline changes created confusion and anxiety. * Leading from the front: leaders with a clinical role were perceived more positively than those operating at a distance from the "shop-floor". * Coordination, collaboration and compromise: departments that accommodated each other's needs fostered productive inter-departmental relationships. * Insecure supply chains: staff took their own measures to ensure PPE availability, including acquisition of items outside NHS supply chains. * Constant communication: rapid methods of personal communication, e.g., WhatsApp were effective, although "WhatsApp fatigue" was endemic. * Balancing skill mix and fatigue: flux in workload required dynamic staff allocation. Underutilised staff groups created frustration and low morale in overworked colleagues. Conclusion(s): The threat to health and society from pandemic events is expected to increase over time.3 We should take this opportunity to gather experiences from those involved in the COVID-19 pandemic to guide future preparations. In early 2020, decision makes in local hospitals were operating with unclear guidance from external agencies. Our data, obtained in the summer of 2021 demonstrates that individual and departmental reflections had already resulted in processes being refined in later waves of COVID-19. Whilst the exact nature of future pandemics will vary, some elements of preparation will remain consistent. We recommend that plans for pandemic management should aim to reduce workload and target the most effective interventions, including by addressing the themes outlined above.

Experiences of patients from minority ethnic backgrounds in the stem cell transplant pathway during COVID-19

Background: Stem cell transplantation is a potentially life-saving intervention for patients with blood cancer. Research suggests that there are existing disparities in access, care and treatment experiences, with patients identifying with a minority ethnic heritage reporting greater disadvantage compared to patients from white British, Irish or Northern European backgrounds. Although we know that the COVID-19 pandemic has been challenging for patients, less is known about the experiences of patients from specific ethnic communities. Method(s): In this research, we undertook 1-1, semi-structured interviews with eight patients within the stem cell transplant pathway who identify with a minority ethnic background. Interviews took place online, between May and November 2021. The interview questions explored views on aspects of the pandemic, including access and experience of care delivery, intervention, mental health and financial impact. Interviews were transcribed verbatim and analysed using reflexive thematic analysis. Result(s): The majority of patient participants were female (75%), and identified with a Black African or Caribbean heritage (75%). Four themes were identified from the data: (1) 'Lack of suitable donors' described the challenges of finding a stem cell donor and the importance of raising awareness of stem cell donation specifically within minority ethnic communities;(2) 'Experiences of care' explored patients' experiences of the healthcare system from pre-diagnosis to post-treatment, including how the COVID-19 pandemic had made some aspects of care easier but acted to disrupt others;(3) 'Intense and unpredictable process' described the nature of living with blood cancer and how this has impacted upon patients' lives including the extent to which they had been able to access and make use of advice to help keep them safe during the pandemic;and (4) 'Coping mechanisms' identified the factors which helped patients' to understand and adjust to living with their diagnosis and the treatment process. Conclusion(s): Patients in the stem cell transplant pathway often experience intense treatment regimens, debilitating symptoms and long hospital stays, which has a significant impact on their lives. Minority ethnic patients describe facing additional challenges in relation to health inequalities both within and outside of the pandemic. Seeking support from family, peers or community organisations can help buffer the negative impacts of living with blood cancer and multiple disadvantages, but such support was more difficult to access during the pandemic. This had psychological consequences for patients who are already within an intense emotional journey. Pandemic recovery plans should address mental health support as a priority.

Enhancer deregulation inTET2-mutant clonal hematopoiesis is associated with increased COVID-19 severity and mortality

Background: COVID-19 causes significant morbidity and mortality, albeit with considerable heterogeneity among affected individuals. It remains unclear which host factors determine disease severity and survival. Given the propensity of clonal hematopoiesis (CH) to promote inflammation in healthy individuals, we investigated its effect on COVID-19 outcomes. Method(s): We performed a multi-omics interrogation of the genome, epigenome, transcriptome, and proteome of peripheral blood mononuclear cells from COVID-19 patients (n=227). We obtained clinical data, laboratory studies, and survival outcomes. We determined CH status and TET2-related DNA methylation. We performed single-cell proteogenomics to understand clonal composition in relation to cell phenotype. We interrogated single-cell gene expression in isolation and in conjunction with DNA accessibility. We integrated these multi-omics data to understand the effect of CH on clonal composition, gene expression, methylation of cis-regulatory elements, and lineage commitment in COVID-19 patients. We performed shRNA knockdowns to validate the effect of one candidate transcription factor in myeloid cell lines. Result(s): The presence of CH was strongly associated with COVID-19 severity and all-cause mortality, independent of age (HR 3.48, 95% CI 1.45-8.36, p=0.005). Differential methylation of promoters and enhancers was prevalent in TET2-mutant, but not DNMT3A-mutant CH. TET2- mutant CH was associated with enhanced classical/intermediate monocytosis and single-cell proteogenomics confirmed an enrichment of TET2 mutations in these cell types. We identified celltype specific gene expression changes associated with TET2 mutations in 102,072 single cells (n=34). Single-cell RNA-seq confirmed the skewing of hematopoiesis towards classical and intermediate monocytes and demonstrated the downregulation of EGR1 (a transcription factor important for monocyte differentiation) along with up-regulation of the lncRNA MALAT1 in monocytes. Combined scRNA-/scATAC-seq in 43,160 single cells (n=18) confirmed the skewing of hematopoiesis and up-regulation of MALAT1 in monocytes along with decreased accessibility of EGR1 motifs in known cis-regulatory elements. Using myeloid cell lines for functional validation, shRNA knockdowns of EGR1 confirmed the up-regulation of MALAT1 (in comparison to wildtype controls). Conclusion(s): CH is an independent prognostic factor in COVID-19 and skews hematopoiesis towards monocytosis. TET2-mutant CH is characterized by differential methylation and accessibility of enhancers binding myeloid transcriptions factors including EGR1. The ensuing loss of EGR1 expression in monocytes causes MALAT1 overexpression, a factor known to promote monocyte differentiation and inflammation. These data provide a mechanistic insight to the adverse prognostic impact of CH in COVID-19.

COVID-associated pernio: Mechanistic insights to an abortive, seronegative SARS-CoV-2 infection

Introduction: SARS-CoV-2 replicates primarily in the airways but generates a systemic immune response mediated by Type I interferons (IFN-I). Pernio is a rare skin manifestation of disorders characterized by excessive IFN-I signalling. Although pernio increased in incidence during the pandemic, the relationship to SARS-CoV-2 remains controversial. Because of the pivotal nature of interferons in COVID-19 outcomes, pernio offers a window to investigate the biology underlying host resiliency to SARS-CoV-2 infection. Method(s): To further assess COVID-associated pernio, we characterized clinical samples from affected patients across 4 waves of the pandemic and investigated mechanistic feasibility in a rodent model. Patients were followed longitudinally with banking of blood and tissue. Golden hamsters were mock-treated or intra-nasally infected with SARS-CoV-2 and harvested at 3-and 30-days post-infection. Result(s): In affected tissue, immunophenotyping utilizing multiplex immunohistochemistry profiled a robust IFN-1 signature characterized by plasmacytoid dendritic cell activation. Viral RNA was detectable in a subset of cases using in situ hybridization for the SARS-CoV-2 S gene transcript. Profiling of the systemic immune response did not reveal a durable type 1 interferon signature. Consistent with previous literature, antibody and T-cell specific responses to SARS-CoV-2 were not detected. Nasopharyngeal SARS-CoV-2 inoculation in hamsters resulted in rapid dissemination of viral RNA and the generation of an IFN-I response that were both detectable in the paws of infected animals. Conclusion(s): Our data support a durable local IFN signature, with direct evidence of viral SARS-CoV-2 RNA in acral skin and suggest that COVID-associated pernio results from an abortive, seronegative SARS-CoV-2 infection.

Dnazymes as a Method for Nipah Henipavirus Detection

Intro: Deoxyribozymes (Dz) are short synthetic DNA oligonucleotides that catalyze the cleavage of a phosphodiester bond between nucleotides in the presence of divalent metal ions. The use of DNAzymes in the in vitro diagnostics increases the specificity and versatility of the analysis. Method(s): We took the well-studied Dz 10-23 with high catalytic activity as the basis of our system. The biosensor is divided into two fragments according to the binary probe principle (Dz1 and Dz2), which consist of target RNA binding sites, a fluorescent substrate (Fsub), and half of the Dz 10-23 catalytic center sequence. Assembly of the Dz 10-23 active center with subsequent Fsub cleavage and registration of a fluorescent signal is possible only if the target RNA is present in the sample. Finding(s): To assess the diagnostic potential of the biosensor, we measured FAM fluorescence in a solution containing synthetic RNA 35 nucleotides long (nip35) corresponding to the NiV target sequence, Fsub labeled with the FAM-BHQ1 and Dz_NiV pair. A mixture of Dz_NiV and Fsub was used as a control. The detection limit of the target RNA reached 5 nM, the signal development time was 30 minutes at a temperature of 37 C . Discussion(s): The specificity of Dz_NiV was evaluated in the presence of synthetic RNAs from six other RNA viruses of similar length: Hendra, Machupo, Sabia, Junin, Guanarito, and SARS-CoV. A fluorescent signal was recorded only in the presence of nip35 in the reaction mixture. The efficiency of Dz_NiV on a long fragment was tested using a plasmid with a cloned target sequence. The site is about 700 b.p. was amplified by PCR, followed by transcription. Conclusion(s): It was developed the highly specific biosensor Dz_NiV for the detection of Nipah virus RNA with a sensitivity limit of 5 nM at 37 C .Copyright © 2023

Self-Care for Rheumatoid Arthritis during the Covid-19 Pandemic: Transferability of Canadian and Uk Experiences

Background/Aims The COVID-19 pandemic posed unique challenges for people worldwide in self-caring for their rheumatoid arthritis (RA). COVID-19 also prompted global changes in public health (e.g., vaccination programs and mask wearing) and rheumatology services (e.g., integrating telehealth with in-person healthcare). To facilitate cross-country learning of how to support people with RA to self-care during and post-pandemic, better understanding of individuals' experiences of self-care in the context of changes in public health and 'integrated' healthcare is needed. Our study aimed to explore transferability in experiences of public health measures and telehealth during COVID- 19 among individuals with RA in Canada and the UK. Methods Between July and October 2022, online focus groups (90 mins) took place with participants living with RA in the UK. Participants were recruited via social media and professional networks (including Versus Arthritis). Each participant received a report >=7days before each focus group, with a request to review in advance. The report contained preliminary findings identified through reflexive thematic analysis of interviews (30-70 mins) with thirty-nine participants with RA in British Columbia, Canada (26-86 years;36 females) between December 2020 and 2021. Nine preliminary themes were identified across three topics: accessing telehealth and in-person healthcare;decision-making around COVID-19 vaccinations and public health measures;and renegotiating 'the self'. The themes guided the focus groups, wherein UK participants shared their perspectives on each theme arising in the Canadian context. Audio-visual recordings were transcribed verbatim, and transcripts were de-identified. Ongoing directed content analysis of focus group data involves a collaborative approach with patient partners. Results Four focus groups involving thirteen participants (44-81 years;11 females) living across the UK were conducted. Participants had lived with RA for between 3-36 years. Canadian experiences typically resonated with UK participants, with some feeling a sense of unity and sadness that challenges were also experienced by others living with RA beyond their national context. Many UK participants supported preferences expressed by Canadian participants for a 'hybrid' healthcare approach to maximise benefits and minimise downsides of telehealth, and in-person consultations post-pandemic. Benefits (e.g., avoiding risk of COVID-19 transmission with telehealth) and disadvantages (e.g., lacking sensitivity/accuracy of in-person assessments) described by Canadian participants also resonated with UKbased participants. Many described how their decision-making on adopting public health measures to maintain their self-care was supported and/or undermined in their local context/community. Conclusion Our findings offer novel insights into the challenges and opportunities experienced by people with RA in their decision-making around public health measures and telehealth during a global pandemic. They also demonstrate some transferability of experiences between the UK and Canada. Insights may serve to inform decision-making for policy and programmes to support self-care (e.g., by integrating telehealth into routine rheumatology practice) across countries during and postpandemic.