Wellcome Open Research

* There are many ways of analyzing the progress of an epidemic, but when it comes to short term forecasting, it is very hard to beat a simple time series regression model. These are good at allowing for the noise in day to day observations, extracting the trend and projecting it forward. * Our regression models are designed to exploit this, using the daily statistics released by PHE and NHSE. These strongly suggest that the tide has turned and that taking one day with the next, the national figures for deaths from this virus will now fall back noticeably, easing the pressure on the NHS and its staff. * There is still a huge range of uncertainty associated with any forecast. The model is currently predicting a total of 113,000 admissions to UK hospitals by the end of April and that 19,000 people will die from the virus in English hospitals by then. There is a 1 in 20 chance that the mortality figures could flatten out more quickly, with around 1,000 more deaths occurring by the end of April. However, there is the same risk that this figure continues to mount, rising to a total of 24,000 by the end of the month. On current trends, the number of deaths in the UK is likely to be 10% higher than the number in England. * Longer term, the impact of the virus will depend critically upon the likely relaxation of the current government strategy of suppression.

ObjectivesTo audit implementation of a local protocol for the treatment of vitamin D deficiency (VDD) among patients hospitalized for Coronavirus Disease 2019 (COVID-19), including an assessment of the prevalence of VDD in these patients, and of potential associations with disease severity and fatality. DesignThis was not a study or clinical trial, but rather a retrospective interim audit (Newcastle-upon-Tyne Hospitals Registration No. 10075) of a local clinical care pathway for hospitalized patients with COVID-19-related illness. The Information (Caldicott) Guardian permitted these data to be shared beyond the confines of our institution. SettingA large tertiary academic NHS Foundation Trust in the North East of England, UK, providing care to COVID-19 patients. ParticipantsOne hundred thirty-four hospitalized patients with documented COVID-19 infection. Main outcome measuresAdherence to local investigation and treatment protocol; prevalence of VDD, and relationship of baseline serum 25(OH)D with markers of COVID-19 severity and inpatient fatality versus recovery. Results55.8% of eligible patients received Colecalciferol replacement, albeit not always loaded as rapidly as our protocol suggested, and no cases of new hypercalcaemia occurred following treatment. Patients admitted to ITU were younger than those managed on medical wards (61.1 years {+/-} 11.8 vs. 76.4 years {+/-} 14.9, p<0.001), with greater prevalence of hypertension, and higher baseline respiratory rate, National Early Warning Score-2 and C-reactive protein level. While mean serum 25(OH)D levels were comparable [i.e. ITU: 33.5 nmol/L {+/-} 16.8 vs. Non-ITU: 48.1 nmol/L {+/-} 38.2, mean difference for Ln-transformed-25(OH)D: 0.14, 95% Confidence Interval (CI) (-0.15, 0.41), p=0.3], only 19% of ITU patients had 25(OH)D levels greater than 50 nmol/L vs. 39.1% of non-ITU patients (p=0.02). However, we found no association with fatality, potentially due to small sample size, limitations of no-trial data and, potentially, the prompt diagnosis and treatment of VDD. ConclusionsSubject to the inherent limitations of observational (non-trial) audit data, analysed retrospectively, we found that patients requiring ITU admission were more frequently vitamin D deficient than those managed on medical wards, despite being significantly younger. Larger prospective studies and/or clinical trials are needed to elucidate the role of vitamin D as a preventive and/or therapeutic strategy for mitigating the effects of COVID-19 infection in patients with VDD. What is already known on this topicO_LIVitamin D deficiency (VDD) is associated with increased risk for acute respiratory tract infections C_LIO_LIA link between VDD and severity of COVID-19 pathophysiology has been proposed C_LIO_LITwo recent (non-peer-reviewed) studies have reported crude associations between VDD in defined geographic populations and COVID-19 severity and mortality C_LI What this study addsO_LIThese data do not arise from a clinical study; rather from an audit of a local replacement protocol for VDD in COVID-19 inpatients in a large UK centre, which found a significantly higher prevalence of VDD among ITU patients compared to non-ITU patients, despite the ITU patients being significantly younger. C_LIO_LIPrompt treatment of VDD following a local protocol did not result in any adverse events, such as hypercalcaemia. C_LIO_LIWhilst by no means conclusive, these data suggest an important association between VDD and COVID-19 severity; hence our report of interim findings in advance of achieving completed outcomes (fatality vs. recovery) for all patients. C_LIO_LIThere is an urgent need for larger studies exploring vitamin D as a potential preventative measure and/or treatment of Covid-19-related illness among individuals with VDD. C_LI

During the current COVID-19 pandemic, it has been suggested that BAME background patients may be disproportionately affected compared to White but few detailed data are available. We took advantage of near real-time hospital data access and analysis pipelines to look at the impact of ethnicity in 1200 consecutive patients admitted between 1st March 2020 and 12th May 2020 to Kings College Hospital NHS Trust in London (UK). Our key findings are firstly that BAME patients are significantly younger and have different co-morbidity profiles than White individuals. Secondly, there is no significant independent effect of ethnicity on severe outcomes (death or ITU admission) within 14-days of symptom onset, after adjustment for age, sex and comorbidities.

IntroductionSince the start of the pandemic SARS-CoV-2 infection has most commonly been confirmed using reverse transcriptase polymerase chain reaction (RT-PCR), with results translated into a binary positive/negative outcomes. Previous studies have found that there is additional useful information in the level of the Cycle threshold (Ct value) of positive cases. Here we characterise variation in Ct values as a proxy for viral loads in more than 3 million test-positive COVID-19 cases in England with the aim of better quantifying the utility of such data. MethodsWe used individual N gene Ct values from symptomatic PCR positive (with Ct value less than 30) Pillar 2 cases in England who self-reported the date of symptom onset, and for whom age, reinfection status, variant status, and the number of vaccines received was available. Those with a positive test result more than 6 days after their reported symptom onset were excluded to mitigate the potential impact of recall bias. We used a generalised additive model, to estimate Ct values empirical mean Ct values for each strata of interest independently as well as to predict Ct values using a model that adjusted for a range of demographic and epidemiological covariates jointly. We present empirical Ct values and compare them to predicted mean Ct values. ResultsWe found that mean Ct values varied by vaccine status, and reinfection status with the number of vaccine doses having little apparent effect. Modelling Ct values as a smooth function of time since onset and other variables struggled to reproduce the individual variation in the data but did match the population-level variation over time relatively well with this being apparently dominated by large differences between variants. Other variation over time was also captured to some degree though their remained several periods where the model could not capture the empirical means with a potential explanation being epidemic phase bias. ConclusionsAnalysing a large dataset of routine Ct values from symptomatic COVID-19 cases in England we found variation based on time since symptom onset, vaccine status, age, and variant. Ct values were highest 1-3 days after symptom onset and differed most due to variant status. We found no clear correlation between previously estimated differences in intrinsic transmissibility and Ct values indicating that this is potentially mediated at least partly by factors other than viral load as estimated using Ct values. We found evidence that a model adjusting for a range of covariates could explain some of the population-level variation over time but systematically underestimated Ct values when incidence was increasing, and overestimated them when incidence was decreasing. This indicates the utility of Ct values from this data source as a tool for surveillance, potentially avoiding some of the biases of aggregated positive counts.

Confirmed Case Data have been widely cited during the current COVID-19 pandemic as an estimate of the spread of the SARS-CoV-2 virus. However, their central role in media, official reports and decision-making may be undeserved and misleading. Previously published Infection Fatality Rates were weighted by age structure in the 50 countries with more reported deaths to obtain country-specific rates. For each country, the number of infections up to the Infection Date (23 days ago = Incubation Period + Onset to Death period) and the present percentage of immune population were estimated using Infection Fatality Rate, the number of reported deaths (which is less prone to undersampling), and projecting back to Infection Date. We then estimated a Detection Index for each country as the percentage of estimated infections that confirmed cases represent. Assuming that detection remains constant after Infection Date, we estimated the number of deaths and the estimated percentage of the population of each country expected to be immune up to 23 days into the future. Estimated Infection Fatality Rates are higher in Europe. In most countries, confirmed cases currently represent less than 30% of estimated infections on Infection Date, and this value decreases with time. Countries with flat curves throughout the pandemic show the lowest immunity percentages and these values seem unlikely to change in the near future, suggesting that they remain vulnerable to new outbreaks. Estimates for some countries with low Infection Fatality Rates suggest a still steep increase in the number of casualties in the next three weeks. Countries that did not control initial outbreaks seem to have reached higher immunity percentages, although mostly still under 5%. We provide the code to monitor the trajectories of these estimates in 178 countries throughout the COVID-19 pandemic.

Background: Sexually transmitted infections (STIs) and vaginal dysbiosis during pregnancy are associated with adverse pregnancy outcomes. Mycoplasma genitalium is the most recent STI implicated but evidence remains limited. The objectives of this study were to investigate 1) the association between M. genitalium infection during pregnancy and gestational age at delivery, preterm birth, miscarriage or stillbirth, and low birth weight and 2) the interaction with vaginal dysbiosis. Methods: We conducted a prospective cohort study in East London, South Africa. We enrolled pregnant women at gestational age <27 weeks, confirmed by ultrasound. We tested vaginal samples using nucleic acid amplification tests for M. genitalium, Chlamydia trachomatis, Neisseria gonorrhoeae, Trichomonas vaginalis, other genital mycoplasmas and Candida spp. We defined vaginal dysbiosis using Gram-stain criteria as a Nugent score 4-10. We used quantile regression to compare the outcome in women with and without M. genitalium across the gestational age distribution, adjusting for prespecified sociodemographic and clinical characteristics and co-occurring organisms. Results: From April 1, 2021 to August 29, 2023, we enrolled 603 women, followed up 584 and obtained pregnancy outcomes for 560 (93%). Median age was 28 years (interquartile range, IQR 24, 33) and 27% of women were living with HIV. M. genitalium was detected in 44/584 (8%, 95% CI 6, 10%) and vaginal dysbiosis in 375/584 (64%) of women. Median gestational age at delivery was 39 weeks +0 days (IQR 37+4, 40+1) in women with and 39 weeks +0 days (37+4, 40+0) in those without M. genitalium. In multivariable models, associations were not observed for any adverse birth outcomes. There was no interaction between M. genitalium and vaginal dysbiosis. Discussion: M. genitalium in pregnancy was not associated with earlier gestational age at delivery or with other adverse birth outcomes. These findings do not support routine testing and treatment for M. genitalium in pregnancy.

This study examined the progress of MRC-funded researchers, with a focus on their success in securing subsequent research income. We report here results from compiling details of the research income of researchers that completed an MRC early career award between 2011 and 2015. Depending on the funding scheme 50-59 per cent of researchers spent on average more than 100,000 GBP per year from new grants secured within three years of finishing their MRC award. In each scheme around 30 per cent of researchers secured less than 50,000 GBP per year research funding in the three years following their MRC award. We apply the same approach to an analysis of progression for the MRC cadre of intramurally-supported Programme Leader Track researchers, and a sample of MRC research grant holders. We also supplement our results with a small number of interviews with researchers that held MRC New Investigator Research Grants. We suggest that these data confirm reasonably good progression rates, with MRC early career schemes demonstrably effective in supporting research careers. They confirm there is a bottleneck with some researchers taking several years to establish their research career, and that expansion of the research base will need balanced new investment across schemes and stages and cannot be driven through growth in early career schemes alone.

Microscopes are essential tools for discoveries on a scale invisible to the unaided human eye. The development of immuno-fluorescence followed by molecular biology techniques and fluorescent fusion proteins have revolutionised the use of optical microscopy in bioscience. The quality of the data produced is dependent upon the sample, its preparation and the instrument used. However, instruments can degrade over time without easily visible changes to the produced images and, in turn, negatively impacts results. By testing instruments and doing comparisons between results over time and between different instruments, problems can be highlighted and corrective action can be taken. Using small fluorescent beads the point spread function (PSF) of the microscope can be recorded and the image resolution measured. Beads were prepared in a concentration matched to the field of view size and dried onto coverslips and mounted on slides. The beads were then imaged as 3D Z-stacks of sufficient size to fully enclose the PSF of the system. This data was uploaded to OMERO and processed using OMERO-metrics, an OMERO plugin developed for this purpose. This paper summarizes the development of workflows and protocols to enable this process, presents the results obtained and demonstrates the detection of significant instrument issues.

Long-read RNA sequencing enables isoform-resolved transcriptomics, but library preparation introduces systematic biases that shape biological interpretation. We benchmarked Oxford Nanopores two protocols--PCR-cDNA and direct RNA--using SKMM2 myeloma cells stimulated with interleukin-6 (IL-6) and ERCC synthetic spike-ins. Direct RNA produced longer, higher-quality reads and more high-confidence isoforms, but showed pronounced 5' coverage loss. PCR-cDNA yielded shorter fragments with 3' underrepresentation, detecting more low-abundance transcripts at reduced confidence. Protocol-specific biases had major consequences: differential expression analysis revealed limited overlap in IL-6-responsive genes, and pathway enrichment was broader in direct RNA. At the isoform level, differential transcript usage was almost entirely protocol-specific, with case studies (e.g. RPL22L1, GRB2, RNF220) illustrating concordance and divergence. ERCC controls confirmed these biases as technical rather than biological. Together, our results show that while both methods provide accurate gene-level quantification, transcript-level conclusions depend critically on protocol choice, highlighting the need for careful selection in long-read transcriptomics.

IntroductionInfectious intestinal disease (IID) is a significant public health concern, with an estimated 17 million cases occurring annually in the UK. However, only a small proportion of cases in the community present to primary health care, and those that do may not be tested for the causative pathogen. The third study of IID in the community (IID3 study) will provide updated estimates of the incidence and aetiology of IID in the UK. This protocol describes the laboratory methods employed by the IID3 study in primary diagnostic and reference laboratories. Methods and analysisStool specimens obtained from symptomatic subjects enrolled in two cohorts recruited prospectively through Primary Care in the UK are tested for 21 pathogens using PCR-based methods, in two separate workflows at Liverpool Clinical Laboratories. The first workflow comprises EntericBio DX, Viral and Clostridium difficile assays (Serosep, Crawley, UK); and the second a panel of assays for detection of Clostridium perfringens, Aeromonas spp., Cyclospora cayetanensis and diarrhoeagenic Escherichia coli based on 3base(R) Technology (Genetic Signatures, Newtown, Australia). Specimens testing positive for bacterial pathogens are subjected to culture. Specimens positive for, and isolates of Salmonella, Campylobacter, Yersinia, Vibrio and Shigella species and diarrhoeagenic E. coli are referred to the Gastrointestinal Bacteria Reference Unit, UK Health Security Agency (UKHSA) for whole genome sequencing and antimicrobial susceptibility testing. The parasites Cryptosporidium, Giardia and Cyclospora are referred for genotyping to the national Cryptosporidium Reference Unit, Public Health Wales. Norovirus positive faecal specimens are referred for Sanger Sequencing-based genotyping to the Enteric Virus Unit, UKHSA. Ethics and disseminationFavourable ethical opinion was granted on the 10th of August 2022 by the East Midlands - Nottingham 1 Research Ethics Committee (IRAS ID 314268, REC Reference 22/EM/0130). The results will be disseminated to key stakeholders including the Food Standards Agency, participating general practices and to the academic community through publications and conference presentations. A real-time breakdown of pathogens identified in faecal samples could be viewed openly on the study Dashboard throughout the surveillance period. Article SummaryO_ST_ABSStrengths and limitations of this studyC_ST_ABS- A total of 5439 stool samples underwent molecular testing for a total of 21 organisms at the Liverpool Clinical Laboratories (LCL), with detailed pathogen characterisation at national reference laboratories. - A commercial Genetic Signatures assay was evaluated in a routine diagnostic setting using commercially-provided pathogen controls, reference isolates and clinical samples, enabling the inclusion of Aeromonas and ETEC PCR targets into the diagnostic panel. - Real-time referrals of PCR-reactive samples were made to UK Health Security Agency (UKHSA) regardless of bacterial culture results, and to Public Health Wales (PHW). - Norovirus PCR-reactive samples were referred to the Enteric Virus Unit at UKHSA for genotyping ahead of Norovirus becoming a notifiable infection on the 6th of April 2025, adding valuable information on Norovirus genotypes circulating in the general population. - A Biobank has been created for residual faecal specimens, forming a rich resource for future studies. - A major challenge was the lack of a harmonised UK-wide digital referral and reporting system, resulting in an inflated manual reporting workload for staff at LCL.

Several studies suggest a lower prevalence of smoking than expected among adults with coronavirus disease (COVID-19). We conducted logistic regression analyses of the UK Biobank prospective study of 0.5 million adults followed for an average of 11 years. Compared to women, men were more likely to be tested and to test positive. In sex-stratified analyses, current smokers had higher adjusted Odds Ratios (OR) for being tested (male OR 1.60, 95%CI 1.32-1.95 and female OR 1.50,1.21-.1.86). Current smokers were more slightly more likely than never smokers to test positive for COVID-19. Further examination of smoking as a risk factor for COVID-19 is required. These must take into account reverse causality, where smokers quit to avoid disease as well as prior diseases.

COVID-19 point prevalence PCR community testing allows disease burden estimation. In a sample of London residents, point prevalence decreased from 2.2% (95%CI 1.4;3.5) in early April (reflecting infection around lockdown implementation) to 0.2% (95%CI 0.03-1.6) in early May (reflecting infection 3-5 weeks into lockdown). Extrapolation from reports of confirmed cases suggest that 5-7.6% of total infections were confirmed by testing during this period. These data complement seroprevalence surveys improving the understanding of transmission in London.

Background Despite numerous randomized controlled trials (RCTs) and systematic reviews (SRs), current sepsis guidelines continue to issue only weak recommendations for corticosteroids. We examined the clinical scope, underlying study pools, and mortality conclusions of SRs evaluating corticosteroids for sepsis. Methods We conducted a meta-research study of SRs on corticosteroids in sepsis (2015 to 2025), extracting SR characteristics, mortality results, and included RCTs. Study-pool overlap was assessed using an SRxRCT inclusion matrix, Jaccard similarity (J), and hierarchical clustering. SRs and RCTs were classified according to standardized Population, Intervention, Comparison, Outcome (PICO) profiles. We explored discordance in short-term mortality conclusions among clinically comparable SRs and potential associations with study-pool composition, target populations, and methodological characteristics. Results Forty-two SRs including 121 unique RCTs were identified. More than half of pairwise SR comparisons shared no RCTs, and only three pairs showed high overlap (J>0.8). SRs addressing similar intervention and target population profiles frequently relied on different study pools. Among 38 SRs with short-term mortality meta-analyses, 15 (39%) reported benefit and 23 (61%) no evidence of effect. Discordance occurred exclusively among SRs evaluating broad, non-specific corticosteroid strategies; conclusions were consistent for hydrocortisone plus fludrocortisone (benefit) and hydrocortisone, ascorbic acid, and thiamine (no evidence of effect). SRs including sepsis +/- shock populations more frequently reported benefit than those restricted to septic shock (62% vs 22%), although estimates were imprecise. No single methodological or clinical factor consistently explained discordance. Conclusions SRs addressing apparently similar clinical questions frequently synthesized different underlying evidence bases and reported discordant conclusions. Guideline developers should therefore consider not only methodological quality and reported PICO, but also whether the RCTs included in an SR adequately represent the intended clinical question. Clinically coherent evidence syntheses may improve the interpretability of pooled treatment effects and support more targeted corticosteroid therapy in sepsis.

Superbugs: A Pop-up Science Shop was a public engagement event in the school summer holidays of 2019, organised by members of Cardiff Universitys School of Medicine. We transformed an empty retail unit in the centre of Wales largest shopping centre into an interactive and immersive microbiology experience. We aimed to facilitate opportunities for two-way dialogue to impart positive impact on the awareness of antibiotic resistance, whilst concurrently evaluating the efficacy of an engagement strategy focused on the utilisation of public spaces to attract public demographics diverse to those who would normally engage with STEM outreach. Over the course of 14 days, we welcomed 6,566 visitors, with 67% attending as part of the natural footfall of the shopping centre. We created 1,625 young Antibiotic Resistance Champions, located in over 200 schools. Through a multi-lateral evaluation strategy, we were able to collect quantitative and qualitative feedback on the success of our delivery model, and the impact on our stakeholders. Herein, we will discuss the evolution of Superbugs from concept, planning and design, to the logistics of delivering an engagement event of this scale. We will focus in particular on the learning outcomes of the project, and how this will shape the future of our Superbugs project, and engagement events beyond. Key MessagesO_LICreating a multi-disciplinary core team is essential to the success of large-scale engagement events as well as the support and development of large numbers of colleagues/volunteers C_LIO_LIUtilising themes of exhibition and gameplay alongside strong fear-empowerment messages is an impactful way to confer positive influence and behaviour around antimicrobial resistance (AMR) and the use of antibiotics C_LIO_LI Pop-up shop is an effective mode of delivery to capture diverse public demographics far beyond those who would traditionally engage with scientific outreach and science engagement. C_LI

The COVID-19 pandemic has spread to almost every country in the world since it started in China in late 2019. Controlling the pandemic requires a multifaceted approach including whole genome sequencing to support public health interventions at local and national levels. One of the most widely used methods for sequencing is the ARTIC protocol, a tiling PCR approach followed by Oxford Nanopore sequencing (ONT) of up to 96 samples at a time. There is a need, however, for a flexible, platform agnostic, method that can provide multiple throughput options depending on changing requirements as the pandemic peaks and troughs. Here we present CoronaHiT, a method capable of multiplexing up to 96 small genomes on a single MinION flowcell or >384 genomes on Illumina NextSeq, using transposase mediated addition of adapters and PCR based addition of barcodes to ARTIC PCR products. We demonstrate the method by sequencing 95 and 59 SARS-CoV-2 genomes for routine and rapid outbreak response runs, respectively, on Nanopore and Illumina platforms and compare to the standard ARTIC LoCost nanopore method. Of the 154 samples sequenced using the three approaches, genomes with [&ge;] 90% coverage (GISAID criteria) were generated for 64.3% of samples for ARTIC LoCost, 71.4% for CoronaHiT-ONT, and 76.6% for CoronaHiT-Illumina and have almost identical clustering on a maximum likelihood tree. In conclusion, we demonstrate that CoronaHiT can multiplex up to 96 SARS-CoV-2 genomes per MinION flowcell and that Illumina sequencing can be performed on the same libraries, which will allow significantly higher throughput. CoronaHiT provides increased coverage for higher Ct samples, thereby increasing the number of high quality genomes that pass the GISAID QC threshold. This protocol will aid the rapid expansion of SARS-CoV-2 genome sequencing globally, to help control the pandemic.

Estimates of population prevalence and genetic diversity of bloodborne viruses in healthy humans are essential to support population-scale monitoring for transfusion transmission risk. In the UK and globally, blood donations are routinely screened for a limited number of high-consequence pathogens, but the full composition of the plasma virome remains to be characterised. Using a novel quantitative targeted metagenomics sequencing approach, we analysed previously unscreened plasma donations collected by NHS Blood and Transplant in England for all major pathogenic and known commensal human bloodborne viruses, and quantified their viral burden. Here we show that in a representative sample of 5,064 UK blood donors in pools of 24 collected over a one-month period, the virome was dominated by a small number of largely persistent species, representing ~11% (12/106) of previously identified human bloodborne viruses. Anelloviruses (TTV, TTMV and TTMDV) was detected in 89.0% of pools, albeit at low read count inconsistent with measured anellovirus viral loads. In contrast, human pegivirus type 1 (HPgV-1), had estimated population prevalence of 3.7% (95% CI 3.0-4.4%), with high read count and complete genome recovery in around one half of positive pools, consistent with high titre in plasma. Estimated prevalences for less common detections included one species of gemykibovirus (0.12%), hepatitis C virus (genotype 1a, 0.04%) and various polyomaviruses and herpesviruses between 0.04% (parvovirus 4, BK polyomavirus) and 0.41% (human herpesvirus 6). Phylogenetic analyses revealed mixed TTV, TTMV and TTMDV populations and almost exclusively genotype 2 HPgV-1, consistent with known genotype distributions in Europe. Our results provide a baseline for describing the healthy plasma virome in UK blood donors.

Practical courses mimic experimental research and may generate valuable data. Yet, data that is generated by students during a course is often lost as there is no centrally organized collection and storage of the data. The loss of data prevents its re-use. To provide access to these data, I present an approach that I call studentsourcing. It collects, aggregates and reuses data that is generated by students in a practical course on cell biology. The course runs annually, and I have recorded the data that was generated by >100 students over 3 years. Two use cases illustrate how the data can be aggregated and re-used either for the scientific record or for teaching. As the data is obtained by different students, in different groups, over different years, it is an excellent opportunity to discuss experimental design and modern data visualization methods such as the superplot. The first use case demonstrates how the data can be presented as an online, interactive dashboard, providing real-time data of the measurements. The second use case shows how central data storage provides a unique opportunity to get precise quantitative data due to the large sample size. Both use cases illustrate how data can be effectively aggregated and re-used. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=169 SRC="FIGDIR/small/561479v2_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@13c8973org.highwire.dtl.DTLVardef@1641e47org.highwire.dtl.DTLVardef@1fff83eorg.highwire.dtl.DTLVardef@2c1ef_HPS_FORMAT_FIGEXP M_FIG C_FIG

IntroductionSeveral viral respiratory infections - notably influenza - are associated with secondary bacterial infection and additional pathology. The extent to which this applies for COVID-19 is unknown. Accordingly, we aimed to define the bacteria causing secondary pneumonias in COVID-19 ICU patients using the FilmArray Pneumonia Panel, and to determine this tests potential in COVID-19 management. MethodsCOVID-19 ICU patients with clinically-suspected secondary infection at 5 UK hospitals were tested with the FilmArray at point of care. We collected patient demographic data and compared FilmArray results with routine culture. ResultsWe report results of 110 FilmArray tests on 94 patients (16 had 2 tests): 69 patients (73%) were male, the median age was 59 yrs; 92 were ventilated. Median hospital stay before testing was 14 days (range 1-38). Fifty-nine (54%) tests were positive, with 141 bacteria detected. Most were Enterobacterales (n=55, including Klebsiella spp. [n= 35]) or Staphylococcus aureus (n=13), as is typical of hospital and ventilator pneumonia. Community pathogens, including Haemophilus influenzae (n=8) and Streptococcus pneumoniae (n=1), were rarer. FilmArray detected one additional virus (Rhinovirus/Enterovirus) and no atypical bacteria. Fewer samples (28 % vs. 54%) were positive by routine culture, and fewer species were reported per sample; Klebsiella species remained the most prevalent pathogens. ConclusionFilmArray had a higher diagnostic yield than culture for ICU COVID-19 patients with suspected secondary pneumonias. The bacteria found mostly were Enterobacterales, S. aureus and P. aeruginosa, as in typical HAP/VAP, but with Klebsiella spp. more prominent. We found almost no viral co-infection. Turnaround from sample to results is around 1h 15 min compared with the usual 72h for culture, giving prescribers earlier data to inform antimicrobial decisions.

BackgroundIn England, free testing for COVID-19 was widely available from early in the pandemic until 1 April 2022. Based on apparent differences in the rate of positive PCR tests at a single laboratory compared to the rest of the laboratory network, we hypothesised that a substantial number of UK PCR tests processed during September and October 2021 may have been incorrectly reported as negative, compared with the rest of the laboratory network. We investigate the epidemiological impact of this incident. MethodsWe estimate the additional number of COVID-19 cases that would have been reported had the sensitivity of the laboratory test procedure not dropped for the period 2 September to 12 October. In addition, by making comparisons between the most affected local areas and comparator populations, we estimate the number of additional infections, cases, hospitalisations and deaths that could have occurred as a result of increased transmission due to the misclassification of tests. ResultsWe estimate that around 39,000 tests may have been incorrectly classified during this period and, as a direct result of this incident, the most affected areas in the South West could have experienced between 6,000 and 34,000 additional reportable cases, with a central estimate of around 24,000 additional reportable cases. Using modelled relationships between key variables, we estimate that this central estimate could have translated to approximately 55,000 additional infections, which means that each incorrect negative test likely led to just over two additional infections. In those same geographical areas, our results also suggest an increased number of admissions and deaths. ConclusionThe incident is likely to have had a measurable impact on cases and infections in the affected areas in the South West of England.

The engagement of the SARS-CoV-2 spike protein with ACE2 is a critical step for viral entry to human cells and accordingly blocking this interaction is a major determinant of the efficacy of monoclonal antibody therapeutics and vaccine-elicited serum antibodies. The emergence of SARS-CoV-2 variants necessitates the development of adaptable assays that can be applied to assess the effectiveness of therapeutics. Through testing of a range of recombinant spike proteins, we have developed a cell based, ACE2/spike protein binding assay that characterises monoclonal anti-spike protein antibodies and neutralising antibodies in donor serum. The assay uses high-content imaging to quantify cell bound spike protein fluorescence. Using spike proteins from the original Wuhan SARS-CoV-2 virus, as well as the delta and omicron variants, we identify differential blocking activity of three monoclonal antibodies directed against the spike receptor binding domain. Importantly, biological activity in the spike binding assay translated to efficacy in a SARS-CoV-2 infection assay. Hence, the spike binding assay has utility to monitor anti-spike antibodies against the major known SARS-CoV-2 variants and is readily adaptable to quantify impact of antibodies against new and emerging SARS-CoV-2 variants.