SARS-CoV-2 surface and air contamination in an acute healthcare setting during the first and second pandemic waves.

BACKGROUND: Surfaces and air in healthcare facilities can be contaminated with SARS-CoV-2. In a previous study, we identified SARS-CoV-2 RNA on surfaces and air in our hospital during the 'first wave' of the COVID-19 pandemic (April 2020). AIM: To explore whether the profile of SARS-CoV-2 surface and air contamination had changed between April 2020 and January 2021. METHODS: A prospective, cross-sectional, observational study in a multisite London hospital. In January 2021, surface and air samples were collected from comparable areas to those sampled in April 2020 comprising six clinical areas and a public area. SARS-CoV-2 was detected using RT-PCR and viral culture. Sampling was additionally undertaken in two wards with only natural ventilation. The ability of the prevalent variants at the time of the study to survive on dry surfaces was evaluated. FINDINGS: No viable virus was recovered from surfaces or air. 5% (14) of 270 surfaces and 4% (1) of 27 air samples were positive for SARS-CoV-2, which was significantly lower than in April 2020 (52% (114) of 218 of surfaces and 48% (13) of 27 air samples (p<0.001, Fisher's Exact Test)). There was no clear difference in the proportion of surfaces and air samples positive for SARS-CoV-2 RNA based on the type of ventilation in the ward. All variants tested survived on dry surfaces for at least 72 hours with a <3-log10 reduction in viable count. CONCLUSION: Our study suggests that enhanced infection prevention measures have reduced the burden of SARS-CoV-2 RNA on surfaces and air in healthcare.

Third primary SARS-CoV-2 mRNA vaccines enhance antibody responses in most patients with haematological malignancies.

SARS-CoV-2 infection, and resulting disease, COVID-19, has a high mortality amongst patients with haematological malignancies. Global vaccine rollouts have reduced hospitalisations and deaths, but vaccine efficacy in patients with haematological malignancies is known to be reduced. The UK-strategy offered a third, mRNA-based, vaccine as an extension to the primary course in these patients. The MARCH database is a retrospective observational study of serological responses in patients with blood disorders. Here we present data on 381 patients with haematological malignancies. By comparison with healthy controls, we report suboptimal responses following two primary vaccines, with significantly enhanced responses following the third primary dose. These responses however are heterogeneous and determined by haematological malignancy sub-type and therapy. We identify a group of patients with continued suboptimal vaccine responses who may benefit from additional doses, prophylactic extended half-life neutralising monoclonal therapies (nMAB) or prompt nMAB treatment in the event of SARS-CoV-2 infection.

Early identification of high-risk individuals for combination monoclonal antibody therapy is feasible by SARS-CoV-2 anti-spike antibody specific lateral flow assay

BackgroundCombination monoclonal antibody therapy has recently been approved for prevention and treatment of severe COVID-19 infection in the UK. Available data suggests benefit is limited to those yet to mount an effective immune response from natural infection or vaccination, but concern exists around ability to make timely assessment of immune status of community-based patients.MethodsHealthcare workers were invited to undergo paired laboratory-based and rapid point-of- care (POC) lateral flow anti-spike antibody testing. Three commercial POC tests were selected to represent currently available testing methods: a split IgM/IgG anti-spike antibody test, an anti-receptor binding domain total antibody test and an anti-spike neutralisation assay. Qualitative POC colourmetric band intensities were independently scored and correlated with quantitative IgG neutralising antibody titres (Abbott Architect SARS-CoV-2 IgG Quant II chemiluminescent microparticle immunoassay [CMIA]). CMIA titres were correlated with the World Health Organisation international reference standard for neutralising antibody. Negative controls were carried out using 2018 pre-pandemic sera and post-pandemic individuals with negative CMIA results (target population).Results190 individuals (median 40 years, IQR 29-49;76.2% female) underwent paired testing, with a further 40 pre-pandemic sera tested. Assays demonstrated high performance characteristics: split IgM/IgG assay sensitivity 96.2% (95%CI 92.4.5–98.5), specificity 100.0% (95%CI 91.8–100.0);anti-receptor binding domain total antibody assay sensitivity 100.0% (95%CI 95.5–100.0), specificity 100.0% (95%CI 69.2–100.0). The neutralising antibody assay had a specificity of 97.0% (95%CI 84.2–99.9%) and strongly correlated with neutralising antibody titre (p<0.001). Probability for matched paired results was significant (McNemar’s p<0.001) while band intensity correlated strongly with neutralising titres (p<0.0001). Positive and negative predictive values for total antibody and neutralising assays were both >99%.ConclusionsPOC assays were found to be reliable predictors of both antibody status and broadly of neutralising antibody titre. Anti-S POC assays have potential to act as suitable alternatives for rapid identification of community patient immune status at presentation.

Clinical and survival differences during separate COVID-19 surges: Investigating the impact of the Sars-CoV-2 alpha variant in critical care patients.

A number of studies have highlighted physiological data from the first surge in critically unwell Covid-19 patients but there is a paucity of data describing emerging variants of SARS-CoV-2, such as B.1.1.7. We compared ventilatory parameters, biochemical and physiological data and mortality between the first and second COVID-19 surges in the United Kingdom, where distinct variants of SARS-CoV-2 were the dominant stain. We performed a retrospective cohort study investigating critically unwell patients admitted with COVID-19 across three tertiary regional ICUs in London, UK. Of 1782 adult ICU patients screened, 330 intubated and ventilated patients diagnosed with COVID-19 were included. In the second wave where B.1.1.7 variant was the dominant strain, patients were had increased severity of ARDS whilst compliance was greater (p<0.05) and d-dimer lower. The 28-day mortality was not statistically significant (1st wave: 42.2% vs 2nd wave: 39.8%). However, when adjusted for key covariates, the hazard ratio for 28-day mortality in those patients with B.1.1.7 was 3.79 (CI 1.04-13.8; p = 0.043) compared to the original strain. During the second surge in the UK, where the COVID-19 variant B.1.1.7 was most prevalent, significantly more patients presented to critical care with severe ARDS. Furthermore, mortality risk was significantly greater in our ICU population during the second wave of the pandemic in those patients with B.1.1.7. As ICUs are experiencing further waves (particularly by the delta (B.1.617.2) variant), we highlight the urgent need for prospective studies describing immunological and pathophysiological differences across novel emerging variants.

Tracking the incidence and risk factors for SARS-CoV-2 infection using historical maternal booking serum samples.

The early transmission dynamics of SARS-CoV-2 in the UK are unknown but their investigation is critical to aid future pandemic planning. We tested over 11,000 anonymised, stored historic antenatal serum samples, given at two north-west London NHS trusts in 2019 and 2020, for total antibody to SARS-CoV-2 receptor binding domain (anti-RBD). Estimated prevalence of seroreactivity increased from 1% prior to mid-February 2020 to 17% in September 2020. Our results show higher prevalence of seroreactivity to SARS-CoV-2 in younger, non-white ethnicity, and more deprived groups. We found no significant interaction between the effects of ethnicity and deprivation. Derived from prevalence, the estimated incidence of seroreactivity reflects the trends observed in daily hospitalisations and deaths in London that followed 10 and 13 days later, respectively. We quantified community transmission of SARS-CoV-2 in London, which peaked in late March / early April 2020 with no evidence of community transmission until after January 2020. Our study was not able to determine the date of introduction of the SARS-CoV-2 virus but demonstrates the value of stored antenatal serum samples as a resource for serosurveillance during future outbreaks.

Early identification of high-risk individuals for monoclonal antibody therapy and prophylaxis is feasible by SARS-CoV-2 anti-spike antibody specific lateral flow assay.

Monoclonal antibody therapy has been approved for prophylaxis and treatment of severe COVID-19 infection. Greatest benefit appears limited to those yet to mount an effective immune response from natural infection or vaccination, but concern exists around ability to make timely assessment of immune status of community-based patients where laboratory-based serodiagnostics predominate. Participants were invited to undergo paired laboratory-based (Abbott Architect SARS-CoV-2 IgG Quant II chemiluminescent microparticle immunoassay) and lateral flow assays (LFA; a split SARS-CoV-2 IgM/IgG and total antibody test) able to detect SARS-CoV-2 anti-spike antibodies. LFA band strength was compared with CMIA titer by log-linear regression. Two hundred individuals (median age 43.5 years, IQR 30-59; 60.5% female) underwent testing, with a further 100 control sera tested. Both LFA band strengths correlated strongly with CMIA antibody titers (P < 0.001). LFAs have the potential to assist in early identification of seronegative patients who may demonstrate the greatest benefit from monoclonal antibody treatment.

Assessing a novel, lab-free, point-of-care test for SARS-CoV-2 (CovidNudge): a diagnostic accuracy study.

BACKGROUND: Access to rapid diagnosis is key to the control and management of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Laboratory RT-PCR testing is the current standard of care but usually requires a centralised laboratory and significant infrastructure. We describe our diagnostic accuracy assessment of a novel, rapid point-of-care real time RT-PCR CovidNudge test, which requires no laboratory handling or sample pre-processing. METHODS: Between April and May, 2020, we obtained two nasopharyngeal swab samples from individuals in three hospitals in London and Oxford (UK). Samples were collected from three groups: self-referred health-care workers with suspected COVID-19; patients attending emergency departments with suspected COVID-19; and hospital inpatient admissions with or without suspected COVID-19. For the CovidNudge test, nasopharyngeal swabs were inserted directly into a cartridge which contains all reagents and components required for RT-PCR reactions, including multiple technical replicates of seven SARS-CoV-2 gene targets (rdrp1, rdrp2, e-gene, n-gene, n1, n2 and n3) and human ribonuclease P (RNaseP) as sample adequacy control. Swab samples were tested in parallel using the CovidNudge platform, and with standard laboratory RT-PCR using swabs in viral transport medium for processing in a central laboratory. The primary analysis was to compare the sensitivity and specificity of the point-of-care CovidNudge test with laboratory-based testing. FINDINGS: We obtained 386 paired samples: 280 (73%) from self-referred health-care workers, 15 (4%) from patients in the emergency department, and 91 (23%) hospital inpatient admissions. Of the 386 paired samples, 67 tested positive on the CovidNudge point-of-care platform and 71 with standard laboratory RT-PCR. The overall sensitivity of the point-of-care test compared with laboratory-based testing was 94% (95% CI 86-98) with an overall specificity of 100% (99-100). The sensitivity of the test varied by group (self-referred healthcare workers 94% [95% CI 85-98]; patients in the emergency department 100% [48-100]; and hospital inpatient admissions 100% [29-100]). Specificity was consistent between groups (self-referred health-care workers 100% [95% CI 98-100]; patients in the emergency department 100% [69-100]; and hospital inpatient admissions 100% [96-100]). Point of care testing performance was similar during a period of high background prevalence of laboratory positive tests (25% [95% 20-31] in April, 2020) and low prevalence (3% [95% 1-9] in inpatient screening). Amplification of viral nucleocapsid (n1, n2, and n3) and envelope protein gene (e-gene) were most sensitive for detection of spiked SARS-CoV-2 RNA. INTERPRETATION: The CovidNudge platform was a sensitive, specific, and rapid point of care test for the presence of SARS-CoV-2 without laboratory handling or sample pre-processing. The device, which has been implemented in UK hospitals since May, 2020, could enable rapid decisions for clinical care and testing programmes. FUNDING: National Institute of Health Research (NIHR) Imperial Biomedical Research Centre, NIHR Health Protection Research Unit in Healthcare Associated Infections and Antimicrobial Resistance at Oxford University in partnership with Public Health England, NIHR Biomedical Research Centre Oxford, and DnaNudge.

Third primary SARS-CoV-2 prophylactic mRNA vaccines enhances antibody responses in the majority of patients with haematological malignancies: Results from the MARCH study (preprint)

SARS-CoV-2 infection, and resulting disease, COVID-19, has a high mortality amongst patients with haematological malignancies. Global vaccine rollouts have successfully reduced hospitalisations and deaths, but the efficacy of vaccination in patients with haematological malignancies is known to be reduced. The UK-strategy offered a third, mRNA-based, vaccine as an extension to the primary course in these patients. Here we quantify serological responses following these vaccines in a cohort of 381 patients with haematological malignancies attending routine haematology outpatient clinics. By comparison with healthy controls, we report suboptimal responses following two primary vaccines, with significantly enhanced responses following the third primary dose. These responses however are heterogeneous and determined by haematological malignancy sub-type and therapy. We identify a group of patients with continued sub-optimal vaccine responses who may benefit from additional doses, as well as early intervention with monoclonal therapies in the event of developing SARS-CoV-2 infection.

Rapid design and implementation of an adaptive pooling workflow for SARS-CoV-2 testing in an NHS diagnostic laboratory: a proof-of-concept study.

Background: Diagnostic laboratories are currently required to provide routine testing of asymptomatic staff and patients as a part of their clinical screening for SARS-CoV-2 infection. However, these cohorts display very different disease prevalence from symptomatic individuals and testing capacity for asymptomatic screening is often limited. Group testing is frequently proposed as a possible solution to address this; however, proposals neglect the technical and operational feasibility of implementation in a front-line diagnostic laboratory. Methods: Between October and December 2020, as a seven-week proof of concept, we took into account scientific, technical and operational feasibility to design and implement an adaptive pooling strategy in an NHS diagnostic laboratory in London (UK). We assessed the impact of pooling on analytical sensitivity and modelled the impact of prevalence on pooling strategy. We then considered the operational constraints to model the potential gains in capacity and the requirements for additional staff and infrastructure. Finally, we developed a LIMS-agnostic laboratory automation workflow and software solution and tested the technical feasibility of our adaptive pooling workflow. Results: First, we determined the analytical sensitivity of the implemented SARS-CoV-2 assay to be 250 copies/mL. We then determined that, in a setting with limited analyser capacity, the testing capacity could be increased by two-fold with pooling, however, in a setting with limited reagents, this could rise to a five-fold increase. These capacity increases could be realized with modest additional resource and staffing requirements whilst utilizing up to 76% fewer plastic consumables and 90% fewer reagents. Finally, we successfully implemented a plate-based pooling workflow and tested 920 patient samples using the reagents that would usually be required to process just 222 samples. Conclusions: Adaptive pooled testing is a scientifically, technically and operationally feasible solution to increase testing capacity in frontline NHS diagnostic laboratories.

Development of a Multiplex Tandem PCR (MT-PCR) Assay for the Detection of Emerging SARS-CoV-2 Variants.

The emergence of variants of SARS-CoV-2 has created challenges for the testing infrastructure. Although large-scale genome sequencing of SARS-CoV-2 has facilitated hospital and public health responses, access to sequencing facilities globally is variable and turnaround times can be significant, so there is a requirement for rapid and cost-effective alternatives. Applying a polymerase chain reaction (PCR)-based single nucleotide polymorphism (SNP) approach enables rapid (<4 h) identification of SARS-CoV-2 lineages from nucleic acid extracts, through the presence or absence of a panel of defined of genomic polymorphisms. For example, the B.1.1.7 lineage ("UK", "Alpha", or "Kent" variant) is characterised by 23 mutations compared to the reference strain, and the most biologically significant of these are found in the S gene. We have developed a SARS-CoV-2 typing assay focused on five positions in the S gene (HV69/70, N501, K417, E484 and P681). This configuration can identify a range of variants, including all the "Variants of Concern" currently designated by national and international public health bodies. The panel has been evaluated using a range of clinical isolates and standardised control materials at four UK hospitals and shows excellent concordance with the known lineage information derived from full sequence analysis. The assay has a turnaround time of about three hours for a set of up to 24 samples and has been utilised to identify emerging variants in a clinical setting.

The Alpha variant was not associated with excess nosocomial SARS-CoV-2 infection in a multi-centre UK hospital study.

OBJECTIVES: Recently emerging SARS-CoV-2 variants have been associated with an increased rate of transmission within the community. We sought to determine whether this also resulted in increased transmission within hospitals. METHODS: We collected viral sequences and epidemiological data of patients with community and healthcare associated SARS-CoV-2 infections, sampled from 16th November 2020 to 10th January 2021, from nine hospitals participating in the COG-UK HOCI study. Outbreaks were identified using ward information, lineage and pairwise genetic differences between viral sequences. RESULTS: Mixed effects logistic regression analysis of 4184 sequences showed healthcare-acquired infections were no more likely to be identified as the Alpha variant than community acquired infections. Nosocomial outbreaks were investigated based on overlapping ward stay and SARS-CoV-2 genome sequence similarity. There was no significant difference in the number of patients involved in outbreaks caused by the Alpha variant compared to outbreaks caused by other lineages. CONCLUSIONS: We find no evidence to support it causing more nosocomial transmission than previous lineages. This suggests that the stringent infection prevention measures already in place in UK hospitals contained the spread of the Alpha variant as effectively as other less transmissible lineages, providing reassurance of their efficacy against emerging variants of concern.

SARS-CoV-2 lineage B.1.1.7 is associated with greater disease severity among hospitalised women but not men: multicentre cohort study.

BACKGROUND: SARS-CoV-2 lineage B.1.1.7 has been associated with an increased rate of transmission and disease severity among subjects testing positive in the community. Its impact on hospitalised patients is less well documented. METHODS: We collected viral sequences and clinical data of patients admitted with SARS-CoV-2 and hospital-onset COVID-19 infections (HOCIs), sampled 16 November 2020 to 10 January 2021, from eight hospitals participating in the COG-UK-HOCI study. Associations between the variant and the outcomes of all-cause mortality and intensive therapy unit (ITU) admission were evaluated using mixed effects Cox models adjusted by age, sex, comorbidities, care home residence, pregnancy and ethnicity. FINDINGS: Sequences were obtained from 2341 inpatients (HOCI cases=786) and analysis of clinical outcomes was carried out in 2147 inpatients with all data available. The HR for mortality of B.1.1.7 compared with other lineages was 1.01 (95% CI 0.79 to 1.28, p=0.94) and for ITU admission was 1.01 (95% CI 0.75 to 1.37, p=0.96). Analysis of sex-specific effects of B.1.1.7 identified increased risk of mortality (HR 1.30, 95% CI 0.95 to 1.78, p=0.096) and ITU admission (HR 1.82, 95% CI 1.15 to 2.90, p=0.011) in females infected with the variant but not males (mortality HR 0.82, 95% CI 0.61 to 1.10, p=0.177; ITU HR 0.74, 95% CI 0.52 to 1.04, p=0.086). INTERPRETATION: In common with smaller studies of patients hospitalised with SARS-CoV-2, we did not find an overall increase in mortality or ITU admission associated with B.1.1.7 compared with other lineages. However, women with B.1.1.7 may be at an increased risk of admission to intensive care and at modestly increased risk of mortality.

Longevity of SARS-CoV-2 immune responses in hemodialysis patients and protection against reinfection.

Patients with end stage kidney disease receiving in-center hemodialysis (ICHD) have had high rates of SARS-CoV-2 infection. Following infection, patients receiving ICHD frequently develop circulating antibodies to SARS-CoV-2, even with asymptomatic infection. Here, we investigated the durability and functionality of the immune responses to SARS-CoV-2 infection in patients receiving ICHD. Three hundred and fifty-six such patients were longitudinally screened for SARS-CoV-2 antibodies and underwent routine PCR-testing for symptomatic and asymptomatic infection. Patients were regularly screened for nucleocapsid protein (anti-NP) and receptor binding domain (anti-RBD) antibodies, and those who became seronegative at six months were screened for SARS-CoV-2 specific T-cell responses. One hundred and twenty-nine (36.2%) patients had detectable antibody to anti-NP at time zero, of whom 127 also had detectable anti-RBD. Significantly, at six months, 71/111 (64.0%) and 99/116 (85.3%) remained anti-NP and anti-RBD seropositive, respectively. For patients who retained antibody, both anti-NP and anti-RBD levels were reduced significantly after six months. Eleven patients who were anti-NP seropositive at time zero, had no detectable antibody at six months; of whom eight were found to have SARS-CoV-2 antigen specific T cell responses. Independent of antibody status at six months, patients with baseline positive SARS-CoV-2 serology were significantly less likely to have PCR confirmed infection over the following six months. Thus, patients receiving ICHD mount durable immune responses six months post SARS-CoV-2 infection, with fewer than 3% of patients showing no evidence of humoral or cellular immunity.

Author Correction: A role for Biofoundries in rapid development and validation of automated SARS-CoV-2 clinical diagnostics.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Humoral and T-cell responses to SARS-CoV-2 vaccination in patients receiving immunosuppression.

OBJECTIVE: There is an urgent need to assess the impact of immunosuppressive therapies on the immunogenicity and efficacy of SARS-CoV-2 vaccination. METHODS: Serological and T-cell ELISpot assays were used to assess the response to first-dose and second-dose SARS-CoV-2 vaccine (with either BNT162b2 mRNA or ChAdOx1 nCoV-19 vaccines) in 140 participants receiving immunosuppression for autoimmune rheumatic and glomerular diseases. RESULTS: Following first-dose vaccine, 28.6% (34/119) of infection-naïve participants seroconverted and 26.0% (13/50) had detectable T-cell responses to SARS-CoV-2. Immune responses were augmented by second-dose vaccine, increasing seroconversion and T-cell response rates to 59.3% (54/91) and 82.6% (38/46), respectively. B-cell depletion at the time of vaccination was associated with failure to seroconvert, and tacrolimus therapy was associated with diminished T-cell responses. Reassuringly, only 8.7% of infection-naïve patients had neither antibody nor T-cell responses detected following second-dose vaccine. In patients with evidence of prior SARS-CoV-2 infection (19/140), all mounted high-titre antibody responses after first-dose vaccine, regardless of immunosuppressive therapy. CONCLUSION: SARS-CoV-2 vaccines are immunogenic in patients receiving immunosuppression, when assessed by a combination of serology and cell-based assays, although the response is impaired compared with healthy individuals. B-cell depletion following rituximab impairs serological responses, but T-cell responses are preserved in this group. We suggest that repeat vaccine doses for serological non-responders should be investigated as means to induce more robust immunological response.

Real-world evaluation of COVID-19 lateral flow device (LFD) mass-testing in healthcare workers at a London hospital; a prospective cohort analysis.

OBJECTIVES: Real-world evaluation of the performance of the Innova lateral flow immunoassay antigen device (LFD) for regular COVID-19 testing of hospital workers. METHODS: This prospective cohort analysis took place at a London NHS Trust. 5076 secondary care healthcare staff participated in LFD testing from 18 November 2020 to21 January 2021. Staff members submitted results and symptoms via an online portal twice weekly. Individuals with positive LFD results were invited for confirmatory SARS CoV-2 PCR testing. The positive predictive value (PPV) of the LFD was measured. Secondary outcome measures included time from LFD result to PCR test and staff symptom profiles. RESULTS: 284/5076 individuals reported a valid positive LFD result, and a paired PCR result was obtained in 259/284 (91.2%). 244 were PCR positive yielding a PPV of 94.21% (244/259, 95% CI 90.73% to 96.43%). 204/259 (78.8%) staff members had the PCR within 36 hours of the LFD test. Symptom profiles were confirmed for 132/244 staff members (54.1%) with positive PCR results (true positives) and 13/15 (86.6%) with negative PCR results (false positives). 91/132 true positives (68.9%) were symptomatic at the time of LFD testing: 65/91 (71.4%) had symptoms meeting the PHE case definition of COVID-19, whilst 26/91 (28.6%) had atypical symptoms. 18/41 (43.9%) staff members who were asymptomatic at the time of positive LFD developed symptoms in the subsequent four days. 9/13 (76.9%) false positives were asymptomatic, 1/13 (7.7%) had atypical symptoms and 3/13 (23.1%) had symptoms matching the PHE case definition. CONCLUSIONS: The PPV of the Innova LFD is high when used amongst hospital staff during periods of high prevalence of COVID-19, yet we find frequent use by symptomatic staff rather than as a purely asymptomatic screening tool. LFD testing does allow earlier isolation of infected workers and facilitates detection of individuals whose symptoms do not qualify for PCR testing.