Humoral responses following SARS-CoV-2 breakthrough infection in COVID-19 vaccinated individuals (preprint)

Numerous studies have shown that a prior SARS-CoV-2 infection can greatly enhance the antibody response to COVID-19 vaccination, with this so called "hybrid immunity" leading to greater neutralization breadth against SARS-CoV-2 variants of concern. However, little is known about how breakthrough infection (BTI) in COVID-19 vaccinated individuals will impact the magnitude and breadth of the neutralizing antibody response. Here, we compared neutralizing antibody responses between unvaccinated and COVID-19 vaccinated individuals (including both AZD1222 and BNT162b2 vaccinees) who have been infected with the delta (B.1.617.2) variant. Rapid production of Spike-reactive IgG was observed in the vaccinated group providing evidence of effective vaccine priming. Overall, potent cross-neutralizing activity against current SARS-CoV-2 variants of concern was observed in the BTI group compared to the infection group. This study provides important insights into population immunity where transmission levels remain high.

Increasing Rates of Intensive Care Unit Bloodstream Infections During the Second COVID-19 Pandemic Wave, Particularly Primary Endogenous Enterococcus faecium, with High Crude Mortality and Associated with Introduction of Immunomodulatory Therapy (preprint)

Background: Reports indicate that COVID-19 patients have more bloodstream infections (BSI) on the intensive care unit (ICU) potentially due to lapses in infection control practice or other factors.Methods: Retrospective single-site study of ICU-BSIs in mechanically ventilated (MV) COVID-19 patients during the first pandemic year. Clinical, demographic and laboratory data including targeted pathogen genome sequencing was analysed during first (March 13th - May 31st 2020) and second (October 1st 2020 - March 15th 2021) pandemic waves.Findings: There were 305 MV-ICU patients in wave one and 440 in wave two with peak occupancy of 113 and 155 patients, respectively. The BSI rate was higher during both waves than pre-pandemic, but more in wave-two than wave-one, particularly during the first 28 days on ICU (14.1 vs. 9.4/1000 bed days; p=0.03) and with E. faecium (3.57 vs. 0.47/1000 bed days; p=0.0067). 22/28 (77%) of E. faecium BSIs had no microbiologically definable focus and the high wave-two rate could not be explained by transmission. Wave-two BSI-patients received more corticosteroids and tocilizumab and had higher crude hospital mortality compared with non-BSI patients (41% vs. 21% p<0.0001), a phenomenon not seen in wave one (32% vs. 27% p=0.551). Interpretation: MV-COVID-19 patients had a greater BSI-burden in wave two associated with the emergence of primary endogenous E. faecium. Identifying mechanisms and causal links between admission COVID-19 disease severity, immunomodulation, BSI and death could help identify new approaches to improving outcomes for COVID-19 patients.

SARS-CoV-2 lineage B.1.1.7 is associated with greater disease severity among hospitalised women but not men (preprint)

Background Severe acute respiratory syndrome coronavirus 2 (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 16/11/2020 - 10/01/2021, 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. Results 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 hazard ratio (HR) for mortality of B.1.1.7 compared to other lineages was 1.01 (95% CI 0.79-1.28, P=0.94) and for ITU admission was 1.01 (95% CI 0.75-1.37, P=0.96). Analysis of sex-specific effects of B.1.1.7 identified increased risk of mortality (HR 1.30, 95% CI 0.95-1.78) and ITU admission (HR 1.82, 95% CI 1.15-2.90) in females infected with the variant but not males (mortality HR 0.82, 95% CI 0.61-1.10; ITU HR 0.74, 95% CI 0.52-1.04). Conclusions 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 to 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.

Antibody longevity and cross-neutralizing activity following SARS-CoV-2 wave 1 and B.1.1.7 infections (preprint)

As SARS-CoV-2 variants continue to emerge globally, a major challenge for COVID-19 vaccination is the generation of a durable antibody response with cross-neutralizing activity against both current and newly emerging viral variants. Cross-neutralizing activity against major variants of concern (B.1.1.7, P.1 and B.1.351) has been observed following vaccination, albeit at a reduced potency, but whether vaccines based on the Spike glycoprotein of these viral variants will produce a superior cross-neutralizing antibody response has not been fully investigated. Here, we used sera from individuals infected in wave 1 in the UK to study the long-term cross-neutralization up to 10 months post onset of symptoms (POS), as well as sera from individuals infected with the B.1.1.7 variant to compare cross-neutralizing activity profiles. We show that neutralizing antibodies with cross-neutralizing activity can be detected from wave 1 up to 10 months POS. Although neutralization of B.1.1.7 and B.1.351 is lower, the difference in neutralization potency decreases at later timepoints suggesting continued antibody maturation and improved tolerance to Spike mutations. Interestingly, we found that B.1.1.7 infection also generates a cross-neutralizing antibody response, which, although still less potent against B.1.351, can neutralize parental wave 1 virus to a similar degree as B.1.1.7. These findings have implications for the optimization of vaccines that protect against newly emerging viral variants.

First and second SARS-CoV-2 waves in inner London: A comparison of admission characteristics and the effects of the B.1.1.7 variant (preprint)

Introduction: A second wave of SARS-CoV-2 infection spread across the UK in 2020 linked with emergence of the more transmissible B.1.1.7 variant. The emergence of new variants, particularly during relaxation of social distancing policies and implementation of mass vaccination, highlights the need for real-time integration of detailed patient clinical data alongside pathogen genomic data. We linked clinical data with viral genome sequence data to compare patients admitted during the first and second waves of SARS-CoV-2 infection. Methods: Clinical, laboratory and demographic data from five electronic health record (EHR) systems was collected for all cases with a positive SARS-CoV-2 RNA test between March 13th 2020 and February 17th 2021. SARS-CoV-2 viral sequencing was performed using Oxford Nanopore Technology. Descriptive data are presented comparing cases between waves, and between cases of B.1.1.7 and non-B.1.1.7 variants. Results: There were 5810 SARS-CoV-2 RNA positive cases comprising inpatients (n=2341), healthcare workers (n=1549), outpatients (n=874), emergency department (ED) attenders not subsequently admitted (n=532), inter-hospital transfers (n=281) and nosocomial cases (n=233). There were two dominant waves of admissions starting from around March 13th and October 20th, both with a temporally aligned rise in nosocomial cases (n=96 in wave one, n=137 in wave two). 1470 SARS-CoV-2 isolates were successfully sequenced, including 216/838 (26%) admitted cases from wave one, 472/1503 (31%) admitted cases in wave two and 121/233 (52%) nosocomial cases. 400/472 (85%) of sequenced isolates from admitted cases in wave two were the B.1.1.7 variant. The first B.1.1.7 variant was identified on 15th November 2020 and increased rapidly to comprise almost all sequenced isolates in January 2021 (n=600/617, 97%). Females made up a larger proportion of admitted cases in wave two (47.2% vs 41.8%, p=0.012), and in those infected with the B.1.1.7 variant compared to non-B.1.1.7 variants (48.0% vs 41.8%, p=0.042). A diagnosis of frailty was less common in wave two (11.5% v 22.8%, p<0.001) and in the group infected with B.1.1.7 (14.5% v 22.4%, p=0.001). There was no difference in severity on admission between waves, as measured by hypoxia at admission (wave one: 64.3% vs wave two: 65.6%, p=0.658). However, a higher proportion of cases infected with the B.1.1.7 variant were hypoxic on admission compared to other variants (70.0% vs 62.5%, p=0.029). Conclusions: Automated EHR data extraction linked with SARS-CoV-2 genome sequence data provides valuable insight into the evolving characteristics of cases admitted to hospital with COVID-19. The proportion of cases with hypoxia on admission was greater in those infected with the B.1.1.7 variant, which supports evidence the B.1.1.7 variant is associated with more severe disease. The number of nosocomial cases was similar in both waves despite introduction of many infection control interventions before wave two, an observation requiring further investigation.

Comparative performance of SARS-CoV-2 lateral flow antigen tests demonstrates their utility for high sensitivity detection of infectious virus in clinical specimens (preprint)

Background: Rapid antigen lateral flow devices (LFDs) are set to become a cornerstone of SARS-CoV-2 mass community testing. However, their reduced sensitivity compared to PCR has raised questions of how well they identify infectious cases. Understanding their capabilities and limitations is therefore essential for successful implementation. To address this, we evaluated six commercial LFDs on the same collection of clinical samples and assessed their correlation with infectious virus culture and cycle threshold (Ct) values. Methods: A head-to-head comparison of specificities and sensitivities was performed on six commercial rapid antigen tests using combined nasal/oropharyngeal swabs, and their limits of detection determined using viral plaque forming units (PFU). Three of the LFDs were selected for a further study, correlating antigen test result with RT-PCR Ct values and positive viral culture in Vero-E6 cells. This included sequential swabs and matched serum samples obtained from four infected individuals with varying disease severities. Detection of antibodies was performed using an IgG/IgM Rapid Test Cassette, and neutralising antibodies by infectious virus assay. Finally, the sensitivities of selected rapid antigen LFTs were assessed in swabs with confirmed B.1.1.7 variant, currently the dominant genotype in the UK. Findings: Most of the rapid antigen LFDs showed a high specificity (>98%), and accurately detected 50 PFU/test (equivalent N1 Ct of 23.7 or RNA copy number of 3x106/ml). Sensitivities of the LFDs performed on clinical samples ranged from 65 to 89%. These sensitivities increased in most tests to over 90% for samples with Cts lower than 25. Positive virus culture was achieved for 57 out of 141 samples, with 80% of the positive cultures from swabs with Cts lower than 23. Importantly, sensitivity of the LFDs increased to over 95% when compared with the detection of infectious virus alone, irrespective of Ct. Longitudinal studies of PCR-positive samples showed that most of the tests identified all infectious samples as positive, but differences in test sensitivities can lead to missed cases in the absence of repeated testing. Finally, test performance was not impacted when re-assessed against swabs positive for the dominant UK variant B.1.1.7. Interpretation: In this comprehensive comparison of antigen LFD and virus infectivity, we demonstrate a clear relationship between Ct values, quantitative culture of infectious virus and antigen LFD positivity in clinical samples. Our data support regular testing of target groups using LFDs to supplement the current PCR testing capacity, to rapidly identify infected individuals in situations where they would otherwise go undetected.

Application of respiratory metagenomics for COVID-19 patients on the intensive care unit to inform appropriate initial antimicrobial treatment and rapid detection of nosocomial transmission (preprint)

Background. Clinical metagenomics (CMg) is being evaluated for translation from a research tool into routine diagnostic service, but its potential to significantly improve management of acutely unwell patients has not been demonstrated. The SARS-CoV-2 pandemic provides impetus to determine that benefit given increased risk of secondary infection and nosocomial transmission by multi-drug resistant (MDR) pathogens linked with expansion of critical care capacity. Methods. Prospective evaluation of CMg using nanopore sequencing was performed on 43 respiratory samples over 14 weeks from a cohort of 274 intubated patients across seven COVID-19 intensive care units. Results. Bacteria or fungi were cultured from 200 (73%) patients, with a predominance of Klebsiella spp. (31%) and C. striatum (7%) amongst other common respiratory pathogens. An 8 hour CMg workflow was 93% sensitive and 81% specific for bacterial identification compared to culture, and reported presence or absence of {beta}-lactam resistance genes carried by Enterobacterales that would modify initial guideline-recommended antibiotics in every case. CMg was also 100% concordant with quantitative PCR for detecting Aspergillus fumigatus (4 positive and 39 negative samples). Single nucleotide polymorphism (SNP)-typing using 24 hour sequence data identified an MDR-K. pneumoniae ST307 outbreak involving 4 patients and an MDR-C. striatum outbreak potentially involving 14 patients across three ICUs. Conclusion. CMg testing for ICU patients provides same-day pathogen detection and antibiotic resistance prediction that significantly improves initial treatment of nosocomial pneumonia and rapidly detects unsuspected outbreaks of MDR-pathogens.

Combined epidemiological and genomic analysis of nosocomial SARS-CoV-2 transmission identifies community social distancing as the dominant intervention reducing outbreaks (preprint)

Many healthcare facilities report SARS-CoV-2 outbreaks but transmission analysis is complicated by the high prevalence of infection and limited viral genetic diversity. The contribution of different vectors to nosocomial infection or the effectiveness of interventions is therefore currently unclear. Detailed epidemiological and viral nanopore sequence data were analysed from 574 consecutive patients with a PCR positive SARS-CoV-2 test between March 13th and March 31st, when the pandemic first impacted on a large, multisite healthcare institution in London. During this time the first major preventative interventions were introduced, including progressive community social distancing (CSD) policies leading to mandatory national lockdown, exclusion of hospital visitors, and introduction of universal surgical facemask-use by healthcare-workers (HCW). Incidence of nosocomial cases, community SARS-CoV-2 cases and infection in a cohort of 228 HCWs followed the same dynamic course, decreasing shortly after introduction of CSD measures and prior to the main hospital-based interventions. We investigated clusters involving nosocomial cases based on overlapping ward-stays during the 14-day incubation period and SARS-CoV-2 genome sequence similarity. Our method placed 63 (79%) of 80 sequenced probable and definite nosocomial cases into 14 clusters containing a median of 4 patients (min 2, max 19) No genetic support was found for the majority of epidemiological clusters (31/44, 70%) and genomics revealed multiple contemporaneous outbreaks within single epidemiological clusters. We included a measure of hospital enrichment compared to community cases to increase confidence in our clusters, which were 1-14 fold enriched. Applying genomics, we could provide a robust estimate of the incubation period for nosocomial transmission, with a median lower bound and upper bound of 6 and 9 days respectively. Six (43%) clusters spanned multiple wards, with evidence of cryptic transmission, and community-onset cases could not be identified in more than half the clusters, particularly on the elective hospital site, implicating HCW as vectors of transmission. Taken together these findings suggest that CSD had the dominant impact on reducing nosocomial transmission by reducing HCW infection.