Unravelling the effect of winter holiday celebrations on SARS-CoV-2 transmission (preprint)

Public holidays have been associated with SARS-CoV-2 incidence surges, although a firm causal link remains to be established. This association is sometimes attributed to events where transmissions occur at a disproportionately high rate, known as superspreading events. Here, we describe a sudden surge in new cases with the Omicron BA.1 strain amongst higher education students in Belgium. Contact tracers classed most of these cases as likely or possibly infected on New Year's Eve, indicating a direct trigger by New Year celebrations. Using a combination of contact tracing and phylogenetic data, we show the limited role of superspreading events in this surge. Finally, the numerous simultaneous transmissions allowed a unique opportunity to determine the distribution of incubation periods of the Omicron strain. Overall, our results indicate that, even under social restrictions, a surge in transmissibility of SARS-CoV-2 can occur when holiday celebrations result in small social gatherings attended simultaneously and communitywide.

Longitudinal analysis of serum neutralization of SARS-CoV-2 Omicron BA.2, BA.4 and BA.5 in patients receiving monoclonal antibodies (preprint)

The emergence of novel Omicron lineages, such as BA.5, may impact the therapeutic efficacy of anti-SARS-CoV-2 neutralizing monoclonal antibodies (mAbs). Here, we evaluated the neutralization and ADCC activity of 6 therapeutic mAbs against Delta, BA.2, BA.4 and BA.5 isolates. The Omicron sub-variants escaped most of the antibodies but remained sensitive to Bebtelovimab and Cilgavimab. Consistent with their shared spike sequence, BA.4 and BA.5 displayed identical neutralization profiles. Sotrovimab was the most efficient at eliciting ADCC. We also analyzed 121 sera from 40 immunocompromised individuals up to 6 months after infusion of 1200 mg of Ronapreve (Imdevimab + Casirivimab), and 300 or 600 mg of Evusheld (Cilgavimab + Tixagevimab). Sera from Ronapreve-treated individuals did not neutralize Omicron subvariants. Evusheld-treated individuals neutralized BA.2 and BA.5, but titers were reduced by 41- and 130-fold, respectively, compared to Delta. A longitudinal evaluation of sera from Evusheld-treated patients revealed a slow decay of mAb levels and neutralization. The decline was more rapid against BA.5. Our data shed light on the antiviral activities of therapeutic mAbs and the duration of effectiveness of Evusheld pre-exposure prophylaxis.

Environmental circulation of adenovirus 40/41 and SARS-CoV-2 in the context of the emergence of acute hepatitis of unknown origin (preprint)

The recent surge of hepatitis of unknown origin in children is hypothesized to be caused by adenovirus 41 and/or SARS-CoV-2 infections. A relatively high proportion of patients testing positive for these viruses concomitantly with the development of acute hepatitis supports this hypothesis. To formally incriminate these viral infections as causative agents of hepatitis, both a plausible physiopathological pathway and supporting epidemiological dynamics in the community need demonstration. In this study, we measured the level of circulation of adenovirus 40/41 and SARS-CoV-2 in the general population of the city of Leuven in Belgium using wastewater monitoring between December 2020 and May 2022 and indoor air sampling in day care centers between November 2021 and May 2022. We also retrospectively analyzed medical records of 12.672 children attending a tertiary hospital draining the same region between January 2019 and April 2022. Our results demonstrate a recent but modest increase in hepatitis of unknown origin concomitant with a surge of circulating adenovirus 41 and SARS-CoV-2 in the general population, including in children under 5.

Early SARS-CoV-2 reinfections within 60 days highlight the need to consider antigenic variations together with duration of immunity in defining retesting policies (preprint)

The emergence of the SARS-CoV-2 Omicron variant, characterized by a significant antigenic diversity compared to the previous Delta variant, had led to a decrease in antibody efficacy in both convalescent and vaccinees sera resulting in high number of reinfections and breakthrough cases worldwide. However, to date, reinfections are defined by the ECDC as two positive tests >/= 60 days apart, influencing retesting policies after an initial positive test in several European countries. In our manuscript, we illustrate by a clinical case supplemental by epidemiological data that early reinfections do occur within 60 days especially in young, unvaccinated individuals. In older patient groups, unvaccinated and patients with a basic vaccination scheme are more vulnerable to reinfections compared to patients who received a first booster vaccine. For this reason, we consider that the duration of protection offered by a previous infection should be reconsidered, in particular when a shift between consecutive SARS-CoV-2 variants occurs.

Comprehensive immunovirological and environmental screening reveals risk factors for fatal COVID-19 during post-vaccination nursing home outbreaks (preprint)

COVID-19 vaccination has resulted in excellent protection against fatal disease, including in the elderly. However, risk factors for post-vaccination fatal COVID-19 are largely unknown. We comprehensively studied three large nursing home outbreaks (20-35% fatal cases) by combining SARS-CoV-2 aerosol monitoring, whole-genome phylogenetic analysis, and immunovirological profiling by digital nCounter transcriptomics. Phylogenetic investigations indicated each outbreak stemmed from a single introduction event, though with different variants (Delta, Gamma, and Mu). SARS-CoV-2 was detected in aerosol samples up to 52 days after the initial infection. Combining demographic, immune and viral parameters, the best predictive models for mortality comprised IFNB1 or age, viral ORF7a and ACE2 receptor transcripts. Comparison with published pre-vaccine fatal COVID-19 signatures and reanalysis of single-cell RNAseq data highlights the unique immune signature in post-vaccine fatal COVID-19 outbreaks. A multi-layered strategy including environmental sampling, immunomonitoring, and early antiviral therapy should be considered to prevent post-vaccination COVID-19 mortality in nursing homes.

Seroneutralization of Omicron BA.1 and BA.2 in patients receiving anti-SARS-CoV-2 monoclonal antibodies. (preprint)

The SARS-CoV-2 Omicron BA.1 variant has been supplanted in many countries by the BA.2 sub-lineage. BA.2 differs from BA.1 by about 21 mutations in its spike. Human anti-spike monoclonalantibodies(mAbs)areusedforpreventionortreatmentofCOVID-19. However, the capacity of therapeutic mAbs to neutralize BA.1 and BA.2 remains poorly characterized. Here, we first compared the sensitivity of BA.1 and BA.2 to neutralization by 9 therapeutic mAbs. In contrast to BA.1, BA.2 was sensitive to Cilgavimab, partly inhibited by Imdevimab and resistant to Adintrevimab and Sotrovimab. Two combinations of mAbs, Ronapreve (Casirivimab + Imdevimab) and Evusheld (Cilgavimab + Tixagevimab), are indicated as a pre-exposure prophylaxis in immunocompromised persons at risk of severe disease. We analyzed sera from 29 such individuals, up to one month after administration of Ronapreve and/or Evusheld. After treatment, all individuals displayed elevated antibody levels in their sera and neutralized Delta with high titers. Ronapreve recipients did not neutralize BA.1 and weakly impaired BA.2. With Evusheld, neutralization of BA.1 and BA.2 was detected in 19 and 29 out of 29 patients, respectively. As compared to Delta, titers were more severely decreased against BA.1 (344-fold) than BA.2 (9-fold). We further report 4 breakthrough Omicron infections among the 29 participants. Therefore, BA.1 and BA.2 exhibit noticeable differences in their sensitivity to therapeutic mAbs. Anti-Omicron activity of Ronapreve, and to a lesser extent that of Evusheld, is reduced in patients sera, a phenomenon associated with decreased clinical efficacy.

The role of airborne transmission in a large single source outbreak of SARS-CoV-2 in a Belgian nursing home in 2020 (preprint)

Abstract Objectives: To better understand the conditions which have led to one of the largest COVID-19 outbreaks in Belgian nursing homes in 2020. Setting: A nursing home in Flanders, Belgium, which experienced a massive outbreak of COVID-19 after a cultural event. An external volunteer who dressed as a legendary figure visited consecutively the 4 living units and tested positive for SARS-CoV-2 the next day. Within days, residents started to display symptoms and the outbreak spread rapidly within the nursing home. Methods: We interviewed key informants and collected standardized data from all residents retrospectively. A batch of 115 positive samples with a Ct value of <37 by qRT-PCR were analysed using whole-genome sequencing. Six months after the outbreak, ventilation assessment of gathering rooms in the nursing home was done using a tracer gas test with calibrated CO2 sensors. Results: Timeline of diagnoses and symptom onsets clearly pointed to the cultural event as the start of the outbreak, with the volunteer as index case. The genotyping of positive samples depicted the presence of one large cluster, suggesting a single source outbreak. The global attack rate among residents was 77% with a significant association between infection and presence at the event. Known risk factors such as short distance to or physical contact with the volunteer, and wearing of a mask during the event were not associated with early infection. The ventilation assessment showed a high background average CO2 level in four main rooms varying from 657 ppm to 846 ppm. Conclusions: Our investigation shows a rapid and widespread single source outbreak of SARS-CoV-2 in a nursing home, in which airborne transmission was the most plausible explanation for the massive intra-facility spread. Our results underscore the importance of ventilation and air quality for the prevention of future outbreaks in closed facilities.

Considerable escape of SARS-CoV-2 variant Omicron to antibody neutralization (preprint)

The SARS-CoV-2 Omicron variant was first identified in November 2021 in Botswana and South Africa 1,2 . It has in the meantime spread to many countries and is expected to rapidly become dominant worldwide. The lineage is characterized by the presence of about 32 mutations in the Spike, located mostly in the N-terminal domain (NTD) and the receptor binding domain (RBD), which may enhance viral fitness and allow antibody evasion. Here, we isolated an infectious Omicron virus in Belgium, from a traveller returning from Egypt. We examined its sensitivity to 9 monoclonal antibodies (mAbs) clinically approved or in development 3 , and to antibodies present in 90 sera from COVID-19 vaccine recipients or convalescent individuals. Omicron was totally or partially resistant to neutralization by all mAbs tested. Sera from Pfizer or AstraZeneca vaccine recipients, sampled 5 months after complete vaccination, barely inhibited Omicron. Sera from COVID-19 convalescent patients collected 6 or 12 months post symptoms displayed low or no neutralizing activity against Omicron. Administration of a booster Pfizer dose as well as vaccination of previously infected individuals generated an anti-Omicron neutralizing response, with titers 5 to 31 fold lower against Omicron than against Delta. Thus, Omicron escapes most therapeutic monoclonal antibodies and to a large extent vaccine-elicited antibodies.

Genetic diversity and evolution of SARS-CoV-2 in Belgium during the first wave outbreak (preprint)

SARS-CoV-2, the causative agent of COVID-19 was first detected in Belgium on 3rd February 2020, albeit the first epidemiological wave started in March and ended in June 2020. One year after the first epidemiological wave hit the country data analyses reveled the temporal and variant distribution of SARS-CoV-2 and its implication with Belgian epidemiological measures. In this study, 766 complete SARS-CoV-2 genomes of samples originating from the first epidemiological were sequenced to characterize the temporal and geographic distribution of the COVID-19 pandemic in Belgium through phylogenetic and variant analysis. Our analysis reveals the presence of the major circulating SARS-CoV-2 clades (G, GH and GR) and lineages circulating in Belgium at that time. Moreover, it contextualizes the density of SARS-CoV-2 cases over time with non-intervention measures taken to prevent the spread of SARS-CoV-2 in Belgium, specific international case imports and the functional implications of the most representative non-synonymous mutations present in Belgium between February to June 2020.

One Year of Laboratory Based COVID-19 Surveillance System in Belgium: Main Indicators and Performance of the Laboratories (preprint)

Background: With the spread of coronavirus disease 2019 (COVID-19), an existing national laboratory based surveillance system was adapted to daily monitor the epidemiological situation of SARS-CoV-2 in the Belgium by following the number of confirmed COVID-19 infections, the number of performed tests and the positivity ratio. We present these main indicators of the surveillance over a one-year period as well as the impact of the performance of the laboratories, regarding speed of processing the samples and reporting results, for surveillance.Methods: We describe the evolution of test capacity, testing strategy and the data collection methods during the first year of the epidemic in Belgium.Results: Between the 1th of March 2020 and the 28th of February 2021, 9,487,470 tests and 773,078 COVID-19 laboratory confirmed cases were reported. Two epidemic waves occurred, with a peak in April and October 2020. The capacity and performance of the laboratories improved continuously during 2020 resulting in a high level performance. Since the end of November 2020 90 to 95% of test results are reported at the latest the day after sampling was performed.Conclusions: Thanks to the effort of all laboratories a performant exhaustive national laboratory based surveillance system to monitor the epidemiological situation of SARS-CoV-2 was set up in Belgium in 2020. On top of expanding the number of laboratories performing diagnostics and significantly increasing the test capacity in Belgium, turnaround times between sampling and testing as well as reporting were optimized over the first year of this pandemic.

Cov-MS: a community-based template assay for clinical MS-based protein detection in Sars-Cov-2 patients (preprint)

Rising population density and global mobility are among the reasons why pathogens such as SARS-CoV-2, the virus that causes COVID-19, spread so rapidly across the globe. The policy response to such pandemics will always have to include accurate monitoring of the spread, as this provides one of the few alternatives to total lockdown. However, COVID-19 diagnosis is currently performed almost exclusively by Reverse Transcription Polymerase Chain Reaction (RT-PCR). Although this is efficient, automatable and acceptably cheap, reliance on one type of technology comes with serious caveats, as illustrated by recurring reagent and test shortages. We therefore developed an alternative diagnostic test that detects proteolytically digested SARS-CoV-2 proteins using Mass Spectrometry (MS). We established the Cov-MS consortium, consisting of fifteen academic labs and several industrial partners to increase applicability, accessibility, sensitivity and robustness of this kind of SARS-CoV-2 detection. This in turn gave rise to the Cov-MS Digital Incubator that allows other labs to join the effort, navigate and share their optimizations, and translate the assay into their clinic. As this test relies on viral proteins instead of RNA, it provides an orthogonal and complementary approach to RT-PCR, using other reagents that are relatively inexpensive and widely available, as well as orthogonally skilled personnel and different instruments. Data are available via ProteomeXchange with identifier PXD022550.