Comparison between enzyme-linked immunospot assay and intracellular cytokine flow cytometry assays for the evaluation of T cell response to SARS-CoV-2 after symptomatic COVID-19.

INTRODUCTION: Evaluation of different cell-based assays for the study of adaptive immune responses against SARS-CoV-2 is crucial for studying long-term and vaccine-induced immunity. METHODS: Enzyme-linked immunospot assay (ELISpot) and intracellular cytokine staining (ICS) using peptide pools spanning the spike protein and nucleoprotein of SARS-CoV-2 were performed in 25 patients who recovered from paucisymptomatic (n = 19) or severe COVID-19 (n = 6). RESULTS: The proportion of paucisymptomatic patients with detectable SARS-CoV-2 T cells was low, as only 44% exhibit a positive T cell response with the ICS and 67% with the ELISpot. The magnitude of SARS-CoV-2 T cell responses was low, both with ICS (median at 0.12% among total T cells) and ELISpot (median at 61 SFCs/million peripheral blood mononuclear cells [PBMC]) assays. Moreover, T cell responses in paucisymptomatic patients seemed lower than among patients with severe disease. In the paucisymptomatic patients, the two assays were well correlated with 76% of concordant responses and a Cohen's kappa of 55. Furthermore, in four patients SARS-CoV-2 T cells were detected by ELISpot but not with ICS. Short-term culture could improve the detection of specific T cells. CONCLUSIONS: In patients who recovered from paucisymptomatic COVID-19, the proportion of detectable anti-SARS-CoV-2 responses and their magnitude seemed lower than in patients with more severe symptoms. The ELISpot appeared to be more sensitive than the ICS assay. Short-term culture revealed that paucisymptomatic patients had nonetheless few SARS-CoV-2 T cells at a very low rate in peripheral blood. These data indicate that various ex-vivo assays may lead to different conclusions about the presence or absence of SARS-CoV-2 T cell immunity.

Educational Setting and SARS-CoV-2 Transmission Among Children With Multisystem Inflammatory Syndrome: A French National Surveillance System.

Background: Multisystem inflammatory syndrome in children (MIS-C) is the most severe form associated with SARS-CoV-2 infection in children. To reduce the spread of SARS-CoV-2 at the population level, educational setting closure have been implemented in many countries. However, the direct benefit of school closure on the MIS-C burden remains to be explored. We aimed to assess the role of educational settings in SARS-CoV-2 transmission among children with MIS-C. Methods: We conducted a French national prospective surveillance of MIS-C, coordinated by Public Health France, from April 2020 to March 2021. During this period, we included all children with MIS-C fulfilling the WHO definition who were reported to Public Health France. For each child, we traced the source of SARS-CoV-2 transmission. The main outcome was the proportion of children with MIS-C, with educational setting-related SARS-CoV-2 infection, during the period of school opening. Results: We included 142 children fulfilling WHO criteria for MIS-C: 104 (70%) cases occurred during school opening periods. In total, 62/104 children (60%, 95%CI [50; 69]) had been contaminated by a household contact and 5/104 in educational settings (5%, 95%CI [2; 11]). Among children with MIS-C occurring during school closure periods, the proportion of household transmission remained similar (66%, 25/38). Conclusion: Children with MIS-C were mainly infected by SARS-CoV-2 within their family environment, and the educational setting played a marginal role in this transmission. This suggests that mitigating school attendance may not reduce substantially the burden of MIS-C.

Systematic Severe Acute Respiratory Syndrome Coronavirus 2 Screening at Hospital Admission in Children: A French Prospective Multicenter Study.

To assess the relevance of systematic severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) screening of all children admitted to hospital, we conducted a prospective multicenter study including 438 consecutive hospitalized children. A symptom-based SARS-CoV-2 testing strategy failed to identify 45% (95% confidence interval, 24%-68%) of hospitalized children infected by SARS-CoV-2. To limit intrahospital transmission, a systematic screening of children admitted to hospital should be considered.

Contribution of Serological Rapid Diagnostic Tests to the Strategy of Contact Tracing in Households Following SARS-CoV-2 Infection Diagnosis in Children.

Background: The contact tracing and isolation of contagious individuals are cornerstones in the control of the COVID-19 pandemic. Strategies to identify household contacts who should be isolated around index children that tested positive for SARS-CoV-2 remain to be clarified. We aimed to compare contact tracing strategies around an index child positive for SARS-CoV-2 using serological rapid diagnostic testing (RDT, chromatography immunoassay). Methods: We conducted a contact tracing study in households of index cases children in the Paris region, France, between May 8 and July 27, 2020. We compared two strategies, one using SARS-CoV-2 reverse transcriptase polymerase chain reaction (RT-PCR) and one combining RT-PCR and serological RDT, initiated once RDT was available. The contacts RT-PCR-/RDT+ were considered to have been previously infected and not requiring quarantine. The primary outcome was the proportion of contacts that could avoid quarantine with the two screening strategies. Results: We included 34 children as index cases. Median age was 7 years. They generated 184 contacts (111 adults, 73 children) tested by RT-PCR: 24/184 (13%) were positive. The strategy combining RDT and RT-PCR was performed in 120/184 contacts (77 adults, 43 children) of 26 index children: 16/120 (13%) were RT-PCR+ and 47/120 (39%) were RDT+. Among the 16 individuals who were RT-PCR+, 14 (87%) were also RDT+. Among the 104 individuals who were RT-PCR-, 33 were RDT+. Hence 33/120 (27%) individuals were not isolated. Conclusions: Following the diagnosis of SARS-CoV-2 infection in children, a strategy combining serological RDT and nasopharyngeal RT-PCR enabled us to identify around one fourth of contacts with past infection and avoid unnecessary quarantine of these individuals.

SARS-CoV-2 transmission among children and staff in daycare centres during a nationwide lockdown in France: a cross-sectional, multicentre, seroprevalence study.

BACKGROUND: The extent to which very young children contribute to the transmission of SARS-CoV-2 is unclear. We aimed to estimate the seroprevalence of antibodies against SARS-CoV-2 in daycare centres that remained open for key workers' children during a nationwide lockdown in France. METHODS: Children and staff who attended one of 22 daycare centres during a nationwide lockdown in France (between March 15 and May 9, 2020) were included in this cross-sectional, multicentre, seroprevalence study. Hospital staff not occupationally exposed to patients with COVID-19, or to children, were enrolled in a comparator group. The primary outcome was SARS-CoV-2 seroprevalence in children, daycare centre staff, and the comparator group. The presence of antibodies against SARS-CoV-2 in capillary whole blood was measured with a rapid chromatographic immunoassay. We computed raw prevalence as the percentage of individuals with a positive IgG or IgM test, and used Bayesian smoothing to account for imperfect sensitivity and specificity of the assay. This study is registered with ClinicalTrials.gov, NCT04413968. FINDINGS: Between June 4 and July 3, 2020, we enrolled 327 children (mean age 1·9 [SD 0·9] years; range 5 months to 4·4 years), 197 daycare centre staff (mean age 40 [12] years), and 164 adults in the comparator group (42 [12] years). Positive serological tests were observed for 14 children (raw seroprevalence 4·3%; 95% CI 2·6-7·1) and 14 daycare centre staff (7·7%; 4·2-11·6). After accounting for imperfect sensitivity and specificity of the assay, we estimated that 3·7% (95% credible interval [95% CrI] 1·3-6·8) of the children and 6·8% (3·2-11·5) of daycare centre staff had SARS-CoV-2 infection. The comparator group fared similarly to the daycare centre staff; nine participants had a positive serological test (raw seroprevalence 5·5%; 95% CI 2·9-10·1), leading to a seroprevalence of 5·0% (95% CrI 1·6-9·8) after accounting for assay characteristics. An exploratory analysis suggested that seropositive children were more likely than seronegative children to have been exposed to an adult household member with laboratory-confirmed COVID-19 (six [43%] of 14 vs 19 [6%] of 307; relative risk 7·1 [95% CI 2·2-22·4]). INTERPRETATION: According to serological test results, the proportion of young children in our sample with SARS-CoV-2 infection was low. Intrafamily transmission seemed more plausible than transmission within daycare centres. Further epidemiological studies are needed to confirm this exploratory hypothesis. FUNDING: Assistance Publique-Hôpitaux de Paris; Mairie de Paris, Conseil Départemental de Seine Saint Denis. TRANSLATIONS: For the French translation of the abstract see Supplementary Materials section.

Emergence of Kawasaki disease related to SARS-CoV-2 infection in an epicentre of the French COVID-19 epidemic: a time-series analysis.

BACKGROUND: Kawasaki disease is an acute febrile systemic childhood vasculitis, which is suspected to be triggered by respiratory viral infections. We aimed to examine whether the ongoing COVID-19 epidemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is associated with an increase in the incidence of Kawasaki disease. METHODS: We did a quasi-experimental interrupted time series analysis over the past 15 years in a tertiary paediatric centre in the Paris region, a French epicentre of the COVID-19 outbreak. The main outcome was the number of Kawasaki disease cases over time, estimated by quasi-Poisson regression. In the same centre, we recorded the number of hospital admissions from the emergency department (2005-2020) and the results of nasopharyngeal multiplex PCR to identify respiratory pathogens (2017-2020). These data were compared with daily hospital admissions due to confirmed COVID-19 in the same region, recorded by Public Health France. FINDINGS: Between Dec 1, 2005, and May 20, 2020, we included 230 patients with Kawasaki disease. The median number of Kawasaki disease hospitalisations estimated by the quasi-Poisson model was 1·2 per month (IQR 1·1-1·3). In April, 2020, we identified a rapid increase of Kawasaki disease that was related to SARS-CoV-2 (six cases per month; 497% increase [95% CI 72-1082]; p=0·0011), starting 2 weeks after the peak of the COVID-19 epidemic. SARS-CoV-2 was the only virus circulating intensely during this period, and was found in eight (80%) of ten patients with Kawasaki disease since April 15 (SARS-CoV-2-positive PCR or serology). A second peak of hospital admissions due to Kawasaki disease was observed in December, 2009 (six cases per month; 365% increase ([31-719]; p=0.0053), concomitant with the influenza A H1N1 pandemic. INTERPRETATION: Our study further suggests that viral respiratory infections, including SAR-CoV-2, could be triggers for Kawasaki disease and indicates the potential timing of an increase in incidence of the disease in COVID-19 epidemics. Health-care providers should be prepared to manage an influx of patients with severe Kawasaki disease, particularly in countries where the peak of COVID-19 has recently been reached. FUNDING: French National Research Agency.

Paediatric multisystem inflammatory syndrome temporally associated with SARS-CoV-2 mimicking Kawasaki disease (Kawa-COVID-19): a multicentre cohort.

BACKGROUND: Current data suggest that COVID-19 is less frequent in children, with a milder course. However, over the past weeks, an increase in the number of children presenting to hospitals in the greater Paris region with a phenotype resembling Kawasaki disease (KD) has led to an alert by the French national health authorities. METHODS: Multicentre compilation of patients with KD in Paris region since April 2020, associated with the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) ('Kawa-COVID-19'). A historical cohort of 'classical' KD served as a comparator. RESULTS: Sixteen patients were included (sex ratio=1, median age 10 years IQR (4·7 to 12.5)). SARS-CoV-2 was detected in 12 cases (69%), while a further three cases had documented recent contact with a quantitative PCR-positive individual (19%). Cardiac involvement included myocarditis in 44% (n=7). Factors prognostic for the development of severe disease (ie, requiring intensive care, n=7) were age over 5 years and ferritinaemia >1400 µg/L. Only five patients (31%) were successfully treated with a single intravenous immunoglobulin (IVIg) infusion, while 10 patients (62%) required a second line of treatment. The Kawa-COVID-19 cohort differed from a comparator group of 'classical' KD by older age at onset 10 vs 2 years (p<0.0001), lower platelet count (188 vs 383 G/L (p<0.0001)), a higher rate of myocarditis 7/16 vs 3/220 (p=0.0001) and resistance to first IVIg treatment 10/16 vs 45/220 (p=0.004). CONCLUSION: Kawa-COVID-19 likely represents a new systemic inflammatory syndrome temporally associated with SARS-CoV-2 infection in children. Further prospective international studies are necessary to confirm these findings and better understand the pathophysiology of Kawa-COVID-19. Trial registration number NCT02377245.