Backgrounds:

Multisystem Inflammatory Syndrome in Children (MIS-C) occurs several weeks after SARS-CoV-2 infection with symptoms including fever, shock and multiorgan failure. Clinical features of MIS-C overlap with Kawasaki Disease (KD), bacterial, and viral infections, making accurate diagnosis challenging. Host genes, measurable through whole blood transcriptomics, are an alternative tool for diagnosing infectious and inflammatory diseases.

Methods

Patients with MIS-C, KD, bacterial, and viral infections were recruited to the EU-funded PERFORM and DIAMONDS studies and the NIH-funded PREVAIL study. Patients were phenotyped using a standardised algorithm. Genome wide RNA sequencing of whole blood was undertaken, and feature selection was performed to identify a diagnostic signature for distinguishing between MIS-C and other infectious and inflammatory conditions. The expression levels of the genes identified were measured using RT-qPCR assays in an independent validation cohort.

Results:

Through feature selection and differential expression analysis, 11 genes with diagnostic potential were identified and taken forward into cross-platform validation using RT-qPCR. With up to 11 genes, it was possible to distinguish between MIS-C vs. KD, bacterial, and viral infections with high accuracy, with an AUC of 92.9% (95% CI: 88.2%-97.6%) in the validation cohort. The diagnostic gene signature retained its high performance when tested within the groups separately in the validation cohort: MIS-C vs. bacterial infections (AUC: 94.6%), vs. viral infections (AUC: 93.1%), and vs. KD (AUC: 89.8%).

Conclusions/Learning Points:

Despite the clinical similarities between MIS-C and other infectious and inflammatory conditions, there are key differences in gene expression profiles that can be used in diagnostic contexts. It will be necessary for the genes reported here to undergo further validation prior to their development into tests with clinical utility.

Backgrounds:

T cell lymphopaenia, and activation are well described features of MIS-C. We evaluated whether T cell exhaustion alongside T cell activation reported in childhood viral infections, occurs in children with a clinical diagnosis of MIS-C.

Methods

Clinical cohort: Children with febrile illnesses and healthy controls were recruited from two tertiary London hospitals between 01/09/2020-31/12/21, as part of the EU-funded DIAMONDS study. 162 participants were included: MIS-C (n=80), COVID-19 pneumonitis (n=7), Kawasaki disease (n=3), severe viral (n=7), and bacterial (n=19) illness, other inflammatory (n=8), paediatric controls (n=13), and adult vaccinated controls (n=25). Eighty-two of 124 patients (66%) required intensive care.

Samples were collected at two time-points: acute illness at admission and convalescence. Qiagen SARS-CoV-2 QuantiFERON kits were used for stimulation of whole blood with mitogen and SARS-CoV-2 antigens, to stimulate CD4+ and CD8+ T cells, and subsequent measurement in the supernatant of IFNγ levels.

Results:

The QuantiFERON assay was performed on 132 samples (acute n=59, convalescence n=19, paediatric controls n=11, adult controls n=25). MIS-C patients had raised baseline IFNγ levels, with lower increase after stimulation with SARS-CoV-2 antigens compared to vaccinated adults. The absolute increase in IFNγ in response to mitogen was lower in MIS-C compared to vaccinated adults, healthy children or bacterial infection. There was recovery in mitogen response in MIS-C in convalescence (Figure 1).

Conclusions/Learning Points:

We report a high baseline IFNγ and low activation by SARS-CoV-2 peptides and mitogen, which suggests T cell exhaustion coexists with features of T cell activation in MIS-C. Further studies on molecular mechanisms of T cell exhaustion are warranted.