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IntroductionThe SARS-CoV-2 pandemic has put significant additional pressure on healthcare systems throughout the world. The identification of at-risk population beyond age, pre-existing medical conditions and socioeconomic status has been the subject of only a small part of the global COVID-19 research so far. To this day, more data is required regarding the association between HLA allele and red blood cell (RBC) antigens expression in regard to SARS-CoV-2 infection susceptibility and virus clearance capability, and COVID-19 susceptibility, severity, and duration. MethodsThe phenotypes for ABO and RhD, and the genotypes for 37 RBC antigens and HLA-A, B, C, DRB1, DQB1 and DPB1 were determined using high throughput platforms (Luminex and Next-generation Sequencing) in 90 Caucasian convalescent plasma donors. The results were compared to expected reference frequencies, local and international databases, and literature. ResultsThe AB group was significantly increased (1.5x, p=0.018) and a non-significant (2.2x, p=0.030) increase was observed for the FY*A allele frequency in the convalescent cohort (N=90) compared to reference frequencies. Some HLA alleles were found significantly overrepresented (HLA-B*44:02, C*05:01, DPB1*04:01, DRB1*04:01 and DRB1*07:01) or underrepresented (A*01:01, B51:01 and DPB1*04:02) in convalescent individuals compared to the local bone marrow registry population. ConclusionOur study of infection-susceptible but non-hospitalized Caucasian COVID-19 patients contributes to the global understanding of host genetic factors associated with SARS-CoV-2 infection susceptibility and severity of the associated disease.
RESUMO
The SARS-CoV-2 virus is the causing agent of the coronavirus disease 2019 (COVID-19) pandemic which is responsible for millions of deaths worldwide. The development of the humoral response to the virus has been the subject of intensive research and development. A flow cytometry-based assay using native full-length SARS-CoV-2 Spike protein expressed in 293T cells was recently proposed as a complementary seropositivity determination assay. The aim of our study was to further develop the flow cytometry assay for potential use as a confirmatory test and to standardize its parameters and results for reliable inter-laboratory use. We have optimized the protocol, established the Receiving Operating Characteristic (ROC) curve and tested reproducibility using pre-COVID plasma samples and convalescent, SARS-CoV-2 individual plasma samples. The flow-based assay was simplified and standardized by cultivating the 293T cells in suspension and expressing results in Mean Equivalent Soluble Fluorochrome (MESF) using an internal antibody positive control. The ROC curve was determined with an area under the curve (AUC) of 0.996 and the assay specificity and sensitivity were established at 100% and 97.7% respectively. Reproducibility was good as determined on multiple cytometers, on different days, and with data acquisition as far as 72h post-staining. The optimized and standardized assay could be used as a high throughput confirmation confirmatory assay in flow cytometry laboratories involved in serological testing.