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Introduction: The current diagnostic methods for SARS-CoV-2 rely on quantitative RT-PCR. However, the presence of viral RNA in samples does not necessarily reflect the presence of an infectious virus. Therefore, the reliable detection of infectious SARS-CoV-2 in clinical samples is necessary to limit viral transmission. Methods: We developed a flow cytometry-based surrogate assay (FlowSA), wherein the presence of infectious SARS-CoV-2 was detected using virus nucleocapsid-specific antibodies. Results: We showed that FlowSA allows the detection of a wide range of viral titers of multiple SARS-CoV-2 variants. Furthermore, the assay was successfully used to detect infectious SARS-CoV-2 in nasopharyngeal swabs from SARS-CoV-2 positive individuals, including those with high Ct values. Notably, FlowSA identified the presence of infectious SARS-CoV-2 in biological specimens that scored negative for cytopathic effect (CPE) in cell culture and would otherwise be considered negative. Conclusion: We propose that FlowSA can be adopted as an alternative to conventional CPE methods for viral diagnostics.

Background: Post-COVID-19 syndrome (PCS) remains a major health issue worldwide, while its pathophysiology is still poorly understood. Systemic oxidative stress (OS) may be involved in PCS, which is reflected by lower circulating free thiols (R–SH, sulfhydryl groups), as they are receptive to rapid oxidation by reactive species. This study aimed to investigate the longitudinal dynamics of serum R–SH after SARS-CoV-2 infection and its association with the development of PCS in individuals with mild COVID-19. Methods: Baseline serum R–SH concentrations were measured and compared between 135 non-hospitalized COVID-19 subjects and 82 healthy controls (HC). In COVID-19 subjects, serum R–SH concentrations were longitudinally measured during the acute disease phase (up to 3 weeks) and at 3, 6, and 12 months of follow-up, and their associations with relevant clinical parameters were investigated, including the development of PCS. Results: Baseline albumin-adjusted serum R–SH were significantly reduced in non-hospitalized COVID-19 subjects as compared to HC (p = 0.041), reflecting systemic OS. In mild COVID-19 subjects, trajectories of albumin-adjusted serum R–SH levels over a course of 12 months were longitudinally associated with the future presence of PCS 18 months after initial infection (b = −9.48, p = 0.023). Conclusion: Non-hospitalized individuals with COVID-19 show evidence of systemic oxidative stress, which is longitudinally associated with the development of PCS. Our results provide a rationale for future studies to further investigate the value of R–SH as a monitoring biomarker and a potential therapeutic target in the development of PCS.

Summary: Background: Lack of specific definitions of clinical characteristics, disease severity, and risk and preventive factors of post-COVID-19 syndrome (PCS) severely impacts research and discovery of new preventive and therapeutics drugs. Methods: This prospective multicenter cohort study was conducted from February 2020 to June 2022 in 5 countries, enrolling SARS-CoV-2 out- and in-patients followed at 3-, 6-, and 12-month from diagnosis, with assessment of clinical and biochemical features, antibody (Ab) response, Variant of Concern (VoC), and physical and mental quality of life (QoL). Outcome of interest was identification of risk and protective factors of PCS by clinical phenotype, setting, severity of disease, treatment, and vaccination status. We used SF-36 questionnaire to assess evolution in QoL index during follow-up and unsupervised machine learning algorithms (principal component analysis, PCA) to explore symptom clusters. Severity of PCS was defined by clinical phenotype and QoL. We also used generalized linear models to analyse the impact of PCS on QoL and associated risk and preventive factors. CT registration number: NCT05097677. Findings: Among 1796 patients enrolled, 1030 (57%) suffered from at least one symptom at 12-month. PCA identified 4 clinical phenotypes: chronic fatigue-like syndrome (CFs: fatigue, headache and memory loss, 757 patients, 42%), respiratory syndrome (REs: cough and dyspnoea, 502, 23%); chronic pain syndrome (CPs: arthralgia and myalgia, 399, 22%); and neurosensorial syndrome (NSs: alteration in taste and smell, 197, 11%). Determinants of clinical phenotypes were different (all comparisons p