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Hospitalized COVID-19 patients exhibit diverse immune responses during acute infection, which are associated with a wide range of clinical outcomes. However, understanding these immune heterogeneities and their links to various clinical complications, especially long COVID, remains a challenge. In this study, we performed unsupervised subtyping of longitudinal multi-omics immunophenotyping in over 1,000 hospitalized patients, identifying two critical subtypes linked to mortality or mechanical ventilation with prolonged hospital stay and three severe subtypes associated with timely acute recovery. We confirmed that unresolved systemic inflammation and T-cell dysfunctions were hallmarks of increased severity and further distinguished patients with similar acute respiratory severity by their distinct immune profiles, which correlated with differences in demographic and clinical complications. Notably, one critical subtype (SubF) was uniquely characterized by early excessive inflammation, insufficient anticoagulation, and fatty acid dysregulation, alongside higher incidences of hematologic, cardiac, and renal complications, and an elevated risk of long COVID. Among the severe subtypes, significant differences in viral clearance and early antiviral responses were observed, with one subtype (SubC) showing strong early T-cell cytotoxicity but a poor humoral response, slower viral clearance, and greater risks of chronic organ dysfunction and long COVID. These findings provide crucial insights into the complex and context-dependent nature of COVID-19 immune responses, highlighting the importance of personalized therapeutic strategies to improve both acute and long-term outcomes.

Chronic viral infections are ubiquitous in humans, with individuals harboring multiple latent viruses that can reactivate during acute illnesses. Recent studies have suggested that SARS-CoV-2 infection can lead to reactivation of latent viruses such as Epstein-Barr Virus (EBV) and cytomegalovirus (CMV), yet, the extent and impact of viral reactivation in COVID-19 and its effect on the host immune system remain incompletely understood. Here we present a comprehensive multi-omic analysis of viral reactivation of all known chronically infecting viruses in 1,154 hospitalized COVID-19 patients, from the Immunophenotyping Assessment in a COVID-19 Cohort (IMPACC) study, who were followed prospectively for twelve months. We reveal significant reactivation of Herpesviridae , Enteroviridae , and Anelloviridae families during acute stage of COVID-19 (0-40 days post-hospitalization), each exhibiting distinct temporal dynamics. We also show that viral reactivation correlated with COVID-19 severity, demographic characteristics, and clinical outcomes, including mortality. Integration of cytokine profiling, cellular immunophenotyping, metabolomics, transcriptomics, and proteomics demonstrated virus-specific host responses, including elevated pro-inflammatory cytokines (e.g. IL-6, CXCL10, and TNF), increased activated CD4+ and CD8+ T-cells, and upregulation of cellular replication genes, independent of COVID-19 severity and SARS-CoV-2 viral load. Notably, persistent Anelloviridae reactivation during convalescence (≥3 months post-hospitalization) was associated with Post-Acute Sequelae of COVID-19 (PASC) symptoms, particularly physical function and fatigue. Our findings highlight a remarkable prevalence and potential impact of chronic viral reactivation on host responses and clinical outcomes during acute COVID-19 and long term PASC sequelae. Our data provide novel immune, transcriptomic, and metabolomic biomarkers of viral reactivation that may inform novel approaches to prognosticate, prevent, or treat acute COVID-19 and PASC., Competing Interests: IMPACC Network Competing Interests The Icahn School of Medicine at Mount Sinai has filed patent applications relating to SARS-CoV-2 serological assays and NDV-based SARS-CoV-2 vaccines which list Florian Krammer as co-inventor. Mount Sinai has spun out a company, Kantaro, to market serological tests for SARS-CoV-2. Florian Krammer has consulted for Merck and Pfizer (before 2020), and is currently consulting for Pfizer, Seqirus, 3rd Rock Ventures, Merck and Avimex. The Krammer laboratory is also collaborating with Pfizer on animal models of SARS-CoV-2. Viviana Simon is a co-inventor on a patent filed relating to SARS-CoV-2 serological assays (the “Serology Assays”). Ofer Levy is a named inventor on patents held by Boston Children’s Hospital relating to vaccine adjuvants and human in vitro platforms that model vaccine action. His laboratory has received research support from GlaxoSmithKline (GSK) and is a co-founder of and advisor to Ovax, Inc. Charles Cairns serves as a consultant to bioMerieux and is funded for a grant from Bill & Melinda Gates Foundation. James A Overton is a consultant at Knocean Inc. Jessica Lasky-Su serves as a scientific advisor of Precion Inc. Scott R. Hutton, Greg Michelloti and Kari Wong are employees of Metabolon Inc. Vicki Seyfert-Margolis is a current employee of MyOwnMed. Nadine Rouphael reports grants or contracts with Merck, Sanofi, Pfizer, Vaccine Company, Quidel, Lilly and Immorna, and has participated on data safety monitoring boards for Moderna, Sanofi, Seqirus, Pfizer, EMMES, ICON, BARDA, Imunon, CyanVac and Micron. Nadine Rouphael has also received support for meetings/travel from Sanofi and Moderna and honoraria from Virology Education. Adeeb Rahman is a current employee of Immunai Inc. Steven Kleinstein is a consultant related to ImmPort data repository for Peraton. Nathan Grabaugh is a consultant for Tempus Labs and the National Basketball Association. Akiko Iwasaki is a consultant for 4BIO, Blue Willow Biologics, Revelar Biotherapeutics, RIGImmune, Xanadu Bio, Paratus Sciences. Monika Kraft receives research funds paid to her institution from NIH, ALA; Sanofi, Astra-Zeneca for work in asthma, serves as a consultant for Astra-Zeneca, Sanofi, Chiesi, GSK for severe asthma; is a co-founder and CMO for RaeSedo, Inc, a company created to develop peptidomimetics for treatment of inflammatory lung disease. Esther Melamed received research funding from Babson Diagnostics and honorarium from Multiple Sclerosis Association of America and has served on the advisory boards of Genentech, Horizon, Teva, and Viela Bio. Carolyn Calfee receives research funding from NIH, FDA, DOD, Roche-Genentech and Quantum Leap Healthcare Collaborative as well as consulting services for Janssen, Vasomune, Gen1e Life Sciences, NGMBio, and Cellenkos. Wade Schulz was an investigator for a research agreement, through Yale University, from the Shenzhen Center for Health Information for work to advance intelligent disease prevention and health promotion; collaborates with the National Center for Cardiovascular Diseases in Beijing; is a technical consultant to Hugo Health, a personal health information platform; cofounder of Refactor Health, an AI-augmented data management platform for health care; and has received grants from Merck and Regeneron Pharmaceutical for research related to COVID-19. Grace A McComsey received research grants from Redhill, Cognivue, Pfizer, and Genentech, and served as a research consultant for Gilead, Merck, Viiv/GSK, and Jenssen. Linda N. Geng received research funding paid to her institution from Pfizer, Inc.

There is currently a lack of biological tools to study the replication cycle and pathogenesis of SARS-CoV-2, the etiological agent of COVID-19. Repurposing the existing tools, including antibodies of SARS-CoV, is an effective way to accelerate the development of therapeutics for COVID-19. Here, we extensively characterized antibodies of the SARS-CoV structural proteins for their cross-reactivity, experimental utility, and neutralization of SARS-CoV-2. We assessed a total of 10 antibodies (six for Spike, two for Membrane, and one for Nucleocapsid and Envelope viral protein). We evaluated the utility of these antibodies against SARS-CoV-2 in a variety of assays, including immunofluorescence, ELISA, biolayer interferometry, western blots, and micro-neutralization. Remarkably, a high proportion of the antibodies we tested showed cross-reactivity, indicating a potentially generalizable theme of cross-reactivity between SARS-CoV and SARS-CoV-2 antibodies. These antibodies should help facilitate further research into SARS-CoV-2 basic biology. Moreover, our study provides critical information about the propensity of SARS-CoV antibodies to cross-react with SARS-CoV-2 and highlights its relevance in defining the clinical significance of such antibodies to improve testing and guide the development of novel vaccines and therapeutics.