Microfluidic immuno-serology assay revealed a limited diversity of protection against COVID-19 in patients with altered immunity

The immune response to SARS-CoV-2 for patients with altered immunity such as hematologic malignancies and autoimmune disease may differ substantially from that in general population. These patients remain at high risk despite wide-spread adoption of vaccination. It is critical to examine the differences at the systems level between the general population and the patients with altered immunity in terms of immunologic and serological responses to COVID-19 infection and vaccination. Here, we developed a novel microfluidic chip for high-plex immuno-serological assay to simultaneously measure up to 50 plasma or serum samples for up to 50 soluble markers including 35 plasma proteins, 11 anti-spike/RBD IgG antibodies spanning all major variants, and controls. Our assay demonstrated the quintuplicate test in a single run with high throughput, low sample volume input, high reproducibility and high accuracy. It was applied to the measurement of 1,012 blood samples including in-depth analysis of sera from 127 patients and 21 healthy donors over multiple time points, either with acute COVID infection or vaccination. The protein association matrix analysis revealed distinct immune mediator protein modules that exhibited a reduced degree of diversity in protein-protein cooperation in patients with hematologic malignancies and patients with autoimmune disorders receiving B cell depletion therapy. Serological analysis identified that COVID infected patients with hematologic malignancies display impaired anti-RBD antibody response despite high level of anti-spike IgG, which could be associated with limited clonotype diversity and functional deficiency in B cells and was further confirmed by single-cell BCR and transcriptome sequencing. These findings underscore the importance to individualize immunization strategy for these high-risk patients and provide an informative tool to monitor their responses at the systems level.

Use of whole genome sequencing to estimate the contribution of immune evasion and waning immunity to decreasing COVID-19 vaccine effectiveness during alpha and delta variant waves

BackgroundThe decline in COVID-19 mRNA vaccine effectiveness (VE) is well established, however the impact of variant-specific immune evasion and waning protection remains unclear. Here, we use whole-genome-sequencing (WGS) to tease apart the contribution of these factors on the decline observed following the introduction of the Delta variant. Further, we evaluate the utility of calendar-period-based variant classification as an alternative to WGS. MethodsWe conducted a test-negative-case-control study among people who received SARS-CoV-2 RT-PCR testing in the Yale New Haven Health System between April 1 and August 24, 2021. Variant classification was performed using WGS and secondarily by calendar-period. We estimated VE as one minus the ratio comparing the odds of infection among vaccinated and unvaccinated people. ResultsOverall, 2,029 cases (RT-PCR positive, sequenced samples) and 343,985 controls (negative RT-PCRs) were included. VE 14-89 days after 2nd dose was significantly higher against WGS-classified Alpha infection (84.4%, 95% confidence interval: 75.6-90.0%) than Delta infection (68.9%, CI: 58.0-77.1%, p-value: 0.013). The odds of WGS-classified Delta infection were significantly higher 90-149 than 14-89 days after 2nd dose (Odds ratio: 1.6, CI: 1.2-2.3). While estimates of VE against calendar-period-classified infections approximated estimates against WGS-classified infections, calendar-period-based classification was subject to outcome misclassification (35% during Alpha period, 4% during Delta period). ConclusionsThese findings suggest that both waning protection and variant-specific immune evasion contributed to the lower effectiveness. While estimates of VE against calendar-period-classified infections mirrored that against WGS-classified infections, our analysis highlights the need for WGS when variants are co-circulating and misclassification is likely. Summary of main pointsUsing whole genome sequencing, we provide direct evidence of waning vaccine effectiveness and variant-specific immune evasion during the Delta wave. Effectiveness estimates against calendar-period-classified infections approximated estimates against WGS-classified infections, however, calendar-period classification was associated with a variant misclassification.

Phenotypes of disease severity in a cohort of hospitalized COVID-19 patients: results from the IMPACC study

BackgroundBetter understanding of the association between characteristics of patients hospitalized with coronavirus disease 2019 (COVID-19) and outcome is needed to further improve upon patient management. MethodsImmunophenotyping Assessment in a COVID-19 Cohort (IMPACC) is a prospective, observational study of 1,164 patients from 20 hospitals across the United States. Disease severity was assessed using a 7-point ordinal scale based on degree of respiratory illness. Patients were prospectively surveyed for 1 year after discharge for post-acute sequalae of COVID-19 (PASC) through quarterly surveys. Demographics, comorbidities, radiographic findings, clinical laboratory values, SARS-CoV-2 PCR and serology were captured over a 28-day period. Multivariable logistic regression was performed. FindingsThe median age was 59 years (interquartile range [IQR] 20); 711 (61%) were men; overall mortality was 14%, and 228 (20%) required invasive mechanical ventilation. Unsupervised clustering of ordinal score over time revealed distinct disease course trajectories. Risk factors associated with prolonged hospitalization or death by day 28 included age [≥] 65 years (odds ratio [OR], 2.01; 95% CI 1.28-3.17), Hispanic ethnicity (OR, 1.71; 95% CI 1.13-2.57), elevated baseline creatinine (OR 2.80; 95% CI 1.63-4.80) or troponin (OR 1.89; 95% 1.03-3.47), baseline lymphopenia (OR 2.19; 95% CI 1.61-2.97), presence of infiltrate by chest imaging (OR 3.16; 95% CI 1.96-5.10), and high SARS-CoV2 viral load (OR 1.53; 95% CI 1.17-2.00). Fatal cases had the lowest ratio of SARS-CoV-2 antibody to viral load levels compared to other trajectories over time (p=0.001). 589 survivors (51%) completed at least one survey at follow-up with 305 (52%) having at least one symptom consistent with PASC, most commonly dyspnea (56% among symptomatic patients). Female sex was the only associated risk factor for PASC. InterpretationIntegration of PCR cycle threshold, and antibody values with demographics, comorbidities, and laboratory/radiographic findings identified risk factors for 28-day outcome severity, though only female sex was associated with PASC. Longitudinal clinical phenotyping offers important insights, and provides a framework for immunophenotyping for acute and long COVID-19. FundingNIH RESEARCH IN CONTEXTO_ST_ABSEvidence before this studyC_ST_ABSWe did a systematic search of the PubMed database from January 1st, 2020 until April 24th, 2022 using the search terms: "hospitalized" AND "SARS-CoV-2" OR "COVID-19" AND "Pro-spective" AND "Antibody" OR "PCR" OR "long term follow up" and applying the following filters: "Multicenter Study" AND "Observational Study". No language restrictions were applied. While clinical, laboratory, and radiographic features associated with severe COVID-19 in hospitalized adults have been described, description of the kinetics of SARS-CoV-2 specific assays available to clinicians (e.g. PCR and binding antibody) and their integration with other variables is scarce for both short and long term follow up. The current literature is comprised of several studies with small sample size, cross-sectional design with laboratory data typically only recorded at a single point in time (e.g., on admission), limited clinical characteristics, variable duration of follow up, single-center setting, retrospective analyses, kinetics of either PCR or antibody testing but not both, and outcomes such as death or, mechanical ventilation that do not allow delineation of variations in clinical course. Added value of this studyIn our large longitudinal multicenter cohort, the description of outcome severity, was not limited to survival versus death, but encompassed a clinical trajectory approach leveraging longitudinal data based on time in hospital, disease severity by ordinal scale based on degree of respiratory illness, and presence or absence of limitations at discharge. Fatal COVID-19 cases had the lowest ratio of antibody to viral load levels over time as compared to non-fatal cases. Integration of PCR cycle threshold and antibody values with demographics, baseline comorbidities, and laboratory/radiographic findings identified additional risk factors for outcome severity over the first 28 days. However, female sex was the only variable associated with persistence of symptoms over time. Persistence of symptoms was not associated with clinical trajectory over the first 28 days, nor with antibody/viral loads from the acute phase. Implications of all the available evidenceThe described calculated ratio (binding IgG/PCR Ct value) is unique compared to other studies, reflecting host pathogen interactions and representing an accessible approach for patient risk stratification. Integration of SARS-CoV-2 viral load and binding antibody kinetics with other laboratory as well as clinical characteristics in hospitalized COVID-19 patients can identify patients likely to have the most severe short-term outcomes, but is not predictive of symptom persistence at one year post-discharge.

Accelerated SARS-CoV-2 intrahost evolution leading to distinct genotypes during chronic infection

The chronic infection hypothesis for novel SARS-CoV-2 variant emergence is increasingly gaining credence following the appearance of Omicron. Here we investigate intrahost evolution and genetic diversity of lineage B.1.517 during a SARS-CoV-2 chronic infection lasting for 471 days (and still ongoing) with consistently recovered infectious virus and high viral loads. During the infection, we found an accelerated virus evolutionary rate translating to 35 nucleotide substitutions per year, approximately two-fold higher than the global SARS-CoV-2 evolutionary rate. This intrahost evolution led to the emergence and persistence of at least three genetically distinct genotypes suggesting the establishment of spatially structured viral populations continually reseeding different genotypes into the nasopharynx. Finally, using unique molecular indexes for accurate intrahost viral sequencing, we tracked the temporal dynamics of genetic diversity to identify advantageous mutations and highlight hallmark changes for chronic infection. Our findings demonstrate that untreated chronic infections accelerate SARS-CoV-2 evolution, ultimately providing opportunity for the emergence of genetically divergent and potentially highly transmissible variants as seen with Delta and Omicron.

Effectiveness of Primary and Booster COVID-19 mRNA Vaccination against Infection Caused by the SARS-CoV-2 Omicron Variant in People with a Prior SARS-CoV-2 Infection

BackgroundThe benefit of vaccination in people who experienced a prior SARS-CoV-2 infection remains unclear. ObjectiveTo estimate the effectiveness of primary (two-dose) and booster (third dose) vaccination against Omicron infection among people with a prior documented infection. DesignTest-negative case-control study. SettingYale New Haven Health System facilities. ParticipantsVaccine eligible people who received SARS-CoV-2 RT-PCR testing between November 1, 2021, and January 31, 2022. MeasurementsWe conducted two analyses, each with an outcome of Omicron BA.1 infection (S-gene target failure defined) and each stratified by prior SARS-CoV-2 infection status. We estimated the effectiveness of primary and booster vaccination. To test whether booster vaccination reduced the risk of infection beyond that of the primary series, we compared the odds among boosted and booster eligible people. ResultsOverall, 10,676 cases and 119,397 controls were included (6.1% and 7.8% occurred following a prior infection, respectively). The effectiveness of primary vaccination 14-149 days after 2nd dose was 36.1% (CI, 7.1% to 56.1%) for people with and 28.5% (CI, 20.0% to 36.2%) without prior infection. The odds ratio comparing boosted and booster eligible people with prior infection was 0.83 (CI, 0.56 to 1.23), whereas the odds ratio comparing boosted and booster eligible people without prior infection was 0.51 (CI, 0.46 to 0.56). LimitationsMisclassification, residual confounding, reliance on TaqPath assay analyzed samples. ConclusionWhile primary vaccination provided protection against BA.1 infection among people with and without prior infection, booster vaccination was only associated with additional protection in people without prior infection. These findings support primary vaccination in people regardless of prior infection status but suggest that infection history should be considered when evaluating the need for booster vaccination. Primary Funding SourceBeatrice Kleinberg Neuwirth and Sendas Family Funds, Merck and Co through their Merck Investigator Studies Program, and the Yale Schools of Public Health and Medicine.

Comparison of infectious SARS-CoV-2 from the nasopharynx of vaccinated and unvaccinated individuals

The frequency of SARS-CoV-2 breakthrough infections in fully vaccinated individuals increased with the emergence of the Delta variant, particularly with longer time from vaccine completion. However, whether breakthrough infections lead to onward transmission remains unclear. Here, we conducted a study involving 125 patients comprised of 72 vaccinated and 53 unvaccinated individuals, to assess the levels of infectious virus in vaccinated and unvaccinated individuals. Quantitative plaque assays showed no significant differences in the titers of virus between these cohorts. However, the proportion of nasopharyngeal samples with culturable virus was lower in the vaccinated patients relative to unvaccinated patients (21% vs. 40%). Finally, time-to-event analysis with Kaplan-Myer curves revealed that protection from culturable infectious virus waned significantly starting at 5 months after completing a 2-dose regimen of mRNA vaccines. These results have important implications in timing of booster dose to prevent onward transmission from breakthrough cases.

De novo emergence of a remdesivir resistance mutation during treatment of persistent SARS-CoV-2 infection in an immunocompromised patient: A case report

SARS-CoV-2 remdesivir resistance mutations have been generated in vitro but have not been reported in patients receiving treatment with the antiviral agent. We present a case of an immunocompromised patient with acquired B-cell deficiency who developed an indolent, protracted course of SARS-CoV-2 infection. Remdesivir therapy alleviated symptoms and produced a transient virologic response, but her course was complicated by recrudescence of high-grade viral shedding. Whole genome sequencing identified a mutation, E802D, in the nsp12 RNA-dependent RNA polymerase, which was not present in pre-treatment specimens. In vitro experiments demonstrated that the mutation conferred a [~]6-fold increase in remdesivir IC50 but resulted in a fitness cost in the absence of remdesivir. Sustained clinical and virologic response was achieved after treatment with casirivimab-imdevimab. Although the fitness cost observed in vitro may limit the risk posed by E802D, this case illustrates the importance of monitoring for remdesivir resistance and the potential benefit of combinatorial therapies in immunocompromised patients with SARS-CoV-2 infection.

Change in Symptoms and Immune Response in People with Post-Acute Sequelae of SARS-Cov-2 Infection (PASC) After SARS-Cov-2 Vaccination

As more people are vaccinated against SARS-CoV-2, many of those already infected are still suffering from Post-Acute Sequelae (PASC). Although there is no current treatment for PASC, reports from patients that the vaccine itself improves, and in some reports, worsens, PASC symptoms may lead to a deeper understanding of the causes of PASC symptoms and viable treatments. As such, we are conducting a study that measures the changes in PASC symptoms after vaccination. We are collecting baseline self-report and biospecimens for immune assays and then are following up with participants to collect the same data at 2-weeks, 6-weeks, and 12-weeks post-vaccination (first dose). Immune assays using blood specimens will include B-cell, T-cell, and myeloid cell panels; evaluation of T-cell responsiveness to SARS-CoV-2 peptides and antigen specific response; autoantibody screening (of IgG, IgM, and IgA antibodies that attack human proteins); and TCR sequencing and antigen mapping of CD8+ T-cells. Mucosal immunity will be measured using saliva specimens. The study aims to provide answers for people with PASC, especially regarding the causes of their symptoms and how the vaccine may affect them, and clues for PASC treatment.

Effectiveness of the CoronaVac vaccine in the elderly population during a P.1 variant-associated epidemic of COVID-19 in Brazil: A test-negative case-control study

ObjectiveTo estimate the effectiveness of the inactivated whole-virus vaccine, CoronaVac, against symptomatic COVID-19 in the elderly population of Sao Paulo State, Brazil during widespread circulation of the Gamma variant. DesignTest negative case-control study. SettingHealth-care facilities in Sao Paulo State, Brazil. Participants43,774 adults aged 70 years or older who were residents of Sao Paulo State and underwent SARS-CoV-2 RT-PCR testing from January 17 to April 29, 2021. 26,433 cases with symptomatic COVID-19 and 17,622 symptomatic, test negative controls were selected into 7,950 matched pairs, according to age, sex, self-reported race, municipality of residence, prior COVID-19 status and date of RT-PCR testing. InterventionVaccination with a two-dose regimen of CoronaVac. Main outcome measuresRT-PCR confirmed symptomatic COVID-19 and COVID-19 associated hospitalizations and deaths. ResultsAdjusted vaccine effectiveness against symptomatic COVID-19 was 18.2% (95% CI, 0.0 to 33.2) in the period 0-13 days after the second dose and 41.6% (95% CI, 26.9 to 53.3) in the period [≥]14 days after the second dose. Adjusted vaccine effectiveness against hospitalisations was 59.0% (95% CI, 44.2 to 69.8) and against deaths was 71.4% (95% CI, 53.7 to 82.3) in the period [≥]14 days after the second dose. Vaccine effectiveness [≥]14 days after the second dose declined with increasing age for the three outcomes, and among individuals aged 70-74 years it was 61.8% (95% CI, 34.8 to 77.7) against symptomatic disease, 80.1% (95% CI, 55.7 to 91.0) against hospitalisations and 86.0% (95% CI, 50.4 to 96.1) against deaths. ConclusionsVaccination with CoronaVac was associated with a reduction in symptomatic COVID-19, hospitalisations and deaths in adults aged 70 years or older in a setting with extensive Gamma variant transmission. However, significant protection was not observed until completion of the two-dose regimen, and vaccine effectiveness declined with increasing age amongst this elderly population. Summary boxesO_ST_ABSWhat is already known on this topicC_ST_ABSRandomised controlled trials (RCT) have yielded varying estimates (51 to 84%) for the effectiveness of the inactivated whole-virus vaccine, CoronaVac, against symptomatic COVID-19. Current evidence is limited on whether CoronaVac is effective against severe disease or death caused by the SARS-CoV-2 variant of concern, Gamma, or in the setting of extensive Gamma variant circulation. More evidence is needed for the real-world effectiveness of CoronaVac and other inactivated vaccines among elderly individuals, a population that was underrepresented in RCTs of these vaccines. What this study addsA two-dose regimen of CoronaVac provides significant protection against symptomatic COVID-19, hospitalisations and deaths among adults [≥]70 years of age in the setting of widespread Gamma variant transmission. Significant protection did not occur until [≥]14 days after administration of the second dose of CoronaVac. The effectiveness of CoronaVac declines with increasing age in the elderly population.

Effectiveness of CoronaVac in the setting of high SARS-CoV-2 P.1 variant transmission in Brazil: A test-negative case-control study

BackgroundSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variant, Gamma, emerged in the city of Manaus in late 2020 during a large resurgence of coronavirus disease (COVID-19), and has spread throughout Brazil. The effectiveness of vaccines in settings with widespread Gamma variant transmission has not been reported. MethodsWe performed a matched test-negative case-control study to estimate the effectiveness of an inactivated vaccine, CoronaVac, in healthcare workers (HCWs) in Manaus, where the Gamma variant accounted for 86% of genotyped SARS-CoV-2 samples at the peak of its epidemic. We performed an early analysis of effectiveness following administration of at least one vaccine dose and an analysis of effectiveness of the two-dose schedule. The primary outcome was symptomatic SARS-CoV-2 infection. FindingsFor the early at-least-one-dose and two-dose analyses the study population was, respectively, 53,176 and 53,153 HCWs residing in Manaus and aged 18 years or older, with complete information on age, residence, and vaccination status. Among 53,153 HCWs eligible for the two-dose analysis, 47,170 (89%) received at least one dose of CoronaVac and 2,656 individuals (5%) underwent RT-PCR testing from 19 January, 2021 to 13 April, 2021. Of 3,195 RT-PCR tests, 885 (28%) were positive. 393 and 418 case- control pairs were selected for the early and two-dose analyses, respectively, matched on calendar time, age, and neighbourhood. Among those who had received both vaccine doses before the RT-PCR sample collection date, the average time from second dose to sample collection date was 14 days (IQR 7-24). In the early analysis, vaccination with at least one dose was associated with a 0.50-fold reduction (adjusted vaccine effectiveness (VE), 49.6%, 95% CI 11.3 to 71.4) in the odds of symptomatic SARS-CoV-2 infection during the period 14 days or more after receiving the first dose. However, we estimated low effectiveness (adjusted VE 36.8%, 95% CI -54.9 to 74.2) of the two-dose schedule against symptomatic SARS-CoV-2 infection during the period 14 days or more after receiving the second dose. A finding that vaccinated individuals were much more likely to be infected than unvaccinated individuals in the period 0-13 days after first dose (aOR 2.11, 95% CI 1.36-3.27) suggests that unmeasured confounding led to downward bias in the vaccine effectiveness estimate. InterpretationEvidence from this test-negative study of the effectiveness of CoronaVac was mixed, and likely affected by bias in this setting. Administration of at least one vaccine dose showed effectiveness against symptomatic SARS-CoV-2 infection in the setting of epidemic Gamma variant transmission. However, the low estimated effectiveness of the two-dose schedule underscores the need to maintain non-pharmaceutical interventions while vaccination campaigns with CoronaVac are being implemented. FundingFundacao Oswaldo Cruz (Fiocruz); Municipal Health Secretary of Manaus Research in ContextO_ST_ABSEvidence before this studyC_ST_ABSWe searched PubMed for articles published from inception of the pandemic until April 3, 2021, with no language restrictions, using the search terms "P.1" AND "vaccine" AND "SARS-CoV-2". Additionally, we searched for "CoronaVac" AND "SARS-CoV-2". Early studies have found plasma from convalescent COVID-19 patients and sera from vaccinated individuals have reduced neutralisation of the SARS-CoV-2 variant, Gamma or P.1, compared with strains isolated earlier in the pandemic. Pfizer BNT162b2 mRNA, Oxford-AstraZeneca ChAdOx1, and CoronaVac are the only vaccines for which such data has been published to date. No studies reported effectiveness of any vaccine on reducing the risk of infection or disease among individuals exposed to P.1 or in settings of high P.1 transmission. Added value of this studyThis study finds that vaccination with CoronaVac was 49.4% (95% CI 13.2 to 71.9) effective at preventing COVID-19 in a setting with likely high prevalence of the Gamma Variant of Concern. However, an analysis of effectiveness by dose was underpowered and failed to find significant effectiveness of the two-dose schedule of CoronaVac (estimated VE 37.1%, 95% CI -53.3 to 74.2). Implications of all the available evidenceThese findings are suggestive for the effectiveness of CoronaVac in healthcare workers in the setting of widespread P.1 transmission but must be strengthened by observational studies in other settings and populations. Based on this evidence, there is a need to implement sustained non-pharmaceutical interventions even as vaccination campaigns continue.

Accuracy of Computable Phenotyping Approaches for SARS-CoV-2 Infection and COVID-19 Hospitalizations from the Electronic Health Record

ObjectiveReal-world data have been critical for rapid-knowledge generation throughout the COVID-19 pandemic. To ensure high-quality results are delivered to guide clinical decision making and the public health response, as well as characterize the response to interventions, it is essential to establish the accuracy of COVID-19 case definitions derived from administrative data to identify infections and hospitalizations. MethodsElectronic Health Record (EHR) data were obtained from the clinical data warehouse of the Yale New Haven Health System (Yale, primary site) and 3 hospital systems of the Mayo Clinic (validation site). Detailed characteristics on demographics, diagnoses, and laboratory results were obtained for all patients with either a positive SARS-CoV-2 PCR or antigen test or ICD-10 diagnosis of COVID-19 (U07.1) between April 1, 2020 and March 1, 2021. Various computable phenotype definitions were evaluated for their accuracy to identify SARS-CoV-2 infection and COVID-19 hospitalizations. ResultsOf the 69,423 individuals with either a diagnosis code or a laboratory diagnosis of a SARS-CoV-2 infection at Yale, 61,023 had a principal or a secondary diagnosis code for COVID-19 and 50,355 had a positive SARS-CoV-2 test. Among those with a positive laboratory test, 38,506 (76.5%) and 3449 (6.8%) had a principal and secondary diagnosis code of COVID-19, respectively, while 8400 (16.7%) had no COVID-19 diagnosis. Moreover, of the 61,023 patients with a COVID-19 diagnosis code, 19,068 (31.2%) did not have a positive laboratory test for SARS-CoV-2 in the EHR. Of the 20 cases randomly sampled from this latter group for manual review, all had a COVID-19 diagnosis code related to asymptomatic testing with negative subsequent test results. The positive predictive value (precision) and sensitivity (recall) of a COVID-19 diagnosis in the medical record for a documented positive SARS-CoV-2 test were 68.8% and 83.3%, respectively. Among 5,109 patients who were hospitalized with a principal diagnosis of COVID-19, 4843 (94.8%) had a positive SARS-CoV-2 test within the 2 weeks preceding hospital admission or during hospitalization. In addition, 789 hospitalizations had a secondary diagnosis of COVID-19, of which 446 (56.5%) had a principal diagnosis consistent with severe clinical manifestation of COVID-19 (e.g., sepsis or respiratory failure). Compared with the cohort that had a principal diagnosis of COVID-19, those with a secondary diagnosis had a more than 2-fold higher in-hospital mortality rate (13.2% vs 28.0%, P<0.001). In the validation sample at Mayo Clinic, diagnosis codes more consistently identified SARS-CoV-2 infection (precision of 95%) but had lower recall (63.5%) with substantial variation across the 3 Mayo Clinic sites. Similar to Yale, diagnosis codes consistently identified COVID-19 hospitalizations at Mayo, with hospitalizations defined by secondary diagnosis code with 2-fold higher in-hospital mortality compared to those with a primary diagnosis of COVID-19. ConclusionsCOVID-19 diagnosis codes misclassified the SARS-CoV-2 infection status of many people, with implications for clinical research and epidemiological surveillance. Moreover, the codes had different performance across two academic health systems and identified groups with different risks of mortality. Real-world data from the EHR can be used to in conjunction with diagnosis codes to improve the identification of people infected with SARS-CoV-2.

Saliva viral load is a dynamic unifying correlate of COVID-19 severity and mortality

While several clinical and immunological parameters correlate with disease severity and mortality in SARS-CoV-2 infection, work remains in identifying unifying correlates of coronavirus disease 2019 (COVID-19) that can be used to guide clinical practice. Here, we examine saliva and nasopharyngeal (NP) viral load over time and correlate them with patient demographics, and cellular and immune profiling. We found that saliva viral load was significantly higher in those with COVID-19 risk factors; that it correlated with increasing levels of disease severity and showed a superior ability over nasopharyngeal viral load as a predictor of mortality over time (AUC=0.90). A comprehensive analysis of immune factors and cell subsets revealed strong predictors of high and low saliva viral load, which were associated with increased disease severity or better overall outcomes, respectively. Saliva viral load was positively associated with many known COVID-19 inflammatory markers such as IL-6, IL-18, IL-10, and CXCL10, as well as type 1 immune response cytokines. Higher saliva viral loads strongly correlated with the progressive depletion of platelets, lymphocytes, and effector T cell subsets including circulating follicular CD4 T cells (cTfh). Anti-spike (S) and anti-receptor binding domain (RBD) IgG levels were negatively correlated with saliva viral load showing a strong temporal association that could help distinguish severity and mortality in COVID-19. Finally, patients with fatal COVID-19 exhibited higher viral loads, which correlated with the depletion of cTfh cells, and lower production of anti-RBD and anti-S IgG levels. Together these results demonstrated that viral load - as measured by saliva but not nasopharyngeal -- is a dynamic unifying correlate of disease presentation, severity, and mortality over time.

Diverse Functional Autoantibodies in Patients with COVID-19

COVID-19 manifests with a wide spectrum of clinical phenotypes that are characterized by exaggerated and misdirected host immune responses1-8. While pathological innate immune activation is well documented in severe disease1, the impact of autoantibodies on disease progression is less defined. Here, we used a high-throughput autoantibody discovery technique called Rapid Extracellular Antigen Profiling (REAP) to screen a cohort of 194 SARS-CoV-2 infected COVID-19 patients and healthcare workers for autoantibodies against 2,770 extracellular and secreted proteins (the "exoproteome"). We found that COVID-19 patients exhibit dramatic increases in autoantibody reactivities compared to uninfected controls, with a high prevalence of autoantibodies against immunomodulatory proteins including cytokines, chemokines, complement components, and cell surface proteins. We established that these autoantibodies perturb immune function and impair virological control by inhibiting immunoreceptor signaling and by altering peripheral immune cell composition, and found that murine surrogates of these autoantibodies exacerbate disease severity in a mouse model of SARS-CoV-2 infection. Analysis of autoantibodies against tissue-associated antigens revealed associations with specific clinical characteristics and disease severity. In summary, these findings implicate a pathological role for exoproteome-directed autoantibodies in COVID-19 with diverse impacts on immune functionality and associations with clinical outcomes.

Clinical Characteristics and Outcomes for 7,995 Patients with SARS-CoV-2 Infection

ObjectiveSevere acute respiratory syndrome virus (SARS-CoV-2) has infected millions of people worldwide. Our goal was to identify risk factors associated with admission and disease severity in patients with SARS-CoV-2. DesignThis was an observational, retrospective study based on real-world data for 7,995 patients with SARS-CoV-2 from a clinical data repository. SettingYale New Haven Health (YNHH) is a five-hospital academic health system serving a diverse patient population with community and teaching facilities in both urban and suburban areas. PopulationsThe study included adult patients who had SARS-CoV-2 testing at YNHH between March 1 and April 30, 2020. Main outcome and performance measuresPrimary outcomes were admission and in-hospital mortality for patients with SARS-CoV-2 infection as determined by RT-PCR testing. We also assessed features associated with the need for respiratory support. ResultsOf the 28605 patients tested for SARS-CoV-2, 7995 patients (27.9%) had an infection (median age 52.3 years) and 2154 (26.9%) of these had an associated admission (median age 66.2 years). Of admitted patients, 1633 (75.8%) had a discharge disposition at the end of the study period. Of these, 192 (11.8%) required invasive mechanical ventilation and 227 (13.5%) expired. Increased age and male sex were positively associated with admission and in-hospital mortality (median age 81.9 years), while comorbidities had a much weaker association with the risk of admission or mortality. Black race (OR 1.43, 95%CI 1.14-1.78) and Hispanic ethnicity (OR 1.81, 95%CI 1.50-2.18) were identified as risk factors for admission, but, among discharged patients, age-adjusted in-hospital mortality was not significantly different among racial and ethnic groups. ConclusionsThis observational study identified, among people testing positive for SARS-CoV-2 infection, older age and male sex as the most strongly associated risks for admission and in-hospital mortality in patients with SARS-CoV-2 infection. While minority racial and ethnic groups had increased burden of disease and risk of admission, age-adjusted in-hospital mortality for discharged patients was not significantly different among racial and ethnic groups. Ongoing studies will be needed to continue to evaluate these risks, particularly in the setting of evolving treatment guidelines.

Longitudinal immunological analyses reveal inflammatory misfiring in severe COVID-19 patients

Recent studies have provided insights into the pathogenesis of coronavirus disease 2019 (COVID-19)1-4. Yet, longitudinal immunological correlates of disease outcome remain unclear. Here, we serially analysed immune responses in 113 COVID-19 patients with moderate (non-ICU) and severe (ICU) disease. Immune profiling revealed an overall increase in innate cell lineages with a concomitant reduction in T cell number. We identify an association between early, elevated cytokines and worse disease outcomes. Following an early increase in cytokines, COVID-19 patients with moderate disease displayed a progressive reduction in type-1 (antiviral) and type-3 (antifungal) responses. In contrast, patients with severe disease maintained these elevated responses throughout the course of disease. Moreover, severe disease was accompanied by an increase in multiple type 2 (anti-helminths) effectors including, IL-5, IL-13, IgE and eosinophils. Unsupervised clustering analysis of plasma and peripheral blood leukocyte data identified 4 immune signatures, representing (A) growth factors, (B) type-2/3 cytokines, (C) mixed type-1/2/3 cytokines, and (D) chemokines that correlated with three distinct disease trajectories of patients. The immune profile of patients who recovered with moderate disease was enriched in tissue reparative growth factor signature (A), while the profile for those with worsened disease trajectory had elevated levels of all four signatures. Thus, we identified development of a maladapted immune response profile associated with severe COVID-19 outcome and early immune signatures that correlate with divergent disease trajectories.

Development and validation of the COVID-19 severity index (CSI): a prognostic tool for early respiratory decompensation

ObjectiveThe goal of this study was to create a predictive model of early hospital respiratory decompensation among patients with COVID-19. DesignObservational, retrospective cohort study. SettingNine-hospital health system within the Northeastern United States. PopulationsAdult patients ([≥] 18 years) admitted from the emergency department who tested positive for SARS-CoV-2 (COVID-19) up to 24 hours after initial presentation. Patients meeting criteria for respiratory critical illness within 4 hours of arrival were excluded. Main outcome and performance measuresWe used a composite endpoint of critical illness as defined by oxygen requirement (greater than 10 L/min by low-flow device, high-flow device, non-invasive, or invasive ventilation) or death within the first 24 hours of hospitalization. We developed models predicting our composite endpoint using patient demographic and clinical data available within the first four hours of arrival. Eight hospitals (n = 932) were used for model development and one hospital (n = 240) was held out for external validation. Area under receiver operating characteristic (AU-ROC), precision-recall curves (AU-PRC), and calibration metrics were used to compare predictive models to three illness scoring systems: Elixhauser comorbidity index, qSOFA, and CURB-65. ResultsDuring the study period from March 1, 2020 to April 27,2020, 1,792 patients were admitted with COVID-19. Six-hundred and twenty patients were excluded based on age or critical illness within the first 4 hours, yielding 1,172 patients in the final cohort. Of these patients, 144 (12.3%) met the composite endpoint within the first 24 hours. We first developed a bedside quick COVID-19 severity index (qCSI), a twelve-point scale using nasal cannula flow rate, respiratory rate, and minimum documented pulse oximetry. We then created a machine-learning gradient boosting model, the COVID-19 severity index (CSI), using twelve additional variables including inflammatory markers and liver chemistries. Both the qCSI (AU-ROC mean [95% CI]: 0.90 [0.85-0.96]) and CSI (AU-ROC: 0.91 [0.86-0.97]) outperformed the comparator models (qSOFA: 0.76 [0.69-0.85]; Elixhauser: 0.70 [0.62-0.80]; CURB-65: AU-ROC 0.66 [0.58-0.77]) on cross-validation and performed well on external validation (qCSI: 0.82, CSI: 0.76, CURB-65: 0.50, qSOFA: 0.59, Elixhauser: 0.61). We find that a qCSI score of 0-3 is associated with a less than 5% risk of critical respiratory illness, while a score of 9-12 is associated with a 57% risk of progression to critical illness. ConclusionsA significant proportion of admitted COVID-19 patients decompensate within 24 hours of hospital presentation and these events are accurately predicted using bedside respiratory exam findings within a simple scoring system.