key: cord-0816811-hsdeesm2
authors: Miller, Emily Happy; Zucker, Jason; Castor, Delivette; Annavajhala, Medini K; Sepulveda, Jorge L; Green, Daniel A; Whittier, Susan; Scherer, Matthew; Medrano, Nicola; Sobieszczyk, Magdalena E; Yin, Michael T; Kuhn, Louise; Uhlemann, Anne-Catrin
title: Pre-test symptom duration and cycle threshold values for SARS-CoV-2 reverse transcription-PCR (RT-PCR) predict COVID-19 mortality
date: 2021-01-04
journal: Open Forum Infect Dis
DOI: 10.1093/ofid/ofab003
sha: 7e9b1bffbf8c6667e1410104b7226e5040867ed7
doc_id: 816811
cord_uid: hsdeesm2

BACKGROUND: The relationship between SARS-CoV-2 viral load and patient symptom duration in both in- and outpatients, and the impact of these factors on patient outcomes, are currently unknown. Understanding these associations is important to clinicians caring for patients with COVID-19. METHODS: We conducted an observational study between March 10–May 30, 2020 at a large quaternary academic medical center in New York City. Patient characteristics, laboratory values, and clinical outcomes were abstracted from the electronic medical records. Of all patients tested for SARS-CoV-2 during this time (N=16,384), there were 5,467 patients with positive tests, of which 4,254 had available Ct values and were included in further analysis. Univariable and multivariable logistic regression models were used to test associations between Ct values, duration of symptoms prior to testing, patient characteristics and mortality. The primary outcome is defined as death or discharge to hospice. RESULTS: Lower Ct values at diagnosis (i.e. higher viral load) were associated with significantly higher mortality among both in- and out-patients. Interestingly, patients with a shorter time since the onset of symptoms to testing had a worse prognosis, with those presenting less than three days from symptom onset having 2-fold increased odds of death. After adjusting for time since symptom onset and other clinical covariates, Ct values remained a strong predictor of mortality. CONCLUSIONS: SARS-CoV-2 RT-PCR Ct value and duration of symptoms are strongly associated with mortality. These two factors add useful information for clinicians to risk stratify patients presenting with COVID-19.

A c c e p t e d M a n u s c r i p t

About 20% of patients with COVID-19 will require hospitalization and a subset will have severe manifestations such as Acute Respiratory Distress Syndrome (ARDS) and a hyperinflammatory state [1] [2] [3] [4] [5] . Much of the morbidity and mortality of COVID-19 has been attributed to the hyperinflammatory state that develops around day 7-10 of infection in a subset of patients and appears to lead to worse outcomes 6 . It is of critical importance to understand which patients presenting to care with COVID-19 will decompensate.

Reverse transcription-polymerase chain reaction (RT-PCR) from nasopharyngeal swab is the most common test for detecting acute SARS-Cov-2 infection. The results of the RT-PCR are reported out as positive or negative, but the cycle threshold (Ct) value has not commonly been reported to providers. Ct values are inversely related to viral loads-the lower the Ct value, the higher the viral load. A correlation between high viral load and disease severity is seen with other respiratory viruses such as Influenza B infection or Rhinovirus infection 7, 8 . Recent studies of SARS-CoV-2 have shown a relationship between lower RT-PCR Ct value and mortality in admitted patients 9, 10 .

However, SARS-CoV-2 viral load is thought to fluctuate over the course of the infection, beginning with the presymptomatic stage. The duration of symptoms may therefore be an important factor that has not been considered yet in the relationship of low Ct values (high viral load) as predictors of outcome.

Understanding Ct values as a viral load proxy, variations in the inpatient and outpatient settings and how the duration of symptoms prior to testing affects the association between Ct values and mortality is critical. Taking advantage of systematically collected data on symptoms, we examined the association between Ct values, symptom duration and mortality in a large cohort of inand outpatients with COVID-19 at New York-Presbyterian Columbia University Irving Medical Center,

A c c e p t e d M a n u s c r i p t Methods:

This observational study was conducted at an academic quaternary care medical center located in Northern Manhattan. Patients of all ages were included who tested positive by SARS-CoV-2 RT-PCR nasopharyngeal test for either viral target between March 10-May 30, 2020. Testing was performed using either high-throughput automated cobas 6800 (Roche Molecular Systems, Branchburg, NJ) or the rapid Xpert Xpress SARS-CoV-2 test on the Infinity platform (Cepheid, Sunnyvale, CA). Both tests detect two viral targets in SARS-CoV-2; cobas 6800 in the ORF1ab nonstructural region and the envelope gene and Xpert Xpress in the nucleocapsid and envelope genes.

Due to its rapid turn-around time, the Xpert Xpress test was used preferentially to screen women presenting to Labor and Delivery and for patients in the Emergency Room. Both tests were used per manufacturer recommendations under FDA Emergency Use Authorization [11] [12] [13] . Per manufacturer recommendations, the limit of detection for a positive test is a cycle threshold level of 40 for the cobas 6800 test and 45 for the Xpert Xpress test. To obtain Ct values, reports were generated directly from the instruments. These were scanned, and a database of Ct values was created using optical character recognition. Of the 5467 patients with positive tests, Ct values could be obtained for 4254. For the purpose of this study, the Ct value for target 1 (ORF1ab for cobas 6800 test, and nucleocapsid for Xpert Xpress) was used for further analysis as these are the SARS-CoV-2 specific targets which do not cross-react with other seasonal coronaviruses.

For all presenting patients, electronic health records data extracted for this analysis included demographics, vital signs, laboratory results, admission, discharge, and transfer dates, medication administrations, procedure codes (current procedural terminology codes) and current and historical international classification of disease (ICD 9 and 10) codes extracted from the clinical data A c c e p t e d M a n u s c r i p t warehouse. The first lab value and vital sign measurement for the index visit were obtained from patient flow sheets. Initial oxygen requirement was measured by oxygen rank severity score which with 0 indicating no supplemental oxygen requirement and 4 indicating need for mechanical ventilation. A subset of consecutive charts was manually reviewed starting with the first patient testing positive for SARS-CoV-2 to our institution. The dataset was further enriched with manually abstracted data entered into a REDCap database that included the date of symptom onset and presenting symptoms. Symptoms and symptom duration were part of the hospitals' COVID-19 admission tools. All data were merged using RStudio.

Before initiation of data collection, approval for the study was obtained from the Columbia University Irving Medical Center Institutional Review Board (IRB). The requirement for obtaining written informed consent was waived by the IRB.

We conducted a cohort analysis to examine the association between the Ct value on the first positive PCR and subsequent patient outcome. The primary endpoint was death or discharge to hospice by the time the final data set was assembled (August 20, 2020). Analyses included both inand out-patients adjusting for demographic factors and co-morbidities (available from the data warehouse). Analyses restricted to inpatients were done adjusting for the data on demographic factors and comorbidities available from the data warehouse as well as for reported duration of symptoms prior to presentation and laboratory parameters, which were largely not available for outpatients.

A c c e p t e d M a n u s c r i p t

Descriptive statistics were reported including counts with percentages, medians and their interquartile ranges (IQR) and Box-and-whisker plots. The Wilcoxon rank-sum test was used to compare groups for continuous variables and chi-squared test for categorical variables. To estimate associations between mortality and other covariates including Ct values, Odds Ratios (OR) and 95% confidence intervals (CI) were calculated using logistic regression. Final multivariable models included all covariates that were associated with the outcome at p<0.05 or which influenced the magnitude of the Ct value associations by more than 10%. Linear regression was used to describe associations between Ct values and duration of symptoms prior to presentation. All statistical analyses were performed in SAS version 9.4 (Cary, NC). Table 1 ). Of the 4,254 patients with positive tests, the majority was obtained on the cobas 6800 (n=3,808) and the remainder on the Xpert Xpress (n=450 A c c e p t e d M a n u s c r i p t 573 (13% of total) requiring ICU level care during admission. Patients who were admitted differed in age, race/ethnicity and frequencies of comorbidities from those who were not admitted (Table 1) .

Among 4254 patients with available Ct values, 542 (13%) met the primary outcome of death or discharge to hospice almost all of whom had been admitted. The small number of non-admitted patients who died were recorded to have died in the emergency room. In admitted patients, who died, the median time to death after positive test was 7 days (IQR 3-17 days 

We then examined predictors of death or discharge to hospice in the 2308 persons with a positive SARS-CoV-2 RT-PCR nasopharyngeal test admitted to the hospital (Table 3 ). This analysis also included laboratory values and the reported duration of symptoms.

In univariable analysis, lower Ct value again was significantly associated with mortality (OR 0.918; 95% CI: 0.904, 0.933). In the multivariable analysis, adjusted for sex, age, level of care, time from symptom onset, and laboratory parameters, Ct value remained significantly associated with mortality (OR 0.937; 95% CI: 0.912 0.963). Interestingly, presentation soon after symptom onset (<3 days) was associated with higher odds of mortality in both univariable and multivariable analysis. A c c e p t e d M a n u s c r i p t

We were able to ascertain the duration of symptoms prior to presentation among 1860 (81%) of the inpatients included in this analysis. Three with reported onset of symptoms only after the test were excluded. The median time from symptom onset to presentation was 6 days (IQR 3-10 days) with 394 (21%) presenting <3 days after symptom onset and 820 (44%), 439 (24%), 105 (6%) and 99 (5%) presenting 3-7 days, 8-14 days, 15-21 days and >21 days after symptom onset, respectively.

Ct values were higher with longer time between onset of symptoms and testing. This pattern was most noticeable among inpatients who survived (n=1,412), with an increase of 0.362 Ct units (95% CI: 0.297-0.427) per day after symptom onset to test; but was also observed in inpatients who died (n=445) with an increase of 0.285 (95% CI: 0.171-0.399) per day (Figure 3 ). Figure 4 shows the median Ct values by time in days after symptom onset to presentation stratified by outcome. There was little increase in the presenting Ct by time since symptom onset until around 3 days, and thereafter Ct values increased with longer time since symptom onset. Table 3 , patients presenting <3 days after symptom onset had a more than 2fold increased odds of death (OR=2.362; 95% CI: 1.857, 3.004) than those presenting later. On a continuous scale in days (>14 days recoded as =15), patients with longer time in days between symptom onset and presentation had better outcomes (OR =0.931; 95% CI: 0.909 0.954).

Categorizing time after symptom onset into <3 days, 4-7 days, 8-14 days and 15+ days, showed odds of death relative to those presenting 8-14 days after symptom onset (group with the lowest odds of death) to be greatest in those presenting <3 days (OR 3.118; 95% CI: 2.251 4.319) but still increased in those presenting 4-7 days (OR 1.510; 95% CI: 1.116 2.043), and with a non-significant trend to be worse if presenting >14 days (OR 1.304 95% CI: 0.851 1.997) (Figure 3) . was most striking in those presenting > 3 days after symptom onset but was still seen in those presenting <3 days after symptom onset (Figure 4 ). Associations between outcome and Ct values and symptom duration prior to testing remained consistent after adjusting for age; older age being a strong predictor of outcome (Table 3 ). Nevertheless, it should be noted that the low rates of outcome in the younger group limited our capacity to fully interrogate effect modification by age.

Here we observed a significant association between lower Ct values (and therefore higher viral load) and mortality in a large cohort of COVID-19 patients. We also found that reported symptom duration prior to testing was an independent predictor of outcomes. These findings provide important new insights into the role of duration of symptoms prior to presentation and its connection with both viral load and patient outcomes. Given the strong association between Ct values and mortality, Ct value on initial SARS-CoV-2 testing may be an important predictor of outcomes, especially for those patients being admitted to the hospital. While Ct value is a strong predictor of poor clinical outcomes, our data does not find a clinically meaningful cut off that could be used for triaging of patients. Duration of symptoms prior to presentation and Ct value on initial SARS-CoV-2 test are independent predictors of mortality and are potentially useful indicators of patient trajectory. As no single value is predictive of a poor outcome, categories of Ct values such as quartiles could be utilized to triage patient's risk for decompensation. Ct values were the lowest A c c e p t e d M a n u s c r i p t (viral load the highest) in the first 3 days of self-reported symptoms. Patients who presented earlier in their course and went on to either die or be discharged to hospice had lower Ct values than those who presented with similar symptom duration and survived.

A correlation between high viral load and mortality has been described in other viral infections, such as Yellow Fever virus and Ebola virus [14] [15] [16] . With Ebola virus and Yellow fever virus, patients who present later to care tend to have higher viral loads and higher mortality. In this study we observe the opposite with COVID-19, where patients who present early with high viral loads have a 2-fold increase risk of death. This is also in line with studies on transmissibility of the virus, which show that risk of transmission is highest in the first few days of symptoms, when viral loads are highest 17, 18 . Therefore, a patient who presents with a low Ct value on their initial test may be on a trajectory to a poor outcome compared to someone who presents with a higher Ct value, even if both patients report similar symptom duration. It is possible that higher viral loads act as a trigger for the hyperinflammatory state seen in severe cases of COVID-19. Based on this data, Ct value and symptom duration could be used in conjunction with other factors that show an association with higher mortality, such as age >75, high IL-6, high LDH and low platelets to help guide the clinician's assessment of risk for poor outcome.

This study fills several important gaps in current knowledge on this topic. First, the role of symptom duration in outcomes and viral load is described for the first time. This information may not be readily available in other datasets but was systematically captured in our COVID-19 admission screens in the medical record. This cohort also includes both inpatients and outpatients as well as adult and pediatric patients. Outpatients and patients who were discharged from the emergency room have been largely excluded from prior studies on the role of Ct values in outcomes. Other strengths of this study include a large cohort with Ct values and definitive outcomes included for most patients. As well as a range of race/ethnicity is represented in this cohort, including Hispanic and African American patients, who are disproportionately affected by the COVID-19 pandemic 19-21 . A c c e p t e d M a n u s c r i p t Several limitations need to be considered. This study was a single quaternary academic medical center during the peak of the major outbreak. This could limit its generalization to other patient populations specifically those with lower circulating amounts of virus. However, as the pandemic continues, and new hot spots emerge this experience will provide useful information for these areas as they navigate their own surges. Data included in the study was limited to what is available in the electronic medical records and errors can exist in both patient recall and/or provider documentation, especially around collection of symptom duration. Even with these issues we were able analyze data from a large cohort of 4,254 patients. Finally, management of patients changed over the course of the epidemic as new data, experimental therapeutics, and clinical trials became available. It is difficult to fully take into account how this constant change of treatment practices may have influenced patient outcomes.

Taken together, this study supports a role for use of Ct values from SARS-CoV-2 testing and history of symptom duration as useful tools to help predict which patients presenting with COVID-19 may have worse outcomes. Patients at particularly high risk for poor outcomes include those presenting early in their disease course with low Ct values. These parameters add value to other variables, such as inflammatory markers, older age and oxygen requirements, that contribute to poor outcomes and should also be considered when attempting to triage patients. Current antiviral treatments for COVID-19, such as remdesivir, offer minimal benefits and have been plagued by availability shortfalls 22, 23 . As more effective antivirals become available, it will be important to target them to patients in the viral phase of the illness when they will likely be most effective. Utilizing Ct values in the context of symptom duration can help providers predict who is at higher risk for decompensation and help guide the use of antivirals. Additional studies are needed to determine all predictors of poor outcomes in COVID-19, however the reporting of Ct values from SARS-CoV-2 RT-PCR testing in conjunction with history of symptom duration can be used by providers as another tool to risk stratify patients and prioritize resources.

A c c e p t e d M a n u s c r i p t M a n u s c r i p t A c c e p t e d M a n u s c r i p t M a n u s c r i p t Ct value indicated cycle threshold from reverse transcription-PCR testing for SARS-CoV-2.

Ct value for initial test. Patients still admitted as of 8/20/2020. Symptom duration prior to test was self-reported by the patient or family member who provided history upon presentation. Days from symptom onset <=0 were excluded. Days from symptom onset >14 were categorized as >15. Figure 4 : Frequency of death or discharge to hospice in 1857 admitted patients by SARS-Co-V-2 cycle threshold (Ct) value quartiles and presentation <3 days or 3 or more days after symptom onset

Characterization and clinical course of 1000 patients with coronavirus disease 2019 in New York: retrospective case series

Epidemiology, clinical course, and outcomes of critically ill adults with COVID-19 in New York City: a prospective cohort study

Characteristics of and Important Lessons From the Coronavirus Disease 2019 (COVID-19) Outbreak in China

Characteristics and Clinical Outcomes of Adult Patients Hospitalized with COVID-19 -Georgia

Clinical features of patients infected with 2019 novel coronavirus in Wuhan

COVID-19 illness in native and immunosuppressed states: A clinical-therapeutic staging proposal

Influenza and rhinovirus viral load and disease severity in upper respiratory tract infections

Correlation of Pandemic (H1N1) 2009 Viral Load with Disease Severity and Prolonged Viral Shedding in Children

Impact of SARS-CoV-2 Viral Load on Risk of Intubation and Mortality Among Hospitalized Patients with Coronavirus Disease

SARS-CoV-2 viral load predicts COVID-19 mortality. The Lancet Respiratory Medicine

Control CfD. CDC 2019-Novel Coronavirus (2019-nCoV) Real-Time RT PCR Diagnostic Panel

Xpert Xpress SARS-CoV-2 Instructions for Use

COVID-19 in Solid Organ Transplant Recipients: Initial Report from the US Epicenter

The Contribution of Ebola Viral Load at Admission and Other Patient Characteristics to Mortality in a Médecins Sans Frontières Ebola Case Management Centre

Predicting Ebola Severity: A Clinical Prioritization Score for Ebola Virus Disease

Predictors of mortality in patients with yellow fever: an observational cohort study. The Lancet Infectious Diseases

SARS-CoV-2 Viral Load in Upper Respiratory Specimens of Infected Patients

Temporal dynamics in viral shedding and transmissibility of COVID-19

Clinical Characteristics of Covid-19 in

Hospitalization and Mortality among Black Patients and White Patients with Covid-19

Presenting Characteristics, Comorbidities, and Outcomes Among 5700 Patients Hospitalized With COVID-19 in the New York City Area

Remdesivir for the Treatment of Covid-19 -Final Report

Repurposed Antiviral Drugs for Covid-19 -Interim WHO Solidarity Trial Results

Intensive Care Unit "#$%&'!()*+,-*.!/+*0!123456"7!8,.*.!!

+*)H! "B!()*+,-*.! 5,M,)

We would like to acknowledge the data in COVID-CARE database based at NYP/Columbia University Irving Medical Center, Division of Infectious Diseases. Funding for MES and JZ: 5UM1AI069470 and supplement to the award. Funding for EHM: 5T32AI100852-08, NIAID.

The authors have no conflicts of interest to declare.

A c c e p t e d M a n u s c r i p t ! !"#$%&'()'*+$,-'./0/%+'