Clinical, Laboratory, and Radiologic Characteristics of Patients With Initial False-Negative Severe Acute Respiratory Syndrome Coronavirus 2 Nucleic Acid Amplification Test Results

Abstract

Background: Concerns about false-negative (FN) severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleic acid amplification tests (NAATs) have prompted recommendations for repeat testing if suspicion for coronavirus disease 2019 (COVID-19) infection is moderate to high. However, the frequency of FNs and patient characteristics associated with FNs are poorly understood.

Methods: We retrospectively reviewed test results from 15 011 adults who underwent ≥1 SARS-CoV-2 NAATs; 2699 had an initial negative NAAT and repeat testing. We defined FNs as ≥1 negative NAATs followed by a positive NAAT within 14 days during the same episode of illness. We stratified subjects with FNs by duration of symptoms before the initial FN test (≤5 days versus >5 days) and examined their clinical, radiologic, and laboratory characteristics.

Results: Sixty of 2699 subjects (2.2%) had a FN result during the study period. The weekly frequency of FNs among subjects with repeat testing peaked at 4.4%, coinciding with peak NAAT positivity (38%). Most subjects with FNs had symptoms (52 of 60; 87%) and chest radiography (19 of 32; 59%) consistent with COVID-19. Of the FN NAATs, 18 of 60 (30%) were performed early (ie, ≤1 day of symptom onset), and 18 of 60 (30%) were performed late (ie, >7 days after symptom onset) in disease. Among 17 subjects with 2 consecutive FNs on NP NAATs, 9 (53%) provided lower respiratory tract (LRT) specimens for testing, all of which were positive.

Conclusions: Our findings support repeated NAATs among symptomatic patients, particularly during periods of higher COVID-19 incidence. The LRT testing should be prioritized to increase yield among patients with high clinical suspicion for COVID-19.

Keywords: COVID-19 testing; coronavirus; false-negative.

Figure 1.

Cohort flow chart demonstrating inclusion and exclusion criteria of subjects in the study. We evaluated subjects who underwent a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleic acid amplification test (NAAT) at Massachusetts General Hospital between March 3 and May 18, 2020 for inclusion in the false-negative test cohort. Subjects with only 1 negative NAAT, an initial positive NAAT at Massachusetts General Hospital (MGH), or documentation of ≥1 prior positive NAATs were not eligible. We excluded subjects with ≥1 repeat SARS-CoV-2 NAATs if all of their NAATs were negative, the initial negative and subsequent positive tests were >14 days apart, a history of symptoms could not be obtained, or 2 reviewers reached consensus that the subject’s discordant tests were performed across separate illness episodes. A total of 60 subjects were included in the false-negative NAAT cohort. *, Initial positive NAAT performed at an outside facility or performed before the study window.

Figure 2.

Weekly proportions of initial positive and false-negative severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleic acid amplification tests (NAATs) during the coronavirus disease 2019 (COVID-19) surge. We examined the percentage of subjects with positive SARS-CoV-2 NAATs (gray bars, left y-axis) among all subjects with SARS-CoV-2 NAATs in relation to the percentage of false-negative (ie, initially negative, repeat-positive) NAATs (red circles, right y-axis) each week throughout the initial COVID-19 surge in Boston, Massachusetts. We calculated the percentage of false-negative tests as the proportion of subjects with an initial negative NAAT who had a subsequent positive NAAT during the same episode of illness among all subjects who had repeat SARS-CoV-2 testing within a 14-day window. Subjects with multiple negative tests before a positive test were counted only once, on the date of their first negative test in the series. (*) indicates weeks with omitted proportion calculations because a 14-day testing window was not available within the study period.

Figure 3.

Cycle threshold (Ct) trajectories categorized by most likely cause of initial false-negative nucleic acid amplification test (NAAT). Among the 60 subjects with false-negative NAATs, we plotted all available severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) NAATs obtained after their initial false-negative test, by number of days postsymptom onset. Tests obtained from a nasopharyngeal (NP) specimen are shown as dots of increasing size based on increasing SARS-CoV-2 viral load. Positive tests are depicted in black (if the Ct value was known) or gray (if the Ct value was unknown). Negative tests are depicted in white. The intervals from the initial negative test to the first positive test, and from the last positive test to any subsequent negative tests, are shown as dotted lines; these intervals signify times of suspected active infection. Intervals between positive tests are shown as solid lines to depict documented active infection. Test patterns were depicted in 5 categories corresponding to the observed viral burden pattern. LRT, lower respiratory tract; URT, upper respiratory tract.