Comparison of Severe Acute Respiratory Syndrome Coronavirus 2 Screening Using Reverse Transcriptase-Quantitative Polymerase Chain Reaction or CRISPR-Based Assays in Asymptomatic College Students

Abstract

Importance: The reopening of colleges and universities in the US during the coronavirus disease 2019 (COVID-19) pandemic is a significant public health challenge. The development of accessible and practical approaches for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection in the college population is paramount for deploying recurrent surveillance testing as an essential strategy for virus detection, containment, and mitigation.

Objective: To determine the prevalence of SARS-CoV-2 in asymptomatic participants in a university community by using CREST (Cas13-based, rugged, equitable, scalable testing), a CRISPR-based test developed for accessible and large-scale viral screening.

Design, setting, and participants: For this cohort study, a total of 1808 asymptomatic participants were screened for SARS-CoV-2 using a CRISPR-based assay and a point-of-reference reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) test. Viral prevalence in self-collected oropharyngeal swab samples collected from May 28 to June 11, 2020, and from June 23 to July 2, 2020, was evaluated.

Exposures: Testing for SARS-CoV-2.

Main outcomes and measures: SARS-CoV-2 status, viral load, and demographic information of the study participants were collected.

Results: Among the 1808 participants (mean [SD] age, 27.3 [11.0] years; 955 [52.8%] female), 732 underwent testing from May to early June (mean [SD] age, 28.4 [11.7] years; 392 [53.6%] female). All test results in this cohort were negative. In contrast, 1076 participants underwent testing from late June to early July (mean [SD] age, 26.6 [10.5] years; 563 [52.3%] female), with 9 positive results by RT-qPCR. Eight of these positive samples were detected by the CRISPR-based assay and confirmed by Clinical Laboratory Improvement Amendments-certified diagnostic testing. The mean (SD) age of the positive cases was 21.7 (3.3) years; all 8 individuals self-identified as students. These metrics showed that a CRISPR-based assay was effective at capturing positive SARS-CoV-2 cases in this student population. Notably, the viral loads detected in these asymptomatic cases resemble those seen in clinical samples, highlighting the potential of covert viral transmission. The shift in viral prevalence coincided with the relaxation of stay-at-home measures.

Conclusions and relevance: These findings reveal a shift in SARS-CoV-2 prevalence in a young and asymptomatic population and uncover the leading edge of a local outbreak that coincided with rising case counts in the surrounding county and the state of California. The concordance between CRISPR-based and RT-qPCR testing suggests that CRISPR-based assays are reliable and offer alternative options for surveillance testing and detection of SARS-CoV-2 outbreaks, as is required to resume operations in higher-education institutions in the US and abroad.

Figure 1.. Detection of Positive Samples by CRISPR-Based and Reverse Transcriptase–Quantitative Polymerase Chain Reaction (RT-qPCR) Assays

A, Distribution of the fluorescence values by cohort and correlation of nucleocapsid sites N1 and N2 signals by CRISPR-based assay. B, Distribution of the 1/quantification cycle (Cq) values by cohort and correlation of N1 and N2 signals detected by RT-qPCR. Cohort 1 underwent testing from May 28 to June 11, 2020; cohort 2, from June 23 to July 2, 2020. The blue dot indicates 1 sample detected by RT-qPCR but not confirmed by the CRISPR-based assay or by a diagnostic test (note the low level of N2 for this sample). The dashed line indicates the detection limit for RT-qPCR (N1, 1/Cq 0.0306; N2, 1/Cq 0.029). AU indicates arbitrary unit.

Figure 2.. Flow Diagram of Sample Collection and Processing

Self-collected oropharyngeal swabs were processed for severe acute respiratory syndrome coronavirus 2 testing using CRISPR-based or reverse transcriptase–quantitative polymerase chain reaction (RT-qPCR) assays. Positive results were confirmed with diagnostic testing in a Clinical Laboratory Improvement Amendments–certified laboratory. Following confirmation, Santa Barbara Cottage Hospital clinicians reported the positive results to the participants and the Santa Barbara County Public Health Department (SBCPHD). CREST indicates Cas13-based, rugged, equitable, scalable testing.

Figure 3.. Viral Loads in Asymptomatic and Confirmed Positive Individuals

A, Viral loads were calculated using the reverse transcriptase–quantitative polymerase chain reaction (RT-qPCR) data for nucleocapsid sites N1 and N2 detection. B, Ribonuclease P (RNaseP) copies were calculated using the RT-qPCR data for this host gene target. Our analyses included the 8 samples with positive test results we detected in cohort 2, which were confirmed by diagnostic testing. Residual clinical samples from patients with known positive test results (n = 6) provided to us by our collaborators at the Santa Barbara County Public Health Department were used as controls. Solid horizontal lines indicate medians. Differences were not significant between N1 asymptomatic and symptomatic samples (P = .95, Mann-Whitney test), N2 asymptomatic and symptomatic samples (P = .50, Mann-Whitney test), or asymptomatic and symptomatic RNaseP samples (P = .95, Mann-Whitney test).

Figure 4.. Daily Prevalence of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) in the Study Population

The dots represent daily SARS-CoV-2 prevalence. The trend line, indicated by the orange line, was calculated by finding the r in a logistic growth model that minimized the error while fixing the percentage prevalence on May 28 to 0.03%. The blue bars represent the cumulative daily number of diagnosed COVID-19 cases in the Goleta and Isla Vista communities based on official data from the Santa Barbara County Public Health Department. The arrow indicates relaxation of stay-at-home measures in the County of Santa Barbara.