Saliva or nasopharyngeal swab samples for detection of SARS-CoV-2

To the editor:

Rapid and accurate diagnostic tests are essential to control the ongoing Covid-19 epidemic. Although the current standard involves testing nasopharyngeal swab samples by quantitative reverse-transcript polymerase chain reaction (RT-QPCR) to detect SARS-Cavi-2, saliva samples may be an alternative diagnostic sample.^1-4 Strict evaluation is required to determine how saliva samples compare with nasopharyngeal swab samples in terms of sensitivity to detection of SARS-CoV-2 during infection.

A total of 70 patients with Covid-19 provided written information to participate in our study (see the Methods section in Supplementary Appendix 1 available with the full text of this letter on NEJM.org). After Kovid-19 was confirmed with a positive nasopharyngeal swab sample at the time of hospital admission, we obtained additional samples from patients during hospitalization. We examined saliva samples collected by patients themselves and nasopharyngeal swabs collected from patients at the same time by health care workers.

Figure 1. Figure 1. RNA titers in SARS-CoV-2 saliva samples and nasopharyngeal swab samples.

Samples of 70 hospital patients diagnosed with Covid-19 were obtained. The panel shows the SARS-CoV-2 RNA titers in the first available nasopharyngeal and saliva samples. The lines indicate similar patient samples. Results were compared using Wilcoxon signed rank test (P <0.001). Panel B showed the percentage of SARS-Cavi-2 in the examination of the first matching nasopharyngeal and saliva samples 1 to 5 days, 6 to 10 days, and 11 or more days (maximum, 53 days) after the diagnosis of Covid. 19. Panel C, according to the days of onset of symptoms, in saliva samples, long lintudinal SARS-CoV-2 RNA. Copies show per milliliter. Each circle represents a different pattern. Dashed lines indicate additional samples of the same patient. The red line indicates a negative saliva sample that was followed by a positive sample in the subsequent collection of the sample. Panel D, according to the days of onset of symptoms, in 97 nasopharyngeal swab specimens, shows litidudinal SARS-Covi-2 RNA copies per milliliter. The red lines indicate negative nasopharyngeal swab specimens where positive swabs appeared in the next collection of samples. Gray area samples in panels C and D indicate that 5610 virus RNA per milliliter of sample. Copies were below the lower limit of the probe, which is at the cycle threshold 38 of our quantitative reverse-transcripts polymerase chain reaction essay targeting SARS-COV. The 2N1 sequence is recommended by the Centers for Disease Control and Prevention. To analyze this data, we have used a linear mixed-effects-regression model (see Supplementary Appendix 1) for the correlation between samples collected from the same person at the same time (i.e. multivariate response) and cross-collected samples. Have a stake. Same patient time (i.e., repetitive measures). All data used to generate this figure, including the raw cycle thresholds, are provided in Supplementary Data 1 in Appendix 2.

Using primer sequences from the Centers for Disease Control and Prevention, we found more copies of SARS-Cavi-2 RNA in saliva samples (log copies per milliliter, 5.58; 95% confidence interval). [CI], 5.09 to 6.07) compared to nasopharyngeal swab samples (log copies per milliliter, 4.93; 95% CI, 4.53 to 5.33) (Figure 1a, And the supplement in Fig. 1. S1). In addition, a higher percentage of saliva samples than nasopharyngeal swab samples was positive for 10 days after COSID-19 diagnosis.Figure 1b). 1 to 5 days after diagnosis, saliva samples were positive in 81% (95% CI, 71 to 96), compared with 71% (95% CI, 67 to 94) in the nasopharyngeal swab sample. These findings suggest that saliva samples and nasopharyngeal swab samples have at least similar sensitivity to SARS-COV-2 detection during hospitalization.

Because the results of testing of nasopharyngeal swab samples to detect SARS-CoV-2 may vary with repeated sampling in individual patients,⁵ We evaluated viral probes in matching samples over time. Levels of SARS-CoV-2 RNA decreased after the onset of symptoms in both saliva samples (initial salivation, .10.11; 95% reliable interval, −0.15 to .00.06).Figure 1c) And nasopharyngeal swab samples (estimated opera, .00.09; 95% reliable interval, .10.13 to .00.05))Figure 1D). In three cases, the negative nasopharyngeal swab sample was performed by a positive swab in the next collection of subsequent samples (Figure 1D); This phenomenon occurs only once with saliva samples (Figure 1c). During the clinical course, we found standard deviations in saliva samples from nasopharyngeal swab samples, 0.98 virus RNA copies; 95% reliable interval, 0.08 to 1.98) saw little difference in the levels of SARS-Cavi-2 RNA. Deviation, copies of 2.01 virus RNA per milliliter; 95% reliable interval, 1.29 to 2.70) (see Supplementary Appendix 1).

Recent studies have shown that SARS-Covi-2 can be detected in the saliva of asymptomatic individuals and outpatients.^{1-3- 1-3} We therefore examined 495 asymptomatic health care workers who provided written information to participate in our prospective study, and we used RT-QPCR for both saliva and nasopharyngeal samples obtained from these individuals. We found SARS-CoV-2 RNA in saliva samples obtained from 13 people who reported no symptoms at or before sample collection. Of these 13 health care workers, 9 matched nasopharyngeal swab samples were collected on the same day, and 7 of these samples tested negative (Fig. S2). Diagnosis in 13 health care workers with positive saliva samples was later confirmed in a diagnostic test of additional nasopharyngeal samples by a laboratory confirmed by CLIA (1988 Clinical Laboratory Improvements).

Variations in the nasopharyngeal sample may be an explanation for the negative outcomes, so observing internal control for appropriate sample collection may provide an alternative assessment technique. In samples collected from patients by health care workers, we found human RNS rather than nasopharyngeal swab samples (standard deviation, standard deviation) of saliva samples (standard deviation, 2.89 ct; 95% CI, 26.53 to 27.69). . Further variation in P. cycle threshold (CT) values ​​was observed. , 2.49 ct; 95% CI, 23.35 to 24.35). When health care workers collected their own samples, we found that RNZ P CT values ​​were even higher in nasopharyngeal swab samples (standard deviation, 2.26 CT; 95% CI, 28.39 to 28.56) than in saliva samples (standard deviation, 1.65 CT; 95). The difference was noticeable. % CI, 24.14 to 24.26) (Fig. S3).

The collection of saliva samples by patients themselves denies the need for direct contact between health care workers and patients. This interaction is the source of the main test interruptions and presents a risk of nasopharyngeal infection. The collection of saliva samples by patients also eliminates the demand for swabs and the supply of personal protective equipment. Given the growing need for testing, our findings support the possibility of saliva samples in the diagnosis of SARS-Cavi-2 infection.

Annie L. Willley, Ph.D.
Yale School Public Health, New Haven, CT
[email protected]

John F. Ournier, MD
Yale School of Medicine, New Haven, CT

Arnau Casanovas-Masana, Ph.D.
Yale School Public Health, New Haven, CT

Melissa Campbell, MD
Maria Tokuama, Ph.D.
Pavitra Vijayakumar, B.A.
Yale School of Medicine, New Haven, CT

Joshua L. Vern Run, Ph.D.
Yale School Public Health, New Haven, CT

Bertie Gang, MD
Yale School of Medicine, New Haven, CT

like this. Catherine Munker, M.S.
Adam J. Moore, M.P.H.
Chantelle BF Vogels, Ph.D.
Mary E. Patron, B.S.
Isabel M. Ott, B.S.
Yale School Public Health, New Haven, CT

Piawen Lu, Ph.D.
Arvind Venkataraman, B.Sc.
Alice Lu-Culligan, B.S.
Jonathan Klein, B.S.
Yale School of Medicine, New Haven, CT

Rebecca Ernest, MPH
Yale School Public Health, New Haven, CT

Michael Simonov, MD
Roopak Dutta, MD, Ph.D.
Ryan Hendoko, MD
Nida Naushad, B.Sc.
Lorenzo R. Sivanan, M.Phil.
Jordan Valdez, B.S.
Yale School of Medicine, New Haven, CT

Elizabeth B. White, AB
Sarah Lepidus, MS
Cheney C. Kalinich, M.P.H.
Yale School Public Health, New Haven, CT

Xiaodong Jiang, MD, Ph.D.
Daniel J. Kim, Abby
Erico Kudo, Ph.D.
Melissa Linon, MS
Tianyang Mao, B.S.
Miu Moriama, Ph.D.
G. E. Oh, MD, Ph.D.
Ansea Park, B.A.
Julio Silva, B.S.
Eric Song, MS
Techhiro Takahashi, MD, Ph.D.
Manabu Taura, Ph.D.
RR-L Weisman, B.A.
Patrick Wong, MS
Yaxin Yang, B.S.
Santos Burmejo, B.S.
Yale School of Medicine, New Haven, CT

Camilla D Audio, MD
Yale New Haven Health, New Haven, CT

Saad B. Omar, MB, BS, Ph.D.
Yale Institute for Global Health, New Haven, CT

Charles S. Della Cruz, MD, Ph.D.
Shelley Farhadian, MD, Ph.D.
Richard A. Martinillo, MD
Akiko Iwasaki, Ph.D.
Yale School of Medicine, New Haven, CT

Nathan D. Grubo, Ph.D.
Albert I. Co, MD
Yale School Public Health, New Haven, CT
[email protected], [email protected]

This letter was dated August 28, 2020, to NEJMRG. Was published on.