key: cord-0956553-qx3pikxx authors: Echavarria, Marcela; Reyes, Noelia S.; Rodriguez, Pamela E.; Ricarte, Carmen; Ypas, Martin; Seoane, Alejandro; Querci, Marcia; Brizio, Marianela; Stryjewski, Martin E.; Carballal, Guadalupe title: Saliva screening of health care workers for SARS-COV-2 detection date: 2022-05-13 journal: Rev Argent Microbiol DOI: 10.1016/j.ram.2022.05.001 sha: c5ea9df579a4dcbed9ed1b69510c0fe44341188a doc_id: 956553 cord_uid: qx3pikxx Health care workers (HCWs) are at high risk for SARS-CoV-2. In addition, pre-symptomatic or asymptomatic transmission accounts for around half of the cases. Saliva testing is an option to detect SARS-CoV-2 infection. To determine the performance of saliva samples for screening, HCWs were tested for SARS-CoV-2 by RT-PCR. Those with a positive result in saliva were tested by nasopharyngeal swabbing for viral RNA detection and blood collection to search for the presence of specific antibodies. In September-October 2020, 100 HCWs were enrolled and followed up. Six subjects (6%) tested positive in saliva. Of them, 5 were positive in a subsequent nasopharyngeal swab and 4 developed signs and symptoms compatible with COVID-19. Among the latter, 3 seroconverted while asymptomatic HCWs remained seronegative. Saliva screening was helpful for identifying SARS-CoV-2 infection in HCWs. This screening permitted rapid personnel isolation avoiding further transmission of the virus in the hospital setting. or asymptomatic transmission accounts for around half of the cases. Saliva testing is an option to detect SARS-CoV-2 infection. To determine the performance of saliva samples for screening, HCWs were tested for SARS-CoV-2 by RT-PCR. Those with a positive result in saliva were tested by nasopharyngeal swabbing for viral RNA detection and blood collection to search for the presence of specific antibodies. In September-October 2020, 100 HCWs were enrolled and followed up. Six subjects (6%) tested positive in saliva. Of them, 5 were positive in a subsequent nasopharyngeal swab and 4 developed signs and symptoms compatible with COVID-19. Among the latter, 3 seroconverted while asymptomatic HCWs remained seronegative. Saliva screening was helpful for identifying SARS-CoV-2 infection in HCWs. This screening permitted rapid personnel isolation avoiding further transmission of the virus in the hospital setting. El personal de salud (PS) tiene un alto riesgo de contraer SARS-CoV-2. La transmisión presintomática/asintomática representa alrededor de la mitad de los casos y el testeo a partir de muestras de saliva puede ser una opción para detectar la infección. Para determinar el rendimiento de estas muestras, 100 voluntarios del PS se sometieron a la detección de SARS-CoV-2 por RT-PCR en muestras de saliva en el período septiembre-octubre de 2020. De aquellos con resultado positivo en saliva se tomaron hisopados nasofaríngeos para detectar ARN viral y muestras de suero para evaluar anticuerpos específicos. Se detectó ARN viral en la saliva de 6 individuos (6%). De ellos, 5 fueron SARS-CoV-2 positivos en hisopado nasofaríngeo y 4 desarrollaron signos y síntomas compatibles con COVID-19. Entre estos últimos, 3 seroconvirtieron, en tanto que los voluntarios asintomáticos permanecieron seronegativos. La muestra de saliva fue útil para identificar la infección por SARS-CoV-2 en esta cohorte del personal de salud y así proceder al rápido aislamiento de los individuos infectados, lo que evitó una mayor transmisión del virus en el ámbito hospitalario. Keyword. Saliva, screening, Health care workers, asymptomatic, PCR, SARS-CoV-2. Palabras clave. Saliva; testeo; personal de salud; asintomático; PCR; SARS-CoV-2 high risk for SARS-CoV-2 infection. Pre-symptomatic or asymptomatic transmission account for around half of the cases. Therefore, increasing capacity and early diagnostic testing would help to reduce the transmission of the virus 5 . Screening approaches are mostly focused on symptomatic HCWs 16 and there are few studies detecting SARS-CoV-2 in pre-symptomatic or asymptomatic HCW 11, 12 . The nasopharyngeal swab (NPS) is the most common respiratory sample utilized for SARS-CoV-2 diagnosis. However, NPS is associated with subjects' discomfort and further exposure to viral aerosols during sample collection 8 . Saliva testing is a non-invasive test to detect SARS-CoV-2 that avoids further HCW exposure. Saliva testing proved to be as sensitive as NPS in detecting SARS-CoV-2 in symptomatic patients 6, 7 . However, the role of saliva testing among asymptomatic HCWs is still to be determined. The objective of this study was to determine the performance of the saliva test to screen for SARS-CoV-2 in HCWs. A descriptive prospective cohort study in HCWs was conducted at CEMIC University Hospital, Buenos Aires, Argentina. Voluntary SARS-CoV-2 testing in saliva was performed in both hospital sites: CEMIC Saavedra and CEMIC Pombo. HCWs were enrolled at 12 different hospital settings. These settings were classified as "high", "middle" or "low" in terms of exposure risk to SARS-CoV-2. High exposure risk settings included: emergency room, intensive care unit (ICU), infectious diseases, and personnel assisting COVID-19 positive inpatients or specimens (technicians, nurses, physicians, physical therapists, laboratory). Middle exposure risk settings included: personnel assisting COVID-19 negative inpatients (technicians, nurses, physicians, physical therapists). Low exposure risk areas An electronic form with personal data and information related to COVID-19 disease or exposure was collected at the time of sampling. Clinical follow-up for any signs or symptoms compatible with COVID-19 was obtained from participants for two weeks from the initial collection of saliva. A nasopharyngeal swab and a blood sample for serology were obtained from participants with positive RT-PCR results for SARS-CoV-2. The study was approved by the CEMIC Ethics Committee (Protocol: 1298/20) and electronic informed consent was obtained from all participants. At study entry, a self-collected saliva sample was obtained for SARS-CoV-2 diagnosis. Participants were instructed on how to collect the sample in a sterile plastic container without viral transport medium. Samples were conserved at 4°C until processed in a biosafety cabinet within 12 hours of arrival. Viscous saliva samples were mechanically disrupted by adding 500 µl viral transport medium (Minimun Essential Medium (Gibco); L-Glutamin 200 mM; HEPES 1N; bovine serum albumin 5% (Sigma) sodium bicarbonate 7.5%; penicillin, streptomycin and amphotericin (pH=7.2). Participants in whom SARS-CoV-2 was detected by PCR in the initial sample of saliva were requested to provide additional saliva samples and NPS. NPS were obtained and placed in a sterile tube conaining 2 ml viral transport media. Nucleic acid was extracted from 100µl saliva sample and eluted in 15µl using manual columns (Quick-RNA TM Viral Kit. Zymo Research CORP.) following the manufacturer's recommendation. One-step real-time multiplex RT-PCR laboratory-optimized was performed, targeting the SARS-CoV-2 E gene and the human RNAse P gene as quality From September 9th to October 13th 2020, 100 asymptomatic/ presymptomatic HCWs were enrolled in the study. Demographic characteristics were described as appropriate. Almost two thirds were female (67%) and the median age was 37 years old (IQR=31-46). Around half of the participants (53%) described having been in contact with COVID-19 patients at some point and 3.8% of these participants reported previous SARS-CoV-2 infection, but were negative in saliva or NPS sample at the time of this screening. Of 100 HCWs, 6 asymptomatic subjects (6%) were positive in saliva (2 physicians, 2 nurses, and 2 administrative personnel, respectively). All positive HCWs belonged to high exposure settings. Upon the initial results in saliva, all these HCWs were separated and licensed from work on the same day of sample collection. NPS from positive patients were mostly obtained within 24-48 hours from saliva testing. Of 6 HCWs with positive saliva, 5 (83%) were positive in subsequent NPS (Table 1) Expanding screening protocols for detecting pre-symptomatic or asymptomatic carriers among HCWs is critical to reduce the spread of SARS-CoV-2 in the hospital setting. Some of the challenges to establish these protocols are related to logistical issues, turnaround times and further exposure to HCW during sampling. Self-collected saliva provides an option to facilitate sample collection, reduce discomfort and minimize HCW exposure. There are few reports utilizing saliva samples in the routine testing of asymptomatic or pre-symptomatic patients, including HCWs 15 . Generally, saliva tests were utilized for follow-up after a positive NPS but not as a primary screening method 13 . In this study, we have used saliva samples as a primary screening method to detect SARS-CoV-2 among HCWs. The incidence of SARS-CoV-2 in pre-symptomatic/asymptomatic HCWs reaching 6% was higher than expected. Most of our positive cases corresponded to HCWs performing work activities in high or middle risk areas. Some authors reported negative results for saliva testing among HCWs 1 while others found percentages ranging from 1.5 to 12.5% using NPS in those asymptomatic 12, 14, 16 . Our screening results in saliva are comparable with studies using NPS. patients using a highly sensitive home brew RT-PCR 6 . In this study, saliva was a useful, valuable and easy tool for SARS-CoV-2 screening and for identifying pre-symptomatic and asymptomatic individuals. Importantly, rapid identification of positive HCWs permitted early licensing avoiding potential SARS-CoV-2 spreaders within the hospital setting. Furthermore, saliva positivity anticipated clinical disease up to 3 days before symptom onset. Long-term SARS-CoV-2 RNA shedding can occur in saliva. In our study, the only patient who gave a discrepant result between NPS and saliva and remained asymptomatic showed a follow-up saliva with a higher Ct value, probably representing prolonged shedding in this sample type 17 . Of 6 positive HCWs, 2 remained asymptomatic. Studies evaluating nasopharyngeal swabs in HCWs demonstrated that 15% to 57% remained asymptomatic 2, 11 . Seroconversion was demonstrated in most of our symptomatic patients. The patient who did not seroconvert even 90 days after symptom onset had a very mild disease. This finding is not unexpected since IgG titers have been associated with disease severity 9 . In addition, lack of seroconversion was previously described in 17% of HCWs infected with SARS-CoV-2 3 . Saliva is an easy to obtain sample that can be self-collected. It is especially convenient for screening in individuals without respiratory symptoms. Furthermore, special attention should be given to asymptomatic individuals who may spread the virus more easily due to higher human interaction than symptomatic patients 10 . Interestingly, previous studies with saliva testing have shown discrepant results. We believe the reason for such discrepancy is related to sample processing and the PCR techniques employed. We have systematically conducted and applied an optimized PCR (home brew). Our technique includes mechanical disruption of the saliva, no addition of any stabilizer or buffer and an increased concentration of magnesium (3.8 mM) in the PCR mix as well as modifications in cycling conditions 6 . sensitivity. Furthermore, this home brew assay incurred less costs than those related to commercial PCR kits (data not shown). This study has several limitations. Our investigation was based on voluntary participation and this may have introduced some voluntary bias. In addition, other populations need to be tested to define the role of saliva to detect SARS-CoV-2 in symptomatic, pre-symptomatic or asymptomatic subjects. Finally, our sample size was not large. However, the number was deemed to be epidemiologically meaningful. Supporting this concept, we were able to detect 6% of HCWs infected with SARS-CoV-2 while they were pre-symptomatic/asymptomatic. Prevalence of positive COVID-19 among asymptomatic health care workers who care patients infected with the novel coronavirus: A retrospective study Occurrence and transmission potential of asymptomatic and presymptomatic SARSCoV-2 infections: A living systematic review and metaanalysis Risk for SARS-CoV-2 infection in healthcare workers Detection of 2019 novel coronavirus (2019-nCoV) by real-time RT-PCR Covid-19: identifying and isolating asymptomatic people helped eliminate virus in Italian village Self-collected saliva for SARS-CoV-2 detection: a prospective study in the emergency room Challenges in use of saliva for detection of SARS CoV-2 RNA in symptomatic outpatients SARS-CoV-2 and saliva as a diagnostic tool: A real possibility. Pesqui Bras Odontopediatria Clin Integr Emergency response for evaluating SARS-CoV-2 immune status, seroprevalence and convalescent plasma in Argentina Asymptomatic transmission of covid-19 Screening of healthcare workers for SARS-CoV-2 highlights the role of asymptomatic carriage in COVID-19 transmission COVID-19: PCR screening of asymptomatic health-care workers at London hospital Saliva samples for detection of SARS-CoV-2 in mildly symptomatic and asymptomatic patients Prevalence of SARS-CoV-2 Infection Among Asymptomatic Health Care Workers in the Greater Routine saliva testing for the identification of silent coronavirus disease 2019 (COVID-19) in healthcare workers Asymptomatic infection by SARS-CoV-2 in healthcare workers: A study in a large teaching hospital in Wuhan, China Viral dynamics of SARS-CoV-2 in saliva from infected patients