Saliva as a Potential Clinical Specimen for Diagnosis of SARS-CoV-2 (preprint)

Background: There is no sign of stopping the spread of corona virus disease- 2019 (COVID-19) pandemic since it has started in December 2019. Rapid and early detection is extremely crucial to slow down the quick spread of the virus and break the human transmission chain. There are very few studies in search of an alternate and convenient diagnostic tool which can substitute nasopharyngeal swab (NPS) specimen for detection of severe acute respiratory syndrome coronavirus- 2 (SARS-CoV-2). We aimed to analyse the comparison and agreement between the feasibility of using the saliva in comparison to NPS for diagnosis of SARS-CoV-2.Methods: A total number of 74 patients were enrolled for this study. We analysed and compared the NPS and saliva specimen collected within 48 h after the symptom onset. We used real time quantitative polymerase chain reaction (RT-qPCR), gene sequencing for the detection and determination SARS-CoV-2 specific genes. Phylogenetic tree was constructed to establish the isolation of viral RNA from saliva. We use Bland-Altman model to identify the agreement between two specimens.Findings: This study shows a lower cycle threshold (CT) mean value for the detection of SARS-CoV-2 ORF1 gene (mean 27.07; 95% CI, 25.62 to 28.52) in saliva methods than that of NPS (mean 28.24; 95% CI, 26.62 to 29.85) specimen although the difference is statistically non-significant (p>0.05). Bland-Altman analysis produces relatively smaller bias and high agreement between these two clinical specimens. Phylogenetic analysis with the RdRp and Spike gene confirmed the presence of SARS-CoV-2 in the saliva samples.Interpretation: In conclusion, our study highlights that saliva represents a promising tool in COVID-19 diagnosis and the collection method would reduce the exposure risk of frontline health workers which is one of major concerns in primary healthcare settings.Funding Statement: ICMR for provided financial grants for this study.Declaration of Interests: The authors declare that they have no competing interests.Ethics Approval Statement: The study was approved by Institutional Ethics Committee and written informed consent was obtained from the study subjects.

Feasibility of Pooled Testing in a Fully Automated High Throughput Platform for Effective Management of SARS-COV-2 in India (preprint)

Introduction: Testing for extreme acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been hit tremendously worldwide, as the scarcity of test kits has now become a major bottleneck, with the need for testing throughput growing. The study assessed the feasibility of pooled testing in the high throughput machine of Roche Cobas 6800 rapidly increasing the testing number for containing the virus spread and management of cases. Methodology: ICMR-RMRC, Bhubaneswar receives samples from various district hospitals from different districts of Odisha. Pooled testing was conducted in 2 methods. Firstly, we adopted the simple two-stage testing algorithm known as Dorfman pooling with minor modification to avoid selection bias and secondly we evaluated the ability of COBAS for detection of a single positive sample within a pool of negative samples.Results: The Cobas 6800 was able to detect the SARS-COV virus in all the samples, however, the amplified RNA reached the threshold later as the number of negative samples increased in the pool. We demonstrated strategies for pooling, which improve test efficiency and while maintaining high sensitivity in a high-throughput system. The comparison of 1410 samples tested individually and in pools of five samples/pool showed that test results were not significantly affected. Implementing the five-sample Dorfman pooling to test 1410 samples, we identified 42 (2.9%) SARS-CoV-2 positive samples, achieving a 3-fold increase in throughput with one-third of the cost.Conclusion: To the best of our knowledge this study is 1st of its kind to assess the feasibility of pooled testing in high throughput machines. As ICMR has always stressed, aggressive timely testing is the standard protocol for containing the virus spread and management of cases, these high throughput machines installed in six states with a high prevalence of Covid-19, can effectively increase the testing capacity by 2-3 fold, adopting the pooled testing strategy for successful management of SARS-CoV-2.Funding: The study was carried out with funding support from Indian Council of Medical Research.Declaration of Interest: The authors declare that they have no competing interests.Ethical Approval Statement: The study was cleared by ICMR-Regional Medical Research Centre ethical committee.