Understanding the Demographic, Clinical, and Real-Time Polymerase Chain Reaction Profiles of COVID-19 Patients in a Tertiary Care Hospital in Northeast India.

Introduction and aims The demographic and clinical profile and dynamics of real-time polymerase chain reaction (RT-PCR) in coronavirus disease 2019 (COVID-19) patients are not well understood. The study aimed to analyze the demographic, clinical, and RT-PCR profiles of COVID-19 patients. Methodology The study was a retrospective, observational study conducted at a COVID-19 care facility, and the study period was from April 2020 to March 2021. Patients with laboratory-confirmed COVID-19 by real-time polymerase chain reaction (RT-PCR) were enrolled in the study. Patients with incomplete details or with only single PCR tests were excluded. Demographic and clinical details and the results of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RT-PCR collected at different time points were retrieved from the records. The statistical software Minitab version 17.1.0 package (Minitab, LLC, State College, PA, USA) and Rstudio version 1.3.959 (Rstudio, Boston, MA, USA) were used for the statistical analysis. Results The mean duration from symptom onset to the last positive RT-PCR was 14.2 ± 4.2 days. The proportions of positive RT-PCR tests were 100%, 40.6%, 7.5%, and 0% at the end of the first, second, third, and fourth weeks of illness. The median duration of days to first negative RT-PCR in the asymptomatic patients was 8 ± 4 days, and 88.2% of asymptomatic patients were RT-PCR-negative within 14 days. A total of 16 symptomatic patients had prolonged positive test results even after three weeks of symptom onset. Older patients were associated with prolonged RT-PCR positivity. Conclusion This study revealed that the average period of RT-PCR positivity from the onset of symptoms is >2 weeks in symptomatic COVID-19 patients. Prolonged observation in the elderly population and repeat RT-PCR before discharge or discontinuation of quarantine is required.

In-house reverse transcriptase polymerase chain reaction for detection of SARS-CoV-2 with increased sensitivity.

As the COVID-19 infection continues to ravage the world, the advent of an efficient as well as the economization of the existing RT-PCR based detection assay essentially can become a blessing in these testing times and significantly help in the management of the pandemic. This study demonstrated an innovative and rapid corroboration of COVID-19 test based on innovative multiplex PCR. An assessment of optimal PCR conditions to simultaneously amplify the SARS-CoV-2 genes E, S and RdRp has been made by fast-conventional and HRM coupled multiplex real-time PCR using the same sets of primers. All variables of practical value were studied by amplifying known target-sequences from ten-fold dilutions of archived positive samples of COVID-19 disease. The multiplexing with newly designed E, S and RdRp primers have shown an efficient amplification of the target region of SARS-CoV-2. A distinct amplification was observed in 37 min using thermal cycler while it took 96 min in HRM coupled real time detection using SYBR green over a wide range of template concentrations. Our findings revealed decent concordance with other commercially available detection kits. This fast HRM coupled multiplex real-time PCR with SYBR green approach offers rapid and sensitive detection of SARS-CoV-2 in a cost-effective manner apart from the added advantage of primer compatibility for use in conventional multiplex PCR. The highly reproducible novel approach can propel extended applicability for developing sustainable commercial product besides providing relief to a resource limited setting.

Laboratory preparedness for SARS-CoV-2 testing in India: Harnessing a network of Virus Research & Diagnostic Laboratories.

Background & objectives: An outbreak of respiratory illness of unknown aetiology was reported from Hubei province of Wuhan, People's Republic of China, in December 2019. The outbreak was attributed to a novel coronavirus (CoV), named as severe acute respiratory syndrome (SARS)-CoV-2 and the disease as COVID-19. Within one month, cases were reported from 25 countries. In view of the novel viral strain with reported high morbidity, establishing early countrywide diagnosis to detect imported cases became critical. Here we describe the role of a countrywide network of VRDLs in early diagnosis of COVID-19. Methods: The Indian Council of Medical Research (ICMR)-National Institute of Virology (NIV), Pune, established screening as well as confirmatory assays for SARS-CoV-2. A total of 13 VRDLs were provided with the E gene screening real-time reverse transcription-polymerase chain reaction (rRT-PCR) assay. VRDLs were selected on the basis of their presence near an international airport/seaport and their past performance. The case definition for testing included all individuals with travel history to Wuhan and symptomatic individuals with travel history to other parts of China. This was later expanded to include symptomatic individuals returning from Singapore, Japan, Hong Kong, Thailand and South Korea. Results: Within a week of standardization of the test at NIV, all VRDLs could initiate testing for SARS-CoV-2. Till February 29, 2020, a total of 2,913 samples were tested. This included both 654 individuals quarantined in the two camps and others fitting within the case definition. The quarantined individuals were tested twice - at days 0 and 14. All tested negative on both occasions. Only three individuals belonging to different districts in Kerala were found to be positive. Interpretation & conclusions: Sudden emergence of SARS-CoV-2 and its potential to cause a pandemic posed an unsurmountable challenge to the public health system of India. However, concerted efforts of various arms of the Government of India resulted in a well-coordinated action at each level. India has successfully demonstrated its ability to establish quick diagnosis of SARS-CoV-2 at NIV, Pune, and the testing VRDLs.