Correction: Comparative analysis of various clinical specimens in detection of SARS-CoV-2 using rRT-PCR in new and follow up cases of COVID-19 infection: Quest for the best choice.

[This corrects the article DOI: 10.1371/journal.pone.0249408.].

Comparative analysis of various clinical specimens in detection of SARS-CoV-2 using rRT-PCR in new and follow up cases of COVID-19 infection: Quest for the best choice.

BACKGROUND: An appropriate specimen is of paramount importance in Real Time reverse transcription-polymerase chain reaction (rRT-PCR) based diagnosis of novel coronavirus (nCoV) disease (COVID-19). Thus, it's pertinent to evaluate various diversified clinical specimens' diagnostic utility in both diagnosis and follow-up of COVID-19. METHODS: A total of 924 initial specimens from 130 COVID-19 symptomatic cases before initiation of treatment and 665 follow up specimens from 15 randomly selected cases comprising of equal number of nasopharyngeal swab (NPS), oropharyngeal swab (OPS), combined NPS and OPS (Combined swab), sputum, plasma, serum and urine were evaluated by rRT-PCR. RESULTS: Demographic analysis showed males (86) twice more affected by COVID-19 than females (44) (p = 0.00001). Combined swabs showed a positivity rate of 100% followed by NPS (91.5%), OPS (72.3%), sputum (63%), while nCoV was found undetected in urine, plasma and serum specimens. The lowest cycle threshold (Ct) values of targeted genes E, ORF1b and RdRP are 10.56, 10.14 and 12.26 respectively and their lowest average Ct values were found in combined swab which indicates high viral load in combined swab among all other specimen types. Analysis of 665 follow-up multi-varied specimens also showed combined swab as the last specimen among all specimen types to become negative, after an average 6.6 (range 4-10) days post-treatment, having lowest (15.48) and average (29.96) Ct values of ORF1b respectively indicating posterior nasopharyngeal tract as primary nCoV afflicted site with high viral load. CONCLUSION: The combined swab may be recommended as a more appropriate specimen for both diagnosis and monitoring of COVID-19 treatment by rRT-PCR for assessing virus clearance to help physicians in taking evidence-based decision before discharging patients. Implementing combined swabs globally will definitely help in management and control of the pandemic, as it is the need of the hour.

Effective pragmatic approach of diagnosis of multidrug-resistant tuberculosis by high-resolution melt curve assay.

Background: Effective management of multidrug-resistant tuberculosis (MDR-TB) requires cost-effective and rapid screening of rifampicin (RIF) and isoniazid (INH) resistance. Accordingly, a highly promising high-resolution melting (HRM) analysis was evaluated in the detection of mutation in rpoB, katG gene and inhA promoter region in Mycobacterium tuberculosis isolates. Methods: A total of 143 M. tuberculosis isolates comprising phenotypically confirmed 94 MDR and 49 sensitive isolates were analyzed by HRM following real-time-polymerase chain reaction in comparison to gold standard of targeted DNA sequencing of rpoB, katG gene and inhA promoter region. Results: HRM correctly identified MDR-TB by rapid and accurate detection of predominantly and infrequently occurring specific single nucleotide polymorphism in rpoB, katG gene and inhA promoter region. rpoB HRM showed sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of 98% each respectively. Predominantly, S531 L/W (TCG → TTG/TGG) mutation accounted for 68.47% of RIF resistance followed by H526Y/R (13.04%, CAC → TAC/CGC), D516Y/V/G (10.86%, GAC → TAC/GTC/GGC), Q513P (4.34%, CAA → CCA), and one rare mutation at codon position L533A (CTG → CGG). Combined KatG and inhA HRM sensitivity, specificity, PPV, and NPV were 90%, 100%, 100%, and 84.48% respectively and detected frequent mutation at codon position S315T/I/N (70%, AGC → ACC, AGC → ACT, AGC → AAC) and rare mutation at codon position T314P (3.3%, ACC → CCC) and 329 (2.2%, GAC → GCC) of katG gene. In inhA, mutations were recorded at mostly promoter position - 15 (10%, C → T) and infrequently at - 8 (3.3%, T → G, T → C). HRM assay limitation noticed in recognizing silent mutation in rpoB as a mutant, nondetection of infrequent mutation S310A in katG, and the inability of detecting mutation outside the targeted region of investigated genes. Conclusion: HRM may prove to be a vital molecular assay in rapid screening of TB cases for early detection of MDR TB, leading to early evidenced-based initiation of antitubercular treatment that will significantly reduce MDR transmission.