ABSTRACT
Coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2, is posing a serious threat to global public health. Reverse transcriptase real-time quantitative polymerase chain reaction (qRT-PCR) is widely used as the gold standard for clinical detection of SARS-CoV-2. Due to technical limitations, the reported positive rates of qRT-PCR assay of throat swab samples vary from 30%-60%. Therefore, the evaluation of alternative strategies to overcome the limitations of qRT-PCR is required. A previous study reported that one-step nested (OSN)-qRT-PCR revealed better suitability for detecting SARS-CoV-2. However, information on the analytical performance of OSN-qRT-PCR is insufficient. In this study, we aimed to analyze OSN-qRT-PCR by comparing it with droplet digital PCR (ddPCR) and qRT-PCR by using a dilution series of SARS-CoV-2 pseudoviral RNA and a quality assessment panel. The clinical performance of OSN-qRT-PCR was also validated and compared with ddPCR and qRT-PCR using specimens from COVID-19 patients. The LoD (copies/ml) of qRT-PCR, ddPCR, and OSN-qRT-PCR were 520.1 (95% CI): 363.23-1145.69) for ORF1ab and 528.1 (95% CI: 347.7-1248.7) for N, 401.8 (95% CI: 284.8-938.3) for ORF1ab and 336.8 (95% CI: 244.6-792.5) for N, and 194.74 (95% CI: 139.7-430.9) for ORF1ab and 189.1 (95% CI: 130.9-433.9) for N, respectively. Of the 34 clinical samples from COVID-19 patients, the positive rates of OSN-qRT-PCR, ddPCR, and qRT-PCR were 82.35% (28/34), 67.65% (23/34), and 58.82% (20/34), respectively. In conclusion, the highly sensitive and specific OSN-qRT-PCR assay is superior to ddPCR and qRT-PCR assays, showing great potential as a technique for detection of SARS-CoV-2 in patients with low viral loads.
ABSTRACT
Novel coronavirus pneumonia (NCP) is an emerging, highly contagious community acquired pneumonia (CAP) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Highly efficient and accurate microbiological laboratory assay is essential to confirm the SARS-CoV-2 infection, rule out other pathogens that can cause CAP, and monitor secondary infections. Here, we enrolled and provide microbiological analysis for 129 suspected and 52 transferred confirmed NCP patients hospitalized in the First Affiliated Hospital of University of Science and Technology of China (USTC) from Jan 21 to Feb 29, 2020. By analyzing the dual swab samples (sputum and pharyngeal) from 129 suspected patients with realtime RT-PCR, we confirmed 33 SARS-CoV-2 infections, with two co-infection cases with adenovirus or rhinovirus. We also used multiplex PCR to detect 13 common respiratory tract pathogens in 96 non-NCP patients, and found that 30 patients (31.25%) were infected with at least one respiratory tract pathogen that may cause CAP. Further, we performed bacterial and fungal cultures as well as fungal serologic tests and found that there is no secondary bacterial/fungal infections in confirmed NCP patients. Our studies suggest that, during the epidemic of NCP in Anhui province, there was a certain proportion of infection and co-infection of other common pathogens of CAP, and the secondary bacterial and fungal infection is not detectable in NCP patients. In comparison with SARS-CoV-2 detection alone, this optimized strategy combining multiple pathogen detection for identification of NCP and other CAP patients as well as cultures and serologic tests for confirmed patients increases the diagnosis efficiency and facilitates the personalized medication.
ABSTRACT
Objective To investigate the feasibility of self‐control materials serum thyroid hormone detected in the clinical labo‐ratory internal quality control in applications .Methods The serum thyroid hormone test results in patients with high‐value ,the ser‐um was mixed with a certain proportion of the healthy population ,dubbed serum control materials ,after aliquots -20 ℃ .Every o‐riginal supporting quality control and simultaneous detection of free thyroxine (FT4) ,three free triiodothyronine (FT3) ,thyroid stimulating hormone (TSH) ,thyroglobulin (TG) ,thyroid‐microglobulin antibody (TMA) ,thyroglobulin antibodies (TGA) and other thyroid hormone six items detected before the appearance of control samples was observed continuously detected six months to observe stability ;used EXCELL do J‐L maps ,statistical mean of each project within six months ,CV and uncontrolled number of observed values in the internal quality control applications .Results Within 6 months after the product was frozen homemade quali‐ty control serum soluble appearance clear and transparent ,no precipitation and turbidity ;1 ,3 ,6 ,10 ,15 ,30 ,45 ,60 ,90 ,120 day of the first magnitude and no significant changes in thyroid hormone ;compared with the original quality control materials ,low CV values . In the six months internal quality control applications ,the cumulative results of each project without 1 ,3 ,6‐month and six months significantly different ;project and six months mean cumulative comparison between the mean drift or change in trend did not ap‐pear ;items and cumulative CV were in control of the allowable range (CV <1/3 times within total variation );6 months of internal quality control work ,runaway alarm :13s ,2 times ,22s ,1 times ,R4s ,1 times .Conclusion Serum thyroid hormone homemade frozen control samples have better stability ,and can replace its import control materials ;internal quality control in their daily activities , through EXCELL test results for J‐L diagram precision can find instability timely corrective treatment to ensure that thyroid func‐tion test results are reliable ,worthy of clinical application .
