Combined Diagnosis of SARS-CoV-2: Rapid Antigen Detection as an Adjunct to Nucleic Acid Detection.

Coronavirus disease 2019 is a serious threat to human life, and early diagnosis and screening can help control the COVID-19 pandemic. The high sensitivity of reverse transcriptase-polymerase chain reaction (RT-PCR) assay is the gold standard for the diagnosis of COVID-19, but there are still some false-negative results. Rapid antigen detection (RAD) is recommended by the World Health Organization (WHO) as a screening method for COVID-19. This review analyzed the characteristics of RDT and found that although the overall sensitivity of RAD was not as high as that of RT-PCR, but RAD was more sensitive in COVID-19 patients within 5 days of the onset of symptoms and in COVID-19 patients with Ct ≤ 25. Therefore, RAD can be used as an adjunct to RT-PCR for screening patients with early COVID-19. Finally, this review provides a combined diagnostic protocol for RAD and nucleic acid testing with the aim of providing a feasible approach for COVID-19 screening.

Characterisation and analysis of linear epitopes corresponding to SARS-CoV-2 outbreak in Jilin Province, China.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its variants have caused hundreds of thousands of deaths and shown serious social influence worldwide. Jilin Province, China, experienced the first wave of the outbreak from December 2020 to February 2021. Here, we analysed the genomic characteristics of the SARS-CoV-2 outbreak in Jilin province using a phylogeographic tree and found that clinical isolates belonged to the B.1 lineage, which was considered to be the ancestral lineage. Several dominant SARS-CoV-2 specific linear B cell epitopes that reacted with the convalescent sera were also analysed and identified using a peptide microarray composed of S, M, and E proteins. Moreover, the serum of convalescent patients infected with SARS-CoV-2 showed neutralising activity against four widely spreading SARS-CoV-2 variants; however, significant differences were observed in neutralising activities against different SARS-CoV-2 variants. These data provide important information on genomic characteristics, linear epitopes, and neutralising activity of SARS-CoV-2 outbreak in Jilin Province, China, which may aid in understanding disease patterns and regional aspects of the pandemic. This article is protected by copyright. All rights reserved.

Comparison of the performance of two real-time fluorescent quantitative PCR kits for the detection of SARS-CoV-2 nucleic acid: a study based on large real clinical samples.

BACKGROUND: The global pandemic of coronavirus disease 2019 (COVID-19) has led to the development of multiple detection kits by national manufacturers for severe acute respiratory syndrome coronavirus 2 viral nucleic acid testing. The purpose of this study is to evaluate the performance of different kits (i.e., Maccura kit and Sansure kit) in real clinical work using clinical samples, which will help with the optimization of the test kits. METHOD: During the past three months (March-May 2022), 1399 pharyngeal swabs from suspected COVID-19 patients have been initially screened using the Maccura kit in Jilin, China, and the test results were verified using the Sansure kit. The cycle threshold (Ct) values generated by the two kits were compared at different viral load levels. Correlation and consistency of the Ct values were investigated using Spearman correlation, Deming regression, and Bland-Altman plots. The cut-off Ct values of the Maccura kit were recalculated by referencing the result of the Sansure kit as a standard. Furthermore, another 163 pharyngeal swabs from suspected COVID-19 patients were collected to verify the new cut-off values. RESULTS: As a result of the Maccura kit testing, 1192 positive cases and 207 suspected COVID-19 cases were verified. After re-examination by the Sansure kit, 1118 positive cases were confirmed. The difference between the Ct values provided by the two kits was statistically significant, except for the N gene at high viral load. The Ct values obtained from the two kits presented a linear positive correlation. The Maccura kit used new cut-off Ct values of 35.00 (ORF1ab gene) and 35.07 (N gene). Based on that, the validation pass rate for the new cut-off Ct values was 91.41%. CONCLUSION: Since the Maccura kit is found to have false positives in actual clinical work, recalculation of the cut-off values can reduce this occurrence. In order to improve the accuracy of the testing, laboratories should use two kits for COVID-19 testing, and the adjusting and optimizing of the kits for their situation are needed.

A complete blood count-based multivariate model for predicting the recovery of patients with moderate COVID-19: a retrospective study.

Many resource-limited countries need an efficient and convenient method to assess disease progression in patients with coronavirus disease 2019 (COVID-19). This study developed and validated a complete blood count-based multivariate model for predicting the recovery of patients with moderate COVID-19. We collected the clinical data and laboratory test results of 86 patients with moderate COVID-19. These data were categorized into two subgroups depending on the laboratory test time. Univariate logistic regression and covariance diagnosis were used to screen for independent factors, and multifactorial logistic regression was used for model building. Data from 38 patients at another hospital were collected for external verification of the model. Basophils (OR 6.372; 95% CI 3.284-12.363), mean corpuscular volume (OR 1.244; 95% CI 1.088-1.422), red blood cell distribution width (OR 2.585; 95% CI 1.261-5.297), and platelet distribution width (OR 1.559; 95% CI 1.154-2.108) could be combined to predict recovery of patients with moderate COVID-19. The ROC curve showed that the model has good discrimination. The calibration curve showed that the model was well-fitted. The DCA showed that the model is clinically useful. Small increases in the above parameters within the normal range suggest an improvement in patients with moderate COVID-19.

Evaluating the role of SARS-CoV-2 target genes based on two nucleic acid assay kits

Background Effective isolation and early treatment of coronavirus disease 2019 (COVID-19) relies on rapid, accurate, and straightforward diagnostic tools. In response to the rapidly increasing number of cases, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) assays for multiple target genes have become widely available in the market. Methods In total, 236 COVID-19 patients with positive results in both RT-qPCR and rapid antigen diagnosis (Ag-RDT) were enrolled in the study. The cycle threshold (Ct) was compared with different onset times and target genes. Comparison between groups was evaluated with the Kruskal-Wallis test and Dunn test. The correlation between target genes was analyzed by Spearman. Results In samples of Ct ≤ 21, Ct was different for the nucleocapsid (N), open reading frame 1ab (ORF1ab), and envelope (E) genes (P < 0.05). Mild COVID-19 patients within 7 days of onset accounted for 67.80% of all enrolled patients. At the above stage, all target genes reached the trough of Ct, and N genes showed lower values than the other target genes. The Ct of the ORF1ab and N gene in asymptomatic patients differed from those of mild patients within 7 days and more than 14 days of onset. The kits used in the study showed strong consistency among target genes, with all correlation coefficients >0.870. Conclusion RT-qPCR confirmed that the N gene performed well in Ct ≤ 21 and samples within 7 days of onset. Ag-RDT was discriminatory for patients within 7 days of onset. This study facilitated early identification and control of COVID-19 prevalence among patients.

Potential Predictive Value of miR-125b-5p, miR-155-5p and Their Target Genes in the Course of COVID-19.

Purpose: This study aimed to provide new biomarkers for predicting the disease course of COVID-19 by analyzing the dynamic changes of microRNA (miRNA) and its target gene expression in the serum of COVID-19 patients at different stages. Methods: Serum samples were collected from all COVID-19 patients at three time points: the acute stage, the turn-negative stage, and the recovery stage. The expression level of miRNA and the target mRNA was measured by Quantitative Real-Time Polymerase Chain Reaction (RT-qPCR). The classification tree model was established to predict the disease course, and the prediction efficiency of independent variables in the model was analyzed using the receiver operating characteristic (ROC) curve. Results: The expression of miR-125b-5p and miR-155-5p was significantly up-regulated in the acute stage and gradually decreased in the turn-negative and recovery stages. The expression of the target genes CDH5, STAT3, and TRIM32 gradually down-regulated in the acute, turn-negative, and recovery stages. MiR-125b-5p, miR-155-5p, STAT3, and TRIM32 constituted a classification tree model with 100% accuracy of prediction and AUC >0.7 for identification and prediction in all stages. Conclusion: MiR-125b-5p, miR-155-5p, STAT3, and TRIM32 could be useful biomarkers to predict the time nodes of the acute, turn-negative, and recovery stages of COVID-19.

The Deubiquitinase USP29 Promotes SARS-CoV-2 Virulence by Preventing Proteasome Degradation of ORF9b.

Ubiquitin signaling is essential for immunity to restrict pathogen proliferation. Due to its enormous impact on human health and the global economy, intensive efforts have been invested in studying severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its interactions with hosts. However, the role of the ubiquitin network in pathogenicity has not yet been explored. Here, we found that ORF9b of SARS-CoV-2 is ubiquitinated on Lys-4 and Lys-40 by unknown E3 ubiquitin ligases and is degraded by the ubiquitin proteasomal system. Importantly, we identified USP29 as a host factor that prevents ORF9b ubiquitination and subsequent degradation. USP29 interacts with the carboxyl end of ORF9b and removes ubiquitin chains from the protein, thereby inhibiting type I interferon (IFN) induction and NF-κB activation. We also found that ORF9b stabilization by USP29 enhanced the virulence of VSV-eGFP and transcription and replication-competent SARS-CoV-2 virus-like-particles (trVLP). Moreover, we observed that the mRNA level of USP29 in SARS-CoV-2 patients was higher than that in healthy people. Our findings provide important evidence indicating that targeting USP29 may effectively combat SARS-CoV-2 infection. IMPORTANCE Coronavirus disease 2019 (COVID-19) is a current global health threat caused by SARS-CoV-2. The innate immune response such as type I IFN (IFN-I) is the first line of host defense against viral infections, whereas SARS-CoV-2 proteins antagonize IFN-I production through distinct mechanisms. Among them, ORF9b inhibits the canonical IκB kinase alpha (IKKɑ)/ß/γ-NF-κB signaling and subsequent IFN production; therefore, discovering the regulation of ORF9b by the host might help develop a novel antiviral strategy. Posttranslational modification of proteins by ubiquitination regulates many biological processes, including viral infections. Here, we report that ORF9b is ubiquitinated and degraded through the proteasome pathway, whereas deubiquitinase USP29 deubiquitinates ORF9b and prevents its degradation, resulting in the enhancement of ORF9b-mediated inhibition of IFN-I and NF-κB activation and the enhancement of virulence of VSV-eGFP and SARS-CoV-2 trVLP.

Early changes in laboratory tests predict liver function damage in patients with moderate coronavirus disease 2019: a retrospective multicenter study.

BACKGROUND: Most patients with coronavirus disease 2019 demonstrate liver function damage. In this study, the laboratory test data of patients with moderate coronavirus disease 2019 were used to establish and evaluate an early prediction model to assess the risk of liver function damage. METHODS: Clinical data and the first laboratory examination results of 101 patients with moderate coronavirus disease 2019 were collected from four hospitals' electronic medical record systems in Jilin Province, China. Data were randomly divided into training and validation sets. A logistic regression analysis was used to determine the independent factors related to liver function damage in patients in the training set to establish a prediction model. Model discrimination, calibration, and clinical usefulness were evaluated in the training and validation sets. RESULTS: The logistic regression analysis showed that plateletcrit, retinol-binding protein, and carbon dioxide combining power could predict liver function damage (P < 0.05 for all). The receiver operating characteristic curve showed high model discrimination (training set area under the curve: 0.899, validation set area under the curve: 0.800; P < 0.05). The calibration curve showed a good fit (training set: P = 0.59, validation set: P = 0.19; P > 0.05). A decision curve analysis confirmed the clinical usefulness of this model. CONCLUSIONS: In this study, the combined model assesses liver function damage in patients with moderate coronavirus disease 2019 performed well. Thus, it may be helpful as a reference for clinical differentiation of liver function damage. Trial registration retrospectively registered.

Screening High-Risk Groups and the General Population for SARS-CoV-2 Nucleic Acids in a Mobile Biosafety Laboratory.

The Severe Acute Respiratory Syndrome coronavirus 2 (SARS-CoV-2) pandemic has challenged public health systems worldwide. Therefore, large-scale testing capacity is extremely important diagnosis and exclusion diagnosis. However, fixed laboratories are limited or far away from remote areas. Fortunately, MBS-Lab is characterized by high mobility and rapid on-site detection of SARS-CoV-2 nucleic acid. MBS-Lab was first used in northern Australia during a melioidosis outbreak in 1997. The MBS-Lab and a well-trained diagnostic team were dispatched to Dongchang District, Tonghua City, Jilin Province, China to assist the SARS-CoV-2 virus screening and diagnosis on January 17, 2021. Altogether, 93,952 oropharyngeal swabs samples were collected and tested among the high-risk groups and the general population in Dongchang District. Two single samples were identified as positive in the second turn screening. In the second turn screening, 3 mixed samples (10 in 1) were identified as positive; 10 mixed samples were identified as positive in the third turn screening. By resampling again, one and four cases were identified as positive, respectively. The positive cases were properly isolated and treated in hospital and avoided to visit family members, friends, colleagues and any other persons. Through this way of large-scale screening, human-human spread of SARS-CoV-2 can be effectively avoided. In addition, all staff members strictly executed multiple safety precautions and reduce exposure risks. In the end, none of the staffs was infected with SARS-CoV-2 virus or other pathogens. As an emergency facility for infectious disease control, the MBS-Lab satisfies the requirements of ports and other remote areas far from fixed laboratories and supplements the capabilities of fixed laboratories.

A Prognostic Model to Predict Recovery of COVID-19 Patients Based on Longitudinal Laboratory Findings.

The temporal change patterns of laboratory data may provide insightful clues into the whole course of COVID-19. This study aimed to evaluate longitudinal change patterns of key laboratory tests in patients with COVID-19, and identify independent prognostic factors by examining the associations between laboratory findings and outcomes of patients. This multicenter study included 56 patients with COVID-19 treated in Jilin Province, China, from January 21, 2020 to March 5, 2020. The laboratory findings, epidemiological characteristics and demographic data were extracted from electronic medical records. The average value of eosinophils and carbon dioxide combining power continued to significantly increase, while the average value of cardiac troponin I and mean platelet volume decreased throughout the course of the disease. The average value of lymphocytes approached the lower limit of the reference interval for the first 5 days and then rose slowly thereafter. The average value of thrombocytocrit peaked on day 7 and slowly declined thereafter. The average value of mean corpuscular volume and serum sodium showed an upward trend from day 8 and day 15, respectively. Age, sex, lactate dehydrogenase, platelet count and globulin level were included in the final model to predict the probability of recovery. The above parameters were verified in 24 patients with COVID-19 in another area of Jilin Province. The risk stratification and management of patients with COVID-19 could be improved according to the temporal trajectories of laboratory tests.

Negative results in nucleic acid test of COVID-19 patients: assessment from the perspective of clinical laboratories.

Since December 2019, there had been an outbreak of coronavirus disease 2019 (COVID-19). Pneumonia caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is prevalent around the world, and the number of infected cases has increased rapidly. Viral nucleic acid test of SARS-CoV-2 can provide direct evidence for rapid diagnosis, disease course monitoring, and therapeutic efficacy. However, in practice, false-negative results in nucleic acid test are common, causing missed diagnoses, which are not conducive to the prevention and control of this outbreak. This article analyzes the possible causes and proposed measures to reduce the false-negative rate of nucleic acid test results and more effectively curb the outbreak.

Laboratory Biosafety Considerations of SARS-CoV-2 at Biosafety Level 2.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the pathogen that causes coronavirus disease 2019 (COVID-19), which was first detected in Wuhan, China. Recent studies have updated the epidemiologic and clinical characteristics of COVID-19 continuously. In China, diagnostic tests and laboratory tests of specimens from persons under investigation are usually performed in a biosafety level 2 environment. Laboratory staff may be at greater risk of exposure due to a higher concentration and invasiveness of emerging pathogens. Current infection prevention strategies are based on lessons learned from severe acute respiratory syndrome, expert judgments, and related regulations. This article summarizes biosafety prevention and control measures performed in severe acute respiratory syndrome coronavirus 2 testing activities and provides practical suggestions for laboratory staff to avoid laboratory-acquired infections in dealing with public health emergencies.

Changes of Laboratory Cardiac Markers and Mechanisms of Cardiac Injury in Coronavirus Disease 2019.

Some patients with coronavirus disease 2019 (COVID-19) show abnormal changes in laboratory myocardial injury markers, suggesting that patients with myocardial injury have a higher mortality rate than those without myocardial injury. This article reviews the possible mechanism of myocardial injury in patients with COVID-19. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) affects the patients with COVID-19 in aspects of direct infection of myocardial injury, specific binding to functional receptors on cardiomyocytes, and immune-mediated myocardial injury. During hospitalization, the monitoring of laboratory myocardial injury markers in patients of COVID-19 should be strengthened.

Mechanism of thrombocytopenia in COVID-19 patients.

Since December 2019, a novel coronavirus has spread throughout China and across the world, causing a continuous increase in confirmed cases within a short period of time. Some studies reported cases of thrombocytopenia, but hardly any studies mentioned how the virus causes thrombocytopenia. We propose several mechanisms by which coronavirus disease 2019 causes thrombocytopenia to better understand this disease and provide more clinical treatment options.