Risk factors for severe acute respiratory syndrome coronavirus 2 RNA environmental contamination in rooms of patients with coronavirus disease 2019.

The risk factors of environmental contamination by SARS-CoV-2 were unknown. We analyzed 1,320 environmental samples obtained from COVID-19 patients for 1 year. The risk factors of contamination of COVID-19 patients' surrounding environment were higher viral load in the respiratory tract and shorter duration from symptom onset to sample collection.

Efficacy of Plant-Made Human Recombinant ACE2 against COVID-19 in a Golden Syrian Hamster Model.

Coronavirus disease 2019 (COVID-19) is a novel infectious respiratory disease caused by SARS-CoV-2. We evaluated the efficacy of a plant-based human recombinant angiotensin-converting enzyme 2 (hrACE2) and hrACE2-foldon (hrACE2-Fd) protein against COVID-19. In addition, we analyzed the antiviral activity of hrACE2 and hrACE2-Fd against SARS-CoV-2 using real-time reverse-transcription PCR and plaque assays. The therapeutic efficacy was detected using the Golden Syrian hamster model infected with SARS-CoV-2. Both hrACE2 and hrACE2-Fd inhibited SARS-CoV-2 by 50% at concentrations below the maximum plasma concentration, with EC50 of 5.8 µg/mL and 6.2 µg/mL, respectively. The hrACE2 and hrACE2-Fd injection groups showed a tendency for decreased viral titers in nasal turbinate tissues on day 3 after virus inoculation; however, this decrease was not detectable in lung tissues. Histopathological examination on day 9 after virus inoculation showed continued inflammation in the SARS-CoV-2 infection group, whereas decreased inflammation was observed in both the hrACE2 and hrACE2-Fd injection groups. No significant changes were observed at other time points. In conclusion, the potential therapeutic efficacy of plant-based proteins, hrACE2 and hrACE2-Fd, against COVID-19 was confirmed in a SARS-CoV-2-inoculated Golden Syrian hamster model. Further preclinical studies on primates and humans are necessary to obtain additional evidence and determine the effectiveness of these therapies.

Prevalence and patterns of major depressive disorder and subthreshold depressive symptoms in south China.

BACKGROUND: Information on major depressive disorder (MDD) and subthreshold depressive symptoms (SDS) is rarely reported in south China. This study examines the prevalence rates and patterns of MDD and SDS of a large representative sample of adult residents in south China. METHODS: The Guangdong Mental Health Survey was conducted on adults (over 18 years) from September to December 2021. Multistage stratified cluster sampling was used and face-to-face interviews were done with a two-stage design by trained lay interviewers and psychiatrists. A total of 16,377 inhabitants were interviewed using standardized assessment tools. Data were weighted to adjust for differential probabilities of selection and differential response. RESULTS: The weighted prevalence rates of MDD and SDS were 2.5 % (95%CI: 2.2 %-2.9 %) and 14.7 % (95%CI: 14.0 %-15.5 %), respectively. Multinomial logistic regression analysis revealed that female, younger age, living in urban area, higher education, unmarried, irregular meal pattern, lack of physical exercise, chronic diseases, irregular napping pattern and short sleep were positively associated with SDS. Besides, female, younger age, unmarried, irregular meal pattern, lack of physical exercise, chronic diseases, short sleep and poor mental health were positively associated with MDD. LIMITATIONS: The cross-sectional nature of the study limited causal inferences. CONCLUSIONS: The prevalence of MDD in Guangdong province in 2021 is higher than in mainland China in 2013. Given the higher prevalence of SDS, and high burden of depression, it also offers valuable opportunities for policymakers and health-care professionals to explore the factors affecting mental health in Guangdong province, especially during the COVID-19 epidemic.

Diagnostic Sensitivity of Saliva and Other Respiratory Tract Samples of SARS-CoV-2 Variants in Patients with COVID-19.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants continue to emerge during the ongoing coronavirus disease 2019 (COVID-19) pandemic. Contrasting studies on the omicron variant have demonstrated higher viral loads in different clinical specimens, which is consistent with its high transmissibility. We investigated the viral load in clinical specimens that were infected with the SARS-CoV-2 wild-type, delta, and omicron variants, and we analyzed the diagnostic accuracy of upper and lower respiratory specimens for these variants. We performed nested reverse transcription (RT)-polymerase chain reaction (PCR), targeting the spike gene and sequencing for variant classification. RT-PCR was performed using upper and lower respiratory specimens, including saliva from 78 COVID-19 patients (wild-type, delta, and omicron variants). A comparison of the sensitivity and specificity, using the area under the receiver operating characteristic curve (AUC) values from the N gene, showed that the omicron variant saliva samples had a higher sensitivity (AUC = 1.000) than did the delta (AUC = 0.875) and the wild-type (AUC = 0.878) variant samples. The sensitivity of the omicron saliva samples was greater than that of the wild-type nasopharynx and sputum samples (P < 0.001). The viral loads of the saliva samples containing the wild-type, delta, and omicron variants were 8.18 × 105, 2.77 × 106, and 5.69 × 105, respectively, which did not differ significantly (P = 0.610). Statistically significant differences were not observed in the saliva viral loads between vaccinated and nonvaccinated patients who were infected with the omicron variant (P = 0.120). In conclusion, omicron saliva samples had higher sensitivity than did wild-type and delta samples, and the viral load did not significantly differ between vaccinated and nonvaccinated patients. Further research is necessary to elucidate the mechanisms underlying the sensitivity differences. IMPORTANCE Owing to the vast heterogeneity of the studies focused on the correlation between the SARS-CoV-2 omicron variant and COVID-19, accurate comparisons of the specificity and sensitivity of samples and associated outcomes are still inconclusive. Moreover, limited information is available on the leading causes of infection and the factors that are associated with the conditions that underlie the spread of infection. Although several studies have contributed important knowledge regarding infectious specimens, the impact of saliva samples remains unknown. This study showed that the sensitivity of the omicron variant saliva samples was higher than that of the wild-type nasopharyngeal and sputum samples. Moreover, neither vaccinated nor nonvaccinated patients who were infected with the omicron variant showed any significant differences in SARS-CoV-2 viral loads. Hence, this study is an important step toward understanding how saliva sample results are correlated with other specimen results, regardless of the vaccination status of patients who are infected with the SARS-CoV-2 omicron variant.

Augmented humoral and cellular immunity against severe acute respiratory syndrome coronavirus 2 after breakthrough infection in kidney transplant recipients who received 3 doses of coronavirus disease 2019 vaccine.

Diminished immune response to coronavirus disease 2019 (COVID-19) vaccines and breakthrough infection (BI) is a major concern for solid organ transplant recipients. Humoral and cellular immune responses of kidney transplant (KT) recipients after a third COVID-19 vaccination were investigated compared to matched health care workers. Anti-severe acute respiratory syndrome coronavirus 2 spike protein antibody and severe acute respiratory syndrome coronavirus 2 specific interferon-gamma releasing assay (IGRA) were assessed. A total of 38 KT recipients, including 20 BI and 18 noninfection, were evaluated. In the KT BI group, antibody titers were significantly increased (median 5 to 724, binding antibody units/mL (P = 0.002) after the third vaccination, but IGRA responses were negligible. After BI, antibody titers increased (median 11 355 binding antibody unit/mL; P < 0.001) and there was a significant increase of IGRA responses to spike proteins (Spike1-Nil, median 0.05 to 0.41 IU/mL; P = 0.009). Antibody titers and IGRA responses were significantly higher in the BI than in the noninfection group after 6 months. Immune responses were stronger in the health care worker than in the KT cohort, but the gap became narrower after BI. In conclusion, KT recipients who experienced BI after 3 COVID-19 vaccinations acquired augmented humoral and cellular immune responses.

Prone positioning improves ventilation-perfusion matching assessed by electrical impedance tomography in patients with ARDS: a prospective physiological study.

BACKGROUND: The physiological effects of prone ventilation in ARDS patients have been discussed for a long time but have not been fully elucidated. Electrical impedance tomography (EIT) has emerged as a tool for bedside monitoring of pulmonary ventilation and perfusion, allowing the opportunity to obtain data. This study aimed to investigate the effect of prone positioning (PP) on ventilation-perfusion matching by contrast-enhanced EIT in patients with ARDS. DESIGN: Monocenter prospective physiologic study. SETTING: University medical ICU. PATIENTS: Ten mechanically ventilated ARDS patients who underwent PP. INTERVENTIONS: We performed EIT evaluation at the initiation of PP, 3 h after PP initiation and the end of PP during the first PP session. MEASUREMENTS AND MAIN RESULTS: The regional distribution of ventilation and perfusion was analyzed based on EIT images and compared to the clinical variables regarding respiratory and hemodynamic status. Prolonged prone ventilation improved oxygenation in the ARDS patients. Based on EIT measurements, the distribution of ventilation was homogenized and dorsal lung ventilation was significantly improved by PP administration, while the effect of PP on lung perfusion was relatively mild, with increased dorsal lung perfusion observed. The ventilation-perfusion matched region was found to increase and correlate with the increased PaO2/FiO2 by PP, which was attributed mainly to reduced shunt in the lung. CONCLUSIONS: Prolonged prone ventilation increased dorsal ventilation and perfusion, which resulted in improved ventilation-perfusion matching and oxygenation. TRIAL REGISTRATION: ClinicalTrials.gov, NCT04725227. Registered on 25 January 2021.

Coronavirus Disease 2019-Associated Coagulopathy.

Coronavirus disease 2019 (COVID-19)-associated coagulopathy is an acute illness characterized by thrombosis with or without hemorrhage after COVID-19 infection. Clinical symptoms of COVID-19-associated coagulopathy can occur at any anatomical site. Various forms of venous thromboembolism, including deep vein thrombosis and pulmonary embolism, are common in acutely ill patients with COVID-19. Laboratory findings, such as D-dimer and platelet counts, can help diagnose COVID-19-associated coagulopathy. Anticoagulation using direct oral anticoagulants and low-molecular-weight heparin is essential for the treatment of COVID-19-associated coagulopathy. Prophylactic anticoagulants are important in preventing COVID-19-associated coagulopathy in patients with severe COVID-19. In particular, the early initiation of prophylactic anticoagulation in patients with COVID-19 can improve survival rates without the risk of serious bleeding events.

Reveal the Mechanisms of Yi-Fei-Jian-Pi-Tang on Covid-19 through Network Pharmacology Approach.

Objectives: The Traditional Chinese Medicine (TCM) formula Yi-Fei-Jian-Pi-Tang (YFJPT) has been demonstrated effective against Corona Virus Disease 2019 (Covid-19). The aim of this article is to make a thorough inquiry about its active constituent as well as mechanisms against Covid-19 via TCM network pharmacology. Methods: All the ingredients of YFJPT are obtained from the pharmacology database of the TCM system. The genes which are associated with the targets are obtained by utilizing UniProt. The herb-target network is built up by utilizing Cytoscape. The target protein-protein interaction network is built by utilizing the STRING database and Cytoscape. The critical targets of YFJPT are explored by Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG). Results: The outcomes show that YFJPT might has 33 therapeutic targets on Covid-19, namely, interleukin 2 (IL2), heme oxygenase 1 (HMOX1), interleukin 4 (IL4), interferon gamma (FNG), α nuclear factor of kappa light polypeptide gene enhancer in Bcells inhibitor, alpha (NFKBIA), nuclear factor-k-gene binding (NFKB), nitric oxide synthase 3 (NOS3), intercellular adhesion molecule 1 (ICAM1), hypoxia inducible factor 1 subunit alpha (HIF1A), mitogen-activated protein kinase 3 (MAPK3), epidermal growth factor receptor (EGFR), interleukin 10 (IL10), jun proto-oncogene (JUN), C-C motif chemokine ligand 2 (CCL2), C-X-C motif chemokine ligand 8 (CXCL8), tumor protein p53 (TP53), interleukin 1 beta (IL1B), AKT serine/threonine kinase 1 (AKT1), tumor necrosis factor (TNF), interleukin 6 (IL6), erb-b2 receptor tyrosine kinase 2 (ERBB2), RELA proto-oncogene (RELA), NF-κB subunit, caspase 8 (CASP8), peroxisome proliferator activated receptor alpha (PPARA), TIMP metallopeptidase inhibitor 1 (TIMP1), transforming growth factor beta 1 (TGFB1), interleukin 1 alpha (IL1A), signal transducer and activator of transcription 1 (STAT1), mitogen-activated protein kinase 8 (MAPK8), myeloperoxidase (MPO), matrix metallopeptidase 3 (MMP3), matrix metallopeptidase 1 (MMP1), and NFE2 like bZIP transcription factor 2 (NFE2L2). The gene enrichment analysis prompts that YFJPT most likely contributes to patients related to Covid-19 by regulating the pathways of cancers. Conclusions: That will lay a foundation for the clinical rational application and further experimental research of YFJPT.

Whole-genome analysis and mutation pattern of SARS-CoV-2 during first and second wave outbreak in Gwangju, Republic of Korea.

To investigate the specific genomic features and mutation pattern, whole and near-complete SARS-CoV-2 genome sequences were analyzed. Clinical samples were collected from 18 COVID-19-positive patients and subjected to nucleic acid purification. Cell culture was performed to extract various SARS-CoV-2 isolates. Whole-genome analysis was performed using next-generation sequencing, and phylogenetic analyses were conducted to determine genetic diversity of the various SARS-CoV-2 isolates. The next-generation sequencing data identified 8 protein-coding regions with 17 mutated proteins. We identified 51 missense point mutations and deletions in 5' and 3' untranslated regions. The phylogenetic analysis revealed that V and GH are the dominant clades of SARS-CoV-2 circulating in the Gwangju region of South Korea. Moreover, statistical analysis confirmed a significant difference between viral load (P < 0.001) and number of mutations (P < 0.0001) in 2 mutually exclusive SARS-CoV-2 clades which indicates frequent genomic alterations in SARS-CoV-2 in patients with high viral load. Our results provide an in-depth analysis of SARS-COV-2 whole genome which we believe, can shed light in the understanding of SARS-COV-2 pathogenesis and mutation pattern which can aid in the development of prevention methods as well as future research into the pathogenesis of SARS-CoV-2 and therapeutic development.

ACE2 and TMPRSS2 Immunolocalization and COVID-19-Related Thyroid Disorder.

Thyroid dysfunction has been reported to be an extrapulmonary symptom of COVID-19. It is important to identify the tissue subset that expresses angiotensin-converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2), which are essential for host infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), in order to understand the viral pathogenesis of COVID-19-related thyroid dysfunction. We investigated the expression and distribution of ACE2- and TMPRSS2-expressing cells in the thyroid gland. RT-PCR and Western blotting were performed on human thyroid follicular cells (Nthy-ori3-1) and rat thyroid tissues to detect the expression levels of ACE and TMPRSS2 mRNA and proteins. We also analyzed the expression patterns of ACE2 and TMPRSS2 in 9 Sprague-Dawley rats and 15 human thyroid tissues, including 5 normal, 5 with Hashimoto's thyroiditis, and 5 with Graves' disease, by immunohistochemistry (IHC) and immunofluorescence. Both ACE2 and TMPRSS2 mRNAs and proteins were detected in the thyroid tissue. However, ACE2 and TMPRSS2 proteins were not expressed in thyroid follicular cells. In IHC, ACE2 and TMPRSS2 were not stained in the follicular cells. No cells co-expressed ACE2 and TMPRSS2. ACE2 was expressed in pericytes between follicles, and TMPRSS2 was mainly stained in the colloid inside the follicle. There was no difference in expression between the normal thyroid, Hashimoto's thyroiditis, and Graves' disease. SARS-CoV-2 does not directly invade the thyroid follicular cells. Whether SARS-CoV-2 infection of pericytes can affect COVID-19-related thyroid dysfunction warrants further study.

Targeted intracellular delivery of Cas13 and Cas9 nucleases using bacterial toxin-based platforms.

Targeted delivery of therapeutic proteins toward specific cells and across cell membranes remains major challenges. Here, we develop protein-based delivery systems utilizing detoxified single-chain bacterial toxins such as diphtheria toxin (DT) and botulinum neurotoxin (BoNT)-like toxin, BoNT/X, as carriers. The system can deliver large protein cargoes including Cas13a, CasRx, Cas9, and Cre recombinase into cells in a receptor-dependent manner, although delivery of ribonucleoproteins containing guide RNAs is not successful. Delivery of Cas13a and CasRx, together with guide RNA expression, reduces mRNAs encoding GFP, SARS-CoV-2 fragments, and endogenous proteins PPIB, KRAS, and CXCR4 in multiple cell lines. Delivery of Cre recombinase modifies the reporter loci in cells. Delivery of Cas9, together with guide RNA expression, generates mutations at the targeted genomic sites in cell lines and induced pluripotent stem cell (iPSC)-derived human neurons. These findings establish modular delivery systems based on single-chain bacterial toxins for delivery of membrane-impermeable therapeutics into targeted cells.

Accuracy of Real-Time Polymerase Chain Reaction in COVID-19 Patients.

Coronavirus disease 2019 (COVID-19) is a mild to severe respiratory illness caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The diagnostic accuracy of the Centers for Disease Control and Prevention (CDC)- or World Health Organization (WHO)-recommended real-time PCR (RT-qPCR) primers in clinical practice remains unproven. We conducted a prospective study on the accuracy of RT-qPCR using an in-house-designed primer set (iNP) targeting the nucleocapsid protein as well as various recommended and commercial primers. The accuracy was assessed by culturing or seroconversion. We enrolled 12 confirmed COVID-19 patients with a total of 590 clinical samples. When a cutoff value of the cycle threshold (Ct) was set to 35, RT-qPCRs with WHO RdRp primers and CDC N1, N2, and N3 primers showed sensitivity of 42.1% to 63.2% and specificity of 90.5% to 100% in sputum, and sensitivity of 65.2% to 69.6% and specificity of 65.2% to 69.6% in nasopharyngeal samples. The sensitivity and specificity of iNP RT-qPCR in sputum and nasopharyngeal samples were 94.8%/100% and 69.6%/100%, respectively. Sputum testing had the highest sensitivity, followed by nasopharyngeal testing (P = 0.0193); self-collected saliva samples yielded better characteristics than oropharyngeal samples (P = 0.0032). Our results suggest that iNP RT-qPCR has better sensitivity and specificity than RT-PCR with WHO (P < 0.0001) or CDC (N1: P = 0.0012, N2: P = 0.0013, N3: P = 0.0012) primers. Sputum RT-qPCR analysis has the highest sensitivity, followed by nasopharyngeal, saliva, and oropharyngeal assays. Our study suggests that considerable improvement is needed for the RT-qPCR WHO and CDC primer sets for detecting SARS-CoV-2. IMPORTANCE Numerous research campaigns have addressed the vast majority of clinical and diagnostic specificity and sensitivity of various primer sets of SARS-CoV2 viral detection. Despite the impressive progress made to resolve the pandemic, there is still a need for continuous and active improvement of primers used for diagnosis in clinical practice. Our study significantly exceeds the scale of previously published research on the specificity and sensitivity of different primers comparing with different specimens and is the most comprehensive to date in terms of constant monitoring of primer sets of current usage. Henceforth, our results suggest that sputum samples sensitivity is the highest, followed by nasopharyngeal, saliva, and oropharyngeal samples. The CDC recommends the use of oropharyngeal specimens, leading to certain discrepancy between the guidelines set forth by the CDC and IDSA. We proved that the oropharyngeal samples demonstrated the lowest sensitivity for the detection of SARS-CoV-2.

Viable Severe Acute Respiratory Syndrome Coronavirus 2 Isolates Exhibit Higher Correlation With Rapid Antigen Assays Than Subgenomic RNA or Genomic RNA.

Background: Rapid identification and effective isolation are crucial for curbing the spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). To meet this requirement, antigen-detection rapid diagnostic tests (Ag-RDTs) are essential. Methods: Between February 2020 and August 2020 we performed a cohort study of patients with confirmed COVID-19. The clinical performance of Ag rapid fluorescence immunoassay (FIA) and Ag Gold was evaluated and compared in parallel with genomic and subgenomic real-time reverse transcription-polymerase chain reaction (rRT-PCR) and cell culture-based assays. Results: In total, 150 samples were tested. Of these, 63 serial samples were obtained from 11 patients with SARS-CoV-2 and 87 from negative controls. Serial respiratory samples were obtained 2 days prior to symptom onset (-2) up to 25 days post-symptom onset. Overall, for rRT-PCR-positive samples (n = 51), the detection sensitivity of Ag rapid FIA and Ag Gold was 74.5% and 53.49%, respectively, with a specificity of 100%; however, for samples with low cycle threshold (Ct) values, Ag rapid FIA and Ag Gold exhibited a sensitivity of 82.61% (Ct ≤ 30, 5.6 log10RNA copies/mL) and 80% (Ct ≤ 25, 6.9 log10RNA copies/mL), respectively. Despite low analytical sensitivity, both Ag-RDTs detected 100% infection in cell culture-positive samples (n = 15) and were highly effective in distinguishing viable samples from those with subgenomic RNA (66.66%). For both Ag-RDTs, all samples that yielded discordant results (rRT-PCR + ve/Ag-RDT -ve) were also negative by culture. Conclusion: The data suggest that Ag-RDTs reliably detect viable SARS-CoV-2; thus, they may serve as an important tool for rapid detection of potentially infectious individuals.

Usefulness of ELISA Using Total Antibody against Plant-Expressed Recombinant Nucleocapsid Protein of SARS-CoV-2.

Here, we aimed to investigate the diagnostic value of a serological assay using the nucleocapsid protein developed for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection and evaluated its performance using three commercial enzyme-linked immunosorbent assays (ELISAs), namely, Standard E 2019 novel coronavirus disease (COVID-19) total antibody (Ab) ELISA (SD Biosensor), and EDI novel coronavirus COVID-19 IgG and IgM ELISA. A recombinant nucleocapsid protein (rNP) was expressed from plants and Escherichia coli for the detection of serum total Ab. We prospectively collected 141 serum samples from 32 patients with reverse transcription-PCR (RT-PCR)-confirmed COVID-19 and determined the sensitivity and dynamics of their total Ab response. Specificity was evaluated using 158 prepandemic samples. To validate the assays, we evaluated the performance using two different cutoff values. The sensitivity and specificity for each assay were as follows: 92.91% and 94.30% (plant-rNP), 83.69% and 98.73% (SD Biosensor), 75.89% and 98.10% (E. coli-rNP), 76.47% and 100% (EDI-IgG), and 80.39% and 80% (EDI-IgM). The plant-based rNP showed the highest sensitivity and area under the receiver operating characteristic (ROC) curve (0.980) among all the assays (P < 0.05). The seroconversion rate for total Ab increased sequentially with disease progression, with a sensitivity of 100% after 10 to 12 days of post-symptom onset (PSO) for both rNP-plant-based and SD Biosensor ELISAs. After 2 weeks of PSO, the seroconversion rates were >80% and 100% for EDI-IgM and EDI-IgG ELISA, respectively. Seroconversion occurred earlier with rNP plant-based ELISA (5 days PSO) compared with E. coli-based (7 days PSO) and SD Biosensor (8 days PSO) ELISA. We determined that rNP produced in plants enables the robust detection of SARS-CoV-2 total Abs. The assay can be used for serosurvey and complementary diagnosis of COVID-19. IMPORTANCE At present, the principal diagnostic methods for COVID-19 comprise the identification of viral nucleic acid by genetic approaches, including PCR-based techniques or next-generation sequencing. However, there is an urgent need for validated serological assays which are crucial for the understanding of immune responses against SARS-CoV-2. In this study, a highly sensitive and specific serological antibody assay was developed for the detection of SARS-CoV-2 with an overall accuracy of 93.56% using a recombinant nucleoprotein expressed from plants.

Emerging Severe Acute Respiratory Syndrome Coronavirus 2 Mutation Hotspots Associated With Clinical Outcomes and Transmission.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the cause of the ongoing coronavirus disease 2019 (COVID-19) pandemic. Understanding the influence of mutations in the SARS-CoV-2 gene on clinical outcomes is critical for treatment and prevention. Here, we analyzed all high-coverage complete SARS-CoV-2 sequences from GISAID database from January 1, 2020, to January 1, 2021, to mine the mutation hotspots associated with clinical outcome and developed a model to predict the clinical outcome in different epidemic strains. Exploring the cause of mutation based on RNA-dependent RNA polymerase (RdRp) and RNA-editing enzyme, mutation was more likely to occur in severe and mild cases than in asymptomatic cases, especially A > G, C > T, and G > A mutations. The mutations associated with asymptomatic outcome were mainly in open reading frame 1ab (ORF1ab) and N genes; especially R6997P and V30L mutations occurred together and were correlated with asymptomatic outcome with high prevalence. D614G, Q57H, and S194L mutations were correlated with mild and severe outcome with high prevalence. Interestingly, the single-nucleotide variant (SNV) frequency was higher with high percentage of nt14408 mutation in RdRp in severe cases. The expression of ADAR and APOBEC was associated with clinical outcome. The model has shown that the asymptomatic percentage has increased over time, while there is high symptomatic percentage in Alpha, Beta, and Gamma. These findings suggest that mutation in the SARS-CoV-2 genome may have a direct association with clinical outcomes and pandemic. Our result and model are helpful to predict the prevalence of epidemic strains and to further study the mechanism of mutation causing severe disease.

Viral Kinetics of Severe Acute Respiratory Syndrome Coronavirus 2 in Patients with Coronavirus Disease 2019.

To determine the relationship between viral kinetics and severity of disease in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, we investigated the viral kinetics and compared the viral loads of patients with coronavirus disease 2019 (COVID-19; the disease caused by SARS-CoV-2), stratified by symptoms and severity. We determined the viral kinetics of 100 patients diagnosed with COVID-19 at Chosun University Hospital between February 2020 and May 2021 and analyzed the differences between asymptomatic, symptomatic, and nonsurvivor patients and between patients who died and those who survived. Clinical samples, comprising respiratory specimens (sputum samples and nasopharynx and oropharynx swab samples), were obtained at different time points of hospitalization, at 1, 3 to 5, 7, 10, 14, and 30 days. SARS-CoV-2 was detected using real-time reverse transcription-PCR (RT-PCR). All three groups, asymptomatic, symptomatic, and deceased patients, had higher numbers of viral copies at symptom onset, and the asymptomatic group had lower numbers of viral copies than the symptomatic or nonsurvivor group. Viral RNA release was detected until 30 days after symptom onset. The virus cleared up earlier in asymptomatic patients than in symptomatic and nonsurvivor patients, and it cleared up earlier in mildly affected patients than in severely affected patients. The cycle threshold values tended to be significantly lower in the group receiving steroids than in the nonsteroid group, even in the low-risk group with a pneumonia severity index of less than 90. The viral loads in patients with COVID-19 were significantly different according to disease severity and steroid use. IMPORTANCE In our study, we analyzed the viral kinetics of COVID-19 patients. Our study reveals differences in viral shedding according to the severity of disease in COVID-19 patients. Viral shedding had a longer duration in severely affected patients, and the cyclic threshold values were lower in the group receiving steroids. This study is expected to be helpful in analyzing the trend of the disease course according to steroid use and severity of SARS-CoV-2 disease.

Eosinophil-mediated lung inflammation associated with elevated natural killer T cell response in COVID-19 patients.

BACKGROUND/AIMS: Coronavirus disease 2019 (COVID-19) is associated with acute respiratory syndrome. The mechanisms underlying the different degrees of pneumonia severity in patients with COVID-19 remain elusive. This study provides evidence that COVID-19 is associated with eosinophil-mediated inflammation. METHODS: We performed a retrospective case series of three patients with laboratory and radiologically confirmed COVID-19 pneumonia admitted to Chosun University Hospital. Demographic and clinical data on inflammatory cell lung infiltration and cytokine levels in patients with COVID-19 were collected. RESULTS: Cytological analysis of sputum, tracheal aspirates, and bronchoalveolar lavage fluid (BALF) samples from all three patients revealed massive infiltration of polymorphonuclear cells (PMNs), such as eosinophils and neutrophils. All sputum and BALF specimens contained high levels of eosinophil cationic proteins. The infiltration of PMNs into the lungs, together with elevated levels of natural killer T (NKT) cells in BALF and peripheral blood samples from patients with severe pneumonia in the acute phase was confirmed by flow cytometry. CONCLUSION: These results suggest that the lungs of COVID-19 patients can exhibit eosinophil-mediated inflammation, together with an elevated NKT cell response, which is associated with COVID-19 pneumonia.

Enhanced eosinophil-mediated inflammation associated with antibody and complement-dependent pneumonic insults in critical COVID-19.

Despite the worldwide effect of the coronavirus disease 2019 (COVID-19) pandemic, the underlying mechanisms of fatal viral pneumonia remain elusive. Here, we show that critical COVID-19 is associated with enhanced eosinophil-mediated inflammation when compared to non-critical cases. In addition, we confirm increased T helper (Th)2-biased adaptive immune responses, accompanying overt complement activation, in the critical group. Moreover, enhanced antibody responses and complement activation are associated with disease pathogenesis as evidenced by formation of immune complexes and membrane attack complexes in airways and vasculature of lung biopsies from six fatal cases, as well as by enhanced hallmark gene set signatures of Fcγ receptor (FcγR) signaling and complement activation in myeloid cells of respiratory specimens from critical COVID-19 patients. These results suggest that SARS-CoV-2 infection may drive specific innate immune responses, including eosinophil-mediated inflammation, and subsequent pulmonary pathogenesis via enhanced Th2-biased immune responses, which might be crucial drivers of critical disease in COVID-19 patients.

The Prevalence of Depression, Anxiety and Associated Factors among the General Public during COVID-19 Pandemic: a Cross-sectional Study in Korea.

BACKGROUND: Since its first case confirmed on January 20, 2020, Korea has been through three waves of the COVID-19 pandemic. Fears of the fourth wave persist as new cases continue to emerge. In such unpredictable times, the mental well-being of people is of crucial importance. This study examined the levels of depression and anxiety and their predictors among the Korean general public in Busan, Korea, during the COVID-19 pandemic. METHODS: We conducted a cross-sectional study via a self-reported questionnaire administered to 2,288 adult residents (aged 19-60 years) of Busan, Korea. Participants' depression and anxiety were assessed using the Korean version of the Patient Health Questionnaire-4 (PHQ-4), which consists of PHQ-2 and Generalized Anxiety Disorder-2 (GAD-2), with the cutoff score of 3. RESULTS: The mean age of the participants was 39.71 years. COVID-19 had several psychosocial impacts on people. It was revealed that 80.3% had restrictions in outside activities, 47.3% reported financial difficulties, and 53.6% had a fear of death or fatal outcome when infected with COVID-19. We performed logistic regression analysis to identify the factors associated with depression and anxiety. A total of 30.7% participants were classified as at risk of depression based on cutoff score of 3 on PHQ-2. Logistic regression analysis revealed that changes in sleep pattern due to COVID-19 were most strongly associated with depression, followed by restrictions in outside activities due to social distancing and increased family conflicts due to COVID-19. Also, 22.6% participants were classified as at risk of anxiety based on a cutoff score of 3 on GAD-2. Analysis revealed that changes in sleep pattern due to COVID-19 were most strongly associated with anxiety, followed by spending a lot of time searching for COVID-19-related information and having a fear of death or fatal outcome when infected with COVID-19. CONCLUSION: The results are alarming; 30.7% had a PHQ-2 score of 3 or higher, indicating depression, and 22.6% had a GAD-2 score of 3 or higher, indicating anxiety. Changes in sleep pattern had the strongest association with both depression and anxiety. Our results can be used to formulate mental health policies tailored to the context of the city. Our findings suggest the high prevalence of depression and anxiety in the society during the COVID-19 pandemic, which places growing importance on early intervention for mental health problems during these times.

Forecasting the COVID-19 transmission in Italy based on the minimum spanning tree of dynamic region network.

BACKGROUND: Italy surpassed 1.5 million confirmed Coronavirus Disease 2019 (COVID-19) infections on November 26, as its death toll rose rapidly in the second wave of COVID-19 outbreak which is a heavy burden on hospitals. Therefore, it is necessary to forecast and early warn the potential outbreak of COVID-19 in the future, which facilitates the timely implementation of appropriate control measures. However, real-time prediction of COVID-19 transmission and outbreaks is usually challenging because of its complexity intertwining both biological systems and social systems. METHODS: By mining the dynamical information from region networks and the short-term time series data, we developed a data-driven model, the minimum-spanning-tree-based dynamical network marker (MST-DNM), to quantitatively analyze and monitor the dynamical process of COVID-19 spreading. Specifically, we collected the historical information of daily cases caused by COVID-19 infection in Italy from February 24, 2020 to November 28, 2020. When applied to the region network of Italy, the MST-DNM model has the ability to monitor the whole process of COVID-19 transmission and successfully identify the early-warning signals. The interpretability and practical significance of our model are explained in detail in this study. RESULTS: The study on the dynamical changes of Italian region networks reveals the dynamic of COVID-19 transmission at the network level. It is noteworthy that the driving force of MST-DNM only relies on small samples rather than years of time series data. Therefore, it is of great potential in public surveillance for emerging infectious diseases.