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1.
EuropePMC; 2021.
Preprint in English | EuropePMC | ID: ppcovidwho-317641

ABSTRACT

SARS-CoV-2, the causative agent of coronavirus disease 19 (COVID 19), is responsible for the ongoing pandemic but still lacks approved antivirals. Repurposing pre-existing FDA approved drugs presents a rapid approach for new therapeutic options. In the present study, we report that three pre-existing FDA-approved drugs, i.e., vapreotide, grazoprevir, and simeprevir, inhibit the replication of SARS-CoV-2 in cells. The E50 values of vapreotide, grazoprevir, and simeprevir against SARS-CoV-2 in Vero E6 cells was 3.98 ± 0.35 μM, 2.08 ± 0.13 μM, and 1.41 ± 0.12 μM, respectively. In vitro biochemical experiments further revealed that vapreotide, grazoprevir, and simeprevir efficiently inhibits the unwinding activity of the Nsp13 helicase of SARS-CoV-2 with IC50 values of ⁓10, ⁓2.5, and ⁓1.25 µM, respectively, providing signs for understanding their antiviral mechanism of action. Given their good safety profiles in their original indications, our study offices new insights in repurposing these drugs alone or in combination with other antivirals in the global fighting against SARS-CoV-2.Funding: This work was financially supported by the Youth Innovation Promotion Association CAS (to H.Y.), and the National Natural Science Foundation of China (No. 31770192 and No. 32070187 to H.Y.).Conflict of Interest: The authors declare no competing interests.

2.
EuropePMC; 2020.
Preprint in English | EuropePMC | ID: ppcovidwho-308350

ABSTRACT

Background: Coronavirus disease 2019 (COVID-19) has emerged as a major global health threat with a great number of deaths worldwide. Acute kidney injury (AKI) is a common complication in patients admitted to the intensive care unit. We aimed toassess the incidence, risk factors and in-hospital outcomes of AKI in COVID-19 patients admitted to intensive care unit Methods: : we conducted a retrospective observational study in intensive care unit of Tongji hospital, which was assigned responsibility for the treatments of severe COVID-19 patients by Wuhan government. The AKI was defined and staged based onKidney Disease: Improving Global Outcomes (KDIGO) criteria. Mild AKI was defined as stage 1, and severe AKI was defined as stage 2 or stage 3. We used logistic regression analysis to evaluate AKI risk factors and the association between AKI and in-hospital mortality. Results: : A total of 150 patients with COVID-19 were included in our study. The median age of patients was 70 (interquartile range, 60-80) years and 62.7% were male. 70 (46.7%) patients developed AKI during hospitalization, corresponding to the 17.3% in stage 1 and 9.3% in stage 2 and 20.0% in stage 3, respectively. Compared to patients without AKI, patients with AKI had higher proportion of mechanical ventilation mortality and higher in-hospital mortality. 95.5% patients with severe AKI received mechanical ventilation and in-hospital mortality was up to 79.5%. Severe AKI was independently associated with high in-hospital mortality (OR: 4.30;95% CI: 1.83-10.10). Logistic regression analysis demonstrated that high serum interleukin-6 (OR: 2.54;95%CI: 1.00-6.42) and interleukin-10 (OR: 3.02;95%CI: 1.17-7.82) were risk factors for severe AKI development. Conclusions: : severe AKI was associated with high in-hospital mortality and inflammatory response may play a role in AKI development in critically ill patients with COVID-19.

3.
EuropePMC; 2020.
Preprint in English | EuropePMC | ID: ppcovidwho-308229

ABSTRACT

Background: :An outbreak of coronavirus disease 2019 (COVID-19) caused by the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) began in Wuhan, Hubei Province, China in December 2019. This study aims to report the clinical characteristics of children COVID-19 in Xiangyang, a city outside of Wuhan within Hubei Province.Methods:We retrospectively investigated the clinical manifestations, Chest CT imaging, and laboratory characteristics of confirmed cases of children with COVID-19 with WHO interim guidance in Xiangyang Central Hospital from Feb 1 to Mar 10, 2020. 10 children cases were confirmed by real-time RT-PCR and were analyzed for epidemiological, demographic, clinical, radiological features and laboratory data. Outcomes were followed up until Mar 10, 2020.Results:6 cases (60%) had never been to Wuhan but closely contacted with family members with confirmed COVID-19, and 4 cases (40%) had made short term trips to Wuhan alone without familial clustering. The most common symptoms were cough (50%) followed by fever (40%), 4 cases (40%) showed asymptomatic characteristics including 2 cases (20%) with abnormal chest computed tomograms (CT) image. 9 cases (90%) were mild type, only 1 case (10%) was moderate type, none of them progressed in severe or critical disease. 4 (40%) cases showed leucopenia but none lymphopenia. Abnormalities on chest CT were detected among 8 cases (80%), 2 of 4 cases without obvious symptoms had abnormal chest CT. Conclusions: Children's infection is mainly caused by family clusters. No transmission to other individuals from children was found in our observation. The clinical manifestations in children with COVID-19 are non-specific with milder symptoms and good outcomes.

4.
Viruses ; 14(2)2022 02 07.
Article in English | MEDLINE | ID: covidwho-1674827

ABSTRACT

A new SARS-CoV-2 variant B.1.1.529 was named by the WHO as Omicron and classified as a Variant of Concern (VOC) on 26 November 2021. Because this variant has more than 50 mutations, including 30 mutations on the spike, it has generated a lot of concerns on the potential impacts of the VOC on COVID-19. Here through ELISA assays using the recombinant RBD proteins with sequences the same to that of SARS-CoV-2 WIV04 (lineage B.1), the Delta variant and the Omicron variant as the coating antigens, the binding capabilities between the RBDs and the antibodies in COVID-19 convalescent sera and vaccine sera after two doses of the inactivated vaccine produced by Sinopharm WIBP are compared with each other. The results showed that the Omicron variant may evade antibodies induced by the ancestral strain and by the inactivated vaccine, with significant reduction in the binding capability of its RBD much greater than that of the Delta variant.


Subject(s)
Antibodies, Viral/metabolism , Binding Sites, Antibody/physiology , COVID-19 Vaccines/immunology , COVID-19/immunology , Convalescence , SARS-CoV-2/immunology , SARS-CoV-2/metabolism , Antibodies, Neutralizing/immunology , Antibodies, Neutralizing/metabolism , Antibodies, Viral/blood , Antibodies, Viral/immunology , Humans , Immune Evasion , Mutation , Neutralization Tests , Vaccines, Inactivated/immunology
5.
Genome Res ; 32(2): 228-241, 2022 02.
Article in English | MEDLINE | ID: covidwho-1642462

ABSTRACT

The pathogenesis of COVID-19 is still elusive, which impedes disease progression prediction, differential diagnosis, and targeted therapy. Plasma cell-free RNAs (cfRNAs) carry unique information from human tissue and thus could point to resourceful solutions for pathogenesis and host-pathogen interactions. Here, we performed a comparative analysis of cfRNA profiles between COVID-19 patients and healthy donors using serial plasma. Analyses of the cfRNA landscape, potential gene regulatory mechanisms, dynamic changes in tRNA pools upon infection, and microbial communities were performed. A total of 380 cfRNA molecules were up-regulated in all COVID-19 patients, of which seven could serve as potential biomarkers (AUC > 0.85) with great sensitivity and specificity. Antiviral (NFKB1A, IFITM3, and IFI27) and neutrophil activation (S100A8, CD68, and CD63)-related genes exhibited decreased expression levels during treatment in COVID-19 patients, which is in accordance with the dynamically enhanced inflammatory response in COVID-19 patients. Noncoding RNAs, including some microRNAs (let 7 family) and long noncoding RNAs (GJA9-MYCBP) targeting interleukin (IL6/IL6R), were differentially expressed between COVID-19 patients and healthy donors, which accounts for the potential core mechanism of cytokine storm syndromes; the tRNA pools change significantly between the COVID-19 and healthy group, leading to the accumulation of SARS-CoV-2 biased codons, which facilitate SARS-CoV-2 replication. Finally, several pneumonia-related microorganisms were detected in the plasma of COVID-19 patients, raising the possibility of simultaneously monitoring immune response regulation and microbial communities using cfRNA analysis. This study fills the knowledge gap in the plasma cfRNA landscape of COVID-19 patients and offers insight into the potential mechanisms of cfRNAs to explain COVID-19 pathogenesis.


Subject(s)
COVID-19 , Cell-Free Nucleic Acids , RNA/blood , COVID-19/blood , COVID-19/genetics , Cell-Free Nucleic Acids/blood , Cytokine Release Syndrome , Humans , SARS-CoV-2
6.
Int J Antimicrob Agents ; 59(1): 106499, 2022 Jan.
Article in English | MEDLINE | ID: covidwho-1587677

ABSTRACT

In a bid to contain the current COVID-19 (coronavirus disease 2019) pandemic, various countermeasures have been applied. To date, however, there is a lack of an effective drug for the treatment of COVID-19. Through molecular modelling studies, simeprevir, a protease inhibitor approved for the management of hepatitis C virus infection, has been predicted as a potential antiviral against SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2), the causative agent of COVID-19. Here we assessed the efficacy of simeprevir against SARS-CoV-2 both in vitro in Vero E6 cells and in vivo in a human angiotensin-converting enzyme 2 (hACE2) transgenic mouse model. The results showed that simeprevir could inhibit SARS-CoV-2 replication in Vero E6 cells with a half-maximal effective concentration (EC50) of 1.41 ± 0.12 µM. In a transgenic hACE2 mouse model of SARS-CoV-2 infection, intraperitoneal administration of simeprevir at 10 mg/kg/day for 3 consecutive days failed to suppress viral replication. These findings collectively imply that simeprevir does not inhibit SARS-CoV-2 in vivo and therefore do not support its application as a treatment against COVID-19 at a dosage of 10 mg/kg/day.


Subject(s)
Angiotensin-Converting Enzyme 2/genetics , Antiviral Agents/pharmacology , Protease Inhibitors/pharmacology , SARS-CoV-2/drug effects , Simeprevir/pharmacology , Virus Replication/drug effects , Animals , Antiviral Agents/therapeutic use , COVID-19/drug therapy , COVID-19/virology , Chlorocebus aethiops , Dose-Response Relationship, Drug , Humans , Male , Mice , Mice, Transgenic , Negative Results , Protease Inhibitors/therapeutic use , Simeprevir/therapeutic use , Vero Cells
7.
2021.
Preprint in English | Other preprints | ID: ppcovidwho-295327

ABSTRACT

To unravel the source of SARS-CoV-2 introduction and the pattern of its spreading and evolution in the United Arab Emirates, we conducted meta-transcriptome sequencing of 1,067 nasopharyngeal swab samples collected between May 9th and Jun 29th, 2020 during the first peak of the local COVID-19 epidemic. We identified global clade distribution and eleven novel genetic variants that were almost absent in the rest of the world defined five subclades specific to the UAE viral population. Cross-settlement human-to-human transmission was related to the local business activity. Perhaps surprisingly, at least 5% of the population were co-infected by SARS-CoV-2 of multiple clades within the same host. We also discovered an enrichment of cytosine-to-uracil mutation among the viral population collected from the nasopharynx, that is different from the adenosine-to-inosine change previously reported in the bronchoalveolar lavage fluid samples and a previously unidentified upregulation of APOBEC4 expression in nasopharynx among infected patients, indicating the innate immune host response mediated by ADAR and APOBEC gene families could be tissue-specific. The genomic epidemiological and molecular biological knowledge reported here provides new insights for the SARS-CoV-2 evolution and transmission and points out future direction on host-pathogen interaction investigation.

8.
J Adv Res ; 36: 201-210, 2022 02.
Article in English | MEDLINE | ID: covidwho-1536631

ABSTRACT

Introduction: The COVID-19 global epidemic caused by severe acute respiratory syndrome coronavirus (SARS-CoV-2) is a great public health emergency. Discovering antiviral drug candidates is urgent for the prevention and treatment of COVID-19. Objectives: This work aims to discover natural SARS-CoV-2 inhibitors from the traditional Chinese herbal medicine licorice. Methods: We screened 125 small molecules from Glycyrrhiza uralensis Fisch. (licorice, Gan-Cao) by virtual ligand screening targeting the receptor-binding domain (RBD) of SARS-CoV-2 spike protein. Potential hit compounds were further evaluated by ELISA, SPR, luciferase assay, antiviral assay and pharmacokinetic study. Results: The triterpenoids licorice-saponin A3 (A3) and glycyrrhetinic acid (GA) could potently inhibit SARS-CoV-2 infection, with EC50 of 75 nM and 3.17 µM, respectively. Moreover, we reveal that A3 mainly targets the nsp7 protein, and GA binds to the spike protein RBD of SARS-CoV-2. Conclusion: In this work, we found GA and A3 from licorice potently inhibit SARS-CoV-2 infection by affecting entry and replication of the virus. Our findings indicate that these triterpenoids may contribute to the clinical efficacy of licorice for COVID-19 and could be promising candidates for antiviral drug development.


Subject(s)
COVID-19 , Glycyrrhiza , Triterpenes , Humans , SARS-CoV-2 , Spike Glycoprotein, Coronavirus , Triterpenes/pharmacology
11.
Infect Drug Resist ; 14: 3175-3181, 2021.
Article in English | MEDLINE | ID: covidwho-1372035

ABSTRACT

OBJECTIVE: The number of children presenting with coronavirus disease 2019 (COVID-19) caused by SARS-CoV-2 infection is increasing, and we aimed to assess the clinical characteristics of pediatric patients with a definite epidemiological history during the early COVID-19 epidemic. METHODS: Retrospective analysis was performed on the clinical data of children admitted to the fever ward of Xiangyang Central Hospital in Hubei province between January 1, 2020 and March 17, 2020. According to definite epidemiological history, patients with SARS-CoV-2 nucleic acid test (NAT) positive detection were grouped as confirmed cases, and patients with two consecutive negative NATs were grouped as suspected cases. We compared the clinical characteristics of the two groups. RESULTS: A total of 47 (47/127, 37%) cases had a definite epidemiological history, of which 32 (68.1%) were suspected, with a median age of 5.5 years (interquartile range [IQR]: 0.7-10.3), and 15 (31.9%) were confirmed, with a median age of 9 years (IQR: 4-14). Statistically significant differences in age, family cluster of infection, and numbers of patients with clinical symptoms and fever (P<0.05) were found between the two groups, but no statistically significant differences in leucocyte and lymphocyte counts were observed (P>0.05). Significant differences were found in the computed tomography (CT) manifestation of ground glass opacity (GGO) between the two groups (P<0.05). CONCLUSION: Children of older age and from family clusters of infection were more easily diagnosed as having COVID-19. GGO changes on chest CT was more likely in confirmed cases. Although obvious clinical manifestations increase our awareness of COVID-19, children without manifestations of fever or cough should not be ignored as they may be asymptomatic carriers.

12.
Pediatr Investig ; : e12282, 2021 Aug 12.
Article in English | MEDLINE | ID: covidwho-1353594

ABSTRACT

IMPORTANCE: The Coronavirus disease 2019 (COVID-19) global pandemic poses a considerable challenge for pediatricians. OBJECTIVE: This study aimed to identify the epidemiological characteristics and clinical features of pediatric patients with COVID-19 in China. METHODS: This multicenter retrospective study included pediatric patients from 46 hospitals in China, covering 12 provinces and two municipalities. Epidemiological, demographic, clinical, laboratory, treatment, and outcome data were analyzed. RESULTS: In total, 211 pediatric patients with COVID-19 were included in this study. The median age was 7.0 years (range: 22 days to 18 years). Approximately 16.3% of the patients exhibited asymptomatic infections, 23.0% had upper respiratory tract infections, and 60.7% had pneumonia, including two with severe pneumonia and one with critical illness. Approximately 78.7% of the pediatric patients occurred in familial clusters. The most three common symptoms or signs at onset in children with COVID-19 were fever (54.5%), cough (49.3%), and pharyngeal congestion (20.8%). Only 17.6% of the patients presented with decreased lymphocyte count, whereas 13.6% had increased lymphocyte count. Among the patients with pneumonia who exhibited abnormal chest computed tomography findings, 18.2% (23/127) of the patients had no other symptoms. Generally, the chest radiographs showed abnormalities that affected both lungs (49.6%); ground-glass opacity (47.2%) was the most common manifestation. The cure and improvement rates were 86.7% (183/211) and 13.3% (28/211), respectively. Only one patient with an underlying condition received invasive mechanical ventilation; none of the patients died. INTERPRETATION: Similar to adults, children of all age groups are susceptible to COVID-19. Fortunately, most pediatric patients have mild symptoms or remain asymptomatic, despite the high incidence of pneumonia. Decreased proportions of white blood cells and lymphocytes are less frequent in children than in adults.

13.
Front Cell Infect Microbiol ; 11: 663884, 2021.
Article in English | MEDLINE | ID: covidwho-1317217

ABSTRACT

Background: The pandemic of Coronavirus Disease 2019 (COVID-19) brings new challenges for pediatricians, especially in the differentiation with non-COVID-19 pneumonia in the peak season of pneumonia. We aimed to compare the clinical characteristics of pediatric patients with COVID-19 and other respiratory pathogens infected pneumonias. Methods: We conducted a multi-center, cross-sectional study of pediatric inpatients in China. Based on pathogenic test results, pediatric patients were divided into three groups, including COVID-19 pneumonia group, Non-COVID-19 viral (NCV) pneumonia group and Non-viral (NV) pneumonia group. Their clinical characteristics were compared by Kruskal-Wallis H test or chi-square test. Results: A total of 636 pediatric pneumonia inpatients, among which 87 in COVID-19 group, 194 in NCV group, and 355 in NV group, were included in analysis. Compared with NCV and NV patients, COVID-19 patients were older (median age 6.33, IQR 2.00-12.00 years), and relatively fewer COVID-19 patients presented fever (63.2%), cough (60.9%), shortness of breath (1.1%), and abnormal pulmonary auscultation (18.4%). The results were verified by the comparison of COVID-19, respiratory syncytial virus (RSV) and influenza A (IFA) pneumonia patients. Approximately 42.5%, 44.8%, and 12.6% of the COVID-19 patients presented simply ground-glass opacity (GGO), simply consolidation, and the both changes on computed tomography (CT) scans, respectively; the proportions were similar as those in NCV and NV group (p>0.05). Only 47.1% of COVID-19 patients had both lungs pneumonia, which was significantly lower than that proportion of nearly 80% in the other two groups. COVID-19 patients presented lower proportions of increased white blood cell count (16.5%) and abnormal procalcitonin (PCT) (10.7%), and a higher proportion of decreased lymphocyte count (44.0%) compared with the other two groups. Conclusion: Majority clinical characteristics of pediatric COVID-19 pneumonia patients were milder than non-COVID-19 patients. However, lymphocytopenia remained a prominent feature of COVID-19 pediatric pneumonia.


Subject(s)
COVID-19 , Pneumonia , Child , China/epidemiology , Cross-Sectional Studies , Humans , Lung/diagnostic imaging , Pneumonia/epidemiology , Retrospective Studies , SARS-CoV-2
14.
Sci Total Environ ; 797: 149085, 2021 Nov 25.
Article in English | MEDLINE | ID: covidwho-1313422

ABSTRACT

The ongoing COVID-19 pandemic has generated a global health crisis that needs well management of not only patients but also environments to reduce SARS-CoV-2 transmission. The gold standard RT-qPCR method is sensitive and rapid to detect SARS-CoV-2 nucleic acid, but does not answer if PCR-positive samples contain infectious virions. To circumvent this problem, we report an SDS-propidium monoazide (PMA) assisted RT-qPCR method that enables rapid discrimination of live and dead SARS-CoV-2 within 3 h. PMA, a photo-reactive dye, can react with viral RNA released or inside inactivated SARS-CoV-2 virions under assistance of 0.005% SDS, but not viral RNA inside live virions. Formation of PMA-RNA conjugates prevents PCR amplification, leaving only infectious virions to be detected. Under optimum conditions, RT-qPCR detection of heat-inactivated SARS-CoV-2 resulted in larger than 9 Ct value differences between PMA-treated and PMA-free groups, while less than 0.5 Ct differences were observed in the detection of infectious SARS-CoV-2 ranging from 20 to 5148 viral particles. Using a cutoff Ct difference of 8.6, this method could differentiate as low as 8 PFU live viruses in the mixtures of live and heat-inactivated virions. Further experiments showed that this method could successfully monitor the natural inactivation process of SARS-CoV-2 on plastic surfaces during storage with comparable results to the gold standard plaque assay. We believe that the culture-free method established here could be used for rapid and convenient determination of infectious SARS-CoV-2 virions in PCR-positive samples, which will facilitate better control of SARS-CoV-2 transmission.


Subject(s)
COVID-19 , SARS-CoV-2 , Azides , Humans , Pandemics , Propidium/analogs & derivatives , Real-Time Polymerase Chain Reaction , Sensitivity and Specificity
15.
Front Med ; 16(1): 102-110, 2022 Feb.
Article in English | MEDLINE | ID: covidwho-1303365

ABSTRACT

Consecutively hospitalized patients with confirmed coronavirus disease 2019 (COVID-19) in Wuhan, China were retrospectively enrolled from January 2020 to March 2020 to investigate the association between the use of renin-angiotensin system inhibitor (RAS-I) and the outcome of this disease. Associations between the use of RAS-I (angiotensin-converting enzyme inhibitor (ACEI) or angiotensin receptor blocker (ARB)), ACEI, and ARB and in-hospital mortality were analyzed using multivariate Cox proportional hazards regression models in overall and subgroup of hypertension status. A total of 2771 patients with COVID-19 were included, with moderate and severe cases accounting for 45.0% and 36.5%, respectively. A total of 195 (7.0%) patients died. RAS-I (hazard ratio (HR)= 0.499, 95% confidence interval (CI) 0.325-0.767) and ARB (HR = 0.410, 95% CI 0.240-0.700) use was associated with a reduced risk of all-cause mortality among patients with COVID-19. For patients with hypertension, RAS-I and ARB applications were also associated with a reduced risk of mortality with HR of 0.352 (95% CI 0.162-0.764) and 0.279 (95% CI 0.115-0.677), respectively. RAS-I exhibited protective effects on the survival outcome of COVID-19. ARB use was associated with a reduced risk of all-cause mortality among patients with COVID-19.


Subject(s)
COVID-19 , Hypertension , Angiotensin Receptor Antagonists/therapeutic use , Angiotensin-Converting Enzyme Inhibitors/therapeutic use , Humans , Hypertension/complications , Hypertension/drug therapy , Renin-Angiotensin System , Retrospective Studies
16.
Sci Rep ; 11(1): 13971, 2021 07 07.
Article in English | MEDLINE | ID: covidwho-1301179

ABSTRACT

To unravel the source of SARS-CoV-2 introduction and the pattern of its spreading and evolution in the United Arab Emirates, we conducted meta-transcriptome sequencing of 1067 nasopharyngeal swab samples collected between May 9th and Jun 29th, 2020 during the first peak of the local COVID-19 epidemic. We identified global clade distribution and eleven novel genetic variants that were almost absent in the rest of the world and that defined five subclades specific to the UAE viral population. Cross-settlement human-to-human transmission was related to the local business activity. Perhaps surprisingly, at least 5% of the population were co-infected by SARS-CoV-2 of multiple clades within the same host. We also discovered an enrichment of cytosine-to-uracil mutation among the viral population collected from the nasopharynx, that is different from the adenosine-to-inosine change previously reported in the bronchoalveolar lavage fluid samples and a previously unidentified upregulation of APOBEC4 expression in nasopharynx among infected patients, indicating the innate immune host response mediated by ADAR and APOBEC gene families could be tissue-specific. The genomic epidemiological and molecular biological knowledge reported here provides new insights for the SARS-CoV-2 evolution and transmission and points out future direction on host-pathogen interaction investigation.


Subject(s)
COVID-19/epidemiology , COVID-19/immunology , Coinfection/epidemiology , Genomics , Immunity, Innate , Mutation , SARS-CoV-2/genetics , Adult , COVID-19/transmission , Cytidine Deaminase/genetics , Female , Gene Expression Profiling , Genome, Viral/genetics , Humans , Male , Middle Aged , Nasopharynx/virology , Organ Specificity , SARS-CoV-2/immunology
17.
Virulence ; 12(1): 1209-1226, 2021 12.
Article in English | MEDLINE | ID: covidwho-1242086

ABSTRACT

New SARS-CoV-2 mutants have been continuously indentified with enhanced transmission ever since its outbreak in early 2020. As an RNA virus, SARS-CoV-2 has a high mutation rate due to the low fidelity of RNA polymerase. To study the single nucleotide polymorphisms (SNPs) dynamics of SARS-CoV-2, 158 SNPs with high confidence were identified by deep meta-transcriptomic sequencing, and the most common SNP type was C > T. Analyses of intra-host population diversity revealed that intra-host quasispecies' composition varies with time during the early onset of symptoms, which implicates viral evolution during infection. Network analysis of co-occurring SNPs revealed the most abundant non-synonymous SNP 22,638 in the S glycoprotein RBD region and 28,144 in the ORF8 region. Furthermore, SARS-CoV-2 variations differ in an individual's respiratory tissue (nose, throat, BALF, or sputum), suggesting independent compartmentalization of SARS-CoV-2 populations in patients. The positive selection analysis of the SARS-CoV-2 genome uncovered the positive selected amino acid G251V on ORF3a. Alternative allele frequency spectrum (AAFS) of all variants revealed that ORF8 could bear alternate alleles with high frequency. Overall, the results show the quasispecies' profile of SARS-CoV-2 in the respiratory tract in the first two months after the outbreak.


Subject(s)
Phylogeny , Polymorphism, Single Nucleotide , Quasispecies , SARS-CoV-2/classification , SARS-CoV-2/genetics , Adult , Aged , Aged, 80 and over , Alleles , COVID-19/virology , Computational Biology , Coronavirus Envelope Proteins/chemistry , Coronavirus Envelope Proteins/genetics , Female , Gene Frequency , Genome, Viral , HEK293 Cells , High-Throughput Nucleotide Sequencing , Humans , Male , Middle Aged , Severity of Illness Index , Young Adult
18.
J Vis Exp ; (169)2021 03 31.
Article in English | MEDLINE | ID: covidwho-1192273

ABSTRACT

Diagnosis of the ongoing SARS-CoV-2 pandemic is a priority for all countries across the globe. Currently, reverse transcription quantitative PCR (RT-qPCR) is the gold standard for SARS-CoV-2 diagnosis as no permanent solution is available. However effective this technique may be, research has emerged showing its limitations in detection and diagnosis especially when it comes to low abundant targets. In contrast, droplet digital PCR (ddPCR), a recent emerging technology with superior advantages over qPCR, has been shown to overcome the challenges of RT-qPCR in diagnosis of SARS-CoV-2 from low abundant target samples. Prospectively, in this article, the capabilities of RT-ddPCR are further expanded by showing steps on how to develop simplex, duplex, triplex probe mix, and quadruplex assays using a two-color detection system. Using primers and probes targeting specific sites of the SARS-CoV-2 genome (N, ORF1ab, RPP30, and RBD2), the development of these assays is shown to be possible. Additionally, step by step detailed protocols, notes, and suggestions on how to improve the assays workflow and analyze data are provided. Adapting this workflow in future works will ensure that the maximum number of targets can be sensitively detected in a small sample significantly improving on cost and sample throughput.


Subject(s)
COVID-19 Nucleic Acid Testing/methods , COVID-19/diagnosis , Reverse Transcriptase Polymerase Chain Reaction/methods , SARS-CoV-2/isolation & purification , DNA Primers , Humans , Pandemics , RNA, Viral/genetics , Reverse Transcription , Sensitivity and Specificity
19.
Cell Discov ; 7(1): 23, 2021 Apr 13.
Article in English | MEDLINE | ID: covidwho-1182823

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a global pandemic of Coronavirus disease 2019 (COVID-19). However, the microbial composition of the respiratory tract and other infected tissues as well as their possible pathogenic contributions to varying degrees of disease severity in COVID-19 patients remain unclear. Between 27 January and 26 February 2020, serial clinical specimens (sputum, nasal and throat swab, anal swab and feces) were collected from a cohort of hospitalized COVID-19 patients, including 8 mildly and 15 severely ill patients in Guangdong province, China. Total RNA was extracted and ultra-deep metatranscriptomic sequencing was performed in combination with laboratory diagnostic assays. We identified distinct signatures of microbial dysbiosis among severely ill COVID-19 patients on broad spectrum antimicrobial therapy. Co-detection of other human respiratory viruses (including human alphaherpesvirus 1, rhinovirus B, and human orthopneumovirus) was demonstrated in 30.8% (4/13) of the severely ill patients, but not in any of the mildly affected patients. Notably, the predominant respiratory microbial taxa of severely ill patients were Burkholderia cepacia complex (BCC), Staphylococcus epidermidis, or Mycoplasma spp. (including M. hominis and M. orale). The presence of the former two bacterial taxa was also confirmed by clinical cultures of respiratory specimens (expectorated sputum or nasal secretions) in 23.1% (3/13) of the severe cases. Finally, a time-dependent, secondary infection of B. cenocepacia with expressions of multiple virulence genes was demonstrated in one severely ill patient, which might accelerate his disease deterioration and death occurring one month after ICU admission. Our findings point to SARS-CoV-2-related microbial dysbiosis and various antibiotic-resistant respiratory microbes/pathogens in hospitalized COVID-19 patients in relation to disease severity. Detection and tracking strategies are needed to prevent the spread of antimicrobial resistance, improve the treatment regimen and clinical outcomes of hospitalized, severely ill COVID-19 patients.

20.
Kidney Dis (Basel) ; 7(2): 111-119, 2021 Mar.
Article in English | MEDLINE | ID: covidwho-1159236

ABSTRACT

BACKGROUND: Coronavirus disease 2019 (COVID-19) has emerged as a major global health threat with a great number of deaths worldwide. Acute kidney injury (AKI) is a common complication in patients admitted to the intensive care unit. We aimed to assess the incidence, risk factors and in-hospital outcomes of AKI in COVID-19 patients admitted to the intensive care unit. METHODS: We conducted a retrospective observational study in the intensive care unit of Tongji Hospital, which was assigned responsibility for the treatments of severe COVID-19 patients by the Wuhan government. AKI was defined and staged based on Kidney Disease: Improving Global Outcomes (KDIGO) criteria. Mild AKI was defined as stage 1, and severe AKI was defined as stage 2 or stage 3. Logistic regression analysis was used to evaluate AKI risk factors, and Cox proportional hazards model was used to assess the association between AKI and in-hospital mortality. RESULTS: A total of 119 patients with COVID-19 were included in our study. The median patient age was 70 years (interquartile range, 59-77) and 61.3% were male. Fifty-one (42.8%) patients developed AKI during hospitalization, corresponding to 14.3% in stage 1, 28.6% in stage 2 and 18.5% in stage 3, respectively. Compared to patients without AKI, patients with AKI had a higher proportion of mechanical ventilation mortality and higher in-hospital mortality. A total of 97.1% of patients with severe AKI received mechanical ventilation and in-hospital mortality was up to 79.4%. Severe AKI was independently associated with high in-hospital mortality (OR: 1.82; 95% CI: 1.06-3.13). Logistic regression analysis demonstrated that high serum interleukin-8 (OR: 4.21; 95% CI: 1.23-14.38), interleukin-10 (OR: 3.32; 95% CI: 1.04-10.59) and interleukin-2 receptor (OR: 4.50; 95% CI: 0.73-6.78) were risk factors for severe AKI development. CONCLUSIONS: Severe AKI was associated with high in-hospital mortality, and inflammatory response may play a role in AKI development in critically ill patients with COVID-19.

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