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2.
Zhejiang Da Xue Xue Bao Yi Xue Ban ; 50(6): 748-754, 2021 Dec 25.
Article in English | MEDLINE | ID: covidwho-1753705

ABSTRACT

To explore the application value of nanopore sequencing technique in the diagnosis and treatment of secondary infections in patients with severe coronavirus disease 2019 (COVID-19). A total of 77 clinical specimens from 3 patients with severe COVID-19 were collected. After heat inactivation, all samples were subjected to total nucleic acid extraction based on magnetic bead enrichment. The extracted DNA was used for DNA library construction, then nanopore real-time sequencing detection was performed. The sequencing data were subjected to Centrifuge software database species matching and R program differential analysis to obtain potential pathogen identification. Nanopore sequencing results were compared with respiratory pathogen qPCR panel screening and conventional microbiological testing results to verify the effectiveness of nanopore sequencing detection. Nanopore sequencing results showed that positive pathogen were obtained in 44 specimens (57.1%). The potential pathogens identified by nanopore sequencing included , , and , et al. , , were also detected in clinical microbiological culture-based detection; was detected in respiratory pathogen screening qPCR panel; was only detected by the nanopore sequencing technique. Comprehensive considerations with the clinical symptoms, the patient was treated with antibiotics against , and the infection was controlled. Nanopore sequencing may assist the diagnosis and treatment of severe COVID-19 patients through rapid identification of potential pathogens.


Subject(s)
COVID-19 , Coinfection , Nanopore Sequencing , Nanopores , COVID-19/diagnosis , Humans , Sequence Analysis, DNA/methods
3.
EuropePMC; 2020.
Preprint in English | EuropePMC | ID: ppcovidwho-325159

ABSTRACT

At least three months have been passed since the outbreak of the severe acute respiratory disease, COVID-19 in Wuhan city, China in December 2019, caused by the infection of a novel coronavirus, SARS-CoV-2. 1,2 . Due to its rapid spread throughout China and abroad, knowledge sharing for both its epidemiology and clinic manifestations is urgently need. Here we analyzed the clinical, molecular and immunological data from 326 confirmed cases of SARS-CoV-2 infection in Shanghai. Genomic sequences assembled from 112 quality samples together with uploaded sequences in Global Initiative on Sharing All Influenza Data (GISAID) showed a stable evolution and suggested two major lineages with differential exposure history during the earliest outbreak in Wuhan. Nevertheless, they exhibited similar virulence and clinical outcomes. Lymphocytopenia, especially the reduced CD4+ and CD8+ T cell counts upon admission, was predictive of disease progression. High level of IL-6 and IL-8 during treatment was observed in severe and critical patients and correlated with decreased lymphocyte count. The determinants of disease severity seemed to stem mostly from host factors such age, lymphocytopenia and its associated cytokine storm whereas viral genetic variation did not significantly affect the outcomes. This comprehensive analysis on the molecular, immunological and clinical data provides a panorama of the key determinants related to the disease outcomes which should be helpful for improving the current combat against this extremely aggressive pandemic.Authors Xiaonan Zhang, Yun Tan, Yun Ling, Gang Lu, Feng Liu, and Zhigang Yi contributed equally to this work.

4.
Sci China Life Sci ; 2021 Dec 24.
Article in English | MEDLINE | ID: covidwho-1596898

ABSTRACT

The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a global crisis. Clinical candidates with high efficacy, ready availability, and that do not develop resistance are in urgent need. Despite that screening to repurpose clinically approved drugs has provided a variety of hits shown to be effective against SARS-CoV-2 infection in cell culture, there are few confirmed antiviral candidates in vivo. In this study, 94 compounds showing high antiviral activity against SARS-CoV-2 in Vero E6 cells were identified from 2,580 FDA-approved small-molecule drugs. Among them, 24 compounds with low cytotoxicity were selected, and of these, 17 compounds also effectively suppressed SARS-CoV-2 infection in HeLa cells transduced with human ACE2. Six compounds disturb multiple processes of the SARS-CoV-2 life cycle. Their prophylactic efficacies were determined in vivo using Syrian hamsters challenged with SARS-CoV-2 infection. Seven compounds reduced weight loss and promoted weight regain of hamsters infected not only with the original strain but also the D614G variant. Except for cisatracurium, six compounds reduced hamster pulmonary viral load, and IL-6 and TNF-α mRNA when assayed at 4 d postinfection. In particular, sertraline, salinomycin, and gilteritinib showed similar protective effects as remdesivir in vivo and did not induce antiviral drug resistance after 10 serial passages of SARS-CoV-2 in vitro, suggesting promising application for COVID-19 treatment.

5.
Nat Commun ; 12(1): 5026, 2021 08 18.
Article in English | MEDLINE | ID: covidwho-1363491

ABSTRACT

Nationwide prospective surveillance of all-age patients with acute respiratory infections was conducted in China between 2009‒2019. Here we report the etiological and epidemiological features of the 231,107 eligible patients enrolled in this analysis. Children <5 years old and school-age children have the highest viral positivity rate (46.9%) and bacterial positivity rate (30.9%). Influenza virus, respiratory syncytial virus and human rhinovirus are the three leading viral pathogens with proportions of 28.5%, 16.8% and 16.7%, and Streptococcus pneumoniae, Mycoplasma pneumoniae and Klebsiella pneumoniae are the three leading bacterial pathogens (29.9%, 18.6% and 15.8%). Negative interactions between viruses and positive interactions between viral and bacterial pathogens are common. A Join-Point analysis reveals the age-specific positivity rate and how this varied for individual pathogens. These data indicate that differential priorities for diagnosis, prevention and control should be highlighted in terms of acute respiratory tract infection patients' demography, geographic locations and season of illness in China.


Subject(s)
Bacteria/isolation & purification , Bacterial Infections/microbiology , Respiratory Tract Infections/microbiology , Respiratory Tract Infections/virology , Virus Diseases/virology , Viruses/isolation & purification , Adolescent , Adult , Bacteria/classification , Bacteria/genetics , Bacterial Infections/epidemiology , Child , Child, Preschool , China/epidemiology , Female , Humans , Infant , Male , Prospective Studies , Respiratory Tract Infections/epidemiology , Seasons , Virus Diseases/epidemiology , Viruses/classification , Viruses/genetics , Young Adult
6.
Cell Discov ; 7(1): 44, 2021 Jun 15.
Article in English | MEDLINE | ID: covidwho-1269383

ABSTRACT

The presence of SARS-CoV-2 mutants, including the emerging variant B.1.1.7, has raised great concerns in terms of pathogenesis, transmission, and immune escape. Characterizing SARS-CoV-2 mutations, evolution, and effects on infectivity and pathogenicity is crucial to the design of antibody therapies and surveillance strategies. Here, we analyzed 454,443 SARS-CoV-2 spike genes/proteins and 14,427 whole-genome sequences. We demonstrated that the early variant B.1.1.7 may not have evolved spontaneously in the United Kingdom or within human populations. Our extensive analyses suggested that Canidae, Mustelidae or Felidae, especially the Canidae family (for example, dog) could be a possible host of the direct progenitor of variant B.1.1.7. An alternative hypothesis is that the variant was simply yet to be sampled. Notably, the SARS-CoV-2 whole-genome represents a large number of potential co-mutations. In addition, we used an experimental SARS-CoV-2 reporter replicon system to introduce the dominant co-mutations NSP12_c14408t, 5'UTR_c241t, and NSP3_c3037t into the viral genome, and to monitor the effect of the mutations on viral replication. Our experimental results demonstrated that the co-mutations significantly attenuated the viral replication. The study provides valuable clues for discovering the transmission chains of variant B.1.1.7 and understanding the evolutionary process of SARS-CoV-2.

7.
Antiviral Res ; 185: 104974, 2021 01.
Article in English | MEDLINE | ID: covidwho-927811

ABSTRACT

Vaccines and antiviral agents are in urgent need to stop the COVID-19 pandemic. To facilitate antiviral screening against SARS-CoV-2 without requirement for high biosafety level facility, we developed a bacterial artificial chromosome (BAC)-vectored replicon of SARS-CoV-2, nCoV-SH01 strain, in which secreted Gaussia luciferase (sGluc) was encoded in viral subgenomic mRNA as a reporter gene. The replicon was devoid of structural genes spike (S), membrane (M), and envelope (E). Upon transfection, the replicon RNA replicated in various cell lines, and was sensitive to interferon alpha (IFN-α), remdesivir, but was resistant to hepatitis C virus inhibitors daclatasvir and sofosbuvir. Replication of the replicon was also sensitive overexpression to zinc-finger antiviral protein (ZAP). We also constructed a four-plasmid in-vitro ligation system that is compatible with the BAC system, which makes it easy to introduce desired mutations into the assembly plasmids for in-vitro ligation. This replicon system would be helpful for performing antiviral screening and dissecting virus-host interactions.


Subject(s)
Antiviral Agents/pharmacology , COVID-19/virology , Chromosomes, Artificial, Bacterial , Replicon/drug effects , SARS-CoV-2/drug effects , SARS-CoV-2/genetics , Adenosine Monophosphate/analogs & derivatives , Adenosine Monophosphate/pharmacology , Alanine/analogs & derivatives , Alanine/pharmacology , Animals , COVID-19/drug therapy , Cell Line , Chlorocebus aethiops , HEK293 Cells , Hepacivirus/drug effects , Hepatitis C/drug therapy , Hepatitis C/virology , Humans , Interferon-alpha/pharmacology , RNA-Binding Proteins/biosynthesis , RNA-Binding Proteins/genetics , Sofosbuvir/pharmacology , Vero Cells , Virus Replication/drug effects
8.
9.
Emerg Microbes Infect ; 9(1): 1988-1996, 2020 Dec.
Article in English | MEDLINE | ID: covidwho-730431

ABSTRACT

ABSTRACT Pandemic SARS-CoV-2 has caused unprecedented mortalities. Vaccine is in urgent need to stop the pandemic. Despite great progresses on SARS-CoV-2 vaccine development, the efficacy of the vaccines remains to be determined. Deciphering the interactions of the viral epitopes with the elicited neutralizing antibodies in convalescent population inspires the vaccine development. In this study, we devised a peptide array composed of 20-mer overlapped peptides of spike (S), membrane (M) and envelope (E) proteins, and performed a screening with 120 COVID-19 convalescent sera and 24 non-COVID-19 sera. We identified five SARS-CoV-2-specific dominant epitopes that reacted with above 40% COVID-19 convalescent sera. Of note, two peptides non-specifically interacted with most of the non-COVID-19 sera. Neutralization assay indicated that only five sera completely blocked viral infection at the dilution of 1:200. By using a peptide-compete neutralizing assay, we found that three dominant epitopes partially competed the neutralization activity of several convalescent sera, suggesting antibodies elicited by these epitopes played an important role in neutralizing viral infection. The epitopes we identified in this study may serve as vaccine candidates to elicit neutralizing antibodies in most vaccinated people or specific antigens for SARS-CoV-2 diagnosis.


Subject(s)
Antibodies, Neutralizing/blood , Antibodies, Viral/blood , Betacoronavirus/immunology , Coronavirus Infections/prevention & control , Epitopes, B-Lymphocyte/immunology , Pandemics/prevention & control , Pneumonia, Viral/prevention & control , Viral Vaccines/immunology , Animals , B-Lymphocytes/immunology , COVID-19 , Cell Line , Chlorocebus aethiops , Coronavirus Infections/diagnosis , Coronavirus Infections/immunology , Coronavirus Infections/therapy , Humans , Immunization, Passive , Pneumonia, Viral/diagnosis , Pneumonia, Viral/immunology , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/immunology , Vero Cells , Viral Envelope Proteins/immunology
10.
Am J Infect Control ; 48(9): 1045-1050, 2020 09.
Article in English | MEDLINE | ID: covidwho-633888

ABSTRACT

BACKGROUND: Humans are generally susceptible to SARS-CoV-2, which has caused a global pandemic of COVID-19. The screening of infected people in the population still mainly depends on clinical symptoms. However, there is limited research on the characteristics of clinical symptoms in different populations, especially in imported cases. METHODS: To retrospectively analyze the clinical data of 494 confirmed COVID-19 patients admitted to a designated hospital in Shanghai from January 20, 2020, to March 31, 2020, we compared the clinical manifestations in different populations and their influencing factors in COVID-19 patients. RESULTS: (1) Of the 494 patients, 453 (91.7%) had different symptoms at admission, and 39 (7.89%) patients were asymptomatic. (2) We compared the symptoms of patients according to different stratifications and found the following results: (a) The proportion of dyspnea was significantly higher in male patients than in female patients (P < .05). (b) The proportions of a stuffy nose, sore throat, and olfactory and gustatory dysfunction were significantly higher in children than in adult patients (P < .05). (c) The proportions of fever, chest tightness, shortness of breath, and fatigue were significantly higher in local cases than in imported cases. In comparison, the proportions of nasal congestion, stuffy nose, sore throat, headache, and olfactory and gustatory dysfunction were significantly lower in imported cases than in imported cases (P < .05). (d) The proportions of chest tightness, shortness of breath, and dyspnea were significantly higher in severely ill patients than in those with mild symptoms (P < .05). (3) Thirty-one asymptomatic patients were significantly younger than symptomatic patients, and they had a higher proportion of imported cases, white blood cell and lymphocyte count levels, and fewer abnormal CT cases than the group of symptomatic patients (P < .05). (4) The number of days since the onset of the disease needed for the symptoms to disappear was associated with the epidemiological history (imported cases), the number of days until the pharyngeal swab nucleic acid test turned negative, the days of hospitalization, the days of onset, and the white blood cell and lymphocyte count levels (P < .05). CONCLUSIONS: The majority of COVID-19 patients (91.7%) had early symptoms, whereas 7.89% of COVID-19 patients were asymptomatic. Younger patients had fewer symptoms, mainly the upper respiratory symptoms, and the illness condition was milder, which was more common in imported cases. Elderly male patients had severe symptoms when admitted. The number of days needed for the patient's symptoms to disappear was closely related to the number of days necessary for the pharyngeal swab nucleic acid test to turn negative.


Subject(s)
Age Factors , Asymptomatic Infections/epidemiology , Betacoronavirus , Coronavirus Infections/pathology , Pneumonia, Viral/pathology , Severity of Illness Index , Adolescent , Adult , Aged , COVID-19 , China/epidemiology , Coronavirus Infections/virology , Female , Humans , Male , Middle Aged , Pandemics , Pneumonia, Viral/virology , Retrospective Studies , SARS-CoV-2 , Young Adult
11.
Nature ; 583(7816): 437-440, 2020 07.
Article in English | MEDLINE | ID: covidwho-326050

ABSTRACT

In December 2019, coronavirus disease 2019 (COVID-19), which is caused by the new coronavirus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), was identified in Wuhan (Hubei province, China)1; it soon spread across the world. In this ongoing pandemic, public health concerns and the urgent need for effective therapeutic measures require a deep understanding of the epidemiology, transmissibility and pathogenesis of COVID-19. Here we analysed clinical, molecular and immunological data from 326 patients with confirmed SARS-CoV-2 infection in Shanghai. The genomic sequences of SARS-CoV-2, assembled from 112 high-quality samples together with sequences in the Global Initiative on Sharing All Influenza Data (GISAID) dataset, showed a stable evolution and suggested that there were two major lineages with differential exposure history during the early phase of the outbreak in Wuhan. Nevertheless, they exhibited similar virulence and clinical outcomes. Lymphocytopenia, especially reduced CD4+ and CD8+ T cell counts upon hospital admission, was predictive of disease progression. High levels of interleukin (IL)-6 and IL-8 during treatment were observed in patients with severe or critical disease and correlated with decreased lymphocyte count. The determinants of disease severity seemed to stem mostly from host factors such as age and lymphocytopenia (and its associated cytokine storm), whereas viral genetic variation did not significantly affect outcomes.


Subject(s)
Betacoronavirus/genetics , Betacoronavirus/pathogenicity , Coronavirus Infections/immunology , Coronavirus Infections/virology , Host-Pathogen Interactions/immunology , Lymphopenia/virology , Pneumonia, Viral/immunology , Pneumonia, Viral/virology , Respiratory Distress Syndrome/virology , Adolescent , Adult , Aged , Aged, 80 and over , Aging , Animals , Asymptomatic Infections/epidemiology , Betacoronavirus/classification , Betacoronavirus/isolation & purification , COVID-19 , China/epidemiology , Cohort Studies , Coronavirus Infections/complications , Coronavirus Infections/epidemiology , Critical Illness/epidemiology , Disease Progression , Evolution, Molecular , Female , Genetic Variation , Genome, Viral/genetics , Hospitalization/statistics & numerical data , Humans , Inflammation Mediators/immunology , Interleukin-6/blood , Interleukin-6/immunology , Interleukin-8/blood , Interleukin-8/immunology , Lymphocyte Count , Lymphopenia/complications , Male , Middle Aged , Pandemics , Phylogeny , Pneumonia, Viral/complications , Pneumonia, Viral/epidemiology , Respiratory Distress Syndrome/complications , SARS-CoV-2 , T-Lymphocytes/cytology , T-Lymphocytes/immunology , Time Factors , Treatment Outcome , Virulence/genetics , Virus Shedding , Young Adult , Zoonoses/transmission , Zoonoses/virology
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