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1.
Eur Radiol Exp ; 4(1): 55, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-33000373

RESUMO

We investigated whether the internal gantry components of our computed tomography (CT) scanner contain severe acute respiratory syndrome 2 (SARS-CoV-2) ribonucleic acid (RNA), bacterial or fungal agents. From 1 to 27 March 2020, we performed 180 examinations of patients with confirmed SARS-CoV-2 infection using a dedicated CT scanner. On 27 March 2020, this CT gantry was opened and sampled in each of the following components: (a) gantry case; (b) inward airflow filter; (c) gantry motor; (d) x-ray tube; (e) outflow fan; (f) fan grid; (g) detectors; and (h) x-ray tube filter. To detect SARS-CoV-2 RNA, samples were analysed using reverse transcriptase-polymerase chain reaction (RT-PCR). To detect bacterial or fungal agents, samples have been collected using "replicate organism detection and counting" contact plates of 24 cm2, containing tryptic soy agar, and subsequently cultured. RT-PCR detected SARS-CoV-2 RNA in the inward airflow filter sample. RT-PCR of remaining gantry samples did not reveal the presence of SARS-CoV-2 RNA. Neither bacterial nor fungal agents grew in the agar-based growth medium after the incubation period. Our data showed that SARS-Cov-2 RNA can be found inside the CT gantry only in the inward airflow filter. All remaining CT gantry components were devoid of SARS-CoV-2 RNA.


Assuntos
Betacoronavirus/isolamento & purificação , Infecções por Coronavirus/virologia , Contaminação de Equipamentos , Pneumonia Viral/virologia , Tomógrafos Computadorizados/virologia , Tomografia Computadorizada por Raios X/instrumentação , Humanos , Pandemias , RNA Viral/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real
2.
Nat Commun ; 11(1): 4906, 2020 09 30.
Artigo em Inglês | MEDLINE | ID: mdl-32999292

RESUMO

The CRISPR-Cas12a RNA-guided complexes have tremendous potential for nucleic acid detection but are limited to the picomolar detection limit without an amplification step. Here, we develop a platform with engineered crRNAs and optimized conditions that enabled us to detect various clinically relevant nucleic acid targets with higher sensitivity, achieving a limit of detection in the femtomolar range without any target pre-amplification step. By extending the 3'- or 5'-ends of the crRNA with different lengths of ssDNA, ssRNA, and phosphorothioate ssDNA, we discover a self-catalytic behavior and an augmented rate of LbCas12a-mediated collateral cleavage activity as high as 3.5-fold compared to the wild-type crRNA and with significant improvement in specificity for target recognition. Particularly, the 7-mer DNA extension to crRNA is determined to be universal and spacer-independent for enhancing the sensitivity and specificity of LbCas12a-mediated nucleic acid detection. We perform a detailed characterization of our engineered ENHANCE system with various crRNA modifications, target types, reporters, and divalent cations. With isothermal amplification of SARS-CoV-2 RNA using RT-LAMP, the modified crRNAs are incorporated in a paper-based lateral flow assay that can detect the target with up to 23-fold higher sensitivity within 40-60 min.


Assuntos
Proteínas de Bactérias/metabolismo , Betacoronavirus/genética , Proteínas Associadas a CRISPR/metabolismo , Endodesoxirribonucleases/metabolismo , Técnicas de Amplificação de Ácido Nucleico/métodos , RNA Viral/isolamento & purificação , Transativadores/metabolismo , Betacoronavirus/isolamento & purificação , Sistemas CRISPR-Cas , Técnicas de Laboratório Clínico , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/virologia , DNA de Cadeia Simples , Pandemias , Pneumonia Viral , RNA Guia/genética , RNA Viral/genética
3.
Colomb Med (Cali) ; 51(2): e4272, 2020 Jun 30.
Artigo em Inglês | MEDLINE | ID: mdl-33012887

RESUMO

In the past four months SARS-CoV-2 has reached most countries in the world. Public health strategies based on widespread testing and proper isolation of positive cases have shown to be helpful to reduce local transmission of SARS-CoV-2. Confirmatory tests, that identify viral RNA, and screening serological tests that identify viral antigens or host antibodies against viral proteins are part of the tools that nations can use to fight infectious disease epidemics. Understanding how each test works can provide insights about their test characteristics and how they can be used for different clinical and public health goals. Testing is a key strategy to reduce viral transmission, not only for this epidemic, but also for others to come.


Assuntos
Técnicas de Laboratório Clínico , Infecções por Coronavirus/diagnóstico , Pneumonia Viral/diagnóstico , Antígenos Virais/sangue , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/prevenção & controle , Humanos , América Latina/epidemiologia , Pandemias/prevenção & controle , Pneumonia Viral/epidemiologia , Pneumonia Viral/prevenção & controle , Saúde Pública , RNA Viral/isolamento & purificação
4.
Zhonghua Liu Xing Bing Xue Za Zhi ; 41(8): 1220-1224, 2020 Aug 10.
Artigo em Chinês | MEDLINE | ID: mdl-32867427

RESUMO

Objective: To understand the epidemiological characteristics of COVID-19 monitoring cases in Yinzhou district based on health big data platform to provide evidence for the construction of COVID-19 monitoring system. Methods: Data on Yinzhou COVID-19 daily surveillance were collected. Information on patients' population classification, epidemiological history, COVID-19 nucleic acid detection rate, positive detection rate and confirmed cases monitoring detection rate were analyzed. Results: Among the 1 595 COVID-19 monitoring cases, 79.94% were community population and 20.06% were key population. The verification rate of monitoring cases was 100.00%. The total percentage of epidemiological history related to Wuhan city or Hubei province was 6.27% in total, and was 2.12% in community population and 22.81% in key population (P<0.001). The total COVID-19 nucleic acid detection rate was 18.24% (291/1 595), and 53.00% in those with epidemiological history and 15.92% in those without (P<0.001).The total positive detection rate was 1.72% (5/291) and the confirmed cases monitoring detection rate was 0.31% (5/1 595). The time interval from the first visit to the first nucleic acid detection of the confirmed monitoring cases and other confirmed cases was statistically insignificant (P>0.05). Conclusions: The monitoring system of COVID-19 based on the health big data platform was working well but the confirmed cases monitoring detection rate need to be improved.


Assuntos
Betacoronavirus , Infecções por Coronavirus/epidemiologia , Pneumonia Viral/epidemiologia , Betacoronavirus/genética , Betacoronavirus/isolamento & purificação , Big Data , China/epidemiologia , Cidades , Surtos de Doenças , Humanos , Pandemias , Vigilância da População , RNA Viral/genética , RNA Viral/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real
5.
Ann Ital Chir ; 91: 235-238, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32877383

RESUMO

The present pandemic caused by the SARS COV-2 coronavirus is still ongoing, although it is registered a slowdown in the spread for new cases. The main environmental route of transmission of SARS-CoV-2 is through droplets and fomites or surfaces, but there is a potential risk of virus spread also in smaller aerosols during various medical procedures causing airborne transmission. To date, no information is available on the risk of contagion from the peritoneal fluid with which surgeons can come into contact during the abdominal surgery on COVID-19 patients. We have investigated the presence of SARS-CoV-2 RNA in the peritoneal cavity of patients affected by COVID-19, intraoperatively and postoperatively. KEY WORDS: Covid-19, Laparotomy, Surgery.


Assuntos
Líquido Ascítico/virologia , Betacoronavirus/isolamento & purificação , Infecções por Coronavirus/transmissão , Transmissão de Doença Infecciosa do Paciente para o Profissional/prevenção & controle , Perfuração Intestinal/cirurgia , Laparotomia , Pandemias , Pneumonia Viral/transmissão , Doenças do Colo Sigmoide/cirurgia , Viremia/transmissão , Aerossóis , Idoso de 80 Anos ou mais , Infecções por Coronavirus/sangue , Infecções por Coronavirus/complicações , Infecções por Coronavirus/prevenção & controle , Estudos Transversais , Divertículo/complicações , Evolução Fatal , Feminino , Humanos , Perfuração Intestinal/sangue , Perfuração Intestinal/complicações , Perfuração Intestinal/virologia , Período Intraoperatório , Nasofaringe/virologia , Pandemias/prevenção & controle , Pneumonia Viral/sangue , Pneumonia Viral/complicações , Pneumonia Viral/prevenção & controle , Período Pós-Operatório , Estudos Prospectivos , RNA Viral/isolamento & purificação , Risco , Soro/virologia , Doenças do Colo Sigmoide/sangue , Doenças do Colo Sigmoide/complicações , Doenças do Colo Sigmoide/virologia , Viremia/virologia
6.
Biomed Res Int ; 2020: 2721381, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32884938

RESUMO

Introduction: Emergency department (ED) triage regarding infection with severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is challenging. During the coronavirus disease 2019 (COVID-19) outbreak in Germany, the diagnostic outcomes of critically ill patients admitted to the resuscitation room in the ED of our academic 754-bed hospital should be analyzed. Methods: All resuscitation room patients between March 1st and April 15th 2020 were included in this retrospective study. Every patient with suspicion of SARS-CoV-2 infection received a pharyngeal swab for real-time polymerase chain reaction (rt-PCR), divided in the clinical subgroups of "highly suspicious for COVID-19" and "COVID-19 as differential diagnosis." All respiratory and infectious symptoms were included as at least "differential diagnosis" as an expanded suspicion strategy. Results: Ninety-five patients were included (trauma n = 14, critically ill n = 81). Of 3 highly suspicious patients, 2 had rt-PCR positive pharyngeal swabs. In 39 patients, COVID-19 was defined as differential diagnosis, and 3 were positive for SARS-CoV-2. Of them, pharyngeal swabs were positive in 1 case, while in 2 cases, only tracheal fluid was rt-PCR positive while the pharyngeal swabs were negative. In one of these 2 cases, chest computed tomography (CT) was also negative for ground-glass opacities but showed a pulmonary abscess and pulmonary embolism. Conclusion: We recommend an expanded suspicion strategy for COVID-19 due to unexpected diagnostic outcomes. Personal protective equipment should be used in every resuscitation room operation due to unexpected cases and initial knowledge gaps. Furthermore, tracheal fluid should be tested for SARS-CoV-2 in every intubated patient due to cases with negative pharyngeal swabs and negative chest CT.


Assuntos
Betacoronavirus , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus/diagnóstico , Pneumonia Viral/diagnóstico , Idoso , Idoso de 80 Anos ou mais , Infecções por Coronavirus/diagnóstico por imagem , Infecções por Coronavirus/epidemiologia , Estado Terminal , Diagnóstico Diferencial , Surtos de Doenças , Serviço Hospitalar de Emergência , Reações Falso-Negativas , Feminino , Alemanha/epidemiologia , Humanos , Masculino , Pessoa de Meia-Idade , Pandemias , Pneumonia Viral/diagnóstico por imagem , Pneumonia Viral/epidemiologia , RNA Viral/genética , RNA Viral/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real , Ressuscitação , Estudos Retrospectivos , Tomografia Computadorizada por Raios X , Triagem
7.
Pneumologie ; 74(9): 615-620, 2020 Sep.
Artigo em Alemão | MEDLINE | ID: mdl-32916744

RESUMO

PATIENT HISTORY AND CLINICAL FINDINGS: A 46-year old construction worker presented at the emergency department with two orthostatic syncopes. The patient complained of prolonged fever and coughs for 7 days which had not improved after oral treatment with sultamicillin for 5 days, prescribed by the patient's general practitioner. Physical examination showed high blood pressure due to previously known hypertension. Other vital signs without pathological findings. Pulmonary auscultation showed basal soft crackling noises of the left lung. FINDINGS AND DIAGNOSIS: Laboratory examination showed increased values for LDH, pro-BNP and CRP and normal values for leucocytes and procalcitonin. Conventional X-Ray of the chest showed bipulmonal lateral atypical infiltrates. After the first PCR turned in negative another PCR-analysis for SARS-CoV-2 of a deep oral swab-sample was performed since the clinical, laboratory and radiological findings were typical for COVID-19. Again, SARS-CoV-2-RNA was not detected. A CT-scan of the chest showed bipulmonal lateral ground-glass attenuation, again typical for COVID-19 associated pneumonia. After a third attempt for a PCR-analysis of a deep oral swab-sample was negative, analysis of a sputum was performed which finally confirmed the diagnosis of COVID-19 associated pneumonia. THERAPY AND COURSE OF EVENTS: The patient was admitted for evaluation of syncopes and suspect of COVID-19 associated pneumonia. The patient was prophylactically isolated while the result of SARS-CoV-2-PCR from a deep oral swab was pending. Suspecting a possible secondary bacterial infection at the beginning, intravenous antibiotic treatment with ampicillin/sulbactam was initiated. While further examinations showed no indication for bacterial infection, antibiotics were discontinued after 3 days. Due to clinical recovery antiviral therapy was not performed after confirming the diagnosis. The patient was discharged 17 days after onset of first symptoms without any requirements for further isolation. CONCLUSION: This casuistic describes a case of COVID-19 associated pneumonia presenting with typical clinical features, laboratory and radiological findings. Detection of viral RNA was not successful from deep oral swab-samples despite repeated attempts. Finally, PCR-analysis of sputum confirmed the diagnosis. Analysis of deeper airway samples (sputum, bronchoalveolar lavage, tracheal secretions) or stool for SARS-CoV-2 should be performed in cases of evident clinical suspicion of COVID-19 and negative PCR results from deep oral swabs.


Assuntos
Infecções por Coronavirus/diagnóstico , Pneumonia Viral/diagnóstico , Betacoronavirus/isolamento & purificação , Técnicas de Laboratório Clínico , Humanos , Pulmão/diagnóstico por imagem , Pessoa de Meia-Idade , Orofaringe/virologia , Pandemias , Reação em Cadeia da Polimerase , RNA Viral/isolamento & purificação , Radiografia Torácica
8.
Indian J Med Res ; 152(1 & 2): 88-94, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32893844

RESUMO

Background & objectives: Public health and diagnostic laboratories are facing huge sample loads for COVID-19 diagnosis by real-time reverse transcription-polymerase chain reaction (RT-PCR). High sensitivity of optimized real-time RT-PCR assays makes pooled testing a potentially efficient strategy for resource utilization when positivity rates for particular regions or groups of individuals are low. We report here a comparative analysis of pooled testing for 5- and 10-sample pools by real-time RT-PCR across 10 COVID-19 testing laboratories in India. Methods: Ten virus research and diagnostic laboratories (VRDLs) testing for COVID-19 by real-time RT-PCR participated in this evaluation. At each laboratory, 100 nasopharyngeal swab samples including 10 positive samples were used to create 5- and 10-sample pools with one positive sample in each pool. RNA extraction and real-time RT-PCR for SARS-CoV-2-specific E gene target were performed for individual positive samples as well as pooled samples. Concordance between individual sample testing and testing in the 5- or 10-sample pools was calculated, and the variation across sites and by sample cycle threshold (Ct) values was analyzed. Results: A total of 110 each of 5- and 10-sample pools were evaluated. Concordance between the 5-sample pool and individual sample testing was 100 per cent in the Ct value ≤30 cycles and 95.5 per cent for Ctvalues ≤33 cycles. Overall concordance between the 5-sample pooled and individual sample testing was 88 per cent while that between 10-sample pool and individual sample testing was 66 per cent. Although the concordance rates for both the 5- and 10-sample pooled testing varied across laboratories, yet for samples with Ct values ≤33 cycles, the concordance was ≥90 per cent across all laboratories for the 5-sample pools. Interpretation & conclusions: Results from this multi-site assessment suggest that pooling five samples for SARS-CoV-2 detection by real-time RT-PCR may be an acceptable strategy without much loss of sensitivity even for low viral loads, while with 10-sample pools, there may be considerably higher numbers of false negatives. However, testing laboratories should perform validations with the specific RNA extraction and RT-PCR kits in use at their centres before initiating pooled testing.


Assuntos
Betacoronavirus/isolamento & purificação , Técnicas de Laboratório Clínico , Infecções por Coronavirus/diagnóstico , Pneumonia Viral/diagnóstico , RNA Viral/isolamento & purificação , Betacoronavirus/genética , Betacoronavirus/patogenicidade , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/genética , Infecções por Coronavirus/virologia , Testes Diagnósticos de Rotina/métodos , Feminino , Humanos , Índia/epidemiologia , Masculino , Pandemias , Pneumonia Viral/epidemiologia , Pneumonia Viral/genética , Pneumonia Viral/virologia , RNA Viral/genética , Reação em Cadeia da Polimerase em Tempo Real/métodos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Testes Sorológicos , Manejo de Espécimes , Carga Viral/genética
12.
Elife ; 92020 09 07.
Artigo em Inglês | MEDLINE | ID: mdl-32894217

RESUMO

Understanding and mitigating SARS-CoV-2 transmission hinges on antibody and viral RNA data that inform exposure and shedding, but extensive variation in assays, study group demographics and laboratory protocols across published studies confounds inference of true biological patterns. Our meta-analysis leverages 3214 datapoints from 516 individuals in 21 studies to reveal that seroconversion of both IgG and IgM occurs around 12 days post-symptom onset (range 1-40), with extensive individual variation that is not significantly associated with disease severity. IgG and IgM detection probabilities increase from roughly 10% at symptom onset to 98-100% by day 22, after which IgM wanes while IgG remains reliably detectable. RNA detection probability decreases from roughly 90% to zero by day 30, and is highest in feces and lower respiratory tract samples. Our findings provide a coherent evidence base for interpreting clinical diagnostics, and for the mathematical models and serological surveys that underpin public health policies.


Assuntos
Betacoronavirus/genética , Betacoronavirus/imunologia , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Pneumonia Viral/imunologia , Pneumonia Viral/virologia , RNA Viral/análise , Anticorpos Antivirais/sangue , Anticorpos Antivirais/isolamento & purificação , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus/sangue , Infecções por Coronavirus/diagnóstico , Ensaio de Imunoadsorção Enzimática , Humanos , Imunoglobulina G/isolamento & purificação , Imunoglobulina M/isolamento & purificação , Pandemias , Pneumonia Viral/sangue , Pneumonia Viral/diagnóstico , RNA Viral/isolamento & purificação
13.
mSphere ; 5(5)2020 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-32878932

RESUMO

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has infected millions within just a few months, causing severe respiratory disease and mortality. Assays to monitor SARS-CoV-2 growth in vitro depend on time-consuming and costly RNA extraction steps, hampering progress in basic research and drug development efforts. Here, we developed a simplified quantitative real-time PCR assay that bypasses viral RNA extraction steps and can monitor SARS-CoV-2 growth from a small amount of cell culture supernatants. In addition, we show that this approach is easily adaptable to numerous other RNA and DNA viruses. Using this assay, we screened the activities of a number of compounds that were predicted to alter SARS-CoV-2 entry and replication as well as HIV-1-specific drugs in a proof-of-concept study. We found that E64D (inhibitor of endosomal proteases cathepsin B and L) and apilimod (endosomal trafficking inhibitor) potently decreased the amount of SARS-CoV-2 RNA in cell culture supernatants with minimal cytotoxicity. Surprisingly, we found that the macropinocytosis inhibitor ethylisopropylamiloride (EIPA) similarly decreased SARS-CoV-2 RNA levels in supernatants, suggesting that entry may additionally be mediated by an alternative pathway. HIV-1-specific inhibitors nevirapine (a nonnucleoside reverse transcriptase inhibitor [NNRTI]), amprenavir (a protease inhibitor), and allosteric integrase inhibitor 2 (ALLINI-2) modestly inhibited SARS-CoV-2 replication, albeit the 50% inhibitory concentration (IC50) values were much higher than that required for HIV-1. Taking the data together, this simplified assay will expedite basic SARS-CoV-2 research, be amenable to mid-throughput screening assays (i.e., drug, CRISPR, small interfering RNA [siRNA], etc.), and be applicable to a broad number of RNA and DNA viruses.IMPORTANCE Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the etiological agent of the coronavirus disease 2019 (COVID-19) pandemic, is continuing to cause immense respiratory disease and social and economic disruptions. Conventional assays that monitor SARS-CoV-2 growth in cell culture rely on costly and time-consuming RNA extraction procedures, hampering progress in basic SARS-CoV-2 research and development of effective therapeutics. Here, we developed a simple quantitative real-time PCR assay to monitor SARS-CoV-2 growth in cell culture supernatants that does not necessitate RNA extraction and that is as accurate and sensitive as existing methods. In a proof-of-concept screen, we found that E64D, apilimod, EIPA, and remdesivir can substantially impede SARS-Cov-2 replication, providing novel insight into viral entry and replication mechanisms. In addition, we show that this approach is easily adaptable to numerous other RNA and DNA viruses. This simplified assay will undoubtedly expedite basic SARS-CoV-2 and virology research and be amenable to use in drug screening platforms to identify therapeutics against SARS-CoV-2.


Assuntos
Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Betacoronavirus/crescimento & desenvolvimento , Técnicas de Cultura de Células/métodos , Infecções por Coronavirus/virologia , Pneumonia Viral/virologia , Reação em Cadeia da Polimerase em Tempo Real/métodos , Betacoronavirus/genética , Betacoronavirus/patogenicidade , Pandemias , RNA Viral/análise , RNA Viral/isolamento & purificação , Replicação Viral/efeitos dos fármacos
14.
Int J Med Sci ; 17(14): 2125-2132, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32922173

RESUMO

Objectives: To present the temporal changes of CT manifestations in COVID-19 patients from a single fangcang shelter hospital and to facilitate the understanding of the disease course. Materials and Methods: This retrospective study included 98 patients (males: females, 43:55, mean year, 49±12 years) with confirmed COVID-19 at Jianghan fangcang shelter hospital admitted between Feb 05, 2020, and Feb 09, 2020, who had initial chest CTs at our hospital. Radiographic features and CT scores were analyzed. Results: A total of 267 CT scans of 98 patients were evaluated. Our study showed a high median total CT score of 7 within the first week from symptom onset, peaked in the 2nd week at 10, followed by persistently high levels of CT score with 9.5, 7 and 7 for the week 3, 4, and >4, respectively, and a prolonged median disease course (30 days, the median interval between the onset of initial symptoms and discharge). Ground-glass opacity (GGO) (58%, 41/71) was the earliest and most frequent finding in week 1. Consolidation (26%, 14/53) and mixed pattern (40%, 21/53) were predominant patterns in 2nd week. GGO and reticular were the main patterns of later phase CT scans in patients with relatively advanced diseases who had longer illness duration (≥4 weeks). Among the 94 CT abnormalities obtained within 3 days from the twice RT-PCR test turned negative, the mixed pattern was mainly presented in patients with disease duration of 2-3 weeks, for GGO and reticular were common during the whole course. Conclusion: Discharged patients from fangcang shelter hospital demonstrated a high extent of lung abnormalities on CT within the first week from symptom onset, peaked at 2nd week, followed by persistence of high levels and a prolonged median disease course. GGO was the predominant pattern in week 1, consolidation and mixed pattern in 2nd week, whereas GGO and reticular patterns in later stages (≥4 weeks).


Assuntos
Betacoronavirus/isolamento & purificação , Técnicas de Laboratório Clínico/estatística & dados numéricos , Infecções por Coronavirus/diagnóstico , Pulmão/diagnóstico por imagem , Pneumonia Viral/diagnóstico , Tomografia Computadorizada por Raios X/estatística & dados numéricos , Adolescente , Adulto , Idoso , Betacoronavirus/genética , China/epidemiologia , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/virologia , Progressão da Doença , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Unidades Móveis de Saúde , Pandemias , Pneumonia Viral/epidemiologia , Pneumonia Viral/virologia , RNA Viral/isolamento & purificação , Estudos Retrospectivos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Índice de Gravidade de Doença , Adulto Jovem
15.
Int J Med Sci ; 17(15): 2373-2378, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32922203

RESUMO

Background: In patients with coronavirus disease 2019 (COVID-19) pneumonia, whether new pulmonary lesions will continue to develop after treatment was unknown. This study aimed to determine whether new pulmonary lesions will develop after treatment in patients with COVID-19 pneumonia, and investigate their CT features and outcomes. Methods: This retrospective study included 56 consecutive patients with confirmed COVID-19 pneumonia from January 20 to March 5, 2020. Their initial and follow-up CT images and clinical data were reviewed. The CT manifestations of primary and newly developed pulmonary lesions and their changes after treatment were mainly evaluated. Results: Among the 56 patients (mean age: 48±15 years, 35 men) with COVID-19 pneumonia, 42 (75.0%) patients developed new pulmonary lesions during treatment. All new lesions developed before the nucleic acid test turned negative. Patients with new lesions were more likely to have lymphopenia (P=0.041) or increased C-reactive protein (CRP) levels (P<0.001) than those without new lesions. Of the 42 patients, 30 (71.4%) patients developed new lesions once, and 12 (28.6%) twice or thrice, which usually appeared when primary lesions were progressing (37, 88.1%) and 1-15 days after treatment. The newly developed lesions were usually multiple (38, 90.5%), distributed in the previously involved (39, 92.9%) or uninvolved (27, 64.3%) lobes, and manifested as ground-glass opacities (GGOs) with consolidation (23, 54.8%) or pure GGOs (19, 45.2%). After their occurrence, the new lesions in most patients (32, 76.2%) showed direct absorption, whereas those in some patients (10, 23.8%) progressed before absorption. Conclusion: During treatment, most patients with COVID-19 pneumonia will develop new pulmonary lesions, which usually manifest as multiple GGOs distributed around the primary lesions or in previously uninvolved lobes, and are subsequently absorbed directly.


Assuntos
Betacoronavirus/isolamento & purificação , Técnicas de Laboratório Clínico/estatística & dados numéricos , Infecções por Coronavirus/mortalidade , Pulmão/diagnóstico por imagem , Pneumonia Viral/mortalidade , Tomografia Computadorizada por Raios X/estatística & dados numéricos , Adulto , Betacoronavirus/genética , Betacoronavirus/patogenicidade , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/terapia , Infecções por Coronavirus/virologia , Progressão da Doença , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Pandemias , Pneumonia Viral/diagnóstico , Pneumonia Viral/terapia , Pneumonia Viral/virologia , RNA Viral/isolamento & purificação , Estudos Retrospectivos
16.
Proc Natl Acad Sci U S A ; 117(39): 24450-24458, 2020 09 29.
Artigo em Inglês | MEDLINE | ID: mdl-32900935

RESUMO

The current severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has had an enormous impact on society worldwide, threatening the lives and livelihoods of many. The effects will continue to grow and worsen if economies begin to open without the proper precautions, including expanded diagnostic capabilities. To address this need for increased testing, we have developed a sensitive reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay compatible with current reagents, which utilizes a colorimetric readout in as little as 30 min. A rapid inactivation protocol capable of inactivating virions, as well as endogenous nucleases, was optimized to increase sensitivity and sample stability. This protocol, combined with the RT-LAMP assay, has a sensitivity of at least 50 viral RNA copies per microliter in a sample. To further increase the sensitivity, a purification protocol compatible with this inactivation method was developed. The inactivation and purification protocol, combined with the RT-LAMP assay, brings the sensitivity to at least 1 viral RNA copy per microliter in a sample. This simple inactivation and purification pipeline is inexpensive and compatible with other downstream RNA detection platforms and uses readily available reagents. It should increase the availability of SARS-CoV-2 testing as well as expand the settings in which this testing can be performed.


Assuntos
Betacoronavirus/isolamento & purificação , Infecções por Coronavirus/diagnóstico , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Amplificação de Ácido Nucleico/métodos , Pneumonia Viral/diagnóstico , Betacoronavirus/genética , Técnicas de Laboratório Clínico/economia , Colorimetria , Infecções por Coronavirus/economia , Infecções por Coronavirus/virologia , Genoma Viral/genética , Humanos , Concentração de Íons de Hidrogênio , Técnicas de Diagnóstico Molecular/economia , Técnicas de Amplificação de Ácido Nucleico/economia , Pandemias , Pneumonia Viral/virologia , Estabilidade de RNA , RNA Viral/química , RNA Viral/genética , RNA Viral/isolamento & purificação , Sensibilidade e Especificidade , Fatores de Tempo , Proteínas Virais/genética , Inativação de Vírus
17.
Nat Commun ; 11(1): 4464, 2020 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-32900994

RESUMO

The SARS-CoV-2 pandemic has shown how a rapid rise in demand for patient and community sample testing can quickly overwhelm testing capability globally. With most diagnostic infrastructure dependent on specialized instruments, their exclusive reagent supplies quickly become bottlenecks, creating an urgent need for approaches to boost testing capacity. We address this challenge by refocusing the London Biofoundry onto the development of alternative testing pipelines. Here, we present a reagent-agnostic automated SARS-CoV-2 testing platform that can be quickly deployed and scaled. Using an in-house-generated, open-source, MS2-virus-like particle (VLP) SARS-CoV-2 standard, we validate RNA extraction and RT-qPCR workflows as well as two detection assays based on CRISPR-Cas13a and RT-loop-mediated isothermal amplification (RT-LAMP). In collaboration with an NHS diagnostic testing lab, we report the performance of the overall workflow and detection of SARS-CoV-2 in patient samples using RT-qPCR, CRISPR-Cas13a, and RT-LAMP. The validated RNA extraction and RT-qPCR platform has been installed in NHS diagnostic labs, increasing testing capacity by 1000 samples per day.


Assuntos
Betacoronavirus/genética , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/virologia , Pneumonia Viral/diagnóstico , Pneumonia Viral/virologia , Betacoronavirus/isolamento & purificação , Bioensaio , Sistemas CRISPR-Cas , Técnicas de Laboratório Clínico/instrumentação , Técnicas de Laboratório Clínico/normas , Humanos , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Amplificação de Ácido Nucleico/métodos , Pandemias , RNA Viral/análise , RNA Viral/genética , RNA Viral/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real , Sensibilidade e Especificidade
18.
Pediatr Infect Dis J ; 39(10): e315-e317, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32932332

RESUMO

In the current study, we report on 4 children with confirmed SARS-CoV-2 infection, of which 3 of them were asymptomatic. These patients had both pharyngeal swabs and anal swabs testing during hospital or after discharge. All the 4 children showed long-time positive viral RNA in the stool specimens after pharyngeal swabs turned negative during the follow-up stage, especially in the asymptomatic children. The positive RNA in stool specimens of asymptomatic children last for more than 54 days after admission or 30 days after discharge.


Assuntos
Infecções Assintomáticas , Betacoronavirus/isolamento & purificação , Infecções por Coronavirus/virologia , Fezes/virologia , Pneumonia Viral/virologia , Betacoronavirus/genética , Criança , Pré-Escolar , Infecções por Coronavirus/patologia , Feminino , Seguimentos , Humanos , Masculino , Pandemias , Faringe/virologia , Pneumonia Viral/patologia , RNA Viral/genética , RNA Viral/isolamento & purificação
19.
ESMO Open ; 5(5)2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32958531

RESUMO

BACKGROUND: Immunosuppression induced by anticancer therapy in a COVID-19-positive asymptomatic patient with cancer may have a devastating effect and, eventually, be lethal. To identify asymptomatic cases among patients receiving active cancer treatment, the Federico II University Hospital in Naples performs rapid serological tests in addition to hospital standard clinical triage for COVID-19 infection. METHODS: From 6 to 17 April 2020, all candidates for chemotherapy, radiotherapy or target/immunotherapy, if negative at the standard clinical triage on the day scheduled for anticancer treatment, received a rapid serological test on peripheral blood for COVID-19 IgM and IgG detection. In case of COVID-19 IgM and/or IgG positivity, patients underwent a real-time PCR (RT-PCR) SARS-CoV-2 test to confirm infection, and active cancer treatment was delayed. RESULTS: Overall 466 patients, negative for COVID-19 symptoms, underwent serological testing in addition to standard clinical triage. The average age was 61 years (range 25-88 years). Most patients (190, 40.8%) had breast cancer, and chemotherapy with or without immunotherapy was administered in 323 (69.3%) patients. Overall 433 (92.9%) patients were IgG-negative and IgM-negative, and 33 (7.1%) were IgM-positive and/or IgG-positive. Among the latter patients, 18 (3.9%), 11 (2.4%) and 4 (0.9%) were IgM-negative/IgG-positive, IgM-positive/IgG-negative and IgM-positive/IgG-positive, respectively. All 33 patients with a positive serological test, tested negative for RT-PCR SARS-CoV-2 test. No patient in our cohort developed symptoms suggestive of active COVID-19 infection. CONCLUSION: Rapid serological testing at hospital admission failed to detect active asymptomatic COVID-19 infection. Moreover, it entailed additional economic and human resources, delayed therapy administrationand increased hospital accesses.


Assuntos
Infecções Assintomáticas , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus/diagnóstico , Imunossupressão/efeitos adversos , Neoplasias/terapia , Pneumonia Viral/diagnóstico , Triagem/normas , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Antineoplásicos Imunológicos/efeitos adversos , Betacoronavirus/genética , Betacoronavirus/imunologia , Betacoronavirus/isolamento & purificação , Quimiorradioterapia/efeitos adversos , Quimiorradioterapia/métodos , Técnicas de Laboratório Clínico/economia , Técnicas de Laboratório Clínico/estatística & dados numéricos , Infecções por Coronavirus/sangue , Infecções por Coronavirus/economia , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Testes Diagnósticos de Rotina/economia , Testes Diagnósticos de Rotina/estatística & dados numéricos , Estudos de Viabilidade , Feminino , Humanos , Imunossupressão/métodos , Masculino , Pessoa de Meia-Idade , Neoplasias/imunologia , Pandemias , Admissão do Paciente/economia , Admissão do Paciente/estatística & dados numéricos , Pneumonia Viral/sangue , Pneumonia Viral/imunologia , Pneumonia Viral/virologia , Guias de Prática Clínica como Assunto , RNA Viral/isolamento & purificação , Reação em Cadeia da Polimerase Via Transcriptase Reversa/economia , Reação em Cadeia da Polimerase Via Transcriptase Reversa/estatística & dados numéricos , Sensibilidade e Especificidade
20.
Rev Soc Bras Med Trop ; 53: e20200619, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32965458

RESUMO

With the large number of individuals infected and recovered from Covid-19, there is intense discussion about the quality and duration of the immunity elicited by SARS-CoV-2 infection, including the possibility of disease recurrence. Here we report a case with strong clinical, epidemiological and laboratorial evidence of, not only reinfection by SARS-CoV-2, but also clinical recurrence of Covid-19.


Assuntos
Infecções por Coronavirus/diagnóstico , Pneumonia Viral/diagnóstico , Recidiva , Anticorpos Antivirais/sangue , Betacoronavirus , Brasil , Feminino , Humanos , Pandemias , RNA Viral/isolamento & purificação , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Adulto Jovem
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