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1.
Euro Surveill ; 25(39)2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-33006300

RESUMO

We found that a single nucleotide polymorphism (SNP) in the nucleoprotein gene of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from a patient interfered with detection in a widely used commercial assay. Some 0.2% of the isolates in the EpiCoV database contain this SNP. Although SARS-CoV-2 was still detected by the other probe in the assay, this underlines the necessity of targeting two independent essential regions of a pathogen for reliable detection.


Assuntos
Betacoronavirus/genética , Técnicas de Laboratório Clínico , Infecções por Coronavirus/diagnóstico , Nucleoproteínas/genética , Pandemias , Pneumonia Viral/diagnóstico , Mutação Puntual , Polimorfismo de Nucleotídeo Único , RNA Viral/genética , Reação em Cadeia da Polimerase em Tempo Real , Proteínas Virais/genética , Sequência de Bases , Betacoronavirus/isolamento & purificação , Técnicas de Laboratório Clínico/métodos , Busca de Comunicante , Infecções por Coronavirus/virologia , Primers do DNA , Erros de Diagnóstico , Reações Falso-Negativas , Feminino , Genes Virais , Humanos , Pessoa de Meia-Idade , Nasofaringe/virologia , Nucleoproteínas/análise , Filogenia , Pneumonia Viral/virologia , Reação em Cadeia da Polimerase em Tempo Real/métodos , Romênia , Doença Relacionada a Viagens , Proteínas Virais/análise
2.
Biomed Res Int ; 2020: 7610678, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33029522

RESUMO

Background: There is a shortage of chemical reagents for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) diagnosis and a surge of SARS-CoV-2 cases, especially in limited-resource settings. Therefore, the combination of an optimal assay kit is necessary. Methods: We compared the ability to screen SARS-CoV-2 among three primer-probe sets in two different master mixes, Invitrogen™ SuperScript™ III One-Step RT-PCR and LightCycler Multiplex RNA Virus Master. Results: The assay with TIB-Molbiol, IDT, and Phu Sa sets for LightCycler Multiplex RNA Virus Master or Invitrogen™ SuperScript™ III One-Step RT-PCR showed positive results from a single reaction of triplicate in the three days of 4.8 copies per reaction. R squared and amplification efficiency were 0.97 and ranged from 107 to 108%, respectively. Conclusions: Our findings indicated that TIB-Molbiol, IDT, and Phu Sa primer-probe sets could be beneficial for the laboratory screening of SARS-CoV-2 by RT-qPCR assay of E gene. There is a need to consider the combination of these reagent sets as a new strategy to increase the testing capacity of screening programs for COVID-19.


Assuntos
Betacoronavirus/genética , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus/diagnóstico , Primers do DNA/genética , Pneumonia Viral/diagnóstico , Sondas RNA/genética , Betacoronavirus/isolamento & purificação , Técnicas de Laboratório Clínico/estatística & dados numéricos , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/virologia , Humanos , Reação em Cadeia da Polimerase Multiplex/métodos , Reação em Cadeia da Polimerase Multiplex/estatística & dados numéricos , Pandemias , Pneumonia Viral/epidemiologia , Pneumonia Viral/virologia , RNA Viral/genética , Reação em Cadeia da Polimerase em Tempo Real/métodos , Reação em Cadeia da Polimerase em Tempo Real/estatística & dados numéricos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/estatística & dados numéricos , Sensibilidade e Especificidade
3.
Am J Case Rep ; 21: e927812, 2020 Oct 03.
Artigo em Inglês | MEDLINE | ID: mdl-33009361

RESUMO

BACKGROUND This is a case report of an immunocompromised patient with a history of non-Hodgkin lymphoma and persistent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection who was seronegative and successfully treated with convalescent plasma. CASE REPORT A 63-year-old woman with a past medical history of non-Hodgkin lymphoma in remission while on maintenance therapy with the anti-CD20 monoclonal antibody, obinutuzumab, tested positive for SARS-CoV-2 via nasopharyngeal reverse transcription polymerase chain reaction (RT-PCR) testing over 12 weeks and persistently tested seronegative for immunoglobulin G (IgG) antibodies using SARS-CoV-2 IgG chemiluminescent microparticle immunoassay technology. During this time, the patient experienced waxing and waning of symptoms, which included fever, myalgia, and non-productive cough, but never acquired severe respiratory distress. She was admitted to our hospital on illness day 88, and her symptoms resolved after the administration of convalescent plasma. CONCLUSIONS As the understanding of the pathogenesis of SARS-CoV-2 continues to evolve, we can currently only speculate about the occurrence of chronic infection vs. reinfection. The protective role of antibodies and their longevity against SARS-CoV-2 remain unclear. Since humoral immunity has an integral role in SARS-CoV-2 infection, various phase 3 vaccine trials are underway. In the context of this pandemic, the present case demonstrates the challenges in our understanding of testing and treating immunocompromised patients.


Assuntos
Anticorpos Monoclonais Humanizados/administração & dosagem , Infecções por Coronavirus/complicações , Infecções por Coronavirus/diagnóstico , Hospedeiro Imunocomprometido , Linfoma não Hodgkin/imunologia , Pneumonia Viral/complicações , Pneumonia Viral/diagnóstico , Antineoplásicos Imunológicos/administração & dosagem , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus/terapia , Feminino , Seguimentos , Humanos , Imunização Passiva/métodos , Linfoma não Hodgkin/complicações , Linfoma não Hodgkin/tratamento farmacológico , Pessoa de Meia-Idade , Pandemias , Reação em Cadeia da Polimerase em Tempo Real/métodos , Testes Sorológicos/métodos , Índice de Gravidade de Doença , Resultado do Tratamento
4.
Rev Peru Med Exp Salud Publica ; 37(2): 203-209, 2020.
Artigo em Espanhol, Inglês | MEDLINE | ID: mdl-32876207

RESUMO

OBJECTIVE: To determine the additional diagnostic performance of a rapid serological test for detection of IgM and IgG antibodies compared to the real-time polymerase chain reaction (RT-PCR) test; for detection of SARS-CoV-2. MATERIALS AND METHODS: A cross-sectional study was carried out including patients hospitalized for COVID-19 in 3 hospitals, health workers exposed to the infection and outpatients who met suspicious case criteria, all of which underwent the molecular test (RT-PCR) and the rapid serological test. The additional diagnostic performance of rapid serological test was evaluated in comparison to molecular tests. Likewise, an approximation was made to the sensitivity and specificity of the rapid serological test. RESULTS: 144 people were included. With the rapid test, 19.4% of positive results were obtained compared to 11.1% in the molecular test (p = 0.03). The rapid serological test detected 21 cases that had been negative by the initial (RT-PCR), providing an additional diagnostic performance of 56.8% compared to the RT-PCR. The additional diagnostic performance was 50.0% during the first week, 70.0% during the second week and 50.0% during the third week of symptom onset. The sensitivity of the rapid serological test was 43.8% and the specificity of 98.9%. CONCLUSIONS: The rapid serological test was able to detect a greater number of cases than those detected by the molecular test especially after the second week of onset of symptoms. It also showed high specificity. It is therefore useful as a complementary test to RT-PCR, especially during the second and third week of illness.


Assuntos
Betacoronavirus/isolamento & purificação , Técnicas de Laboratório Clínico , Infecções por Coronavirus/diagnóstico , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Pneumonia Viral/diagnóstico , Adulto , Infecções por Coronavirus/imunologia , Estudos Transversais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Pandemias , Pneumonia Viral/imunologia , Reação em Cadeia da Polimerase em Tempo Real/métodos , Sensibilidade e Especificidade , Testes Sorológicos/métodos
5.
J Clin Virol ; 131: 104581, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32889496

RESUMO

INTRODUCTION: During the first month of the SARS-CoV-2 outbreak, rapid development of PCR-based diagnostic tests became a global priority so that timely diagnosis, isolation, and contact tracing could minimize the advancing pandemic surge. Designing these tests for broad, long-term detection was complicated by limited information about the novel virus' genome sequence and how it might mutate during global spread and adaptation to humans. METHODS: We assessed eight widely adopted lab developed PCR tests for SARS-CoV-2 against 15,001 SARS-CoV-2 genome sequences. Using a custom bioinformatic pipeline called PCR_strainer, we identified all mismatches and sequence variants in genome locations targeted by 15 sets of primer/probe oligonucleotides from these assays. RESULTS: For 12 out of 15 primer/probe sets, over 98 % of SARS-CoV-2 genomes had no mismatches. Two primer/probe sets contained a single mismatch in the reverse primer that was present in over 99 % of genomes. One primer/probe set targeted a location with extensive polymorphisms with 23 sequence observed variants at the forward primer location. One of these variants, which contains three nucleotide mismatches, arose in February as part of the emergence of a viral clade and was present in 18.8 % of the genomes we analyzed. DISCUSSION: Most early PCR diagnostic tests for SARS-CoV-2 remain inclusive of circulating viral diversity, but three assays with extensive mismatches highlight assay design challenges for novel pathogens and provide valuable lessons for PCR assay design during future outbreaks. Our bioinformatics pipeline is also presented as a useful general-purpose tool for assessing PCR diagnostics assays against circulating strains.


Assuntos
Betacoronavirus/genética , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus/diagnóstico , Oligonucleotídeos/genética , Pneumonia Viral/diagnóstico , Reação em Cadeia da Polimerase em Tempo Real/métodos , Biologia Computacional , Simulação por Computador , Infecções por Coronavirus/virologia , Genoma Viral , Humanos , Pandemias , Pneumonia Viral/virologia , RNA Viral , Sensibilidade e Especificidade
6.
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
8.
J Bras Nefrol ; 42(2 suppl 1): 4-8, 2020 Aug 26.
Artigo em Inglês, Português | MEDLINE | ID: mdl-32877490

RESUMO

The Covid-19 pandemic brought several challenges to the healthcare system: diagnosis, treatment and measures to prevent the spread of the disease. With the greater availability and variety of diagnostic tests, it is essential to properly interpret them. This paper intends to help dialysis units concerning the use of clinical criteria and diagnostic tests for decision making regarding the discontinuation of isolation of patients with suspected or confirmed Covid-19, as well as the return to work activities for employees with suspected or confirmed Covid-19.


Assuntos
Betacoronavirus , Técnicas de Laboratório Clínico/normas , Infecções por Coronavirus/diagnóstico , Nefrologia/normas , Pneumonia Viral/diagnóstico , Diálise Renal , Retorno ao Trabalho , Algoritmos , Brasil , Lista de Checagem , Tomada de Decisão Clínica , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus/epidemiologia , Humanos , Doenças Profissionais/diagnóstico , Pandemias , Isolamento de Pacientes , Pneumonia Viral/epidemiologia , Reação em Cadeia da Polimerase em Tempo Real/métodos , Reação em Cadeia da Polimerase em Tempo Real/normas , Sociedades Médicas/normas , Unidade Hospitalar de Urologia/normas
9.
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
10.
BMC Infect Dis ; 20(1): 657, 2020 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-32894079

RESUMO

BACKGROUND: Tuberculosis is still a significant diagnostic and therapeutic challenge with high proportion of smear- and culture- negative incidences worldwide. The conventional diagnostic tests are time-consuming and have a low sensitivity. Digital PCR is a novel technology which can detect target sequences with relatively low abundance and obtain the absolute copy numbers of the targets. METHODS: We assessed the accuracy of dPCR in TB diagnosis using more than 250 specimens, and for the first time, we selected M.tuberculosis-specific IS1081 in addition to widely used IS6110 as the amplification targets for dPCR. The quantification of target DNA was calculated using QuantaSoft Version 1.7.4.0917 (BioRad), and SPSS version 13.0 software (SPSS Inc., Chicago, IL, USA) was used for statistical analyses. RESULTS: IS6110-dPCR was more sensitive than IS1081, with the sensitivity and specificity accounting for 40.6 and 93.4% respectively. When we classified the TB patients by personal factors for high copy number of M.tuberculosis derived DNA in plasma: bilateral TB, extrapulmonary TB and disseminated TB, the sensitivity of both IS6110- and IS1081- dPCR was the highest in patients with disseminated TB (IS6110, 100%; IS1081, 68.8%), while their sensitivity was a bit higher in patients with extrapulmonary TB (IS6110, 50.0%; IS1081, 39.3%) than that in bilateral TB (IS6110, 43.3%; IS1081, 33.3%). Compared with traditional TB diagnostic tests, joint detection IS6110 & IS1081-dPCR was not as sensitive as smear microscope or mycobacterial culture, but it was higher than IS6110 qPCR (p < 0.05) and was able to detect 47.4% of smear-negative TB patients. CONCLUSION: Our study suggested that plasma IS6110-dPCR is a rapid, moderate accurate and less-invasive method to detect M.tuberculosis DNA in plasma of TB patients and IS6110 & IS1081-dPCR has a potential to aid diagnosis of smear-negative TB.


Assuntos
Elementos de DNA Transponíveis/genética , DNA Bacteriano/genética , Mycobacterium tuberculosis/genética , Reação em Cadeia da Polimerase em Tempo Real/métodos , Tuberculose Pulmonar/diagnóstico , Adolescente , Adulto , Idoso , DNA Bacteriano/sangue , Confiabilidade dos Dados , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Sensibilidade e Especificidade , Adulto Jovem
11.
Acta Biomed ; 91(3): e2020003, 2020 09 07.
Artigo em Inglês | MEDLINE | ID: mdl-32921701

RESUMO

BACKGROUND: The COVID-19 outbreak is now a pandemic disease reaching as much as 210 countries worldwide with more than 2.5 million infected people and nearly 200.000 deaths. Amplification of viral RNA by RT-PCR represents the gold standard for confirmation of infection, yet it showed false-negative rates as large as 15-20% which may jeopardize the effect of the restrictive measures taken by governments. We previously showed that several hematological parameters were significantly different between COVID-19 positive and negative patients. Among them aspartate aminotransferase and lactate dehydrogenase had predictive values as large as 90%. Thus a combination of RT-PCR and blood tests could reduce the false-negative rate of the genetic test. METHODS: We retrospectively analyzed 24 patients showing multiple and inconsistent RT-PCR, test during their first hospitalization period, and compared the genetic tests results with their AST and LDH levels. RESULTS: We showed that when considering the hematological parameters, the RT-PCR false-negative rates were reduced by almost 4-fold. CONCLUSIONS: The study represents a preliminary work aiming at the development of strategies that, by combining RT-PCR tests with routine blood tests, will lower or even abolish the rate of RT-PCR false-negative results and thus will identify, with high accuracy, patients infected by COVID-19.


Assuntos
Betacoronavirus/genética , Infecções por Coronavirus/diagnóstico , Pandemias , Pneumonia Viral/diagnóstico , RNA Viral/análise , Reação em Cadeia da Polimerase em Tempo Real/métodos , Adulto , Idoso , Idoso de 80 Anos ou mais , Aspartato Aminotransferases/sangue , Biomarcadores/sangue , Infecções por Coronavirus/sangue , Infecções por Coronavirus/epidemiologia , Diagnóstico Diferencial , Reações Falso-Negativas , Feminino , Seguimentos , Testes Hematológicos/métodos , Humanos , Itália/epidemiologia , L-Lactato Desidrogenase/sangue , Masculino , Pessoa de Meia-Idade , Pneumonia Viral/sangue , Pneumonia Viral/epidemiologia , Reprodutibilidade dos Testes , Estudos Retrospectivos
12.
Acta Biomed ; 91(3): e2020025, 2020 09 07.
Artigo em Inglês | MEDLINE | ID: mdl-32921721

RESUMO

BACKGROUND AND AIM OF THE WORK: The ongoing pandemic has elicited an increasing interest regarding the SARS-CoV-2 viral RNA detection in saliva specimens rather than through nasopharyngeal swabs. Our aim was to conduct a meta-analysis on the sensitivity and specificity of SARS-CoV-2 viral RNA detection through RT-qPCR based on salivary specimens compared to conventional nasopharyngeal swabs. METHODS: We reported our meta-analysis according to the PRISMA statement. We searched Pubmed, Embase, and pre-print archive medRxiv.og for eligible studies published up to June 1st, 2020. Raw data included true/false positive and negative tests, and the total number of tests. Sensitivity and specificity data were calculated for every study, and then pooled in a random-effects model. Heterogeneity was assessed using the I2 measure. Reporting bias was assessed by means of funnel plots and regression analysis. RESULTS: The systematic review eventually retrieved 14 studies including a total of 15 estimates, the were included in quantitative synthesis. We found a pooled specificity of 97.7% (95%CI 93.8-99.2) and a pooled sensitivity of 83.4% (95%CI 73.1-90.4), with an overall agreement assessed by means of Cohen's kappa equals to 0.750, 95%CI 0.62-0.88 (i.e. moderate agreement), with high heterogeneity and risk of reporting bias. CONCLUSIONS: In conclusion, diagnostic tests based on salivary specimens are somewhat reliable, but relatively few studies have been carried out. Moreover, such studies are characterized by low numbers and low sample power. Therefore, the of salivary samples is currently questionable for clinical purposes and cannot substitute other more conventional RT-qPCR based on nasopharyngeal swabs.


Assuntos
Betacoronavirus/genética , Infecções por Coronavirus/diagnóstico , Nasofaringe/virologia , Pandemias , Pneumonia Viral/diagnóstico , RNA Viral/análise , Reação em Cadeia da Polimerase em Tempo Real/métodos , Saliva/virologia , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/virologia , Humanos , Pneumonia Viral/epidemiologia , Pneumonia Viral/virologia
13.
Perit Dial Int ; 40(5): 499-503, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32869702

RESUMO

The pandemic of respiratory disease (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is life-threatening in peritoneal dialysis (PD) patients. In PD patients with systemic viral infections, peritoneal effluent may be theoretically contaminated. We searched for the presence of SARS-CoV-2 genetic material by real-time reverse transcriptase-polymerase chain reaction assays in serial PD effluents of three PD infected patients. Nasopharyngeal swabs obtained at admission showed high viral load in all three patients, whereas none of the PD effluent specimen tested positive, even after dialysate concentration. Those results support at most a very low SARS-CoV-2 dissemination risk by the peritoneal effluent of PD patients. Imposing special disposal procedures, such as the instillation of hypochlorite in the drainage bags to prevent viral spread to health-care workers, are probably not required.


Assuntos
Líquido Ascítico/virologia , Infecções por Coronavirus/epidemiologia , Falência Renal Crônica/terapia , Diálise Peritoneal/métodos , Pneumonia Viral/epidemiologia , Síndrome Respiratória Aguda Grave/epidemiologia , Adulto , Técnicas de Laboratório Clínico , Infecções por Coronavirus/diagnóstico , Feminino , Humanos , Falência Renal Crônica/diagnóstico , Falência Renal Crônica/epidemiologia , Masculino , Pessoa de Meia-Idade , Pandemias , Pneumonia Viral/diagnóstico , Reação em Cadeia da Polimerase em Tempo Real/métodos , Amostragem , Sensibilidade e Especificidade , Síndrome Respiratória Aguda Grave/diagnóstico , Carga Viral
14.
BMC Infect Dis ; 20(1): 670, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32933492

RESUMO

BACKGROUND: The 2014/15 Ebola outbreak in West Africa resulted in 11,000 deaths and massive strain on local health systems, and the ongoing outbreak in Democratic Republic of Congo has afflicted more than 3000 people. Accurate, rapid Ebola diagnostics suitable for field deployment would enable prompt identification and effective response to future outbreaks, yet remain largely unavailable. The purpose of this study was to assess the accuracy of three novel rapid diagnostic tests (RDTs): an Ebola, an Ebola-Malaria, and a Fever Panel test that includes Ebola, all from a single manufacturer. METHODS: We evaluated the three RDTs in 109 Ebola-positive and 96 Ebola-negative stored serum samples collected during the outbreak in Guinea in 2014/15, and tested by real-time polymerase chain reaction (RT-PCR). Sensitivity, specificity, and overall percent agreement were calculated for each RDT using RT-PCR as a reference standard, stratified by Ct value ranges. RESULTS: All tests performed with high accuracy on samples with low Ct value (high viral load). The Fever Panel test performed with the highest accuracy, with a sensitivity of 89.9% and specificity of 90.6%. The Ebola and Ebola-Malaria tests performed comparably to each other: sensitivity was 77.1 and 78% respectively, and specificity was 91.7% for the Ebola test and 95.8% for the Ebola-Malaria test. CONCLUSIONS: This study evaluated the accuracy of three novel rapid diagnostic tests for Ebola. The tests may have significant public health relevance, particularly the Fever Panel test, which detects seven pathogens including Ebola. Given limitations to the study resulting from uncertain sample quality, further evaluation is warranted. All tests performed with highest accuracy on samples with low Ct value (high viral load), and the data presented here suggests that these RDTs may be useful for point-of-care diagnosis of cases in the context of an outbreak. Restrictions to their use in non-severe Ebola cases or for longitudinal monitoring, when viral loads are lower, may be appropriate. Highlighting the challenge in developing and evaluating Ebola RDTs, there were concerns regarding sample integrity and reference testing, and there is a need for additional research to validate these assays.


Assuntos
Doença pelo Vírus Ebola/diagnóstico , Sistemas Automatizados de Assistência Junto ao Leito , Surtos de Doenças , Ebolavirus/genética , Ebolavirus/isolamento & purificação , Guiné/epidemiologia , Doença pelo Vírus Ebola/epidemiologia , Doença pelo Vírus Ebola/virologia , Humanos , RNA Viral/análise , RNA Viral/metabolismo , Kit de Reagentes para Diagnóstico , Reação em Cadeia da Polimerase em Tempo Real/métodos , Sensibilidade e Especificidade
15.
Am J Case Rep ; 21: e927304, 2020 Sep 26.
Artigo em Inglês | MEDLINE | ID: mdl-32978364

RESUMO

BACKGROUND This case series describes 5 patients with SARS-CoV-2 infection and COVID-19 in Ecuador who had been treated with hydroxychloroquine for systemic lupus erythematosus (SLE) prior to their COVID-19 illness. CASE REPORT Case #1 reports a 29-year-old woman who had been treated with 200 mg of hydroxychloroquine per day for 1 year and presented with flu-like symptoms, chest pain, fever, odynophagia, asthenia, dry cough, and chills. Case #2 was a 34-year-old woman whose treatment for SLE included 200 mg of hydroxychloroquine per day since 2017. She arrived at the clinic with a dry cough, asthenia, and myalgias. Case #3 was a 24-year-old woman who had been using 200 mg of hydroxychloroquine per day since 2010. She presented with asthenia, myalgias, headaches, hypogeusia, and anosmia. Case #4 was a 39-year-old woman taking 200 mg of hydroxychloroquine every day for SLE who presented with dyspnea, chest pain, odynophagia, hypogeusia, anosmia, diarrhea, and fever. Case #5 was a 46-year-old woman who had been taking 200 mg of hydroxychloroquine since 2019. She came to our hospital complaining of chest pain, fever, and dyspnea. In all 5 patients, SARS-CoV-2 infection was confirmed with a nasopharyngeal SARS-CoV-2 reverse transcription-polymerase chain reaction (RT-PCR) test using the Cepheid/GeneXpert system. CONCLUSIONS All 5 of our patients with SLE who were taking hydroxychloroquine presented with SARS-CoV-2 infection and symptoms of COVID-19. This case series provides support for a lack of prevention of COVID-19 by hydroxychloroquine.


Assuntos
Infecções por Coronavirus/prevenção & controle , Hidroxicloroquina/administração & dosagem , Lúpus Eritematoso Sistêmico/tratamento farmacológico , Pandemias/prevenção & controle , Pneumonia Viral/prevenção & controle , Adulto , Técnicas de Laboratório Clínico , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/epidemiologia , Dispneia/diagnóstico , Dispneia/etiologia , Equador , Serviço Hospitalar de Emergência , Feminino , Febre/diagnóstico , Febre/etiologia , Humanos , Pessoa de Meia-Idade , Pandemias/estatística & dados numéricos , Pneumonia Viral/diagnóstico , Pneumonia Viral/epidemiologia , RNA Viral/análise , Reação em Cadeia da Polimerase em Tempo Real/métodos , Medição de Risco , Amostragem , Falha de Tratamento , Adulto Jovem
16.
PLoS One ; 15(9): e0236311, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32898153

RESUMO

Since SARS-CoV-2-based disease (COVID-19) spreads as a pandemic, the necessity of a highly sensitive molecular diagnosis that can drastically reduce false negatives reverse transcription PCR (rtPCR) results, raises as a major clinical need. Here we evaluated the performance of a ddPCR-based assay to quantify SARS-CoV-2 titer in 55 suspected COVID-19 cases with negative rtPCR results thanks to in-house ddPCR assay (targeting RdRp and host RNaseP). Samples were collected at ASST-GOM Niguarda between February and May 2020 at hospital admission. Clinical and imaging data were obtained for clinical staging and definition of disease severity. Patients were mainly female (45.5%) with a median age of 73 (57-84) years. ddPCR-based assay detected SARS-CoV-2 genome in nasopharyngeal samples of 19 (34.5%) patients (median viral-load: 128 copies/mL, IQR: 72-345). In 15 of them (78.9%), chest CT showed a classical COVID-19 bilateral interstitial pneumonia; 14 patients (73.7%) showed severe COVID-19 manifestations. ddPCR did not identify any trace of SARS-CoV-2 genome in the respiratory samples of the remaining 36 patients. The serological assay performed in a subgroup of 34 patients at the later stage of illness (from 3 days to 90 days after) confirmed the presence of SARS-CoV-2 antibodies in all patients tested positive for SARS-CoV-2 in ddPCR (100%). Contrariwise, negative tests were observed in 95.0% ddPCR negative patients (P<0.001). Thanks to a ddPCR-based assay, we achieved a rapid and accurate SARS-CoV-2 diagnosis in rtPCR-negative respiratory samples of individuals with COVID-19 suspect, allowing the rapid taking care and correct management of these patients.


Assuntos
Betacoronavirus/genética , Infecções por Coronavirus/diagnóstico , Nasofaringe/virologia , Pneumonia Viral/diagnóstico , RNA Viral/análise , Reação em Cadeia da Polimerase em Tempo Real/métodos , Idoso , Idoso de 80 Anos ou mais , Betacoronavirus/isolamento & purificação , Infecções por Coronavirus/patologia , Infecções por Coronavirus/virologia , Feminino , Humanos , Limite de Detecção , Masculino , Pessoa de Meia-Idade , Pandemias , Pneumonia Viral/patologia , Pneumonia Viral/virologia , RNA Viral/metabolismo , Índice de Gravidade de Doença , Carga Viral
17.
Viruses ; 12(9)2020 08 27.
Artigo em Inglês | MEDLINE | ID: mdl-32867108

RESUMO

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) compromises the ability of military forces to fulfill missions. At the beginning of May 2020, 22 out of 70 Belgian soldiers deployed to a military education and training center in Maradi, Niger, developed mild COVID-19 compatible symptoms. Immediately upon their return to Belgium, and two weeks later, all seventy soldiers were tested for SARS-CoV-2 RNA (RT-qPCR) and antibodies (two immunoassays). Nine soldiers had at least one positive COVID-19 diagnostic test result. Five of them exhibited COVID-19 symptoms (mainly anosmia, ageusia, and fever), while four were asymptomatic. In four soldiers, SARS-CoV-2 viral load was detected and the genomes were sequenced. Conventional and genomic epidemiological data suggest that these genomes have an African most recent common ancestor and that the Belgian military service men were infected through contact with locals. The medical military command implemented testing of all Belgian soldiers for SARS-CoV-2 viral load and antibodies, two to three days before their departure on a mission abroad or on the high seas, and for specific missions immediately upon their return in Belgium. Some military operational settings (e.g., training camps in austere environments and ships) were also equipped with mobile infectious disease (COVID-19) testing capacity.


Assuntos
Betacoronavirus/isolamento & purificação , Infecções por Coronavirus/epidemiologia , Militares/estatística & dados numéricos , Pneumonia Viral/epidemiologia , Adulto , Bélgica/epidemiologia , Técnicas de Laboratório Clínico , Infecções por Coronavirus/diagnóstico , Surtos de Doenças , Humanos , Masculino , Epidemiologia Molecular , Níger/epidemiologia , Pandemias , Pneumonia Viral/diagnóstico , Reação em Cadeia da Polimerase em Tempo Real/métodos , Testes Sorológicos , Carga Viral , Adulto Jovem
18.
Am J Case Rep ; 21: e925931, 2020 Sep 27.
Artigo em Inglês | MEDLINE | ID: mdl-32980852

RESUMO

BACKGROUND The worldwide spread of the severe acute respiratory syndrome-coronavirus-2 (SARS-COV-2) has created unprecedented situations for healthcare professionals and healthcare systems. Although infection with this virus is considered the main health problem currently, other diseases are still prevalent. CASE REPORT This report describes a 59-year-old man who presented with symptoms of dyspnea and fever that were attributed to Covid-19 infection. His clinical condition deteriorated and further examinations revealed a subjacent severe aortic regurgitation due to acute infective endocarditis. Surgical treatment was successful. CONCLUSIONS The results of diagnostic tests for Covid-19 should be re-evaluated whenever there are clinical mismatches or doubts, as false-positive Covid-19 test results can occur. Clinical interpretation should not be determined exclusively by the Covid-19 pandemic. This case report highlights the importance of using validated and approved serological and molecular testing to detect infection with SARS-CoV-2, and to repeat tests when there is doubt about presenting symptoms.


Assuntos
Insuficiência da Valva Aórtica/cirurgia , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus/diagnóstico , Diagnóstico Tardio , Endocardite/complicações , Endocardite/diagnóstico , Pneumonia Viral/diagnóstico , Anticorpos Antivirais/análise , Insuficiência da Valva Aórtica/complicações , Insuficiência da Valva Aórtica/diagnóstico , Procedimentos Cirúrgicos Cardíacos/métodos , Infecções por Coronavirus/complicações , Estado Terminal , Progressão da Doença , Dispneia/diagnóstico , Dispneia/etiologia , Endocardite/virologia , Reações Falso-Positivas , Febre/diagnóstico , Febre/etiologia , Humanos , Masculino , Pessoa de Meia-Idade , Pandemias , Pneumonia Viral/complicações , Prognóstico , Reação em Cadeia da Polimerase em Tempo Real/métodos , Medição de Risco , Resultado do Tratamento
19.
Am J Case Rep ; 21: e927662, 2020 Sep 29.
Artigo em Inglês | MEDLINE | ID: mdl-32991573

RESUMO

BACKGROUND There is no evidence-based treatment for coronavirus disease 2019 (COVID-19). We report the case of a 63-year-old woman with SARS-CoV-2 infection who developed severe COVID-19 pneumonia and was treated with convalescent plasma. CASE REPORT A 63-year-old woman who presented with severe and prolonged course of COVID-19 disease (fever up to 39.4°C, persistent cough, and dyspnea) received a convalescent plasma transfusion, which led to complete recovery. The diagnosis was confirmed by RT-PCR testing using the CFX96 Real-Time System (Bio-Rad, USA) from nasopharyngeal swabs. In laboratory tests, an increase in acute-phase parameters was observed. Chest computed tomography (CT) showed abnormalities typical for COVID-19. On days 9 and 11 of the disease, she received the convalescent plasma prepared from a single plasmapheresis donation from a male donor. This male donor was qualified as a convalescent plasma donor according to Polish guidelines, which are compliant with European guidelines. He donated plasma at the Regional Centre for Transfusion Medicine in Bialystok, Poland. The therapy with convalescent plasma led to clinical improvement and normalization of inflammatory parameters. CONCLUSIONS This report presents a case of severe COVID-19 pneumonia in a 63-year-old woman who was given supportive treatment with convalescent plasma. Ongoing clinical trials will determine whether convalescent plasma therapy is an effective treatment for SARS-CoV-2 infection.


Assuntos
Transfusão de Componentes Sanguíneos/métodos , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/terapia , Pneumonia Viral/terapia , Síndrome Respiratória Aguda Grave/terapia , Feminino , Seguimentos , Humanos , Pessoa de Meia-Idade , Pandemias , Pneumonia Viral/diagnóstico , Pneumonia Viral/diagnóstico por imagem , RNA Viral/análise , Reação em Cadeia da Polimerase em Tempo Real/métodos , Medição de Risco , Síndrome Respiratória Aguda Grave/diagnóstico , Resultado do Tratamento
20.
Am J Case Rep ; 21: e926785, 2020 Sep 24.
Artigo em Inglês | MEDLINE | ID: mdl-32970653

RESUMO

BACKGROUND In corona virus disease 2019 (COVID-19), which emerged in December 2019 and is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), most case presentations have been related to the respiratory tract. Several recent studies reveal that angiotensin-converting enzyme 2 (ACE2), which was found in the target cells of the virus, is highly expressed in the lungs, small bowel, and vasculature. CASE REPORT A 29-year-old male construction worker from India presented with left-sided colicky abdominal pain. He tested positive for infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) by reverse transcription-polymerase chain reaction (RT-PCR). Isolated superior mesenteric vein thrombosis was diagnosed by CT (computed tomography) scan. He was managed by anti-coagulants and clinically improved. CONCLUSIONS This case report indicates that isolated venous thrombosis of the abdominal vessels without concurrent arterial thrombosis can be a complication of the hyper-coagulability state in COVID-19 patients. Hence, early evaluation of abdominal vessels in covid-19 patients who present with any abdominal symptoms should be considered, especially when found to have an elevated D-dimer level, as early treatment of thrombosis with low-molecular-weight heparin can have a significant impact on the therapeutic outcome.


Assuntos
Anticoagulantes/administração & dosagem , Infecções por Coronavirus/complicações , Oclusão Vascular Mesentérica/diagnóstico por imagem , Pneumonia Viral/complicações , Trombose Venosa/diagnóstico por imagem , Trombose Venosa/tratamento farmacológico , Dor Abdominal/diagnóstico , Dor Abdominal/etiologia , Adulto , Técnicas de Laboratório Clínico/métodos , Indústria da Construção , Infecções por Coronavirus/diagnóstico , Humanos , Índia , Masculino , Oclusão Vascular Mesentérica/tratamento farmacológico , Oclusão Vascular Mesentérica/virologia , Veias Mesentéricas , Pandemias , Pneumonia Viral/diagnóstico , Radiografia Torácica/métodos , Reação em Cadeia da Polimerase em Tempo Real/métodos , Medição de Risco , Índice de Gravidade de Doença , Tomografia Computadorizada por Raios X/métodos , Resultado do Tratamento , Trombose Venosa/complicações
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