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Sci Rep ; 12(1): 586, 2022 01 12.
Artigo em Inglês | MEDLINE | ID: mdl-35022478


The performance of a laboratory-developed IgG/IgA flow cytometry-based immunoassay (FCI) using Jurkat T cells stably expressing full-length native S protein was compared against Elecsys electrochemiluminiscent (ECLIA) Anti-SARS-CoV-2 S (Roche Diagnostics, Pleasanton, CA, USA), and Liaison SARS-CoV-2 TrimericS IgG chemiluminiscent assay (CLIA) (Diasorin S.p.a, Saluggia, IT) for detection of SARS-CoV-2-specific antibodies. A total of 225 serum/plasma specimens from 120 acute or convalescent COVID-19 individuals were included. Overall, IgG/IgA-FCI yielded the highest number of positives (n = 179), followed by IgA-FCI (n = 177), Roche ECLIA (n = 175), IgG-FCI (n = 172) and Diasorin CLIA (n = 154). For sera collected early after the onset of symptoms (within 15 days) IgG/IgA-FCI also returned the highest number of positive results (52/72; 72.2%). Positive percent agreement between FCI and compared immunoassays was highest for Roche ECLIA, ranging from 96.1 (IgG/IgA-FCI) to 97.7% (IgG-FCI), whereas negative percent agreement was higher between FCI and Diasosin CLIA, regardless of antibody isotype. The data suggest that FCI may outperform Roche ECLIA and Diasorin CLIA in terms of clinical sensitivity for serological diagnosis of SARS-CoV-2 infection.

Anticorpos Antivirais/sangue , Citometria de Fluxo/métodos , Imunoensaio/métodos , SARS-CoV-2/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Anticorpos Antivirais/imunologia , Teste Sorológico para COVID-19 , Humanos , Células Jurkat , Estudos Retrospectivos , Sensibilidade e Especificidade
Eur J Clin Microbiol Infect Dis ; 40(9): 1925-1932, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-33876385


We optimized and prospectively evaluated a simple MALDI-TOF MS-based method for direct detection of third-generation oxymino-cephalosporin resistance (3rd CephR) in Escherichia coli and Klebsiella spp. from blood cultures (BC). In addition, we assessed the performance of a lateral flow immunochromatographic assay (LFIC) for detecting extended-spectrum ß-lactamases (ESBL) (NG-Test CTX-M MULTI assay) using bacterial pellets from BC. A total of 168 BCs from unique patients were included. A pre-established volume of BC flagged as positive was transferred in brain heart infusion with or without ceftriaxone (2 mg/ml). After 2-h incubation, intact bacterial pellets were used for MALDI-TOF MS testing. Identification of bacterial species (index score > 2) in the presence of CRO was considered marker of 3rd CephR. The LFIC assay was evaluated in 141 BC. Bacteremia episodes were caused by E. coli (n = 115) or Klebsiella spp. (n = 53). A total of 49 strains were 3rd CephR by broth microdilution, of which 41 were ESBL producers, seven expressed ESBL and OXA-48 type D carbapenemase, and one harbored a plasmid-mediated AmpC. The MALDI-TOF MS method yielded four very major errors (false susceptibility) and two major errors (false resistance). The overall sensitivity of the assay was 91.8% and the specificity 98.3%. Concordance between the LFIC assay and the MALDI-TOF MS method for detection of ESBL-mediated 3rd CephR was 100%. Both evaluated methods may prove useful for early adjustment of empirical therapy in patients with E. coli and Klebsiella spp. bloodstream infections. Whether their use has a beneficial impact on patient outcomes is currently under investigation.

Antibacterianos/farmacologia , Bacteriemia/microbiologia , Hemocultura/métodos , Cefalosporinas/farmacologia , Escherichia coli/efeitos dos fármacos , Klebsiella/efeitos dos fármacos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/normas , Adulto , Idoso , Idoso de 80 Anos ou mais , Antibacterianos/uso terapêutico , Cefalosporinas/uso terapêutico , Infecções por Escherichia coli/sangue , Infecções por Escherichia coli/tratamento farmacológico , Feminino , Humanos , Imunoensaio/normas , Infecções por Klebsiella/sangue , Infecções por Klebsiella/tratamento farmacológico , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Estudos Prospectivos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos
Eur J Clin Microbiol Infect Dis ; 40(3): 485-494, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33404891


Whether antibody levels measured by commercially available enzyme or chemiluminescent immunoassays targeting the SARS-CoV-2 spike (S) protein can act as a proxy for serum neutralizing activity remains to be established for many of these assays. We evaluated the degree of correlation between neutralizing antibodies (NtAb) binding the SARS-CoV-2 spike (S) protein and SARS-CoV-2-S-IgG levels measured by four commercial immunoassays in sera drawn from hospitalized COVID-19 patients. Ninety sera from 51 hospitalized COVID-19 patients were tested by a pseudotyped virus neutralization assay, the LIAISON SARS-CoV-2 S1/S2 IgG, the Euroimmun SARS-CoV-2 IgG ELISA, the MAGLUMI 2019-nCoV IgG, and the COVID-19 ELISA IgG assays. Overall, the results obtained with the COVID-19 ELISA IgG test showed the highest agreement with the NtAb assay (κ, 0.85; 95% CI, 0.63-1). The most sensitive tests were the pseudotyped virus NtAb assay and the COVID-19 ELISA IgG assay (92.2% for both). Overall, the degree correlation between antibody titers resulting in 50% virus neutralization (NtAb50) in the pseudotyped virus assay and SARS-CoV-2 IgG levels was strong for the Euroimmun SARS-CoV-2 IgG ELISA (rho = 0.73) and moderate for the remaining assays (rho = 0.48 to 0.59). The kinetic profile of serum NtAb50 titers could not be reliably predicted by any of the SARS-CoV-2 IgG immunoassays. The suitability of SARS-CoV-2-S-IgG commercial immunoassays for inferring neutralizing activity of sera from hospitalized COVID-19 patients varies widely across tests and is influenced by the time of sera collection after the onset of symptoms.

Anticorpos Neutralizantes/sangue , Teste Sorológico para COVID-19/métodos , COVID-19/diagnóstico , Imunoensaio/métodos , SARS-CoV-2/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Adulto , Idoso , Anticorpos Antivirais/sangue , COVID-19/sangue , Feminino , Hospitalização , Humanos , Imunoglobulina G/sangue , Cinética , Masculino , Pessoa de Meia-Idade , Testes de Neutralização , Sensibilidade e Especificidade
J Med Virol ; 93(2): 1141-1144, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-32706420


Knowledge of the precise timing of SARS-CoV-2 infection may be of clinical and epidemiological relevance. The presence of low-avidity IgGs has conventionally been considered an indicator of recent infection. Here, we carried out qualitative assessment of SARS-CoV-2-specific antibody avidity using an urea (6M) dissociation test performed on a lateral flow immunochromatographic IgG/IgM device. We included a total of 76 serum specimens collected from 57 COVID-19 patients, of which 39 tested positive for both IgG and IgM and 37 only for IgG. Sera losing IgG reactivity after urea treatment (n = 28) were drawn significantly earlier (P = .04) after onset of symptoms than those which preserved it (n = 48). This assay may be helpful to estimate the time of acquisition of infection in patients with mild to severe COVID-19.

Anticorpos Antivirais/sangue , Afinidade de Anticorpos , Teste Sorológico para COVID-19 , COVID-19/diagnóstico , Imunoensaio , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Pesquisa Qualitativa , SARS-CoV-2/imunologia
J Med Virol ; 93(4): 2301-2306, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33236799


Assessment of commercial severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) immunoassays for their capacity to provide reliable information on sera neutralizing activity is an emerging need. We evaluated the performance of two commercially available lateral flow immunochromatographic assays (LFIC; Wondfo SARS-CoV-2 Antibody test and the INNOVITA 2019-nCoV Ab test) in comparison with a SARS-CoV-2 neutralization pseudotyped assay for coronavirus disease 2019 (COVID-19) diagnosis in hospitalized patients and investigate whether the intensity of the test band in LFIC associates with neutralizing antibody (NtAb) titers. Ninety sera were included from 51 patients with moderate to severe COVID-19. A green fluorescent protein (GFP) reporter-based pseudotyped neutralization assay (vesicular stomatitis virus coated with SARS-CoV-2 spike protein) was used. Test line intensity was scored using a 4-level scale (0 to 3+). The overall sensitivity of LFIC assays was 91.1% for the Wondfo SARS-CoV-2 Antibody test, 72.2% for the INNOVITA 2019-nCoV IgG, 85.6% for the INNOVITA 2019-nCoV IgM, and 92.2% for the NtAb assay. Sensitivity increased for all assays in sera collected beyond day 14 after symptoms onset (93.9%, 79.6%, 93.9%, and 93.9%, respectively). Reactivities equal to or more intense than the positive control line (≥2+) in the Wondfo assay had a negative predictive value of 100% and a positive predictive value of 96.4% for high NtAb50 titers (≥1/160). Our findings support the use of LFIC assays evaluated herein, particularly the Wondfo test, for COVID-19 diagnosis. We also find evidence that these rapid immunoassays can be used to predict high SARS-CoV-2-S NtAb50 titers.

Anticorpos Neutralizantes/sangue , Teste para COVID-19/métodos , COVID-19/imunologia , Imunoensaio/métodos , SARS-CoV-2/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , COVID-19/sangue , COVID-19/diagnóstico , COVID-19/virologia , Proteínas de Fluorescência Verde , Humanos , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Glicoproteína da Espícula de Coronavírus/imunologia
Clin Microbiol Infect ; 27(3): 472.e7-472.e10, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33189872


OBJECTIVES: To our knowledge no previous study has assessed the performance of a rapid antigen diagnostic immunoassay (RAD) conducted at the point of care (POC). We evaluated the Panbio™ COVID-19 Ag Rapid Test Device for diagnosis of coronavirus 2019 disease (COVID-19) in symptomatic patients (n = 412) attending primary healthcare centres. METHODS: RAD was performed immediately after sampling following the manufacturer's instructions (reading at 15 min). RT-PCRs were carried out within 24 h of specimen collection. Samples displaying discordant results were processed for culture in Vero E6 cells. Presence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in cell cultures was confirmed by RT-PCR. RESULTS: Out of 412 patients, 43 (10.4%) tested positive by RT-PCR and RAD, and 358 (86.9%) tested negative by both methods; discordant results (RT-PCR+/RAD-) were obtained in 11 patients (2.7%). Overall specificity and sensitivity of rapid antigen detection (RAD) was 100% (95%CI 98.7-100%) and 79.6% (95%CI 67.0-88.8%), respectively, taking RT-PCR as the reference. Overall RAD negative predictive value for an estimated prevalence of 5% and 10% was 99% (95%CI 97.4-99.6%) and 97.9% (95%CI 95.9-98.9), respectively. SARS-CoV-2 could not be cultured from specimens yielding RT-PCR+/RAD- results (n = 11). CONCLUSION: The Panbio™ COVID-19 Ag Rapid Test Device performed well as a POC test for early diagnosis of COVID-19 in primary healthcare centres. More crucially, the data suggested that patients with RT-PCR-proven COVID-19 testing negative by RAD are unlikely to be infectious.

Teste Sorológico para COVID-19/métodos , COVID-19/diagnóstico , Testes Imediatos , SARS-CoV-2/isolamento & purificação , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Instituições de Assistência Ambulatorial , Antígenos Virais/análise , Teste de Ácido Nucleico para COVID-19 , Criança , Pré-Escolar , Diagnóstico Precoce , Feminino , Humanos , Imunoensaio , Lactente , Masculino , Pessoa de Meia-Idade , Nasofaringe/virologia , Kit de Reagentes para Diagnóstico , SARS-CoV-2/genética , SARS-CoV-2/imunologia , Sensibilidade e Especificidade , Adulto Jovem