ABSTRACT
ObjectivesThere is an imperative need to determine the durability of adaptive immunity to SARS-CoV-2. We enumerated SARS-CoV-2-reactive CD4+ and CD8+ T cells targeting S1 and M proteins and measured RBD-specific serum IgG over a period of 2-6 months after symptoms onset in a cohort of subjects who had recovered from severe clinical forms of COVID-19. MethodsWe recruited 58 patients (38 males and 20 females; median age, 62.5 years), who had been hospitalized with bilateral pneumonia, 60% with one or more comorbidities. IgG antibodies binding to SARS-CoV-2 RBD were measured by ELISA. SARS-CoV-2-reactive CD69+-expressing-IFN{gamma}-producing-CD4+ and CD8+ T cells were enumerated in heparinized whole blood by flow cytometry for ICS. ResultsDetectable SARS-CoV-2-S1/M-reactive CD69+-IFN-{gamma} CD4+ and CD8+ T cells were displayed in 17 (29.3%) and 6 (10.3%) subjects respectively, at a median of 84 days after onset of symptoms (range, 58-191 days). Concurrent comorbidities increased the risk (OR, 3.15; 95% CI, 1.03-9.61; P=0.04) of undetectable T-cell responses in models adjusted for age, sex and hospitalization ward. Twenty-one out of the 35 patients (60%) had detectable RBD-specific serum IgGs at a median of 118 days (range, 60 to 145 days) after symptoms onset. SARS-CoV-2 RBD-specific IgG serum levels were found to drop significantly over time. ConclusionA relatively limited number of subjects who developed severe forms of COVID-19 had detectable SARS-CoV-2-S1/M IFN{gamma} CD4+ and CD8+ T cells at midterm after clinical diagnosis. Our data also indicated that serum levels of RBD-specific IgGs decline over time, becoming undetectable in some patients.
ABSTRACT
BackgroundWhether 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. ObjectivesTo evaluate 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. Patients and MethodsNinety sera from 51 hospitalized COVID-19 patients were assayed 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. ResultsOverall, the results obtained with the COVID-19 ELISA IgG test showed the highest agreement with the NtAb assay ({kappa}, 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. ConclusionsThe 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.
ABSTRACT
PurposeAssessment of commercial 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 COVID-19 diagnosis in hospitalized patients, and investigate whether the intensity of the test band in LFIC associates with neutralizing antibody (NtAb) titers. Patients and MethodsNinety 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+). ResultsOverall 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). ConclusionsOur 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.
ABSTRACT
BackgroundThe involvement of SARS-CoV-2 antibodies in mediating immunopathogenetic events in COVID-19 patients has been suggested. By using several experimental approaches, we investigated the potential association between SARS-CoV-2 IgGs recognizing the spike (S) protein receptor-binding domain (RBD), neutralizing antibodies (NtAb) targeting S, and COVID-19 severity. Patients and MethodsThis unicenter, retrospective, observational study included 51 hospitalized patients (24 at the intensive care unit; ICU). A total of 93 sera from these patients collected at different time points from the onset of symptoms were analyzed. SARS-CoV-2 RBD IgGs were quantitated by ELISA and NtAb50 titers were measured in a GFP reporter-based pseudotyped virus platform. Demographic and clinical data, complete blood counts, as well as serum levels of ferritin, Dimer-D, C reactive protein (CRP), lactose dehydrogenase (LDH), and interleukin-6 (IL-6) were retrieved from clinical charts. ResultsThe overall correlation between levels of both antibody measurements was good (Rho=0.79; P=0<0.001). SARS-CoV-2 RBD IgG and NtAb50 levels in sera collected up to day 30 after the onset of symptoms were comparable between ICU and non-ICU patients (P=>0.1). The percentage of patients who exhibited high NtAb50 titers ([≥] 160) was similar (P=0.20) in ICU (79%) and non-ICU (60%) patients. Four ICU patients died; two of these achieved NtAb50 titers [≥] 1/160 while the other two exhibited a 1/80 titer. Very weak (Rho=>0.0-<0.2) or weak (Rho=>0.2-<0.4) correlations were observed between anti-RBD IgGs, NtAb50, and serum levels pro-inflammatory biomarkers. ConclusionsThe data presented herein do not support an association between SARS-CoV-2 RBD IgG or NtAb50 levels and COVID-19 severity.
ABSTRACT
Real-time reverse transcription polymerase-chain reaction (RT-PCR) is the mainstay of Covid-19 diagnosis. False-negative RT-PCR results may hamper clinical management of patients and hinder the adoption of epidemiological measures to control the pandemic. The current study was aimed at assessing whether amplification of {beta}-glucoronidase (GUSB) gene would help estimate the accuracy of SARS-CoV-2 RT-PCR negative results in upper respiratory tract (URT) specimens. URT specimens that tested negative by SARS-CoV-2 RT-PCR displayed higher GUSB RT-PCR cycle thresholds (CT) (P=0.070) than those testing positive (median, 30.7; range, 27.0-40.0, and median 29.7; range 25.5-36.8, respectively), this reflecting poorer cellularity. Receiver operating characteristic (roc) curve analysis indicated that a CT threshold of 31.2 discriminated best between positive and negative SARS CoV-2 RT-PCRs (area under a curve, 0.66; 95% CI, 0.50-0.81; P=0.08). This cut-off yielded a true negative ratio of 89% and accuracy of 70%. The data suggested that amplification of the GUSB gene by RT-PCR may help to appraise the accuracy of negative SARS-CoV-2 RT-PCR results in patients in whom Covid-19 is eventually diagnosed.
ABSTRACT
There is scarce information on the frequency of co-detection of respiratory pathogens (RP) in patients with Covid-19. Documentation of coinfections in Covid-19 pneumonia patients may be relevant for appropriate clinical and therapeutic management of patients. Between March 4th and March 28th, 2020, a total of 183 adult patients testing positive by SARS CoV-2 RT-PCR on respiratory specimens were hospitalized with interstitial pneumonia at our center, of whom 103 were tested for other RP by a multiplexed PCR assay. Three patients had a positive result for either one (n=2; Coronavirus HKU1 or Mycoplasma pneumoniae) or two targets (n=1; Influenza virus A (H3) and Respiratory syncytial virus B). Twenty-three patients testing negative by SARS CoV-2 RT-PCR and presentig with clinical, laboratory findings and imaging compatibe with Covid-19 pneumonia underwent RP screening. Of these, 6 (26%) had a positive result for a single RP. Our data indicate that despite the apparent rarity of coinfections in patients with Covid-19 pneumonia, routine testing for RP should be advised, since agents for which specific therapy can be prescribed may be detected.
ABSTRACT
Noroviruses infect persons of all ages, often causing epidemic outbreaks of acute gastroenteritis as well as sporadic cases. The application of novel molecular methods to the diagnosis of norovirus infections is now revealing their real impact. Molecular epidemiology studies have identified the most common viral genotypes responsible for human infections. Norovirus gastroenteritis is usually mild and of short duration, although the disease can also be severe, especially in the elderly, or may become chronic, as occurred in the immunocompromised patients. Several factors have been identified regarding the differential susceptibility to norovirus infection among individuals, consisting of several histo-blood antigens (ABO, Lewis and secretor) that are involved in the binding process of noroviruses to the enterocytes. The expression of these antigens in humans is genetically encoded, and shows a high polymorphism, which combined with the genetic diversity of noroviruses, makes the virus-host relationship rather complex. The diagnosis of norovirus infections is not performed routinely in many laboratories, but those involved in epidemiological surveillance have identified norovirus strains that evolve sequentially over time, similarly to Influenza viruses.
