Clinical assessment of SNIBE Maglumi SARS-CoV-2 antigen fully-automated chemiluminescent immunoassay.

OBJECTIVES: Given that SARS-CoV-2 antigen tests will represent a pillar for supporting or surrogating molecular testing in the endemic period, we report here the clinical performance of the new SNIBE Maglumi SARS-CoV-2 antigen fully-automated chemiluminescent immunoassay (MAG-CLIA SARS-CoV-2 Ag). METHODS: The study population consisted of 181 subjects (mean age 61 ± 21 years; 92 females) undergoing coronavirus disease 2019 (COVID-19) testing at the local diagnostic facility, from December 2022 to February 2023. Routine diagnostic practice involved the collection of a double nostril nasopharyngeal swab, analyzed in duplicate with SARS-CoV-2 antigen (MAG-CLIA SARS-CoV-2 Ag) and molecular (Altona Diagnostics RealStar SARS-CoV-2 RT-PCR Kit) tests. RESULTS: A significant Spearman's correlation was found between MAG-CLIA SARS-CoV-2 Ag and mean Ct values of SARS-CoV-2 E and S genes (r=-0.95; p<0.001). In all nasopharyngeal samples, the area under the curve (AUC) of MAG-CLIA SARS-CoV-2 Ag was 0.86 (95% CI, 0.81-0.90), with 0.71 sensitivity and 1.00 specificity at 7 ng/L cut-off, increasing to 0.98 (95% CI, 0.96-1.00) AUC and 0.96 sensitivity (with 0.97 specificity) in high viral load samples. When SARS-CoV-2 N protein concentration was replaced with raw instrumental readings (i.e., relative light units [RLU]), the AUC in all samples increased to 0.94. A RLU value of 945 was associated with 88.4% accuracy, 0.85 sensitivity, 0.95 specificity, 0.77 negative predictive value (NPV) and 0.97 positive predictive value (PPV), respectively. CONCLUSIONS: We found satisfactory analytical performance of MAG-CLIA SARS-CoV-2 Ag, which could be used as surrogate of molecular testing for identifying high viral load samples. Broadening the reportable range of values may generate even better performance.

Assessment of humoral and cellular immunity after bivalent BNT162b2 vaccination and potential association with reactogenicity.

OBJECTIVES: This study investigated the feasibility and clinical value of using a novel, automated and high-throughput SARS-CoV-2 Interferon Gamma Release Assay (IGRA), combined with total anti-SARS-CoV-2 antibodies assessment, for evaluating the immune response after bivalent BNT162b2 vaccination. METHODS: A cohort of healthcare workers, who already underwent primary vaccination and boosting with monovalent BNT162b2 vaccine, received a booster dose of the new BNT162b2 bivalent formulation. Blood samples were taken immediately before vaccination (T0) and 1 month afterwards (T1). Humoral and cellular immunity were assayed with Roche Elecsys Anti-SARS-CoV-2 and Roche Elecsys IGRA SARS-CoV-2, respectively. RESULTS: The study population consisted of 51 subjects (median age: 43 years; 51% females). Total anti-SARS-CoV-2 antibodies and IGRA SARS-CoV-2 values increased at T1 from 9,050 to 25,000 BAU/mL (p<0.001), and from 0.44 to 0.78 IU/mL (p=0.385), accounting for median increase of 2.0 and 1.6 folds, respectively. Increased T1 values of total anti-SARS-CoV-2 antibodies and IGRA SARS-CoV-2 were recorded in 100% and 68.6% subjects, respectively. In those with baseline values below the median, post-vaccine levels displayed larger increases of 3.3 and 5.1 folds for anti-SARS-CoV-2 total antibodies and IGRA SARS-CoV-2, respectively. The variation of total anti-SARS-CoV-2 antibodies was inversely associated with their T0 values (r=-0.97; p<0.001), whilst that of IGRA SARS-CoV-2 was inversely associated with its T0 value (r=-0.58; p<0.001). No other signifcant associations were found with demographical or clinical variables, including side effects. CONCLUSIONS: The bivalent BNT162b2 vaccine booster enhances humoral and cellular immunity against SARS-CoV-2, especially in recipients with lower baseline biological protection.

Real-world assessment of the clinical performance of COVID-VIRO ALL IN rapid SARS-CoV-2 antigen test.

OBJECTIVES: Since the external validation of severe acute respiratory syndrome coronavirus 2 antigen rapid diagnostic tests (SARS-CoV-2 RDT-Ags) is a necessary requisite before they can be introduced into routine clinical practice, this study reports the results of a real-world assessment of the clinical performance of the new COVID-VIRO ALL IN device. METHODS: The study population consisted in 165 outpatients (median age: 43 years, range: 14-68 years; 66.1% females) who had paired nasal and nasopharyngeal samples collected upon hospital presentation. The samples were concomitantly tested with the AAZ-LMB COVID-VIRO ALL IN SARS-CoV-2 RDT-Ag and with Cepheid Xpert Xpress SARS-CoV-2 real-time reverse transcription polymerase chain reaction (RT-PCR). RESULTS: The number of subjects with positive RT-PCR results (i.e., mean Ct value <45) was 116 (70.3%), 109 (66.1%) and 86 (52.1%) with mean Ct values <37 and <30, respectively. In all RT-PCR positive samples, COVID-VIRO ALL IN displayed 78.8% agreement, 0.698 sensitivity, 1.000 specificity, 0.583 negative predictive value (NPV) and 1.000 positive predictive value (PPV) compared to RT-PCR. The median Ct value of samples testing positive with COVID-VIRO ALL IN was significantly lower than those testing negative (22.8 vs. 32.2; p<0.001). In samples with high viral load (i.e., Ct value <30), COVID-VIRO ALL IN displayed 92.1% agreement, 0.895 sensitivity, 0.949 specificity, 0.983 NPV and 0.951 PPV compared to RT-PCR. CONCLUSIONS: Although the diagnostic performance of COVID-VIRO ALL IN do not exactly match those of the manufacturer, its high NPV in high viral load samples would enable fast-track and rapid identification of highly contagious subjects.

Effect of BNT162b2 booster dose on anti-SARS-CoV-2 spike trimeric IgG antibodies in seronegative individuals.

OBJECTIVES: We provide here an updated analysis of an ongoing serosurveillance study, presenting data on the effect of a third dose of Pfizer/BioNTech BNT162b2 vaccine on serum anti-SARS-CoV-2 IgG antibodies. METHODS: We tested baseline SARS-CoV-2 seronegative healthcare workers undergoing primary vaccination with the mRNA-based COVID-19 Comirnaty vaccine, followed by administration of homologous vaccine booster (third dose). Venous blood was collected before either dose of primary vaccination, at 1, 3 and 6 months afterwards, as well as before and 1 month after receiving the vaccine booster. The serum concentration of anti-SARS-CoV-2 IgG was assayed with DiaSorin Trimeric spike IgG immunoassay. RESULTS: The final study population included 53 SARS-CoV-2 seronegative healthcare workers (median age 46 years; 60% females). A first peak of anti-SARS-CoV-2 spike trimeric IgG values was reached 1 month after completing primary vaccination, after which the levels gradually declined until before receiving the vaccine booster. A second peak of anti-SARS-CoV-2 spike trimeric IgG concentration was observed 1 month after receiving the vaccine booster dose (8,700 kBAU/L), which was 39-fold higher than before receiving the vaccine booster (221 kBAU/L; p<0.001), but was also nearly threefold higher compared to values seen at the first peak (2,990 kBAU/L; p<0.001). The rate of subjects with protective anti-SARS-CoV-2 spike trimeric IgG values (i.e., >264 kBAU/L) increased from 47.2% to 100% after 1 month from vaccine booster. CONCLUSIONS: These results support current policies fostering COVID-19 vaccine boosters to reinforce humoral immunity against SARS-CoV-2.

Comparative longitudinal variation of total IgG and IgA anti-SARS-CoV-2 antibodies in recipients of BNT162b2 vaccination.

Objectives: This article aims to summarize the 6-month variation of a vast array of anti-SARS-CoV-2 antibodies in recipients of BNT162b2 mRNA-based vaccination. Methods: The study population consisted of 84 baseline SARS-CoV-2 seronegative healthcare employees (median age 45 years, 53.6% females), receiving mRNA-based BNT162b2 primary vaccination cycle. Blood was collected before the first and second BNT162b2 vaccine doses, as well as 1, 3 and 6 months afterwards. The serum titers of the following anti-SARS-CoV-2 antibodies were assayed: total anti-RBD (receptor binding domain), anti-spike trimeric IgG, anti-RBD IgG and anti-spike S1 IgA. Results: All antibodies' levels peaked 1 month after vaccination, but then displayed a considerable decrease. The median rates of 6-month decline were -95% for IgG anti-SARS-CoV-2 RBD, -85% for IgG anti-SARS-CoV-2 trimeric spike, -73% for IgA anti-SARS-CoV-2 S1 and -56% for total anti-SARS-CoV-2 RBD antibodies, respectively. The median time of seronegativization was estimated at 579 days for total anti-SARS-CoV-2 RBD antibodies, 271 days for IgG anti-SARS-CoV-2 trimeric spike, 264 days for IgG anti-SARS-CoV-2 RBD and 208 days for IgA anti-SARS-CoV-2 S1, respectively. The rate of seropositive subjects declined from 98-100% at the peak to 50-100% after 6 months. The inter-individual variation of anti-SARS-CoV-2 antibodies reduction at 6 months was 3-44% from the peak. Conclusions: The results of this longitudinal serosurvey demonstrate that the titer of anti-SARS-CoV-2 antibodies declined 6 months after BNT162b2 vaccination, with median time of IgG/IgA seronegativization estimated between 7 and 9 months, thus supporting the opportunity of administering vaccine boosters approximately 5 to 6 months after the last dose of the primary vaccination cycle.

Clinical Assessment of the DiaSorin LIAISON SARS-CoV-2 Ag Chemiluminescence Immunoassay.

BACKGROUND: Due to the large volume of tests needed in a relatively short time for screening and diagnosing severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, antigen immunoassays may provide a potential supplement to molecular testing. This study was aimed to assess the clinical preference of DiaSorin LIAISON SARS-CoV-2 Ag chemiluminescence immunoassay. METHODS: An upper respiratory specimen was collected in a series of patients referred to the Laboratory Medicine service of Pederzoli Hospital (Peschiera del Garda, Verona, Italy) for screening or diagnosis of SARS-CoV-2 infection. Nasopharyngeal samples were assayed with DiaSorin LIAISON SARS-CoV-2 Ag test and Altona Diagnostics RealStar® SARS-CoV-2 RT-PCR Kit. RESULTS: The final study population consisted of 421 patients (median age, 48 years; 227 women), 301 (71.5%) with positive result of molecular testing, and 126 (29.9%) with cycle threshold (Ct) values of both E and S genes <29.5, thus reflecting higher infectivity. The area under the curve of DiaSorin LIAISON SARS-CoV-2 Ag test 0.82 (95% CI, 0.79-0.86) for sample positivity and 0.98 for higher sample infectivity (95% CI, 0.97 to 0.99). The optimal cut-off for sample positivity was 82 TCID50/mL (0.78 sensitivity, 0.73 specificity and 77% diagnostic accuracy), whilst that for identifying samples associated with a high infective risk was 106 TCID50/mL (0.94 sensitivity, 0.96 specificity and 95% diagnostic accuracy). CONCLUSION: The performance of this chemiluminescence immunoassay would not permit it to replace molecular testing for diagnosing SARS-CoV-2, but may enable rapid and efficient detection of subjects with high SARS-CoV-2 viral load, who are responsible for the largest proportion of infectious clusters.

Clinical assessment of the Roche SARS-CoV-2 rapid antigen test.

OBJECTIVES: Novel point-of-care antigen assays present a promising opportunity for rapid screening of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections. The purpose of this study was the clinical assessment of the new Roche SARS-CoV-2 Rapid Antigen Test. METHODS: The clinical performance of Roche SARS-CoV-2 Rapid Antigen Test was evaluated vs. a reverse transcription polymerase chain reaction (RT-PCR) laboratory-based assay (Seegene AllplexTM2019-nCoV) in nasopharyngeal swabs collected from a series of consecutive patients referred for SARS-CoV-2 diagnostics to the Pederzoli Hospital (Peschiera del Garda, Verona, Italy) over a 2-week period. RESULTS: The final study population consisted of 321 consecutive patients (mean age, 46 years and IQR, 32-56 years; 181 women, 56.4%), with 149/321 (46.4%) positive for SARS-CoV-2 RNA via the Seegene AllplexTM2019-nCoV Assay, and 109/321 (34.0%) positive with Roche SARS-CoV-2 Rapid Antigen Test, respectively. The overall accuracy of Roche SARS-CoV-2 Rapid Antigen Test compared to molecular testing was 86.9%, with 72.5% sensitivity and 99.4% specificity. Progressive decline in performance was observed as cycle threshold (Ct) values of different SARS-CoV-2 gene targets increased. The sensitivity was found to range between 97-100% in clinical samples with Ct values <25, between 50-81% in those with Ct values between 25 and <30, but low as 12-18% in samples with Ct values between 30 and <37. CONCLUSIONS: The clinical performance of Roche SARS-CoV-2 Rapid Antigen Test is excellent in nasopharyngeal swabs with Ct values <25, which makes it a reliable screening test in patients with high viral load. However, mass community screening would require the use of more sensitive techniques.

Real-world assessment of Fluorecare SARS-CoV-2 Spike Protein Test Kit.

Objectives: Since commercial SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) antigen rapid detection tests (Ag-RDTs) display broad diagnostic efficiency, this study aimed to evaluate the clinical performance of Fluorecare SARS-CoV-2 Spike Protein Test Kit in a real-life scenario. Methods: The study population consisted of a series of patients undergoing SARS-Cov-2 diagnostic testing at Pederzoli Hospital of Peschiera del Garda (Verona, Italy). A nasopharyngeal swab was collected upon hospital admission and assayed with molecular (Altona Diagnostics RealStar® SARSCoV-2 RT-PCR Kit) and antigen (Fluorecare SARS-CoV-2 Spike Protein Test Kit) tests. Results: The study population consisted of 354 patients (mean age, 47 ± 20 years; 195 women, 55.1%), 223 (65.8%) positive at molecular testing. A significant correlation was found between Fluorecare SARS-CoV-2 Spike Protein Test Kit and Altona (both S and E genes: r=-0.75; p<0.001). The cumulative area under the curve in all nasopharyngeal samples was 0.68. At ≥1.0 S/CO manufacturer's cut-off, the sensitivity, specificity, negative and positive predictive values were 27.5, 99.2, 41.5 and 98.5%, respectively. Considerable improvement of sensitivity was observed as Ct values decreased, becoming 66.7% in samples with mean Ct values <30, 90.5% in those with mean Ct values <25, up to 100% in those with mean Ct values <20. Conclusions: The modest sensitivity and negative predictive value of Fluorecare SARS-CoV-2 Spike Protein Test Kit makes unadvisable to use this assay as surrogate of molecular testing for definitively diagnosing SARS-CoV-2 infection, though its suitable sensitivity at high viral load could make it a reliable screening test for patients with higher infective potential.