scholarly journals Comparative Evaluation of SARS-CoV-2 IgG Assays in India

2020 ◽  
Author(s):  
◽  
Shinjini Bhatnagar
Keyword(s):  
Comparative Evaluation ◽  
Spike Protein ◽  
Clinical Samples ◽  
Plasma Samples ◽  
Rt Pcr ◽  
High Agreement ◽  
Asymptomatic Individuals ◽  
Low Prevalence ◽  
Sera Panel

IgG immunoassays have been developed and used widely for clinical samples and serosurveys for SARS-CoV-2. We compared the performance of three immunoassays, an in-house RBD assay, and two commercial assays, the Diasorin LIAISON SARS-CoV-2 IgG CLIA which detects antibodies against S1/S2 domains of the Spike protein and the Zydus Kavach assay based on inactivated virus using a well-characterized sera-panel. 379 sera/plasma samples from RT-PCR positive individuals >20 days of illness in symptomatic or RT-PCR positivity in asymptomatic individuals and 184 pre-pandemic samples were used. The sensitivity of the assays were 84.7, 82.6 and 75.7 respectively for RBD, LIAISON and Kavach. Kavach and the in-house RBD ELISA showed a specificity of 99.5% and 100%, respectively. The RBD and LIAISON (S1/S2) assays showed high agreement (94.7%;95%CI:92.0,96.6) and were able to correctly identify more positives than Kavach. All three assays are suitable for serosurveillance studies, but in low prevalence sites, estimation of exposure may require adjustment based on our findings.

scholarly journals Rapid and Sensitive Detection of SARS-CoV-2 Infection Using Quantitative Peptide Enrichment LC-MS/MS Analysis

2021 ◽  
Author(s):  
Andreas Hober ◽  
Tran-Minh Khue Hua ◽  
Dominic Foley ◽  
Thomas McDonald ◽  
Johannes P.C. Vissers ◽  
...  
Keyword(s):  
Large Scale ◽  
Virus Detection ◽  
Viral Proteins ◽  
Clinical Samples ◽  
Molecular Testing ◽  
Rt Pcr ◽  
Proteomics Analysis ◽  
Peptide Enrichment ◽  
Asymptomatic Individuals ◽  
Detection Technologies

Reliable, robust, large-scale molecular testing for SARS-CoV-2 is essential for monitoring the ongoing Covid-19 pandemic. We have developed a scalable analytical approach to detect viral proteins based on peptide immunoaffinity enrichment combined with liquid chromatography - mass spectrometry (LC-MS). This is a multiplexed strategy, based on targeted proteomics analysis and read-out by LC-MS, capable of precisely quantifying and confirming the presence of SARS-CoV-2 in PBS swab media from combined throat/nasopharynx/saliva samples. The results reveal that the levels of SARS-CoV-2 measured by LC-MS correlate well with their corresponding RT-PCR readout. The analytical workflow shows similar turnaround times as regular RT-PCR instrumentation with quantitative readout of viral proteins corresponding to cycle thresholds (Ct) equivalents ranging from 21 to 34. Using RT-PCR as a reference, we demonstrate that the LC-MS-based method has 100% estimated specificity and 83.3% estimated sensitivity when analyzing clinical samples collected from asymptomatic individuals. These results suggest that a scalable analytical method based on LC-MS has a place in future pandemic preparedness centers to complement current virus detection technologies.

scholarly journals Evaluation of real-time nucleic acid sequence-based amplification for detection of Chikungunya virus in clinical samples

2009 ◽  
Vol 58 (9) ◽  
pp. 1168-1172 ◽  
Author(s):  
J.-N. Telles ◽  
K. Le Roux ◽  
P. Grivard ◽  
G. Vernet ◽  
A. Michault
Keyword(s):  
Nucleic Acid ◽  
Real Time ◽  
Chikungunya Virus ◽  
False Negative ◽  
Nucleic Acid Sequence ◽  
Clinical Samples ◽  
Acid Extraction ◽  
Plasma Samples ◽  
Rt Pcr ◽  
Nucleic Acid Extraction

The Chikungunya virus (CHIKV) is a member of the genus Alphavirus that is transmitted to humans by Aedes mosquitoes. In 2005 and 2006, the Indian Ocean island of La Réunion was hit with an unprecedented CHIKV fever outbreak that infected 300 000 people. In the present study, we describe the evaluation of real-time nucleic acid sequence-based amplification (RT-NASBA) for the detection of CHIKV in clinical samples. A co-extracted and co-amplified chimerical CHIKV RNA sequence was used as an internal control to eliminate false-negative results. The detection threshold of the assay was determined from quantified CHIKV-positive plasma, and estimated to be 200 copies per NASBA reaction. The specificity of the assay was determined using blast analyses and non-cross-reactivity using an O'nyong-nyong virus culture and 250 CHIKV RT-PCR-negative plasma samples. A 100 % specificity was found and no invalid result was obtained, showing the good quality of the nucleic acid extraction. The assay was then evaluated using 252 CHIKV-positive RT-PCR plasma samples. The samples were all tested positive, including those with low viral load. This evaluation showed that the RT-NASBA is a rapid (5 h from sample nucleic acid extraction to detection), sensitive, specific and reliable method for the routine diagnosis of CHIKV in clinical samples.

scholarly journals Prospective analytical performance evaluation of the QuickNavi™-COVID19 Ag for asymptomatic individuals

2021 ◽  
Author(s):  
Yoshihiko Kiyasu ◽  
Yuto Takeuchi ◽  
Yusaku Akashi ◽  
Daisuke Kato ◽  
Miwa Kuwahara ◽  
...  
Keyword(s):  
Real Time ◽  
Diagnostic Performance ◽  
False Positives ◽  
Clinical Samples ◽  
Lower Sensitivity ◽  
Antigen Test ◽  
Rt Pcr ◽  
Asymptomatic Group ◽  
Asymptomatic Volunteers ◽  
Asymptomatic Individuals

AbstractIntroductionAntigen testing may help screen for and detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections in asymptomatic individuals. However, limited data regarding the diagnostic performance of antigen tests for this group are available.MethodsWe used clinical samples to prospectively evaluate the analytical and clinical performance of the antigen test QuickNavi™-COVID19 Ag. This study was conducted at a PCR center between October 7, 2020 and January 9, 2021. Two nasopharyngeal samples per patient were obtained with flocked swabs; one was used for the antigen test, and the other for real-time reverse transcription PCR (RT-PCR). The diagnostic performance of the antigen test was compared between asymptomatic and symptomatic patients, and the RT-PCR results were used as a reference.ResultsAmong the 1,934 collected samples, SARS-CoV-2 was detected by real-time RT-PCR in 188 (9.7%); 76 (40.4%) of these samples were from asymptomatic individuals. Over half of the total samples (1,073; 55.5%) were obtained from asymptomatic volunteers. The sensitivity of the antigen test was significantly lower for asymptomatic group than for symptomatic patients (67.1% vs 89.3%, p < 0.001). The specificity was 100% for both groups, and no false positives were observed among all 1,934 samples. The median Ct value for the asymptomatic group was significantly higher than that of the symptomatic group (24 vs 20, p < 0.001).ConclusionsThe QuickNavi™-COVID19 Ag showed a lower sensitivity for asymptomatic group than for symptomatic patients. However, its specificity was consistently high, and no false positives were found in this study.

scholarly journals Impact of Pool Testing in Detection of Asymptomatic Patients with COVID-19

2020 ◽  
Vol 52 (1) ◽  
pp. e15-e16 ◽  
Author(s):  
David L Smalley ◽  
Patricia M Cisarik ◽  
James Grantham ◽  
William Cloud ◽  
R Brock Neil ◽  
...  
Keyword(s):  
Single Patient ◽  
Rt Pcr ◽  
Detection Capability ◽  
Chain Reaction ◽  
Asymptomatic Patients ◽  
Alternative Approaches ◽  
Polymerase Chain ◽  
Asymptomatic Individuals ◽  
Low Prevalence ◽  
The Impact

Abstract Objective During the current pandemic, COVID-19 has been detected in patients using real-time reverse transcriptase-polymerase chain reaction (RT-PCR) that confirms the presence of SARS-CoV-2 RNA. The demand for increased testing, particularly for asymptomatic individuals required alternative approaches to single-patient RT-PCR testing, such as pooling. Methods This study explored the impact of dilution on the detectability of SARS-CoV-2 in asymptomatic patients using RT-PCR and demonstrated that pooling can be effective in low prevalence populations. Results The RT-PCR results for the 3:1, 5:1, and 7:1 aliquot samples showed little differences in CT values, confirming detection capability at these dilutions. Conclusion Based on the results of the present study, a pooled approach with up to 5:1 sample aliquots and using the current RT-PCR methodology likely will detect SARS CoV2 RNA among asymptomatic patients.

scholarly journals Rapid Screening for SARS-CoV-2 Variants of Concern in Clinical and Environmental Samples Using Nested RT-PCR Assays Targeting Key Mutations of the Spike Protein

2021 ◽  
Author(s):  
G. La Rosa ◽  
P. Mancini ◽  
G. Bonanno Ferraro ◽  
C. Veneri ◽  
M. Iaconelli ◽  
...  
Keyword(s):  
Spike Protein ◽  
Rapid Screening ◽  
Clinical Samples ◽  
Correct Identification ◽  
Potential Contribution ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Field Samples ◽  
Pcr Assays ◽  
Viral Isolates

SUMMARYNew SARS-CoV-2 mutations are constantly emerging, raising concerns of increased transmissibility, virulence or escape from host immune response.We describe a nested RT-PCR assay (∼1500 bps) to detect multiple key spike protein mutations distinctive of the major known circulating SARS-CoV-2 variants, including the three Variants of Concern (VOCs) 20I/501Y.V1 (United Kingdom), 20H/501Y.V2 (South Africa), and 20J/501Y.V3 (Brazil), as well as the 20E.EU1 variant (Spain), the CAL.20C recently identified in California, and the mink-associated variant (GR, lineage B.1.1.298). Prior to application to field samples, the discriminatory potential of this PCR assay was explored using GISAID and Nextclade. To extend variant detection to challenging matrices such as sewage, where the amplification of long fragments is problematic, two short nested RT-PCR assays (∼300 bps) were also designed, targeting portions of the region spanned by the long nested assay.The three newly-designed assays were then tested on field samples, including 7 fully-sequenced viral isolates from swab samples and 34 urban wastewater samples, some of which collected in areas where circulation of VOCs had been reported.The long assay successfully amplified all the previously characterized viral isolates, allowing the correct identification of variants 20I/501Y.V1 and 20E.EU1 present in the panel. The sequences obtained using the short assays were consistent with those obtained with the long assay. Mutations characteristic of VOCs (UK and Brazilian variant) and of other variant (Spanish) were detected in sewage samples. To our knowledge, this is the first evidence of the presence of sequences harboring key mutations of 20I/501Y.V1 and 20J/501Y.V3 in urban wastewaters, highlighting the potential contribution of wastewater surveillance to explore SARS-CoV-2 diversity.The developed nested RT-PCR assays can be used as an initial rapid screening test to select clinical samples containing mutations of interest. This can speed up diagnosis and optimize resources since it allows full genome sequencing to be done only on clinically relevant specimens. The assays can be also employed for a rapid and cost-effective detection of VOCs or other variants in sewage for the purposes of wastewater-based epidemiology. The approach proposed here can be used to better understand SARS-CoV-2 variant diversity, geographic distribution and impact worldwide.

scholarly journals Rapid and sensitive detection of SARS-CoV-2 infection using quantitative peptide enrichment LC-MS analysis

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Andreas Hober ◽  
Khue Hua Tran-Minh ◽  
Dominic Foley ◽  
Thomas McDonald ◽  
Johannes PC Vissers ◽  
...  
Keyword(s):  
Large Scale ◽  
Limit Of Detection ◽  
Virus Detection ◽  
Viral Proteins ◽  
Clinical Samples ◽  
Molecular Testing ◽  
Rt Pcr ◽  
Percent Agreement ◽  
Peptide Enrichment ◽  
Asymptomatic Individuals

Reliable, robust, large-scale molecular testing for SARS-CoV-2 is essential for monitoring the ongoing Covid-19 pandemic. We have developed a scalable analytical approach to detect viral proteins based on peptide immunoaffinity enrichment combined with liquid chromatography - mass spectrometry (LC-MS). This is a multiplexed strategy, based on targeted proteomics analysis and read-out by LC-MS, capable of precisely quantifying and confirming the presence of SARS-CoV-2 in PBS swab media from combined throat/nasopharynx/saliva samples.<br />The results reveal that the levels of SARS-CoV-2 measured by LC-MS correlate well with their corresponding RT-PCR readout (r=0.79). The analytical workflow shows similar turnaround times as regular RT-PCR instrumentation with a quantitative readout of viral proteins corresponding to cycle thresholds (Ct) equivalents ranging from 21 to 34. Using RT-PCR as a reference, we demonstrate that the LC-MS-based method has 100% negative percent agreement (estimated specificity) and 95% positive percent agreement (estimated sensitivity) when analyzing clinical samples collected from asymptomatic individuals with a Ct within the limit of detection of the mass spectrometer (Ct ≤30). These results suggest that a scalable analytical method based on LC-MS has a place in future pandemic preparedness centers to complement current virus detection technologies.

scholarly journals Gynecologic Surgery during the COVID-19 Pandemic: Is Universal Screening Mandatory?

2021 ◽  
Vol 2021 ◽  
pp. 1-5
Author(s):  
Wilasinee Areeruk ◽  
Keerati Chiengthong ◽  
Somsook Santibenchakul ◽  
Shina Oranratanaphan ◽  
Tarinee Manchana
Keyword(s):  
Electronic Medical Records ◽  
Medical Records ◽  
Tertiary Care Hospital ◽  
Tertiary Care ◽  
Healthcare Personnel ◽  
Gynecologic Surgery ◽  
Care Hospital ◽  
Rt Pcr ◽  
Asymptomatic Individuals ◽  
Low Prevalence

This study is aimed at evaluating the results of the universal preoperative screening for COVID-19 in gynecologic cases operated on during its outbreak in a tertiary care hospital in Bangkok, Thailand. A retrospective descriptive study was done on all patients who underwent elective or emergency gynecologic surgeries during the pandemic period in Thailand (April 15 to June 5, 2020). The COVID-19 screening results by symptom-based screening, risk-based screening, and RT-PCR for COVID-19 were collected from the electronic medical records. Among 129 patients who underwent gynecologic surgeries, none had a positive RT-PCR for COVID-19. Symptom-based screening found no patients with positive symptoms for COVID-19. Risk-based screening found 4 patients (3.1%) who were in contact with suspected or confirmed COVID-19 cases and 4 patients (3.1%) who were healthcare personnel. In conclusion, routine preoperative RT-PCR for COVID-19 may need to be reconsidered among asymptomatic individuals in a low-prevalence country during the well-controlled COVID-19 situation. Larger studies are required to ascertain the benefit of universal preoperative COVID-19 testing.

Inactivation of Factor XIa in Vivo: Studies in Chimpanzees and in Humans

1996 ◽  
Vol 76 (04) ◽  
pp. 549-555 ◽  
Author(s):  
Walter A Wuillemin ◽  
C Erik Hack ◽  
Wim K Bleeker ◽  
Bart J Biemond ◽  
Marcel Levi ◽  
...  
Keyword(s):  
Antithrombin Iii ◽  
Life Time ◽  
In Vivo Studies ◽  
Clinical Samples ◽  
Plasma Samples ◽  
C1 Inhibitor ◽  
Elimination Kinetics ◽  
Factor Xia ◽  
Best Parameter

SummaryC1-inhibitor (C1Inh), antithrombin III (ATIII), α1-antitrypsin (a1AT), and α2-antiplasmin (a2AP) are known inhibitors of factor XIa (FXIa). However, their precise contribution to FXIa inactivation in vivo is not known. We investigated FXIa inactivation in chimpanzees and assessed the contribution of these inhibitors to FXIa inactivation in patients with presumed FXI activation.Chimpanzees were infused with FXIa and the various FXIa-FXIa inhibitor complexes formed were measured. Most of FXIa was complexed to C1Inh (68%), followed by a2AP (13%), a1AT (10%), and ATIII (9%). Analysis of the plasma elimination kinetics revealed a half-life time of clearance (t1/2) for the FXIa-FXIa inhibitor complexes of 95 to 104 min, except for FXIa-a1AT, which had a t1/2 of 349 min. Due to this long t1/2, FXIa-a1AT complexes were predicted to show the highest levels in plasma samples from patients with activation of FXI. This was indeed shown in patients with disseminated intravascular coagulation, recent myocardial infarction or unstable angina pectoris. We conclude from this study that in vivo C1Inh is the predominant inhibitor of FXIa, but that FXIa-a1 AT complexes due to their relatively long t1/2 may be the best parameter to assess FXI activation in clinical samples.

scholarly journals Replicative Deformed Wing Virus Found in the Head of Adults from Symptomatic Commercial Bumblebee (Bombus terrestris) Colonies

2021 ◽  
Vol 8 (7) ◽  
pp. 117
Author(s):  
Giovanni Cilia ◽  
Laura Zavatta ◽  
Rosa Ranalli ◽  
Antonio Nanetti ◽  
Laura Bortolotti
Keyword(s):  
Bombus Terrestris ◽  
The Body ◽  
Rt Pcr ◽  
Deformed Wing Virus ◽  
The United Kingdom ◽  
Target Organs ◽  
Specific Organ ◽  
Virus Localization ◽  
Juvenile Stages ◽  
Asymptomatic Individuals

The deformed wing virus (DWV) is one of the most common honey bee pathogens. The virus may also be detected in other insect species, including Bombus terrestris adults from wild and managed colonies. In this study, individuals of all stages, castes, and sexes were sampled from three commercial colonies exhibiting the presence of deformed workers and analysed for the presence of DWV. Adults (deformed individuals, gynes, workers, males) had their head exscinded from the rest of the body and the two parts were analysed separately by RT-PCR. Juvenile stages (pupae, larvae, and eggs) were analysed undissected. All individuals tested positive for replicative DWV, but deformed adults showed a higher number of copies compared to asymptomatic individuals. Moreover, they showed viral infection in their heads. Sequence analysis indicated that the obtained DWV amplicons belonged to a strain isolated in the United Kingdom. Further studies are needed to characterize the specific DWV target organs in the bumblebees. The result of this study indicates the evidence of DWV infection in B. terrestris specimens that could cause wing deformities, suggesting a relationship between the deformities and the virus localization in the head. Further studies are needed to define if a specific organ could be a target in symptomatic bumblebees.

scholarly journals Automated Western immunoblotting detection of anti-SARS-CoV-2 serum antibodies

2021 ◽  
Author(s):  
Sophie Edouard ◽  
Rita Jaafar ◽  
Nicolas Orain ◽  
Philippe Parola ◽  
Philippe Colson ◽  
...  
Keyword(s):  
Negative Control ◽  
Cross Reactivity ◽  
Spike Protein ◽  
Rt Pcr ◽  
Substantial Agreement ◽  
Western Immunoblotting ◽  
Elisa Kit ◽  
Line Test ◽  
Serologic Assay ◽  
Control Samples

AbstractELISA and chemiluminescence serological assays for COVID-19 are currently incorporating only one or two SARS-CoV-2 antigens. We developed an automated Western immunoblotting as a complementary serologic assay for COVID-19. The JessTM Simple Western system, an automated capillary-based assay, was used, incorporating an inactivated SARS-CoV-2 lineage 20a strain as the source of antigen, and total immunoglobulins (IgG, IgM, IgA) detection. In total, 602 sera were tested including 223 from RT-PCR-confirmed COVID-19 patients, 76 from patients diagnosed with seasonal HCoVs and 303 from coronavirus-negative control sera. We also compared this assay with the EUROIMMUN® SARS-CoV-2 IgG ELISA kit. Among 223 sera obtained from RT-PCR-confirmed COVID-19 patients, 180/223 (81%) exhibited reactivity against the nucleocapsid and 70/223 (31%) against the spike protein. Nucleocapsid reactivity was further detected in 9/76 (14%) samples collected from patients diagnosed with seasonal HCoVs and in 15/303 (5%) coronavirus-negative control samples. In the subset of sera collected more than 2 weeks after the onset of symptoms, the sensitivity was 94% and the specificity 93%, the latter value probably reflecting cross-reactivity of SARS-CoV-2 with other coronaviruses. The automated Western immunoblotting presented a substantial agreement (90%) with the compared ELISA (Cohen’s Kappa=0.64). Automated Western immunoblotting may be used as a second line test to monitor exposure of people to HCoVs including SARS-CoV-2.

Sign in / Sign up

Export Citation Format

Share Document