Enhancing the sensitivity of rapid antigen detection test (RADT) of different SARS-CoV-2 variants and lineages using fluorescence-labeled antibodies and a fluorescent meter.

RT-qPCR is considered the gold standard for diagnosis of COVID-19; however, it is laborious, time-consuming, and expensive. RADTs have evolved recently as relatively inexpensive methods to address these shortcomings, but their performance for detecting different SARS-COV-2 variants remains limited. RADT test performance could be enhanced using different antibody labeling and signal detection techniques. Here, we aimed to evaluate the performance of two antigen RADTs for detecting different SARS-CoV-2 variants: (i) the conventional colorimetric RADT (Ab-conjugated with gold beads); and (ii) the new Finecare™ RADT (Ab-coated fluorescent beads). Finecare™ is a meter used for the detection of a fluorescent signal. 187 frozen nasopharyngeal swabs collected in Universal transport (UTM) that are RT-qPCR positive for different SARS-CoV-2 variants were selected, including Alpha (n = 60), Delta (n = 59), and Omicron variants (n = 108). Sixty flu and 60 RSV-positive samples were included as negative controls (total sample number = 347). The conventional RADT showed sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of 62.4% (95%CI: 54-70), 100% (95%CI: 97-100), 100% (95%CI: 100-100), and 58% (95%CI: 49-67), respectively. These measurements were enhanced using the Finecare™ RADT: sensitivity, specificity, PPV, and NPV were 92.6% (95%CI: 89.08-92.3), 96% (95%CI: 96-99.61), 98% (95%CI: 89-92.3), and 85% (95%CI: 96-99.6) respectively. The sensitivity of both RADTs could be greatly underestimated because nasopharyngeal swab samples collected UTM and stored at -80 °C were used. Despite that, our results indicate that the Finecare™ RADT is appropriate for clinical laboratory and community-based surveillance due to its high sensitivity and specificity.

Neutralizing antibodies against SARS-CoV-2 are higher but decline faster in mRNA vaccinees compared to individuals with natural infection.

BACKGROUND: Waning protection against emerging SARS-CoV-2 variants by pre-existing antibodies elicited due to current vaccination or natural infection is a global concern. Whether this is due to the waning of immunity to SARS-COV-2 remains unclear. AIM: We aimed to investigate the dynamics of antibody isotype responses among vaccinated naïve (VN) and naturally infected (NI) individuals. METHODS: We followed up antibody levels in COVID-19 mRNA-vaccinated subjects without prior infection (VN, n = 100) in two phases: phase-I (P-I) at ~ 1.4 and phase-II (P-II) at ~ 5.3 months. Antibody levels were compared to those of unvaccinated and naturally infected subjects (NI, n = 40) at ~ 1.7 (P-1) and 5.2 (P-II) months post-infection. Neutralizing antibodies (NTAb), anti-S-RBD-IgG, -IgM, and anti-S-IgA isotypes were measured. RESULTS: The VN group elicited significantly greater antibody responses (p < 0.001) than the NI group at P-I, except for IgM. In the VN group, a significant waning in antibody response was observed in all isotypes. There was about ~ a 4-fold decline in NTAb levels (p < 0.001), anti-S-RBD-IgG (~5-folds, p < 0.001), anti-S-RBD-IgM (~6-folds, p < 0.001), and anti-S1-IgA (2-folds, p < 0.001). In the NI group, a significant but less steady decline was notable in S-RBD-IgM (~2-folds, p < 0.001), and a much smaller but significant difference in NTAb (<2-folds, p < 0.001) anti-S-RBD IgG (<2-folds, p = 0.005). Unlike the VN group, the NI group mounted a lasting anti-S1-IgA response with no significant decline. Anti-S1-IgA, which were ~ 3 folds higher in VN subjects compared to NI in P-1 (p < 0.001), dropped to almost the same levels, with no significant difference observed between the two groups in P-II. CONCLUSION: While double-dose mRNA vaccination boosted antibody levels, vaccinated individuals' 'boost' was relatively short-lived.

Challenges in Laboratory Diagnosis of the Novel Coronavirus SARS-CoV-2.

The recent outbreak of the Coronavirus disease 2019 (COVID-19) has quickly spread worldwide since its discovery in Wuhan city, China in December 2019. A comprehensive strategy, including surveillance, diagnostics, research, clinical treatment, and development of vaccines, is urgently needed to win the battle against COVID-19. The past three unprecedented outbreaks of emerging human coronavirus infections at the beginning of the 21st century have highlighted the importance of readily available, accurate, and rapid diagnostic technologies to contain emerging and re-emerging pandemics. Real-time reverse transcriptase-polymerase chain reaction (rRT-PCR) based assays performed on respiratory specimens remain the gold standard for COVID-19 diagnostics. However, point-of-care technologies and serologic immunoassays are rapidly emerging with high sensitivity and specificity as well. Even though excellent techniques are available for the diagnosis of symptomatic patients with COVID-19 in well-equipped laboratories; critical gaps still remain in screening asymptomatic people who are in the incubation phase of the virus, as well as in the accurate determination of live viral shedding during convalescence to inform decisions for ending isolation. This review article aims to discuss the currently available laboratory methods and surveillance technologies available for the detection of COVID-19, their performance characteristics and highlight the gaps in current diagnostic capacity, and finally, propose potential solutions. We also summarize the specifications of the majority of the available commercial kits (PCR, EIA, and POC) for laboratory diagnosis of COVID-19.

Effect of multiple freeze-thaw cycles on the detection of anti-SARS-CoV-2 IgG antibodies.

Several studies have investigated the effect of repeated freeze-thaw (F/T) cycles on RNA detection for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). However, no data are available regarding the effect of repeated F/T cycles on SARS-CoV-2 antibody detection in serum. We investigated the effect of multiple F/T cycles on anti-SARS-CoV-2 IgG detection using an ELISA test targeting the nucleocapsid antibodies. Ten positive and 1 negative SARS-CoV-2 IgG sera from 11 participants, in replicates of 5, were subjected to a total of 16 F/T cycles and stored at 4 °C until tested by ELISA. Statistical analysis was performed to test for F/T cycle effect. None of the 10 positive sera became negative after 16 F/T cycles. There was no significant difference in the OD average reading between the first and last F/T cycles, except for one serum with a minimal decline in the OD. The random effect linear regression of log (OD) on the number of cycles showed no significant trend, with a slope consistent with zero (B=-0.0001; 95 % CI -0.0008; 0.0006; P-value=0.781). These results suggest that multiple F/T cycles had no effect on the ability of the ELISA assay to detect SARS-CoV-2 IgG antibodies.

Epidemiology of SARS-CoV2 in Qatar's primary care population aged 10 years and above.

BACKGROUND: There is an urgent need to elucidate the epidemiology of the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV2) and characterize its potential impact. Investing in characterising the SARS-CoV2 will help plan and improve the response to the pandemic. Furthermore, it will help identify the most efficient ways of managing the pandemic, avoiding public health policies and interventions that may be unduly restrictive of normal activity or unnecessarily costly. This paper describes the design and reports findings of a population based epidemiological study undertaken to characterise SARS-CoV2 in Qatar using limited resources in a timely manner. METHODS: Asymptomatic individuals ≥10 years registered with Qatar's publicly funded primary health provider were eligible. A stratified random sampling technique was utilized to identify the study sample. Participants were invited to an appointment where they completed a questionnaire and provided samples for polymerase chain reaction and Immunoglobulin M and G immunoassay tests. Data collected were analyzed to calculate point and period prevalence by sociodemographic, lifestyle and clinical characteristics. RESULTS: Of 18,918 individuals invited for the study, 2084 participated (response rate 10.8%). The overall point prevalence and period prevalence were estimated to be 1.6% (95% CI 1.1-2.2) and 14.6% (95% CI 13.1-16.2) respectively. Period prevalence of SARS-CoV2 infection was not considerably different across age groups (9.7-19.8%). It was higher in males compared to females (16.2 and 12.7% respectively). A significant variation was observed by nationality (7.1 to 22.2%) and municipalities (6.9-35.3%). CONCLUSIONS: The study provides an example of a methodologically robust approach that can be undertaken in a timely manner with limited resources. It reports much-needed epidemiological data about the spread of SARS-CoV2. Given the low prevalence rates, majority of the population in Qatar remains susceptible. Enhanced surveillance must continue to be in place, particularly due to the large number of asymptomatic cases observed. Robust contact tracing and social distancing measures are key to prevent future outbreaks.

A review of novel coronavirus disease (COVID-19): based on genomic structure, phylogeny, current shreds of evidence, candidate vaccines, and drug repurposing.

Coronavirus disease (COVID-19) pandemic is instigated by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). As of March 13, 2021, more than 118.9 million cases were infected with COVID-19 worldwide. SARS-CoV-2 is a positive-sense single-stranded RNA beta-CoV. Most COVID-19 infected individuals recover within 1-3 weeks. Nevertheless, approximately 5% of patients develop acute respiratory distress syndrome and other systemic complications, leading to death. Structural genetic analyses of SARS-CoV-2 have shown genomic resemblances but a low evolutionary correlation to SARS-CoV-1 responsible for the 2002-2004 outbreak. The S glycoprotein is critical for cell adhesion and the entrance of the virus into the host. The process of cell entry uses the cellular receptor named angiotensin-converting enzyme 2. Recent evidence proposed that the CD147 as a SARS-CoV-2's potential receptor. The viral genome is mainly held by two non-structural proteins (NSPs), ORF1a and ORF1ab, along with structural proteins. Although NSPs are conserved among the ßCoVs, mutations in NSP2 and NSP3 may play critical roles in transmitting the virus and cell tropism. To date, no specific/targeted anti-viral treatments exist. Notably, more than 50 COVID-19 candidate vaccines in clinical trials, and a few being administered. Preventive precautions are the primary strategy to limit the viral load transmission and spread, emphasizing the urgent need for developing significant drug targets and vaccines against COVID-19. This review provides a cumulative overview of the genomic structure, transmission, phylogeny of SARS-CoV-2 from Indian clusters, treatment options, updated discoveries, and future standpoints for COVID-19. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-021-02749-0.

Diagnostic Efficiency of Three Fully Automated Serology Assays and Their Correlation with a Novel Surrogate Virus Neutralization Test in Symptomatic and Asymptomatic SARS-COV-2 Individuals.

To support the deployment of serology assays for population screening during the COVID-19 pandemic, we compared the performance of three fully automated SARS-CoV-2 IgG assays: Mindray CL-900i® (target: spike [S] and nucleocapsid [N]), BioMérieux VIDAS®3 (target: receptor-binding domain [RBD]) and Diasorin LIAISON®XL (target: S1 and S2 subunits). A total of 111 SARS-CoV-2 RT-PCR- positive samples collected at ≥ 21 days post symptom onset, and 127 pre-pandemic control samples were included. Diagnostic performance was assessed in correlation to RT-PCR and a surrogate virus-neutralizing test (sVNT). Moreover, cross-reactivity with other viral antibodies was investigated. Compared to RT-PCR, LIAISON®XL showed the highest overall specificity (100%), followed by VIDAS®3 (98.4%) and CL-900i® (95.3%). The highest sensitivity was demonstrated by CL-900i® (90.1%), followed by VIDAS®3 (88.3%) and LIAISON®XL (85.6%). The sensitivity of all assays was higher in symptomatic patients (91.1-98.2%) compared to asymptomatic patients (78.4-80.4%). In correlation to sVNT, all assays showed excellent sensitivities (92.2-96.1%). In addition, VIDAS®3 demonstrated the best correlation (r = 0.75) with the sVNT. The present study provides insights on the performance of three fully automated assays, which could help diagnostic laboratories in the choice of a particular assay according to the intended use.

Severe acute respiratory syndrome coronavirus-2 natural animal reservoirs and experimental models: systematic review.

The current severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) outbreak has been rapidly spreading worldwide, causing serious global concern. The role that animal hosts play in disease transmission is still understudied and researchers wish to find suitable animal models for fundamental research and drug discovery. In this systematic review, we aimed to compile and discuss all articles that describe experimental or natural infections with SARS-CoV-2, from the initial discovery of the virus in December 2019 through to October 2020. We systematically searched four databases (Scopus, PubMed, Science Direct and Web of Science). The following data were extracted from the included studies: type of infection (natural or experimental), age, sample numbers, dose, route of inoculation, viral replication, detection method, clinical symptoms and transmission. Fifty-four studies were included, of which 34 were conducted on animal reservoirs (naturally or experimentally infected), and 20 involved models for testing vaccines and therapeutics. Our search revealed that Rousettus aegyptiacus (fruit bats), pangolins, felines, mink, ferrets and rabbits were all susceptible to SARS-CoV-2, while dogs were weakly susceptible and pigs, poultry, and tree shrews were not. In addition, virus replication in mice, mink, hamsters and ferrets resembled subclinical human infection, so these animals might serve as useful models for future studies to evaluate vaccines or antiviral agents and to study host-pathogen interactions. Our review comprehensively summarized current evidence on SARS-CoV-2 infection in animals and their usefulness as models for studying vaccines and antiviral drugs. Our findings may direct future studies for vaccine development, antiviral drugs and therapeutic agents to manage SARS-CoV-2-caused diseases.

Performance evaluation of five ELISA kits for detecting anti-SARS-COV-2 IgG antibodies.

OBJECTIVES: To evaluate and compare the performances of five commercial ELISA assays (EDI, AnshLabs, Dia.Pro, NovaTec, and Lionex) for detecting anti-SARS-CoV-2 IgG. METHODS: Seventy negative control samples (collected before the COVID-19 pandemic) and samples from 101 RT-PCR-confirmed SARS-CoV-2 patients (collected at different time points from symptom onset: ≤7, 8-14 and >14 days) were used to compare the sensitivity, specificity, agreement, and positive and negative predictive values of each assay with RT-PCR. A concordance assessment between the five assays was also conducted. Cross-reactivity with other HCoV, non-HCoV respiratory viruses, non-respiratory viruses, and nuclear antigens was investigated. RESULTS: Lionex showed the highest specificity (98.6%; 95% CI 92.3-99.8), followed by EDI and Dia.Pro (97.1%; 95% CI 90.2-99.2), NovaTec (85.7%; 95% CI 75.7-92.1), then AnshLabs (75.7%; 95% CI 64.5-84.2). All ELISA kits cross-reacted with one anti-MERS IgG-positive sample, except Lionex. The sensitivity was low during the early stages of the disease but improved over time. After 14 days from symptom onset, Lionex and NovaTec showed the highest sensitivity at 87.9% (95% CI 72.7-95.2) and 86.4% (95% CI 78.5-91.7), respectively. The agreement with RT-PCR results based on Cohen's kappa was as follows: Lionex (0.89) > NovaTec (0.70) > Dia.Pro (0.69) > AnshLabs (0.63) > EDI (0.55). CONCLUSION: The Lionex and NovaLisa IgG ELISA kits, demonstrated the best overall performance.