Commercialized kits to assess T-cell responses against SARS-CoV-2 S peptides. A pilot study in health care workers.

Background: It is crucial to assess the levels of protection generated by natural infection or SARS-CoV-2 vaccines, mainly in individuals professionally exposed and in vulnerable groups. Measuring T-cell responses may complement antibody tests currently in use as correlates of protection. Our aim was to assess the feasibility of a validated assay of T-cell responses. Methods: Twenty health-care-workers (HCW) were included. Antibody test to SARS-CoV-2 N and S-proteins in parallel with a commercially available whole-blood-interferon-gamma-release-assay (IGRA) to S-peptides and two detection methods, CLIA and ELISA were determined. Results: IGRA test detected T-cell responses in naturally exposed and vaccinated HCW already after first vaccination dose. The correlation by the two detection methods was very high (R > 0.8) and sensitivity and specificity ranged between 100 and 86% and 100-73% respectively. Even though there was a very high concordance between specific antibody levels and the IGRA assay in the ability to detect immune response to SARS-CoV-2, there was a relatively low quantitative correlation. In the small group primed by natural infection, one vaccine dose was sufficient to reach immune response plateau. IGRA was positive in one, with Ig(S) antibody negative vaccinated immunosuppressed HCW illustrating another advantage of the IGRA-test. Conclusion: Whole-blood-IGRA-tests amenable to automation and constitutes a promising additional tool for measuring the state of the immune response to SARS-CoV-2; they are applicable to large number of samples and may become a valuable correlate of protection to COVID-19, particularly for vulnerable groups at risk of being re-exposed to infection, as are health-care-workers.

Introducción: Es fundamental evaluar los niveles de protección inmune en infectados o tras la vacunación frente a SARS-CoV-2. La cuantificación de la respuesta inmune celular T puede complementar la determinación de anticuerpos. Evaluamos la viabilidad de un ensayo comercial validado de respuesta celular T específica frente a SARS-CoV-2. Métodos: Se incluyeron veinte trabajadores sanitarios (TS). Medimos anticuerpos contra las proteínas N y S de SARS-CoV-2 y realizamos el ensayo de liberación de interferón-gamma (IFNγ) en sangre completa (IGRA) frente a péptidos de la proteína S. IFNγ se determinó mediante dos métodos de detección: CLIA y ELISA. Resultados: IGRA detectó respuesta celular T en TS tanto infectados como vacunados. La correlación de los dos métodos de detección de IFNγ fue muy alta (R >0,8) y la sensibilidad y la especificidad variaron entre 100 y 86% y 100-73% respectivamente. Hubo una concordancia muy alta entre los niveles de anticuerpos específicos y el ensayo IGRA aunque la correlación cuantitativa fue relativamente baja. En el grupo de infectados, una dosis de vacuna fue suficiente para alcanzar el «plateau¼ de respuesta inmune. IGRA fue claramente positivo en un profesional vacunado inmunosuprimido que presentaba anticuerpos contra la proteína S negativos. Conclusiones: IGRA frente a péptidos de la proteína-S es susceptible de automatización y constituye una herramienta prometedora para medir la respuesta inmune celular frente a SARS-CoV-2; es aplicable a un gran número de muestras y puede servir para valorar la protección, particularmente en los grupos vulnerables en riesgo de volver a exponerse a la infección, como los TS.

Application of newly developed SARS-CoV2 serology test along with real-time PCR for early detection in health care workers and on-time plasma donation.

BACKGROUND: As the daily number of coronavirus infection disease 19 (COVID19) patients increases, the necessity of early diagnosis becomes more obvious. In this respect, we aimed to develop a serological test for specifically detecting anti-SARS-CoV2 antibodies. METHODS: We collected serum and saliva samples from 609 individuals who work at TBZMED affiliated hospitals in Tabriz, Iran, from April to June of 2020. Real-time PCR technique was used to detect SARS-CoV-2 genome using specific primers. An enzyme linked immunosorbent assay (ELISA) test was designed based on virus nucleocapsid (N), spike (S) and its receptor binding domain (RBD) protein, and the collected sera were subjected to IgM and/or IgG analysis. RESULT: Real-time PCR results showed that 66 people were infected with the SARS-CoV-2. Our designed ELISA kit showed 93.75% and 98% of sensitivity and specificity, respectively. In this study, 5.74% of participants had specific IgG against RBD, whereas the percentage for IgM positive individuals was 5.58%. Approximately the same results were observed for S protein. The number of positive participants for NP increased further, and the results of this antigen showed 7.38% for IgG and 7.06% for IgM. CONCLUSION: The ELISA test beside real-time PCR could provide a reliable serologic profile for the status of the disease progress and early detection of individuals. More importantly, it possesses the potential to identify the best candidates for plasma donation according to the antibody titers.

Current state of diagnostic, screening and surveillance testing methods for COVID-19 from an analytical chemistry point of view.

Since December 2019, we have been in the battlefield with a new threat to the humanity known as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In this review, we describe the four main methods used for diagnosis, screening and/or surveillance of SARS-CoV-2: Real-time reverse transcription polymerase chain reaction (RT-PCR); chest computed tomography (CT); and different complementary alternatives developed in order to obtain rapid results, antigen and antibody detection. All of them compare the highlighting advantages and disadvantages from an analytical point of view. The gold standard method in terms of sensitivity and specificity is the RT-PCR. The different modifications propose to make it more rapid and applicable at point of care (POC) are also presented and discussed. CT images are limited to central hospitals. However, being combined with RT-PCR is the most robust and accurate way to confirm COVID-19 infection. Antibody tests, although unable to provide reliable results on the status of the infection, are suitable for carrying out maximum screening of the population in order to know the immune capacity. More recently, antigen tests, less sensitive than RT-PCR, have been authorized to determine in a quicker way whether the patient is infected at the time of analysis and without the need of specific instruments.

Development of in-house ELISAs for the detection of anti-SARS­CoV­2 RBD and N IgG and IgM antibodies in biological samples.

By the end of year 2019, the new virus SARS-CoV-2 appeared, causing the Coronavirus Disease 2019 (COVID-19), and spread very fast globally. A continuing need for diagnostic tools is a must to contain its spread. Till now, the gold standard method, the reverse transcription polymerase chain reaction (RT-PCR), is the precise procedure to detect the virus. However, SARS-CoV-2 may escape RT-PCR detection for several reasons. The development of well-designed, specific and sensitive serological test like enzyme immunoassay (EIA) is needed. This EIA can stand alone or work side by side with RT-PCR. In this study, we developed several EIAs including plates that are coated with either specially designed SARS-CoV-2 nucleocapsid or surface recombinant proteins. Each protein type can separately detect anti-SARS-CoV-2 IgM or IgG antibodies. For each EIAs, the cut-off value, specificity and sensitivity were determined utilizing RT-PCR confirmed Covid-19 and pre-pandemic healthy and other viruses-infected sera. Also, the receiver operator characteristic (ROC) analysis was performed to define the specificities and sensitivities of the optimized assay. The in-house EIAs were validated by comparing against commercial EIA kits. All in-house EIAs showed high specificity (98-99%) and sensitivity (97.8-98.9%) for the detection of IgG/IgM against RBD and N proteins of SARS-CoV-2. From these results, the developed Anti-RBD and anti-N IgG and IgM antibodies EIAs can be used as a specific and sensitive tool to detect SARS-CoV-2 infection, calculate the burden of disease and case fatality rates.

Longitudinal evaluation of the Abbott ARCHITECT SARS-CoV-2 IgM and IgG assays in a pediatric population.

BACKGROUND: Clinical laboratory testing has been an essential part of COVID-19 management. Serology can provide valuable information regarding a patient's exposure to virus, and may have a larger role to play as vaccines becomes available. Limited data is available on the serological response in pediatric patients. Here we investigate the use of one manufacturer's commercial assays for detecting IgM and IgG in an exclusively pediatric population. METHODS: Abbott SARS-CoV-2 IgM and IgG assays were performed on an Abbott ARCHITECT i1000. For specificity studies, we tested 78 patient specimens collected before the COVID-19 pandemic, and 66 specimens from patients who tested negative for SARS-CoV-2 nucleic acid amplification test (NAAT) during the COVID-19 pandemic. For sensitivity we tested 181 specimens from 41 patients with a positive NAAT result. Precision data was acquired for 20 days. RESULTS: For IgM, the highest qualitative positive agreement with molecular results was observed to be 15-30 days after a positive NAAT result or after symptom onset. For IgG, the highest positive agreement was 31-60 days after a positive NAAT result or 61-90 days after the start of symptoms. IgM started to decline 30 days after NAAT results and faded by 90 days. IgG started to decrease 60 days after a positive NAAT result. CONCLUSION: The Abbott IgM and IgG assays have negative agreements of 98.7-100% relative to NAAT results. The IgM and IgG levels assayed by these methods start to decline months after positive molecular results and onset of symptoms in a pediatric population.

Potential compound from herbal food of Rhizoma Polygonati for treatment of COVID-19 analyzed by network pharmacology: Viral and cancer signaling mechanisms.

Rhizoma Polygonati (huangjing in Chinese, ) is a medicine food homology herb used as a component of traditional Chinese medicine treating COVID-19 in the current pandemic emergency in China but the mechanisms remain elusive. Here using TCMSP and Swiss Target Prediction databases to sort out the potential targets of the main chemical components and GenCLiP3, NCBI, and GeneCard databases to search for COVID-19 related targets, the chemical compound-target-pathway network was analyzed. Each component was molecularly docked with host cell target angiotensin converting enzyme II, SARS-CoV-2 targets Spike protein, RNA-dependent RNA polymerase, or 3CL hydrolase. Our results showed a higher affinity of the compound diosgenin and (+)-Syringaresinol-O-beta-D-glucoside binding to the three SARS-CoV-2 proteins compared to the other compounds tested. Thus, our data suggest that potential compounds in Rhizoma Polygonati may act on different targets with viral and cancer related signaling and have a great potential in treatment of COVID-19.

Natural medicinal plant products as an immune-boosters: A possible role to lessen the impact of Covid-19.

Transmissible Covid-19, caused by novel corona virus since last of 2019 has outspread widely until now. Where, India was the second most affected country and 3rd in mortality rate. In world ancient history, medicinal plants were played a crucial role to cure several diseases. In present study, we show some novel natural medicinal plant metabolites as the potential inhibitors against papain-like protease (PLpro), main protease (Mpro) and RNA-dependent RNA polymerase (RdRp), transmembrane proteinase Serine 2 (TMPRSS2) and angiotensin converting enzyme-2 (ACE-2) of Covid-19. Plant metabolites were having been proven to inhibit SARS-CoVs, which also actively walkable against Covid-19.