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1.
Front Immunol ; 13: 1020159, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36248791

RESUMO

The emergency of new SARS-CoV-2 variants that feature increased immune escape marks an urgent demand for better vaccines that will provide broader immunogenicity. Here, we evaluated the immunogenic capacity of vaccine candidates based on the recombinant trimeric spike protein (S) of different SARS-CoV-2 variants of concern (VOC), including the ancestral Wuhan, Beta and Delta viruses. In particular, we assessed formulations containing either single or combined S protein variants. Our study shows that the formulation containing the single S protein from the ancestral Wuhan virus at a concentration of 2µg (SW2-Vac 2µg) displayed in the mouse model the highest IgG antibody levels against all the three (Wuhan, Beta, and Delta) SARS-CoV-2 S protein variants tested. In addition, this formulation induced significantly higher neutralizing antibody titers against the three viral variants when compared with authorized Gam-COVID-Vac-rAd26/rAd5 (Sputnik V) or ChAdOx1 (AstraZeneca) vaccines. SW2-Vac 2µg was also able to induce IFN-gamma and IL-17, memory CD4 populations and follicular T cells. Used as a booster dose for schedules performed with different authorized vaccines, SW2-Vac 2µg vaccine candidate also induced higher levels of total IgG and IgG isotypes against S protein from different SARS-CoV-2 variants in comparison with those observed with homologous 3-dose schedule of Sputnik V or AstraZeneca. Moreover, SW2-Vac 2µg booster induced broadly strong neutralizing antibody levels against the three tested SARS-CoV-2 variants. SW2-Vac 2µg booster also induced CD4+ central memory, CD4+ effector and CD8+ populations. Overall, the results demonstrate that SW2-Vac 2 µg is a promising formulation for the development of a next generation COVID-19 vaccine.


Assuntos
Vacinas contra COVID-19 , COVID-19 , Animais , Anticorpos Neutralizantes , Anticorpos Antivirais , COVID-19/prevenção & controle , Humanos , Imunoglobulina G , Interleucina-17 , Camundongos , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus/genética
2.
Front Immunol ; 13: 992370, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36225925

RESUMO

The COVID-19 pandemic has particularly affected older adults residing in nursing homes, resulting in high rates of hospitalisation and death. Here, we evaluated the longitudinal humoral response and neutralising capacity in plasma samples of volunteers vaccinated with different platforms (Sputnik V, BBIBP-CorV, and AZD1222). A cohort of 851 participants, mean age 83 (60-103 years), from the province of Buenos Aires, Argentina were included. Sequential plasma samples were taken at different time points after vaccination. After completing the vaccination schedule, infection-naïve volunteers who received either Sputnik V or AZD1222 exhibited significantly higher specific anti-Spike IgG titers than those who received BBIBP-CorV. Strong correlation between anti-Spike IgG titers and neutralising activity levels was evidenced at all times studied (rho=0.7 a 0.9). Previous exposure to SARS-CoV-2 and age <80 years were both associated with higher specific antibody levels. No differences in neutralising capacity were observed for the infection-naïve participants in either gender or age group. Similar to anti-Spike IgG titers, neutralising capacity decreased 3 to 9-fold at 6 months after initial vaccination for all platforms. Neutralising capacity against Omicron was between 10-58 fold lower compared to ancestral B.1 for all vaccine platforms at 21 days post dose 2 and 180 days post dose 1. This work provides evidence about the humoral response and neutralising capacity elicited by vaccination of a vulnerable elderly population. This data could be useful for pandemic management in defining public health policies, highlighting the need to apply reinforcements after a complete vaccination schedule.


Assuntos
COVID-19 , Idoso , Idoso de 80 Anos ou mais , Anticorpos Antivirais , Argentina/epidemiologia , COVID-19/epidemiologia , COVID-19/prevenção & controle , ChAdOx1 nCoV-19 , Humanos , Imunoglobulina G , Pandemias , SARS-CoV-2 , Vacinação
3.
Front Immunol ; 13: 909995, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36263055

RESUMO

Background: Recent studies have shown the presence of SARS-CoV-2-specific antibodies in the milk of breastfeeding mothers vaccinated with mRNA and convalescent. However, limited information is available in lactating women receiving other vaccine platforms used in developing countries, such as the inactivated SARS-CoV-2 vaccine BBIBP-CorV (Sinopharm) and the non-replicating adenovirus vaccines Sputnik V (Gamaleya Institute) and ChAdOx1-S (Oxford AstraZeneca). Methods: Here, we evaluated anti-SARS-CoV-2 IgG and IgA levels in both serum and milk samples using a longitudinal and a cross-sectional cohort of 208 breastfeeding vaccinated women from Argentina with or without previous SARS-CoV-2 infection. Results: The analysis showed that IgA levels remain constant in serum and milk of breastfeeding mothers between the first and second doses of vector-based vaccines (Sputnik V and ChAdOx1-S). After the second dose, anti-spike IgA was found positive in 100% of the serum samples and in 66% of breastmilk samples. In addition, no significant differences in milk IgA levels were observed in participants receiving BBIBP-CorV, Sputnik V or ChAdOx1-S. IgG levels in milk increased after the second dose of vector-based vaccines. Paired longitudinal samples taken at 45 and 120 days after the second dose showed a decrease in milk IgG levels over time. Study of IgA levels in serum and milk of vaccinated naïve of infection and vaccinated-convalescent breastfeeding participants showed significantly higher levels in vaccinated-convalescent than in participants without previous infection. Conclusion: This study is relevant to understand the protection against SARS-CoV-2 by passive immunity in newborns and children who are not yet eligible to receive vaccination.


Assuntos
Vacinas contra Adenovirus , COVID-19 , Vacinas Virais , Recém-Nascido , Criança , Humanos , Feminino , Vacinas contra COVID-19 , SARS-CoV-2 , Leite Humano , Estudos Transversais , Lactação , COVID-19/prevenção & controle , Anticorpos Antivirais , Imunoglobulina G , Imunoglobulina A , RNA Mensageiro
4.
Cell Rep Med ; 3(8): 100706, 2022 08 16.
Artigo em Inglês | MEDLINE | ID: mdl-35926505

RESUMO

Heterologous vaccination against coronavirus disease 2019 (COVID-19) provides a rational strategy to rapidly increase vaccination coverage in many regions of the world. Although data regarding messenger RNA (mRNA) and ChAdOx1 vaccine combinations are available, there is limited information about the combination of these platforms with other vaccines widely used in developing countries, such as BBIBP-CorV and Sputnik V. Here, we assess the immunogenicity and reactogenicity of 15 vaccine combinations in 1,314 participants. We evaluate immunoglobulin G (IgG) anti-spike response and virus neutralizing titers and observe that a number of heterologous vaccine combinations are equivalent or superior to homologous schemes. For all cohorts in this study, the highest antibody response is induced by mRNA-1273 as the second dose. No serious adverse events are detected in any of the schedules analyzed. Our observations provide rational support for the use of different vaccine combinations to achieve wide vaccine coverage in the shortest possible time.


Assuntos
COVID-19 , Vacinas Virais , Vacina de mRNA-1273 contra 2019-nCoV , Anticorpos Antivirais , COVID-19/prevenção & controle , Humanos , Imunização , RNA Mensageiro/genética , SARS-CoV-2 , Vacinação
7.
J Infect Dis ; 226(10): 1717-1720, 2022 Nov 11.
Artigo em Inglês | MEDLINE | ID: mdl-35723970

RESUMO

Infection with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variant is usually asymptomatic or mild and appears to be poorly immunogenic at least in unvaccinated individuals. Here, we found that health care workers vaccinated with 2 doses of Sputnik V and a booster dose of ChAdOx1 mount a vigorous neutralizing-antibody response after Omicron breakthrough infection.


Assuntos
Formação de Anticorpos , COVID-19 , Humanos , SARS-CoV-2 , COVID-19/prevenção & controle , Vacinação , Anticorpos Neutralizantes , Anticorpos Antivirais
8.
Nucleic Acids Res ; 50(12): 6968-6979, 2022 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-35736223

RESUMO

The non-structural protein 3 helicase (NS3h) is a multifunctional protein that is critical in RNA replication and other stages in the flavivirus life cycle. NS3h uses energy from ATP hydrolysis to translocate along single stranded nucleic acid and to unwind double stranded RNA. Here we present a detailed mechanistic analysis of the product release stage in the catalytic cycle of the dengue virus (DENV) NS3h. This study is based on a combined experimental and computational approach of product-inhibition studies and free energy calculations. Our results support a model in which the catalytic cycle of ATP hydrolysis proceeds through an ordered sequential mechanism that includes a ternary complex intermediate (NS3h-Pi-ADP), which evolves releasing the first product, phosphate (Pi), and subsequently ADP. Our results indicate that in the product release stage of the DENV NS3h a novel open-loop conformation plays an important role that may be conserved in NS3 proteins of other flaviviruses as well.

9.
Front Vet Sci ; 9: 790058, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35310416

RESUMO

SARS-CoV-2 reverse zoonosis, particularly to domestic animals, and the potential role of infected animals in perpetuating the spread of the virus is an issue of increasing concern. In this case report, we identified the natural infection of two cats by SARS-CoV-2, in Argentina, whose owner had been previously infected by SARS-CoV-2. Viral genetic material was detected in feline oropharyngeal (OP) and rectal (R) swab by RT-qPCR, and sequence analysis revealed that the virus infecting the owner and one cat were genetically similar. The alpha variant (B.1.1.7 lineage) was identified with a unique additional mutation, strongly suggesting human-to-cat route of transmission. This study reinforces the One Health concept and the importance of integrating human, animal, and environmental perspectives to promptly address relevant health issues.

11.
Methods Mol Biol ; 2409: 99-117, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-34709638

RESUMO

It has become increasingly evident that unveiling the mechanisms of virus entry, assembly, and virion release is fundamental for identifying means for preventing viral spread and controlling viral disease. Due to virus mobility and structural and/or functional heterogeneity among viral particles, high spatiotemporal resolution single-virus/single-particle techniques are required to capture the behavior of viral particles inside infected cells.In this chapter, we present fluorescence imaging analysis methods for studying the mobility of fluorescently labeled dengue virus (DENV) proteins in live infected cells. Some of the most recent Fluorescence Fluctuation Spectroscopy (FFS) methods will be presented and, in particular, the pair Correlation Functions (pCF) approach will be discussed. The pCF method does not require individual molecule isolation, as in a particle-tracking experiment, to capture single viral protein behavior. In this regard, image acquisition is followed by the spatiotemporal cross-correlation function at increasing time delays, yielding a quantitative view of single-particle mobility in intact live infected cells.We provide a general overview and a practical guidance for the implementation of advanced FFS techniques, and the pair Correlation Functions analysis, as quantitative tools to reveal insights into previously unreported DENV mechanisms. We expect this protocol report will serve as an incentive for further applying correlation imaging studies in virology research.


Assuntos
Vírus da Dengue , Dengue , Capsídeo , Proteínas do Capsídeo , Humanos , Vírion , Internalização do Vírus
12.
Sci Rep ; 11(1): 24415, 2021 12 24.
Artigo em Inglês | MEDLINE | ID: mdl-34952906

RESUMO

Flaviviruses are major human disease-causing pathogens, including dengue virus (DENV), Zika virus, yellow fever virus and others. DENV infects hundreds of millions of people per year around the world, causing a tremendous social and economic burden. DENV capsid (C) protein plays an essential role during genome encapsidation and viral particle formation. It has been previously shown that DENV C enters the nucleus in infected cells. However, whether DENV C protein exhibits nuclear export remains unclear. By spatially cross-correlating different regions of the cell, we investigated DENV C movement across the nuclear envelope during the infection cycle. We observed that transport takes place in both directions and with similar translocation times (in the ms time scale) suggesting a bidirectional movement of both C protein import and export.Furthermore, from the pair cross-correlation functions in cytoplasmic or nuclear regions we found two populations of C molecules in each compartment with fast and slow mobilities. While in the cytoplasm the correlation times were in the 2-6 and 40-110 ms range for the fast and slow mobility populations respectively, in the cell nucleus they were 1-10 and 25-140 ms range, respectively. The fast mobility of DENV C in cytoplasmic and nuclear regions agreed with the diffusion coefficients from Brownian motion previously reported from correlation analysis. These studies provide the first evidence of DENV C shuttling from and to the nucleus in infected cells, opening new venues for antiviral interventions.


Assuntos
Proteínas do Capsídeo/ultraestrutura , Vírus da Dengue/ultraestrutura , Dengue/virologia , Transporte Ativo do Núcleo Celular , Animais , Linhagem Celular , Cricetinae
13.
J Biol Chem ; 297(3): 101059, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34375636

RESUMO

Dengue virus (DENV) and Zika virus (ZIKV) capsid proteins efficiently recruit and surround the viral RNA at the endoplasmic reticulum (ER) membrane to yield nascent viral particles. However, little is known either about the molecular mechanisms by which multiple copies of capsid proteins assemble into nucleocapsids (NCs) or how the NC is recruited and wrapped by the ER membrane during particle morphogenesis. Here, we measured relevant interactions concerning this viral process using purified DENV and ZIKV capsid proteins, membranes mimicking the ER lipid composition, and nucleic acids in in vitro conditions to understand the biophysical properties of the RNA genome encapsidation process. We found that both ZIKV and DENV capsid proteins bound to liposomes at liquid-disordered phase regions, docked exogenous membranes, and RNA molecules. Liquid-liquid phase separation is prone to occur when positively charged proteins interact with nucleic acids, which is indeed the case for the studied capsids. We characterized these liquid condensates by measuring nucleic acid partition constants and the extent of water dipolar relaxation, observing a cooperative process for the formation of the new phase that involves a distinct water organization. Our data support a new model in which capsid-RNA complexes directly bind the ER membrane, seeding the process of RNA recruitment for viral particle assembly. These results contribute to our understanding of the viral NC formation as a stable liquid-liquid phase transition, which could be relevant for dengue and Zika gemmation, opening new avenues for antiviral intervention.


Assuntos
Proteínas do Capsídeo/metabolismo , Vírus da Dengue/metabolismo , Dengue/virologia , Membranas Intracelulares/virologia , Bicamadas Lipídicas/metabolismo , RNA Viral/metabolismo , Infecção por Zika virus/virologia , Zika virus/metabolismo , Capsídeo/metabolismo , Proteínas do Capsídeo/genética , Vírus da Dengue/genética , Retículo Endoplasmático/virologia , Humanos , Lipossomos , RNA Viral/genética , Zika virus/genética
14.
Cell Rep Med ; 2(8): 100359, 2021 08 17.
Artigo em Inglês | MEDLINE | ID: mdl-34308389

RESUMO

Massive vaccination offers great promise for halting the global COVID-19 pandemic. However, the limited supply and uneven vaccine distribution create an urgent need to optimize vaccination strategies. We evaluate SARS-CoV-2-specific antibody responses after Sputnik V vaccination of healthcare workers in Argentina, measuring IgG anti-spike titers and neutralizing capacity after one and two doses in a cohort of naive or previously infected volunteers. By 21 days after receiving the first dose of the vaccine, 94% of naive participants develop spike-specific IgG antibodies. A single Sputnik V dose elicits higher antibody levels and virus-neutralizing capacity in previously infected individuals than in naive ones receiving the full two-dose schedule. The high seroconversion rate after a single dose in naive participants suggests a benefit of delaying administration of the second dose to increase the number of people vaccinated. The data presented provide information for guiding public health decisions in light of the current global health emergency.


Assuntos
Vacinas contra COVID-19/imunologia , COVID-19/prevenção & controle , SARS-CoV-2/imunologia , Vacinas Sintéticas/imunologia , Animais , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Argentina/epidemiologia , COVID-19/imunologia , Chlorocebus aethiops , Células HEK293 , Pessoal de Saúde , Humanos , Pandemias , SARS-CoV-2/patogenicidade , Soroconversão , Glicoproteína da Espícula de Coronavírus/imunologia , Vacinação , Vacinas , Células Vero
15.
Medicina (B.Aires) ; 81(2): 135-142, June 2021. graf
Artigo em Inglês | LILACS | ID: biblio-1287262

RESUMO

Abstract Most countries in Latin America have already reported thousands of confirmed cases and vulnerable populations are the most affected by the coronavirus disease 2019 (COVID-19) pandemic. Preventive measures such as hygiene, social distancing, and isolation, essential to stop the spread of coronavirus, are difficult to accomplish for vulnerable populations due to their living conditions. Seroepidemiological surveys are assets to measure the transmission for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Until July 1st, the incidence rate of SARS-CoV-2 infection in Barrio Padre Mugica, one of the largest slums in Buenos Aires City, was 5.9%. This study aimed to establish the prevalence of SARS-CoV-2 antibodies immunoglobulin G (IgG) immediately after the outbreak, and to identify neighbourhood, household and individual factors associated with seroconversion. The prevalence based on IgG was 53.4% (95% CI 52.8% to 54.1%). For each polymerase chain reaction (RT-qPCR) confirmed case, nine people tested IgG positive, indicating a high rate of undetected (probably asymptomatic) infections. Hence, the high rate of undiagnosed people suggests that clinical criteria and epidemiological nexus should be considered. The high seroprevalence observed in the context of an intense epidemic in a vulnerable area might serve as a reference to other countries. This study contributes to future decision making by understanding population immunity against SARS-CoV2 and its relation to living conditions and foccus that comprehensive biosocial, household-level interventions are needed.


Resumen Muchos países de América Latina han informado miles de casos confirmados y las poblaciones vulnerables son las más afectadas por la pandemia de la enfermedad por coronavirus 2019 (COVID-19). Las medidas preventivas como la higiene, el distanciamiento social y el aislamiento, fundamentales para frenar la propagación del coronavirus, son difíciles de lograr en estas poblaciones debido a sus condiciones de vida. Los estudios seroepidemiológicos son de gran utilidad para medir la transmisión del síndrome respiratorio agudo severo coronavirus 2 (SARS-CoV-2). Hasta el 1 de julio, la tasa de incidencia de la infección por SARS-CoV-2 en el Barrio Padre Mugica, uno de los barrios marginales más grandes de la ciudad de Buenos Aires, era del 5.9%. Este estudio tuvo como objetivo estimar la prevalencia de anticuerpos inmunoglobulina G (IgG) para SARS-CoV-2 inmediatamente después del brote, e identificar factores del barrio, hogar e individuales asociados con la seroconversión. La prevalencia basada en IgG fue del 53.4% (IC del 95%: 52.8% a 54.1%). Para cada caso confirmado por reacción en cadena de la polimerasa (RT-qPCR), nueve personas dieron positivo en IgG, lo que indica una alta tasa de infecciones no detectadas y probablemente asintomáticas. La alta tasa de personas no diagnosticadas sugiere que se deben considerar los criterios clínicos y el nexo epidemiológico. La alta seroprevalencia observada en el contexto de una intensa epidemia en una zona vulnerable podría servir de referencia a otros países. Este estudio contribuye a la toma de decisiones futuras al comprender la inmunidad de la población contra el SARS-CoV2 en su relación con las condiciones de vida y por su enfoque en la necesidad de intervenciones integrales a nivel del hogar.


Assuntos
Humanos , Áreas de Pobreza , COVID-19 , RNA Viral , Estudos Soroepidemiológicos , SARS-CoV-2 , Anticorpos Antivirais
16.
Medicina (B Aires) ; 81(2): 135-142, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33906129

RESUMO

Most countries in Latin America have already reported thousands of confirmed cases and vulnerable populations are the most affected by the coronavirus disease 2019 (COVID-19) pandemic. Preventive measures such as hygiene, social distancing, and isolation, essential to stop the spread of coronavirus, are difficult to accomplish for vulnerable populations due to their living conditions. Seroepidemiological surveys are assets to measure the transmission for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Until July 1st, the incidence rate of SARS-CoV-2 infection in Barrio Padre Mugica, one of the largest slums in Buenos Aires City, was 5.9%. This study aimed to establish the prevalence of SARS-CoV-2 antibodies immunoglobulin G (IgG) immediately after the outbreak, and to identify neighbourhood, household and individual factors associated with seroconversion. The prevalence based on IgG was 53.4% (95% CI 52.8% to 54.1%). For each polymerase chain reaction (RT-qPCR) confirmed case, nine people tested IgG positive, indicating a high rate of undetected (probably asymptomatic) infections. Hence, the high rate of undiagnosed people suggests that clinical criteria and epidemiological nexus should be considered. The high seroprevalence observed in the context of an intense epidemic in a vulnerable area might serve as a reference to other countries. This study contributes to future decision making by understanding population immunity against SARS-CoV2 and its relation to living conditions and foccus that comprehensive biosocial, household-level interventions are needed.


Muchos países de América Latina han informado miles de casos confirmados y las poblaciones vulnerables son las más afectadas por la pandemia de la enfermedad por coronavirus 2019 (COVID-19). Las medidas preventivas como la higiene, el distanciamiento social y el aislamiento, fundamentales para frenar la propagación del coronavirus, son difíciles de lograr en estas poblaciones debido a sus condiciones de vida. Los estudios seroepidemiológicos son de gran utilidad para medir la transmisión del síndrome respiratorio agudo severo coronavirus 2 (SARS-CoV-2). Hasta el 1 de julio, la tasa de incidencia de la infección por SARS-CoV-2 en el Barrio Padre Mugica, uno de los barrios marginales más grandes de la ciudad de Buenos Aires, era del 5.9%. Este estudio tuvo como objetivo estimar la prevalencia de anticuerpos inmunoglobulina G (IgG) para SARS-CoV-2 inmediatamente después del brote, e identificar factores del barrio, hogar e individuales asociados con la seroconversión. La prevalencia basada en IgG fue del 53.4% (IC del 95%: 52.8% a 54.1%). Para cada caso confirmado por reacción en cadena de la polimerasa (RT-qPCR), nueve personas dieron positivo en IgG, lo que indica una alta tasa de infecciones no detectadas y probablemente asintomáticas. La alta tasa de personas no diagnosticadas sugiere que se deben considerar los criterios clínicos y el nexo epidemiológico. La alta seroprevalencia observada en el contexto de una intensa epidemia en una zona vulnerable podría servir de referencia a otros países. Este estudio contribuye a la toma de decisiones futuras al comprender la inmunidad de la población contra el SARS-CoV2 en su relación con las condiciones de vida y por su enfoque en la necesidad de intervenciones integrales a nivel del hogar.


Assuntos
COVID-19 , Áreas de Pobreza , Anticorpos Antivirais , Humanos , RNA Viral , SARS-CoV-2 , Estudos Soroepidemiológicos
17.
mBio ; 12(1)2021 02 16.
Artigo em Inglês | MEDLINE | ID: mdl-33593976

RESUMO

The global coronavirus disease 2019 (COVID-19) pandemic has mobilized efforts to develop vaccines and antibody-based therapeutics, including convalescent-phase plasma therapy, that inhibit viral entry by inducing or transferring neutralizing antibodies (nAbs) against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike glycoprotein (CoV2-S). However, rigorous efficacy testing requires extensive screening with live virus under onerous biosafety level 3 (BSL3) conditions, which limits high-throughput screening of patient and vaccine sera. Myriad BSL2-compatible surrogate virus neutralization assays (VNAs) have been developed to overcome this barrier. Yet, there is marked variability between VNAs and how their results are presented, making intergroup comparisons difficult. To address these limitations, we developed a standardized VNA using CoV2-S pseudotyped particles (CoV2pp) based on vesicular stomatitis virus bearing the Renilla luciferase gene in place of its G glycoprotein (VSVΔG); this assay can be robustly produced at scale and generate accurate neutralizing titers within 18 h postinfection. Our standardized CoV2pp VNA showed a strong positive correlation with CoV2-S enzyme-linked immunosorbent assay (ELISA) results and live-virus neutralizations in confirmed convalescent-patient sera. Three independent groups subsequently validated our standardized CoV2pp VNA (n > 120). Our data (i) show that absolute 50% inhibitory concentration (absIC50), absIC80, and absIC90 values can be legitimately compared across diverse cohorts, (ii) highlight the substantial but consistent variability in neutralization potency across these cohorts, and (iii) support the use of the absIC80 as a more meaningful metric for assessing the neutralization potency of a vaccine or convalescent-phase sera. Lastly, we used our CoV2pp in a screen to identify ultrapermissive 293T clones that stably express ACE2 or ACE2 plus TMPRSS2. When these are used in combination with our CoV2pp, we can produce CoV2pp sufficient for 150,000 standardized VNAs/week.IMPORTANCE Vaccines and antibody-based therapeutics like convalescent-phase plasma therapy are premised upon inducing or transferring neutralizing antibodies that inhibit SARS-CoV-2 entry into cells. Virus neutralization assays (VNAs) for measuring neutralizing antibody titers (NATs) are an essential part of determining vaccine or therapeutic efficacy. However, such efficacy testing is limited by the inherent dangers of working with the live virus, which requires specialized high-level biocontainment facilities. We therefore developed a standardized replication-defective pseudotyped particle system that mimics the entry of live SARS-CoV-2. This tool allows for the safe and efficient measurement of NATs, determination of other forms of entry inhibition, and thorough investigation of virus entry mechanisms. Four independent labs across the globe validated our standardized VNA using diverse cohorts. We argue that a standardized and scalable assay is necessary for meaningful comparisons of the myriad of vaccines and antibody-based therapeutics becoming available. Our data provide generalizable metrics for assessing their efficacy.


Assuntos
COVID-19/diagnóstico , COVID-19/imunologia , SARS-CoV-2/imunologia , SARS-CoV-2/patogenicidade , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/metabolismo , Anticorpos Antivirais/metabolismo , Ensaio de Imunoadsorção Enzimática , Humanos , Testes de Neutralização
18.
PLoS Pathog ; 17(1): e1009161, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33444413

RESUMO

We report the emergency development and application of a robust serologic test to evaluate acute and convalescent antibody responses to SARS-CoV-2 in Argentina. The assays, COVIDAR IgG and IgM, which were produced and provided for free to health authorities, private and public health institutions and nursing homes, use a combination of a trimer stabilized spike protein and the receptor binding domain (RBD) in a single enzyme-linked immunosorbent assay (ELISA) plate. Over half million tests have already been distributed to detect and quantify antibodies for multiple purposes, including assessment of immune responses in hospitalized patients and large seroprevalence studies in neighborhoods, slums and health care workers, which resulted in a powerful tool for asymptomatic detection and policy making in the country. Analysis of antibody levels and longitudinal studies of symptomatic and asymptomatic SARS-CoV-2 infections in over one thousand patient samples provided insightful information about IgM and IgG seroconversion time and kinetics, and IgM waning profiles. At least 35% of patients showed seroconversion within 7 days, and 95% within 45 days of symptoms onset, with simultaneous or close sequential IgM and IgG detection. Longitudinal studies of asymptomatic cases showed a wide range of antibody responses with median levels below those observed in symptomatic patients. Regarding convalescent plasma applications, a protocol was standardized for the assessment of end point IgG antibody titers with COVIDAR with more than 500 plasma donors. The protocol showed a positive correlation with neutralizing antibody titers, and was used for clinical trials and therapies across the country. Using this protocol, about 80% of convalescent donor plasmas were potentially suitable for therapies. Here, we demonstrate the importance of providing a robust and specific serologic assay for generating new information about antibody kinetics in infected individuals and mitigation policies to cope with pandemic needs.


Assuntos
COVID-19/virologia , SARS-CoV-2/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Adulto , Idoso , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Formação de Anticorpos , Argentina/epidemiologia , COVID-19/epidemiologia , Ensaio de Imunoadsorção Enzimática/métodos , Feminino , Humanos , Imunoglobulina G/imunologia , Imunoglobulina M/imunologia , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Pandemias , SARS-CoV-2/isolamento & purificação , Estudos Soroepidemiológicos
19.
N Engl J Med ; 384(7): 619-629, 2021 02 18.
Artigo em Inglês | MEDLINE | ID: mdl-33232588

RESUMO

BACKGROUND: Convalescent plasma is frequently administered to patients with Covid-19 and has been reported, largely on the basis of observational data, to improve clinical outcomes. Minimal data are available from adequately powered randomized, controlled trials. METHODS: We randomly assigned hospitalized adult patients with severe Covid-19 pneumonia in a 2:1 ratio to receive convalescent plasma or placebo. The primary outcome was the patient's clinical status 30 days after the intervention, as measured on a six-point ordinal scale ranging from total recovery to death. RESULTS: A total of 228 patients were assigned to receive convalescent plasma and 105 to receive placebo. The median time from the onset of symptoms to enrollment in the trial was 8 days (interquartile range, 5 to 10), and hypoxemia was the most frequent severity criterion for enrollment. The infused convalescent plasma had a median titer of 1:3200 of total SARS-CoV-2 antibodies (interquartile range, 1:800 to 1:3200). No patients were lost to follow-up. At day 30 day, no significant difference was noted between the convalescent plasma group and the placebo group in the distribution of clinical outcomes according to the ordinal scale (odds ratio, 0.83; 95% confidence interval [CI], 0.52 to 1.35; P = 0.46). Overall mortality was 10.96% in the convalescent plasma group and 11.43% in the placebo group, for a risk difference of -0.46 percentage points (95% CI, -7.8 to 6.8). Total SARS-CoV-2 antibody titers tended to be higher in the convalescent plasma group at day 2 after the intervention. Adverse events and serious adverse events were similar in the two groups. CONCLUSIONS: No significant differences were observed in clinical status or overall mortality between patients treated with convalescent plasma and those who received placebo. (PlasmAr ClinicalTrials.gov number, NCT04383535.).


Assuntos
Anticorpos Neutralizantes/sangue , COVID-19/terapia , Imunoglobulina G/sangue , Pneumonia Viral/terapia , SARS-CoV-2/imunologia , Idoso , Idoso de 80 Anos ou mais , Transfusão de Componentes Sanguíneos , COVID-19/complicações , COVID-19/mortalidade , Progressão da Doença , Método Duplo-Cego , Feminino , Hospitalização , Humanos , Imunização Passiva , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Pneumonia Viral/etiologia , Pneumonia Viral/mortalidade , Índice de Gravidade de Doença
20.
Rev. argent. salud publica ; 12(supl.1): 14-14, oct. 2020. graf
Artigo em Espanhol | LILACS-Express | LILACS | ID: biblio-1155725

RESUMO

INTRODUCCIÓN en el contexto de pandemia por el nuevo coronavirus (COVID-19), la situación del personal de salud (PS) constituye un foco de interés, tanto por su alta exposición como por la posibilidad de convertirse en diseminadores de la infección en la comunidad. Estos trabajadores enfrentan un riesgo laboral de morbimortalidad sin precedentes. El objetivo fue estimar la seroprevalencia de infección por el nuevo coronavirus (SARS-CoV-2) en el PS de la Región Sanitaria VIII, provincia de Buenos Aires, durante junio de 2020. MÉTODOS se utilizó un diseño transversal. Se realizó un muestreo probabilístico por conglomerados bietápico. Se recabaron datos a partir de un cuestionario autoadministrado y una muestra de sangre para determinación de anticuerpos. Se utilizó el test COVIDAR IgG e IgM®. RESULTADOS se incluyeron 738 trabajadores de la salud, la tasa de respuesta general fue del 73,80 %. El 71,83% fueron mujeres, el 46,39% tenía entre 35 y 49 años. Enfermeros y médicos representaron más de la mitad del personal. El 75,86% refirió usar siempre el equipo de protección personal. El 5,61% tuvo contacto estrecho con un caso confirmado de COVID-19. El 4,60% tenía un hisopado nasofaríngeo previo, con resultado negativo. Se encontraron cinco trabajadores con IgG positiva para SARS-CoV-2 (cuatro mujeres y un varón) e IgM negativa. La edad media de los casos fue de 35 años, dos fueron asintomáticos, en ninguno se había tomado muestra de hisopado. La seroprevalencia general fue de 0,75%, sin diferencias significativas entre estratos. DISCUSIÓN la seroprevalencia hallada fue baja, con una gran proporción de trabajadores susceptibles a la infección. Se refuerza la necesidad de complementar las estrategias de vigilancia epidemiológica pasiva con el monitoreo serológico en personal de salud.


INTRODUCTION in the context of the new coronavirus (COVID-19) pandemic, the situation of health care workers (HCW) constitutes a focus of interest, due to their high exposure and the possibility of becoming disseminators of the infection in the community These workers face an unprecedented occupational risk of morbidity and mortality The aim of this study was to estimate the seroprevalence of the new coronavirus (SARS-CoV-2) infection in health workers of the Sanitary Region VIII, at province of Buenos Aires during June 2020. METHODS a cross-sectional design was used. A probabilistic sampling by two-stage conglomerates was carried out. Data were collected from a self-administered questionnaire and a blood sample for antibody identification. The COVIDAR IgG and IgM® test was used. RESULTS 738 health workers were included; the overall response rate was 73.80%. 71.83% of that were women; 46.39% were between 35 and 49 years of age. Nurses and physicians accounted for more than half of the staff. 75.86% of people claimed to always use personal protective equipment. 5.61% of people had close contact with a confirmed case of COVID-19.4.60% of people had previously had a nasopharyngeal swab with a negative result. Five workers had positive IgG for SARS-CoV-2 (four women and one man), with negative IgM. The mean age of the cases was 35 years old; two of them were asymptomatic; neither of them had a swab sample taken. The overall seroprevalence was 0.75%, with no significant differences between strata. DISCUSSION the seroprevalence found was low; indicating a large proportion of workers was susceptible to infection. We stress the need to complement passive epidemiological surveillance strategies with serological monitoring in health workers.

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