Studying the Humoral Response against SARS-CoV-2 in Cuban COVID-19 Recovered Patients.

Understanding the immune response generated by SARS-CoV-2 is critical for assessing efficient therapeutic protocols and gaining insights into the durability of protective immunity. The current work was aimed at studying the specific humoral responses against SARS-CoV-2 in Cuban COVID-19 convalescents. We developed suitable tools and methods based on ELISA methodology, for supporting this evaluation. Here, we describe the development of an ELISA for the quantification of anti-RBD IgG titers in a large number of samples and a similar test in the presence of NH4SCN as chaotropic agent for estimating the RBD specific antibody avidity. Additionally, a simple and rapid ELISA based on antibody-mediated blockage of the binding RBD-ACE2 was implemented for detecting, as a surrogate of conventional test, the levels of anti-RBD inhibitory antibodies in convalescent sera. In a cohort of 273 unvaccinated convalescents, we identified higher anti-RBD IgG titer (1 : 1,330, p < 0.0001) and higher levels of inhibitory antibodies blocking RBD-ACE2 binding (1 : 216, p < 0.05) among those who had recovered from severe illness. Our results suggest that disease severity, and not demographic features such as age, sex, and skin color, is the main determinant of the magnitude and neutralizing ability of the anti-RBD antibody response. An additional paired longitudinal assessment in 14 symptomatic convalescents revealed a decline in the antiviral antibody response and the persistence of neutralizing antibodies for at least 4 months after the onset of symptoms. Overall, SARS-CoV-2 infection elicits different levels of antibody response according to disease severity that declines over time and can be monitored using our homemade serological assays.

Jamaican fruit bats mount a stable and highly neutralizing antibody response after bat influenza virus infection.

Antibodies are an essential component of the antiviral response in many species, but to date, there is no compelling evidence that bats are capable of eliciting a robust humoral immunity, including neutralizing antibodies. Here, we report that infection of Jamaican fruit bats with the bat influenza A virus H18N11 elicits a rapid and stable humoral immune response with a strong neutralizing capacity, associated with no detectable viral shedding after repeat challenge infection. Thus, the neutralizing antibody response of bats might play an important role in the bat immunity.

Development of an indirect ELISA for the serologic detection of bovine viral diarrhea virus based on E2 antigen sub-genotypes 1b, 1e, and 1d.

Bovine viral diarrhea virus (BVDV) causes ongoing economic losses to cattle industries, directly through reduced herd performance or indirectly through control program costs. ELISA assays, one of the most widely used techniques due to their ease of implementation, have been a valuable tool for mass surveillance and detection of BVDV. In this study, we developed a new indirect ELISA (rE2-ELISA) for serologic detection of BVDV. The assay considers three recombinant E2 protein subtypes as antigens, allowing serologic diagnosis of BVDV-1b (high prevalence worldwide), BVDV-1d and 1e (high prevalence in southern Chile) sub-genotypes. Recombinant E2 (rE2) proteins were successfully expressed in stably transfected CHO cells. Conditions for rE2 ELISAs were established after determining appropriate concentrations of antigen, blocking agent, secondary antibody, and serum dilutions to achieve maximum discrimination between positive and negative serum samples. The developed rE2-ELISA showed a sensitivity of 92.86% and a specificity of 98.33%. Clinical testing of 180 serum samples from herds in southern Chile showed high accuracy (kappa > 0.8) compared to the commercial BVDV Total Ab kit (IDEXX), with 95.37% positive and 87.5% negative predictive value. In addition, the rE2 ELISA has shown the capability to detect anti-BVDV antibodies from naturally infected animals with sub-genotypes 1b, 1e, or undetermined. These results indicate that the developed indirect ELISA could serve as a valid, and efficient alternative for identifying BVDV-infected animals, thus contributing to the success of disease control and eradication programs.

Efficacy of convalescent plasma in hospitalized COVID-19 patients: findings from a controlled trial.

The COVID-19 pandemic has driven the search for alternative therapies, including convalescent plasma, historically used in infectious diseases. Despite results in other diseases, its effectiveness against COVID-19 remains uncertain with conflicting results in clinical trials. A pragmatic, single-center, prospective, and open randomized controlled trial was carried out in a hospital in Brazil, with the aim of evaluating the impact of convalescent plasma on the clinical improvement of patients hospitalized with COVID-19. The World Health Organization (WHO) ordinal scale was used to measure clinical improvement, focusing on the reduction in disease severity by up to 2 points, while antibody and C-reactive protein levels were monitored over time. After hospital admission, participants were randomized 1:1 to receive convalescent plasma and standard treatment or to be part of the control group with standard treatment. Follow-up was carried out on days 1, 3, 7, 14 and/or at discharge. From January 14 to April 4, 2022, 38 patients were included, but 3 were excluded due to protocol deviations, resulting in a total of 35 patients: 19 in the control group and 16 in the plasma group. There was no significant difference in clinical improvement between the convalescent plasma group and the control group, nor in secondary outcomes. The study had limitations due to the small number of patients and limited representation of COVID-19 cases. Broader investigations are needed to integrate therapies into medical protocols, both for COVID-19 and other diseases. Conducting randomized studies is challenging due to the complexity of medical conditions and the variety of treatments available.

Detection of Neutralizing Antibodies in Serum Samples Using a SARS-CoV-2 Pseudotyped Virus Assay.

Conventional live virus research on severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causal agent of coronavirus disease-19 (COVID-19), requires Biosafety Level 3 (BSL-3) facilities. SARS-CoV-2 pseudotyped viruses have emerged as valuable tools in virology, mimicking the entry process of the SARS-CoV-2 virus into human cells by expressing its spike glycoprotein in a surrogate system using recombinant plasmids. One significant application of this tool is in functional assays for the evaluation of neutralizing antibodies. Pseudotyped viruses have the advantage of being competent for only a single cycle of infection, providing better safety and versatility and allowing them to be studied in BSL-2 laboratories. Here, we describe three protocols for the detection of SARS-CoV-2 neutralizing antibodies through a pseudotyped virus assay. First, SARS-CoV-2 S pseudotyped viruses (PV SARS-CoV-2 S) are produced using a Moloney murine leukemia virus (MuLV) three-plasmid system. The plasmids are designed to express the GagPol packing proteins, enhanced green fluorescent protein (eGFP) as a readout system, and the SARS-CoV-2 S protein modified to remove the endoplasmic reticulum retention domain and to improve infection. Next, the internalization of PV SARS-CoV-2 S protein in human embryonic kidney 293T (HEK-293T) cells overexpressing angiotensin-converting enzyme 2 (HEK-293T-ACE2) is confirmed by fluorescence microscopy and quantified using flow cytometry. Finally, PV SARS-CoV-2 S is used to screen neutralizing antibodies in serum samples from convalescent COVID-19 patients; it can also be used for studying the cell entry mechanisms of different SARS-CoV-2 variants, evaluating antiviral agents, and designing vaccines. © 2024 Wiley Periodicals LLC. Basic Protocol 1: Generation of PV SARS-CoV-2 S pseudotyped virus Basic Protocol 2: Assay of PV SARS-CoV-2 S internalization in target cells. Basic Protocol 3: Detection of neutralizing antibodies in serum samples.

Overcoming Limited Access to Virus Infection Rapid Testing: Development of a Lateral Flow Test for SARS-CoV-2 with Locally Available Resources.

The COVID-19 pandemic highlighted testing inequities in developing countries. Lack of lateral flow test (LFT) manufacturing capacity was a major COVID-19 response bottleneck in low- and middle-income regions. Here we report the development of an open-access LFT for SARS-CoV-2 detection comparable to commercial tests that requires only locally available supplies. The main critical resource is a locally developed horse polyclonal antibody (pAb) whose sensitivity and selectivity are greatly enhanced by affinity purification. We demonstrate that these Abs can perform similarly to commercial monoclonal antibodies (mAbs), as well as mAbs and other pAbs developed against the same antigen. We report a workflow for test optimization using nasopharyngeal swabs collected for RT-qPCR, spiked with the inactivated virus to determine analytical performance characteristics as the limit of detection, among others. Our final prototype showed a performance similar to available tests (sensitivity of 83.3% compared to RT-qPCR, and 90.9% compared to commercial antigen tests). Finally, we discuss the possibility and the challenges of utilizing affinity-purified pAbs as an alternative for the local development of antigen tests in an outbreak context and as a tool to address inequalities in access to rapid tests.

Do immunosuppressive treatments influence immune responses against adenovirus-based COVID-19 vaccines in patients with multiple sclerosis? An Argentine multicenter study.

Introduction: There are no reports in LATAM related to longitudinal humoral and cellular response to adenovirus based COVID-19 vaccines in people with Multiple Sclerosis (pwMS) under different disease modifying therapies (DMTs) and neutralization of the Omicron and Wuhan variants of SARS-COV-2. Methods: IgG anti- SARS-COV-2 spike titer were measured in a cohort of 101 pwMS under fingolimod, dimethyl fumarate, cladribine and antiCD20, as well as 28 healthy controls (HC) were measured 6 weeks after vaccination with 2nd dose (Sputnik V or AZD1222) and 3nd dose (homologous or heterologous schedule). Neutralizing capacity was against Omicron (BA.1) and Wuhan (D614G) variants and pseudotyped particles and Cellular response were analyzed. Results: Multivariate regression analysis showed anti-cd20 (ß= -,349, 95% CI: -3655.6 - -369.01, p=0.017) and fingolimod (ß=-,399, 95% CI: -3363.8 - -250.9, p=0.023) treatments as an independent factor associated with low antibody response (r2 adjusted=0.157). After the 2nd dose we found a correlation between total and neutralizing titers against D614G (rho=0.6; p<0.001; slope 0.8, 95%CI:0.4-1.3), with no differences between DMTs. Neutralization capacity was lower for BA.1 (slope 0.3, 95%CI:0.1-0.4). After the 3rd dose, neutralization of BA.1 improved (slope: 0.9 95%CI:0.6-1.2), without differences between DMTs. A fraction of pwMS generated anti-Spike CD4+ and CD8+ T cell response. In contrast, pwMS under antiCD20 generated CD8+TNF+IL2+ response without differences with HC, even in the absence of humoral response. The 3rd dose significantly increased the neutralization against the Omicron, as observed in the immunocompetent population. Discussion: Findings regarding humoral and cellular response are consistent with previous reports.

Immunologic mediators profile in COVID-19 convalescence.

SARS-CoV-2 caused the pandemic situation experienced since the beginning of 2020, and many countries faced the rapid spread and severe form of the disease. Mechanisms of interaction between the virus and the host were observed during acute phase, but few data are available when related to immunity dynamics in convalescents. We conducted a longitudinal study, with 51 healthy donors and 62 COVID-19 convalescent patients, which these had a 2-month follow-up after symptoms recovery. Venous blood sample was obtained from all participants to measure blood count, subpopulations of monocytes, lymphocytes, natural killer cells and dendritic cells. Serum was used to measure cytokines, chemokines, growth factors, anti-N IgG and anti-S IgG/IgM antibodies. Statistic was performed by Kruskal-Wallis test, and linear regression with days post symptoms and antibody titers. All analysis had confidence interval of 95%. Less than 35% of convalescents were anti-S IgM+, while more than 80% were IgG+ in D30. Anti-N IgG decreased along time, with loss of seroreactivity of 13%. Eosinophil count played a distinct role on both antibodies during all study, and the convalescence was orchestrated by higher neutrophil-to-lymphocyte ratio and IL-15, but initial stages were marked by increase in myeloid DCs, B1 lymphocytes, inflammatory and patrolling monocytes, G-CSF and IL-2. Later convalescence seemed to change to cytotoxicity mediated by T lymphocytes, plasmacytoid DCs, VEGF, IL-9 and CXCL10. Anti-S IgG antibodies showed the longest perseverance and may be a better option for diagnosis. The inflammatory pattern is yet present on initial stage of convalescence, but quickly shifts to a reparative dynamic. Meanwhile eosinophils seem to play a role on anti-N levels in convalescence, although may not be the major causative agent. We must highlight the importance of immunological markers on acute clinical outcomes, but their comprehension to potentialize adaptive system must be explored to improve immunizations and further preventive policies.

Sputnik V-Induced Antibodies against SARS-CoV-2 Variants during the Dissemination of the Gamma Variant in Venezuela.

The COVID-19 pandemic was characterized by the emergence and succession of SARS-CoV-2 variants able to evade the antibody response induced by natural infection and vaccination. To evaluate the IgG reactivity and neutralizing capacity of the serum of individuals vaccinated with Sputnik V (105 volunteers vaccinated) against different viral variants. IgG reactivity to the Spike protein (S) was evaluated by ELISA. A plaque reduction neutralization test was performed using different viral variant isolates. At 42 days post-vaccination, the frequency of recognition and reactivity to the S protein of the Omicron variant was lower compared to that of the other variants. In general, a higher average neutralization titer was seen against the ancestral variant compared to the variants, especially Omicron. However, some sera exhibited a higher neutralization titer to the Gamma variant compared to the ancestral variant, suggesting unapparent exposure during the clinical trial. Antibodies induced by Sputnik V can recognize, persist, and neutralize SARS-CoV-2 variants, with Omicron being the one that best evades this response. These results represent a unique report on the humoral response induced by a globally lesser-studied vaccine in terms of efficacy and immune escape, offering insights into developing vaccines targeting unknown coronaviruses.

HCoV-NL63 and HCoV-HKU1 seroprevalence and its relationship with the clinical features of COVID-19 patients from Villavicencio, Colombia / Seroprevalencia de HCoV-NL63 y HCoV-HKU1 y su relación con las características clínicas de pacientes con COVID-19 de Villavicencio, Colombia

INTRODUCTION: Due to the cross-reactivity between SARS-CoV-2 and common human coronaviruses, previous infections with these viruses could contribute to serological or cellular cross-protection against severe COVID-19. However, protective immunity may not develop, or pre-existing immunity could increase COVID-19 severity. OBJECTIVE: To determine the seroprevalence of IgG antibodies against HCoV-NL63 and HCoV-HKU1 and correlate previous exposure with COVID-19 signs in patients from Villavicencio. MATERIALS AND METHODS: A cross-sectional retrospective study was conducted. ELISA technique was used to search for IgG antibodies against HCoV-NL3 and HCoV-HKU1 in patients with positive RT-qPCR results for SARS-CoV-2. Patients were grouped according to COVID-19 clinical characteristics in four groups: group 1: asymptomatic (n = 23); group 2: hospitalized (n = 24); group 3: intensive care units (n = 24), and group 4: dead (n = 22). RESULTS: The overall seroprevalence of IgG antibodies against HCoV was 74.2% (n = 69; 95% CI: 65.3-83.1), with 66.7% of HCoV-NL63 (n = 62; 95% CI: 57,1-76,2), and 25.8% of HCoV-HKU1 (n = 24; 95% CI: 16,9-34,7). Based on crosstab analysis, prior exposure to HCoV-NL63 was associated with protection against severe COVID-19 (p = 0.042; adjusted OR = 0.159; 95% CI: 0.027-0.938), and previous coinfection of HCoV-NL63 and HCoVHKU1 was considered a positive association to severe COVID-19 (p = 0.048; adjusted OR = 16.704; 95% CI: 1.020 - 273.670). CONCLUSION: To our knowledge, this is the first study addressing seroprevalence of HCoV IgG antibodies in Colombia and Latin America. Previous exposure to HCoV-NL63 could protect against severe COVID-19, whereas patients with underlying HCoV-NL63 and HCoVHKU1 coinfection could be hospitalized with severe signs of COVID-19.

Introducción: Debido a la reactividad cruzada entre SARS-CoV-2 y los coronavirus humanos comunes, las infecciones previas con estos virus podrían contribuir a la protección cruzada serológica o celular contra la COVID-19 grave. Sin embargo, la inmunidad protectora puede no desarrollarse o la inmunidad preexistente podría generar COVID-19 grave. Objetivo: Determinar la seroprevalencia de anticuerpos IgG frente a HCoV-NL63 y HCoVHKU1, y correlacionar su previa exposición con los signos de COVID-19 en pacientes de Villavicencio. Materiales y métodos: Se realizó un estudio retrospectivo observacional analítico y transversal. Se utilizó la técnica ELISA para buscar anticuerpos IgG contra HCoV-NL3 y HCoV-HKU1 en pacientes con resultado positivo de RT-qPCR para SARS-CoV-2. Los pacientes se agruparon según los signos de COVID-19 en cuatro grupos: grupo 1: asintomáticos (n = 23); grupo 2: hospitalizados (n = 24); grupo 3: unidad de cuidados intensivos (n = 24), y grupo 4: fallecidos (n = 22). Resultados: La seroprevalencia general de IgG anti-HCoV fue de 74.2 % (n = 69; IC95%: 65,3-83,1), con 66,7 % de HCoV-NL63 (n = 62; IC95% :57,1-76,2) y 25,8 % de HCoV-HKU1 (n = 24; [IC95%:16,9-34,7). Según el análisis de las tablas de contingencia, la exposición previa a HCoV-NL63 se asoció con protección de una COVID-19 grave (p = 0,042; OR ajustado = 0,159; IC95%: 0,027-0,938) y la previa coinfección de HCoV-NL63 y HCoV-HKU1 se asoció con padecimiento de signos clínicos graves por COVID-19 (p = 0,048; OR ajustado = 16,704; IC95%: 1,020- 73,670). Conclusión: Según la literatura revisada hasta la fecha, este es el primer estudio sobre la seroprevalencia de anticuerpos IgG de HCoV en Colombia y Latinoamérica. La exposición previa a HCoV-NL63 podría proteger contra la COVID-19 grave, mientras que los pacientes con coinfección subyacente de HCoV-NL63 y HCoV-HKU1 podrían resultar hospitalizados con signos graves de COVID-19.

(Re-)Emergence of Oropouche Virus (OROV) Infections: Systematic Review and Meta-Analysis of Observational Studies.

Oropouche Virus (OROV; genus of Orthobunyavirus) is the causal agent of Oropouche Fever (OF). Due to the lack of specific signs and symptoms and the limited availability of diagnostic tests, the actual epidemiology of OROV infections and OF has been extensively disputed. In this systematic review with meta-analysis, a literature search was carried out in PubMed, Scopus, EMBASE, and MedRxiv in order to retrieve relevant articles on the documented occurrence of OROV infections. Pooled detection rates were then calculated for anti-OROV antibodies and virus detection (i.e., viral RNA detected by viral cultures and/or real-time polymerase chain reaction [RT-qPCR]). Where available, detection rates for other arboviruses (i.e., Dengue [DENV], Chikungunya [CHKV], and Zika Virus [ZIKV]) were calculated and compared to those for OROV. A total of 47 studies from South America and the Caribbean were retrieved. In individuals affected by febrile illness during OROV outbreaks, a documented prevalence of 0.45% (95% confidence interval [95%CI] 0.16 to 1.12) for virus isolation, 12.21% (95%CI 4.96 to 27.09) for seroprevalence (including both IgM and IgG class antibodies), and 12.45% (95%CI 3.28 to 37.39) for the detection of OROV-targeting IgM class antibodies were eventually documented. In the general population, seroprevalence was estimated to be 24.45% (95%CI 7.83 to 55.21) for IgG class antibodies. The OROV detection rate from the cerebrospinal fluids of suspected cases of viral encephalitis was estimated to be 2.40% (95%CI 1.17 to 5.03). The occurrence of OROV infections was consistently lower than that of DENV, CHKV, and ZIKV during outbreaks (Risk Ratio [RR] 24.82, 95%CI 21.12 to 29.16; RR 2.207, 95%CI 1.427 to 3.412; and RR 7.900, 95%CI 5.386 to 11.578, respectively) and in the general population (RR 23.614, 95%CI 20.584 to 27.129; RR 3.103, 95%CI 2.056 to 4.685; and RR 49.500, 95%CI 12.256 to 199.921, respectively). In conclusion, our study stresses the possibly high underestimation of OROV prevalence in the general population of South America, the potential global threat represented by this arbovirus infection, and the potential preventive role of a comprehensive "One Health approach".

Influence of previous COVID-19 exposure and vaccine type (CoronaVac, ChAdOx1 nCov-19 or BNT162b2) on antibody and cytokine (Th1 or Th2) responses.

To achieve global herd immunity, widespread vaccination is the most effective strategy. Vaccines stimulate the immune system, generating cytokines and chemokines, isotype antibodies, and neutralizing antibodies; all these molecules collectively provide a more comprehensive characterization of the immune response post-vaccination. We conducted a longitudinal study in northwestern Mexico, involving 120 individuals before vaccination and after the first dose of the SARS-CoV-2 vaccine, and 46 individuals after their second dose. Our findings reveal that antibody levels stabilize over time; cytokine levels generally increase following the first dose but decrease after the second dose and higher than normal levels in IgG1 and IgG3 concentrations are present. Most of the innate cytokines determined in this study were higher after the first dose of the vaccine. Regardless of previous infection history, this finding suggests that the first dose of the vaccine is crucial and may stimulate immunity by enhancing the innate immune response. Conversely, increased levels of IL-4, indicative of a Th2 response, were found in individuals without prior exposure to the virus and in those vaccinated with CoronaVac. These results suggest that the immune response to COVID-19 vaccines is multi-faceted, with preexisting immunity potentiating a more robust innate response. Vaccine type plays a critical role, with genetic vaccines favoring a Th1 response and inactivated vaccines like CoronaVac skewing toward a Th2 profile.

Assessment of immune responses to a Comirnaty® booster following CoronaVac® vaccination in healthcare workers.

BACKGROUND: The immunological response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and immunisation is variable. OBJECTIVES: To describe the humoral immune response by correlating IgA and IgG antibodies with NAbs titration following CoronaVac® immunisation and an mRNA (Comirnaty®) booster among healthcare workers (HCWs) and to compare the cytokine and interleukin profiles between HCWs vaccinated with CoronaVac and coronavirus disease 2019 (COVID-19) infected patients. METHODS: Samples from 133 HCWs collected at 20 (T1) and 90 (T2) days after CoronaVac immunisation and 15 (T3) days after a booster dose with the Comirnaty vaccine were analysed for IgA and IgG EIA and neutralisation assay. Cytokine levels from vaccinated individuals at T1 day and COVID-19 patients were compared. FINDINGS: Neutralising antibodies (NAbs) were observed in 81.7% of participants at T1, but only 49.2% maintained detectable NAbs after 90 days. The booster dose increased NAbs response in all participants. The cytokines with the highest levels post-vaccination were IL-6 and MCP-1. The MCP-1, IL-18, and IFN- γ levels were higher in COVID-19 patients than in vaccinated HCWs, while IL-22 levels increased in the vaccinated HCWs group. MAIN CONCLUSIONS: The neutralisation titres in the T2 samples decreased, and antibody levels detected at T2 showed a more significant reduction than the neutralisation. The higher IL-22 expression in immunised individuals compared to those with COVID-19 suggests that IL-22 may be beneficial in protecting against severe disease.

Neutralizing antibodies in milk and blood of lactating women vaccinated for SARS-CoV-2: a systematic review.

OBJECTIVE: To compare the presence of neutralizing antibodies against SARS-CoV-2 found in the breast milk and blood of vaccinated lactating women with those not vaccinated. DATA SOURCE: The study was registered in the International Prospective Register of Systematic Reviews (PROSPERO) under CRD42021287554 and followed the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. Cohort, case-control, and cross-sectional studies that evaluated antibodies against SARS-CoV-2 in the milk and blood of vaccinated mothers and had as control group unvaccinated mothers were eligible. Health Sciences Descriptors (DeCs), Medical Subject Headings (MeSH) and Emtree descriptors were used for the Virtual Health Library (VHL), Medical Literature Analysis and Retrieval System Online (Medline/Pubmed), and Embase databases, respectively. In the Web of Science and Scopus, the strategy was adapted. No restrictions on the publication period and language were set. DATA SYNTHESIS: The search identified 233 records, of which 128 duplicates and 101 papers that did not meet the inclusion criteria were excluded. Hence, four cohort studies were eligible. Nursing mothers vaccinated with the Pfizer-BioNTech and Moderna vaccines showed antibodies against SARS-CoV-2 in their blood and breast milk. CONCLUSIONS: Vaccinated lactating women had higher levels of immunoglobulin G (IgG) and A (IgA) in serum and breast milk than unvaccinated women.

Diagnosis of viral infectious diseases through sensitive detection of human serum antibodies using a modified label-free electrochemical biosensor.

Rapid virus identification is crucial for preventing outbreaks. The COVID-19 pandemic has highlighted the critical nature of rapid virus detection. Here, we designed a label-free electrochemical biosensor modified with gold nanoparticles (AuNPs) to detect IgG antibodies from human serum, enabling rapid point-of-care diagnostics. AuNPs were synthesized and characterized. A multivariate optimization was carried out to determine the optimal condition for functionalizing AuNPs with anti-IgG. Subsequently, using a glassy carbon electrode (GCE), a modified AuNPs/GCE electrochemical biosensor was developed for IgG detection. The results indicated that AuNPs displayed a spherical morphology with a size distribution of 19.54 nm. Additionally, the zeta potential was recorded at -7.84 mV. Central composite design (CCD) analysis determined the optimal conditions for functionalizing AuNPs to be an anti-IgG concentration of 320 µg mL-1, a temperature of 25 °C, and pH of 7.4. The characterization study confirmed the successful synthesis and functionalization of AuNPs. Through electrochemical impedance spectroscopy measurement, the biosensor demonstrated a limit of detection (LOD) of 0.2 ng mL-1 and limit of quantification (LOQ) of 0.8 ng mL-1. Furthermore, tests in real samples showed the interaction between IgG antibodies in serum samples and AuNPs/GCE, confirming the biosensor's ability to detect and quantify IgG in clinical samples.

Hantavirus: Preliminary assessment of lateral flow immune assay prototypes to detect IgM and IgG antibodies.

Three lateral flow immunoassay prototypes developed to detect IgM, IgG and IgM/IgG antibodies against Hantavirus were evaluated. A total of 163 samples were tested: 10 from Hantavirus patients, 103 from related diseases, and 50 from healthy controls. The prototypes exhibited 100 % sensitivity, 97.5 % to 99.3 % specificity, indicating promising improved diagnosis.

Homologous equivalence study of immunogenicity after third dose of Covid-19 vaccine (recombinant) with an interval of six months after the second dose, comparing the interval of eight and 12 weeks between the first two doses.

BACKGROUND: In response to the coronavirus disease 2019 (Covid-19) pandemic, Brazil authorised the Astra Zeneca/Fiocruz vaccine in January 2021. As the Delta variant emerged in May 2021, interval between vaccine doses was adjusted. By September 2021, the Brazilian National Immunisation Program recommended a booster dose for individuals over 70, and later expanded the recommendation to all adults. OBJECTIVES: Assess the equivalence of IgG antibody response against the Covid-19 S protein before and approximately 28 days after the third dose of a Covid-19 recombinant vaccine. Two groups received initial two doses with intervals of eight and 12 weeks. METHODS: This is a phase IV clinical study, uncontrolled, non-randomised. The study proposes calculating the ratio of geometric means titres (GMT) 28 days after the third dose, with a target ratio of confidence interval (CI) between 0.77 and 1.3. FINDINGS: In the primary endpoint, there was no equivalence between the eight- and 12-week intervals with a slight variation favouring the eight-week group. Post-third dose, both groups showed increases titres at 28 days, three months, six months and 12 months. Both groups responded similarly to Delta and Omicron BA.1, with a more significant increase for Delta. MAIN CONCLUSIONS: The study showed strong and consistent immune response in all age groups receiving the Covid-19 recombinant vaccine. Third dose elicited an increase in GMT by at least three times aligned with Ministry of Health strategies emphasising Bio-Manguinhos crucial role in pandemic control in the country.

Enhanced Assessment of Cross-Reactive Antigenic Determinants within the Spike Protein.

Despite successful vaccination efforts, the emergence of new SARS-CoV-2 variants poses ongoing challenges to control COVID-19. Understanding humoral responses regarding SARS-CoV-2 infections and their impact is crucial for developing future vaccines that are effective worldwide. Here, we identified 41 immunodominant linear B-cell epitopes in its spike glycoprotein with an SPOT synthesis peptide array probed with a pool of serum from hospitalized COVID-19 patients. The bioinformatics showed a restricted set of epitopes unique to SARS-CoV-2 compared to other coronavirus family members. Potential crosstalk was also detected with Dengue virus (DENV), which was confirmed by screening individuals infected with DENV before the COVID-19 pandemic in a commercial ELISA for anti-SARS-CoV-2 antibodies. A high-resolution evaluation of antibody reactivity against peptides representing epitopes in the spike protein identified ten sequences in the NTD, RBD, and S2 domains. Functionally, antibody-dependent enhancement (ADE) in SARS-CoV-2 infections of monocytes was observed in vitro with pre-pandemic Dengue-positive sera. A significant increase in viral load was measured compared to that of the controls, with no detectable neutralization or considerable cell death, suggesting its role in viral entry. Cross-reactivity against peptides from spike proteins was observed for the pre-pandemic sera. This study highlights the importance of identifying specific epitopes generated during the humoral response to a pathogenic infection to understand the potential interplay of previous and future infections on diseases and their impact on vaccinations and immunodiagnostics.

Co-administration of recombinant BCG and SARS-CoV-2 proteins leads to robust antiviral immunity.

SARS-CoV-2 is the causative virus of COVID-19, which has been responsible for millions of deaths worldwide since its discovery. After its emergence, several variants have been identified that challenge the efficacy of the available vaccines. Previously, we generated and evaluated a vaccine based on a recombinant Bacillus Calmette-Guérin (rBCG) expressing the nucleoprotein (N) of SARS-CoV-2 (rBCG-N-SARS-CoV-2). This protein is a highly immunogenic antigen and well conserved among variants. Here, we tested the administration of this vaccine with recombinant N and viral Spike proteins (S), or Receptor Binding Domain (RBD-Omicron variant), plus a booster with the recombinant proteins only, as a novel and effective strategy to protect against SARS-CoV-2 variants. METHODS: BALB/c mice were immunized with rBCG-N-SARS-CoV-2 and recombinant SARS-CoV-2 proteins in Alum adjuvant, followed by a booster with recombinant proteins to assess the safety and virus-specific cellular and humoral immune responses against SARS-CoV-2 antigens. RESULTS: Immunization with rBCG-N-SARS-CoV-2 + recombinant proteins as a vaccine was safe and promoted the activation of CD4+ and CD8+ T cells that recognize SARS-CoV-2 N, S, and RBD antigens. These cells were able to secrete cytokines with an antiviral profile. This immunization strategy also induced robust titers of specific antibodies against N, S, and RBD and neutralizing antibodies of SARS-CoV-2. CONCLUSIONS: Co-administration of the rBCG-N-SARS-CoV-2 vaccine with recombinant SARS-CoV-2 proteins could be an effective alternative to control particular SARS-CoV-2 variants. Due to its safety and capacity to induce virus-specific immune responses, we believe the rBCG-N-SARS-CoV-2 + Proteins vaccine could be an attractive candidate to protect against this virus, especially in newborns.

Comparison of Adverse Events and Antibody Responses Among Different COVID-19 Vaccination Schedules.

Global investment in developing COVID-19 vaccines has been substantial, but vaccine hesitancy has emerged due to misinformation. Concerns about adverse events, vaccine shortages, dosing confusion, mixing vaccines, and access issues contribute to hesitancy. Initially, the WHO recommended homologous vaccination (same vaccine for both doses), but evolving factors led to consideration of heterologous vaccination (different vaccines). The study compared reactogenicity and antibody response for both viral protein spike (S) and nucleocapsid (N) in 205 participants who received three vaccination regimens: same vaccine for all doses (Pfizer), two initial doses of the same vaccine (CoronaVac or AstraZeneca), and a Pfizer booster. ChAdOx1 and BNT162b2 vaccines were the most reactogenic vaccines, while CoronaVac vaccine was the least. ChAdOx1 and BNT162b2 achieved 100% of S-IgG seropositivity with one dose, while CoronaVac required two doses, emphasizing the importance of the second dose in achieving complete immunization across the population with different vaccine regimes. Pfizer recipients showed the highest S-IgG antibody titers, followed by AstraZeneca recipients, both after the first and second doses. A third vaccine dose was essential to boost the S-IgG antibodies and equalize the antibody levels among the different vaccine schedules. CoronaVac induced N-IgG antibodies, while in the Pfizer and AstraZeneca groups, they were induced by a natural infection, reinforcing the role of N protein as a biomarker of infection.