Inactivated and Immunogenic SARS-CoV-2 for Safe Use in Immunoassays and as an Immunization Control for Non-Clinical Trials.

Successful SARS-CoV-2 inactivation allows its safe use in Biosafety Level 2 facilities, and the use of the whole viral particle helps in the development of analytical methods and a more reliable immune response, contributing to the development and improvement of in vitro and in vivo assays. In order to obtain a functional product, we evaluated several inactivation protocols and observed that 0.03% beta-propiolactone for 24 h was the best condition tested, as it promoted SARS-CoV-2 inactivation above 99.99% and no cytopathic effect was visualized after five serial passages. Moreover, RT-qPCR and transmission electron microscopy revealed that RNA quantification and viral structure integrity were preserved. The antigenicity of inactivated SARS-CoV-2 was confirmed by ELISA using different Spike-neutralizing monoclonal antibodies. K18-hACE2 mice immunized with inactivated SARS-CoV-2, formulated in AddaS03TM, presented high neutralizing antibody titers, no significant weight loss, and longer survival than controls from a lethal challenge, despite RNA detection in the oropharyngeal swab, lung, and brain. This work emphasizes the importance of using different techniques to confirm viral inactivation and avoid potentially disastrous contamination. We believe that an efficiently inactivated product can be used in several applications, including the development and improvement of molecular diagnostic kits, as an antigen for antibody production as well as a control for non-clinical trials.

Chikungunya, Zika and Dengue seroprevalence rates among pregnant women in a hospital of southeastern Brazil / Soroprevalência de chikungunya, zika e dengue em gestantes de um hospital do sudeste do Brasil / Seroprevalencia de chikungunya, zika y dengue en mujeres embarazadas en un hospital del sureste de Brasil

Background and objectives: For decades, dengue outbreaks have been affecting vast territories of the Americas. In 2010's decade, Chikungunya and Zika virus (CHIKV and ZIKV) emerged as new arboviruses in the region. While several seroprevalence rates have been reported for dengue virus (DENV) infection in Brazil, serological surveys for the latest are scarce. We aimed to evaluate the seroprevalence of CHIKV, ZIKV, and DENV infections in pregnant women at admission to a public maternity hospital of Nova Iguaçu, state of Rio de Janeiro. Methods: A simple questionnaire was applied, containing limited demographic, obstetric, and clinical data, alongside with blood collection. Different commercial test kits, based on enzyme-linked immunosorbent assay (ELISA), were used. Results: Among 349 pregnant women enrolled from July to December 2017, there was a 28.4% seroreactivity for CHIKV, 47.2% for ZIKV, and 88.8% for DENV. Conclusion: These findings reflect the high dengue endemicity scenario and suggest a significant reach of the recent outbreaks of ZIKV and CHIKV infections in the region.(AU)

Justificativas e objetivos: Há décadas, surtos de dengue afetam vastos territórios das Américas. Na década de 2010, os vírus Chikungunya e Zika (CHIKV e ZIKV) surgiram como arbovírus emergentes na região. Embora diversas taxas de soroprevalência tenham sido relatadas para a infecção pelo vírus da dengue (DENV) no Brasil, pesquisas sorológicas para chikungunya e zika são escassas. Objetivou-se avaliar a soroprevalência das infecções por CHIKV, ZIKV e DENV em gestantes admitidas em uma maternidade pública de Nova Iguaçu, estado do Rio de Janeiro. Métodos: Foi aplicado um questionário simples, contendo dados demográficos, obstétricos e clínicos limitados, sendo realizada coleta de sangue na mesma visita. Diferentes kits de teste comerciais, baseados em ensaio imunoenzimático (ELISA), foram utilizados. Resultados: De 349 gestantes recrutadas de julho a dezembro de 2017, houve sororreatividade de 28,4% para CHIKV, 47,2% para ZIKV e 88,8% para DENV. Conclusão: Esses achados refletem o cenário de alta endemicidade da dengue e sugerem um alcance significativo dos surtos recentes causados por ZIKV e CHIKV na região.(AU)

Justificación y objetivos: Durante décadas, los brotes de dengue han afectado a vastos territorios de las Américas. En la década de 2010, los virus Chikungunya y Zika (CHIKV y ZIKV) surgieron como arbovirus emergentes en la región. Aunque se han reportadas varias tasas de seroprevalencia para la infección por el virus del dengue (DENV) en Brasil, la investigación serológica para el chikungunya y el Zika es escasa. Este estudio tuvo como objetivo evaluar la seroprevalencia de infecciones por CHIKV, ZIKV y DENV en mujeres embarazadas ingresadas en una maternidad pública en Nova Iguaçu, estado de Rio de Janeiro. Métodos: Se aplicó un sencillo cuestionario, que contenía datos demográficos, obstétricos y clínicos limitados, y se extrajo sangre en la misma visita. Se utilizaron diferentes kits de prueba comerciales basados en el ensayo inmunoabsorbente ligado a enzimas (ELISA). Resultados: De 349 mujeres embarazadas reclutadas de julio a diciembre de 2017, hubo serorreactividad de 28,4% para CHIKV, 47,2% para ZIKV y 88,8% para DENV. Conclusión: Estos hallazgos reflejan el escenario de alta endemicidad para el dengue y sugieren una variedad significativa de brotes recientes causados por ZIKV y CHIKV en la región.(AU)

Two-Step In Vitro Model to Evaluate the Cellular Immune Response to SARS-CoV-2.

The cellular immune response plays an important role in COVID-19, caused by SARS-CoV-2. This feature makes use of in vitro models' useful tools to evaluate vaccines and biopharmaceutical effects. Here, we developed a two-step model to evaluate the cellular immune response after SARS-CoV-2 infection-induced or spike protein stimulation in peripheral blood mononuclear cells (PBMC) from both unexposed and COVID-19 (primo-infected) individuals (Step1). Moreover, the supernatants of these cultures were used to evaluate its effects on lung cell lines (A549) (Step2). When PBMC from the unexposed were infected by SARS-CoV-2, cytotoxic natural killer and nonclassical monocytes expressing inflammatory cytokines genes were raised. The supernatant of these cells can induce apoptosis of A549 cells (mock vs. Step2 [mean]: 6.4% × 17.7%). Meanwhile, PBMCs from primo-infected presented their memory CD4+ T cells activated with a high production of IFNG and antiviral genes. Supernatant from past COVID-19 subjects contributed to reduce apoptosis (mock vs. Step2 [ratio]: 7.2 × 1.4) and to elevate the antiviral activity (iNOS) of A549 cells (mock vs. Step2 [mean]: 31.5% × 55.7%). Our findings showed features of immune primary cells and lung cell lines response after SARS-CoV-2 or spike protein stimulation that can be used as an in vitro model to study the immunity effects after SARS-CoV-2 antigen exposure.

Safety and immunogenicity of 17DD attenuated yellow fever vaccine in howler monkeys (Alouatta spp.).

BACKGROUND: Alouatta spp. are highly susceptible to yellow fever (YF) infection and develop an often fatal disease. The threat posed by an outbreak started in 2016 leads us to investigate vaccination as a potential tool in preventing YF in non-human primates (NHP). METHODS: Susceptible howler monkeys were immunized with three different concentrations of the human Brazilian commercial YF17DD vaccine. Post-vaccination viremia/RNAemia, immunogenicity, and safety were characterized. RESULTS: The vaccine did not produce YF clinical manifestations in any of the NHPs. After immunization, all animals seroconverted demonstrating the ability of the YF vaccine to induce humoral response in Alouatta species. CONCLUSIONS: The present work has demonstrated the safe and immunogenic profile of the existing YF 17DD vaccine in howler monkeys. This knowledge may support further studies with other susceptible monkey species and provide a possible solution for controlling epizootics and preventing the devastation of endangered species.

Chikungunya, Zika and Dengue seroprevalence rates among pregnant women in a hospital of southeastern Brazil / Soroprevalência de chikungunya, zika e dengue em gestantes de um hospital do sudeste do Brasil

Justificativas e objetivos: Há décadas, surtos de dengue afetam vastos territórios das Américas. Na década de 2010, os vírus Chikungunya e Zika (CHIKV e ZIKV) surgiram como arbovírus emergentes na região. Embora diversas taxas de soroprevalência tenham sido relatadas para a infecção pelo vírus da dengue (DENV) no Brasil, pesquisas sorológicas para chikungunya e zika são escassas. Objetivou-se avaliar a soroprevalência das infecções por CHIKV, ZIKV e DENV em gestantes admitidas em uma maternidade pública de Nova Iguaçu, estado do Rio de Janeiro. Métodos: Foi aplicado um questionário simples, contendo dados demográficos, obstétricos e clínicos limitados, sendo realizada coleta de sangue na mesma visita. Diferentes kits de teste comerciais, baseados em ensaio imunoenzimático (ELISA), foram utilizados. Resultados: De 349 gestantes recrutadas de julho a dezembro de 2017, houve sororreatividade de 28,4% para CHIKV, 47,2% para ZIKV e 88,8% para DENV. Conclusão: Esses achados refletem o cenário de alta endemicidade da dengue e sugerem um alcance significativo dos surtos recentes causados por ZIKV e CHIKV na região.

Survey on Non-Human Primates and Mosquitoes Does not Provide Evidences of Spillover/Spillback between the Urban and Sylvatic Cycles of Yellow Fever and Zika Viruses Following Severe Outbreaks in Southeast Brazil.

In the last decade, Flaviviruses such as yellow fever (YFV) and Zika (ZIKV) have expanded their transmission areas. These viruses originated in Africa, where they exhibit both sylvatic and interhuman transmission cycles. In Brazil, the risk of YFV urbanization has grown, with the sylvatic transmission approaching the most densely populated metropolis, while concern about ZIKV spillback to a sylvatic cycle has risen. To investigate these health threats, we carried out extensive collections and arbovirus screening of 144 free-living, non-human primates (NHPs) and 5219 mosquitoes before, during, and after ZIKV and YFV outbreaks (2015-2018) in southeast Brazil. ZIKV infection was not detected in any NHP collected at any time. In contrast, current and previous YFV infections were detected in NHPs sampled between 2017 and 2018, but not before the onset of the YFV outbreak. Mosquito pools screened by high-throughput PCR were positive for YFV when captured in the wild and during the YFV outbreak, but were negative for 94 other arboviruses, including ZIKV, regardless of the time of collection. In conclusion, there was no evidence of YFV transmission in coastal southeast Brazil before the current outbreak, nor the spread or establishment of an independent sylvatic cycle of ZIKV or urban Aedes aegypti transmission of YFV in the region. In view of the region's receptivity and vulnerability to arbovirus transmission, surveillance of NHPs and mosquitoes should be strengthened and continuous.

Protocolo operacional de extubação na unidade de terapia intensiva adulta de um hospital de ensino da região norte do estado do Ceará / Operational protocol of extubation in the adult intensive therapy unit of a hospital of teaching of the northern region of the state of Ceará / Protocolo operacional de extubre en la unidad de terapia intensiva adulta de un hospital de la enseñanza de la región norte del estado del Ceará

Objetivo: analisar um Protocolo Operacional de Extubação na Unidade de Terapia Intensiva (UTI) Adulta de um Hospital de Ensino da região Norte do estado do Ceará. Método: Estudo longitudinal de caráter descritivo com abordagem quantitativa. A coleta de dados foi realizada por meio da aplicação de uma ficha de avaliação e um protocolo de extubação com sua aplicação e análise, no período de agosto a outubro de 2018, análise estatística se deu através do uso do teste qui-quadrado, ao nível de 5% de significância, a pesquisa foi aprovado pelo Comitê de Ética em Pesquisa Envolvendo Seres Humanos sob o número CAAE 89038418.6.0000.8109, sob o Parecer Nº 2.749.337. Resultado: o uso de um protocolo de extubação resultou em uma redução das taxas de falha de extubação, diminuindo os índices que levam a estas falhas demonstrando a eficácia do mesmo. Conclusão: necessidade de novas pesquisas, abordando um número maior de indivíduos, uso de educação permanente para a equipe multiprofissional, com a aplicação de protocolos pela equipe.(AU)

Objective: to analyze an Operational Protocol for Extubation in the Adult Intensive Care Unit (ICU) of a Teaching Hospital in the northern region of the state of Ceará. Method: Longitudinal descriptive study with quantitative approach. Data collection was performed through the application of an evaluation form and an extubation protocol with its application and analysis, in the period from August to October 2018, statistical analysis was done using the chi-square test, at the level of 5% significance, the research was approved by the Research Ethics Committee Involving Human Beings under the number CAAE 89038418.6.0000.8109, under Opinion No. 2,749,337. Results: The use of an extubation protocol resulted in a reduction in the rates of extubation failure, reducing the indices that lead to these failures demonstrating the efficacy of extubation. Conclusion: need for new research, addressing a greater number of individuals, use of permanent education for the multiprofessional team, with the application of protocols by the team.(AU)

Objetivo: analizar un Protocolo Operativo de Extubación en la Unidad de Terapia Intensiva (UTI) Adulta de un Hospital de Enseñanza de la región Norte del estado de Ceará. Método: Estudio longitudinal de carácter descriptivo con abordaje cuantitativo. La recolección de datos fue realizada por medio de la aplicación de una ficha de evaluación y un protocolo de extubación con su aplicación y análisis, en el período de agosto a octubre de 2018, el análisis estadístico se dio a través del uso del test chi-cuadrado, al nivel de 5% de significancia, la encuesta fue aprobada por el Comité de Ética en Investigación Envolviendo Seres Humanos bajo el número CAAE 89038418.6.0000.8109, bajo el Dictamen Nº 2.749.337. Resultado: el uso de un protocolo de extubación resultó en una reducción de las tasas de fallo de extubación, disminuyendo los índices que llevan a estas fallas demostrando la eficacia del mismo. Conclusión: necesidad de nuevas investigaciones, abordando un número mayor de individuos, uso de educación permanente para el equipo multiprofesional, con la aplicación de protocolos por el equipo.(AU)