Primer registro de Aedes vittatus en la provincia de Santiago de Cuba / First record of Aedes vittatus in Santiago de Cuba province

Aedes vittatus, vector que transmite el virus de la fiebre amarilla, el dengue, el zika y el chikungunya, se detectó por primera vez en enero de 2020 en la localidad de Ramón de las Yaguas del municipio Santiago de Cuba, lo que constituyó el primer registro en la provincia del mismo nombre. Este hallazgo significó una alerta para el resto del país y conllevó a investigar sobre su bioecología y competencia vectorial. A tales efectos se realizó el presente estudio con el objetivo de informar lo relacionado con la presencia de esta especie invasora, la distribución espacial y los sitios de cría para adoptar decisiones preventivas y de control.

Aedes vittatus, vector that transmits the yellow fever virus, dengue, zika and chikungunya, was detected for the first time in January, 2020 in the town of Ramón de las Yaguas town from Santiago de Cuba municipality, what constituted the first record in the province with the same name. This finding meant an alert for the rest of the country and led to investigate on its bioecology and vectorial competence. To such effects the present study was carried out with the objective of informing everything related to the presence of this invading species, the space distribution and breeding sites to adopt preventive and control decisions.

Yellow fever vaccination before and during the covid-19 pandemic in Brazil

ABSTRACT OBJECTIVE To analyze the number of yellow fever vaccine doses administered before and during the covid-19 pandemic in Brazil. METHODS This is an ecological, time series study based on data from the National Immunization Program. Differences between the median number of yellow fever vaccine doses administered in Brazil and in its regions before (from April/2019 to March/2020) and after (from April/2020 to March/2021) the implementation of social distancing measures in the country were assessed via the Mann-Whitney test. Prais-Winsten regression models were used for time series analyses. RESULTS We found a reduction in the median number of yellow fever vaccine doses administered in Brazil and in its regions: North (-34.71%), Midwest (-21.72%), South (-63.50%), and Southeast (-34.42%) (p < 0.05). Series showed stationary behavior in Brazil and in its five regions during the covid-19 pandemic (p > 0.05). Brazilian states also showed stationary trends, except for two states which recorded an increasing trend in the number of administered yellow fever vaccine doses, namely: Alagoas State (before: β = 64, p = 0.081; after: β = 897, p = 0.039), which became a yellow fever vaccine recommendation zone, and Roraima State (before: β = 68, p = 0.724; after: β = 150, p = 0.000), which intensified yellow fever vaccinations due to a yellow fever case confirmation in a Venezuelan State in 2020. CONCLUSION The reduced number of yellow fever vaccine doses administered during the covid-19 pandemic in Brazil may favor the reemergence of urban yellow fever cases in the country.

Seroprevalence of arboviruses in Ecuador: Implications for improved surveillance / Seroprevalencia de arbovirus en Ecuador: implicaciones para mejorar su vigilancia

Abstract | Introduction: Arthropod-borne viruses (arboviruses) cause morbidity and mortality in humans and domestic animals worldwide. The percentage of population immunity or susceptibility to these viruses in Ecuador is unknown. Objectives: To investigate the proportion of Ecuadorian populations with IgG antibodies (Abs) (past exposure/immunity) and IgM Abs (current exposure) against flaviviruses and alphaviruses and to study the activity of these viruses in Ecuador. Materials and methods: During 2009-2011, we conducted a serosurvey for selected arboviruses in humans (n=1,842), equines (n=149), and sentinel hamsters (n=84) at two coastal locations and one in the Amazon basin (Eastern Ecuador) using enzyme-linked immunosorbent assay and hemagglutination inhibition test. Results: From 20.63% to 63.61% of humans showed IgG-antibodies for the flaviviruses: Dengue virus (DENV), yellow fever virus (YFV) Saint Louis encephalitis virus, and West Nile virus (WNV); from 4.67% to 8.63% showed IgG-Abs for the alphaviruses: Venezuelan equine encephalitis virus, eastern equine encephalitis virus, and western equine encephalitis virus. IgM-Abs were found for DENV and WNV. Equines and hamsters showed antibodies to alphaviruses in all locations; two hamsters seroconverted to YFV in the Amazonia. Conclusions: The results show a YFV vaccination history and suggest the activity of arboviruses not included in the current surveillance scheme. Enhanced arbovirus and mosquito surveillance, as well as continued YFV vaccination and evaluation of its coverage/ effectiveness, are recommended.

Resumen | Introducción. Los virus transmitidos por artrópodos (arbovirus) causan morbilidad y mortalidad en humanos y animales domésticos mundialmente. Se desconoce el porcentaje de inmunidad o vulnerabilidad de la población ecuatoriana ante estos virus. Objetivos. Investigar la proporción de poblaciones ecuatorianas con anticuerpos IgG (exposición o inmunidad pasada) y anticuerpos IgM (exposición reciente) contra flavivirus y alfavirus, e investigar su actividad en Ecuador. Materiales y métodos. Entre 2009 y 2011, se llevó a cabo una encuesta serológica para arbovirus en humanos (n=1.842), equinos (n=149) y hámsters centinela (n=84) en dos localidades costeras y en una en la Amazonía, utilizando la prueba ELISA (Enzyme-Linked ImmunoSorbent Assay) y la prueba de inhibición de la hemaglutinación. Resultados. Entre el 20,63 y el 63,61 % de los humanos registraron IgG contra el virus del dengue (DENV), el de la fiebre amarilla (YFV), el de la encefalitis de San Luis y el del Nilo Occidental (WNV); entre 4,67 y 8,63 % tenían IgG para los virus de la encefalitis equina venezolana, de la encefalitis equina del este y de la encefalitis equina del oeste. Se encontró IgM para DENV y WNV. En los equinos y en los hámsters se encontraron anticuerpos contra alfavirus en todas las localidades muestreadas; dos hámsters mostraron seroconversión a YFV en la Amazonía. Conclusiones. Los resultados del estudio evidenciaron los antecedentes de vacunación contra el YFV y sugieren la actividad de arbovirus no incluidos en el esquema de vigilancia actual. Se recomienda ampliar la vigilancia de arbovirus y mosquitos, continuar con la vacunación contra el YFV, y evaluar su cobertura y efectividad.

Steroidal saponins from the roots of Solanum sisymbriifolium Lam. (Solanaceae) have inhibitory activity against dengue virus and yellow fever virus

Dengue is the most important arthropod-borne viral disease worldwide. Infection with any of the four dengue virus (DENV) serotypes can be asymptomatic or lead to disease with clinical symptoms ranging from undifferentiated and self-limiting fever to severe dengue disease, which can be fatal in some cases. Currently, no specific antiviral compound is available for treating DENV. The aim of this study was to identify compounds in plants from Paraguayan folk medicine with inhibitory effects against DENV. We found high virucidal activity (50% maximal effective concentration (EC50) value of 24.97 µg/mL) against DENV-2 in the ethanolic extract of the roots of Solanum sisymbriifolium Lam. (Solanaceae) without an evident cytotoxic effect on Vero E6 cells. Three saponins isolated from the root extract showed virucidal effects (EC50 values ranging from 24.9 to 35.1 µg/mL) against DENV-2. Additionally, the saponins showed inhibitory activity against yellow fever virus (EC50 values ranging from 126 to 302.6 µg/mL), the prototype virus of the Flavivirus genus, suggesting that they may also be effective against other members of this genus. Consequently, these saponins may be lead compounds for the development of antiviral agents.

Seasonal population dynamics of the primary yellow fever vector Haemagogus leucocelaenus (Dyar & Shannon) (Diptera: Culicidae) is mainly influenced by temperature in the Atlantic Forest, southeast Brazil

BACKGROUND Southeast Brazil has recently experienced a Yellow Fever virus (YFV) outbreak where the mosquito Haemagogus leucocelaenus was a primary vector. Climatic factors influence the abundance of mosquito vectors and arbovirus transmission. OBJECTIVES We aimed at describing the population dynamics of Hg. leucocelaenus in a county touched by the recent YFV outbreak. METHODS Fortnightly egg collections with ovitraps were performed from November 2012 to February 2017 in a forest in Nova Iguaçu, Rio de Janeiro, Brazil. The effects of mean temperature and rainfall on the Hg. leucocelaenus population dynamics were explored. FINDINGS Hg. leucocelaenus eggs were continuously collected throughout the study, with a peak in the warmer months (December-March). The climatic variables had a time-lagged effect and four weeks before sampling was the best predictor for the positivity of ovitraps and total number of eggs collected. The probability of finding > 50% positive ovitraps increased when the mean temperature was above 24ºC. The number of Hg. leucocelaenus eggs expressively increase when the mean temperature and accumulated precipitation surpassed 27ºC and 100 mm, respectively, although the effect of rainfall was less pronounced. MAIN CONCLUSIONS Monitoring population dynamics of Hg. leucocelaenus and climatic factors in YFV risk areas, especially mean temperature, may assist in developing climate-based surveillance procedures to timely strengthening prophylaxis and control.

Experimental yellow fever virus infection in the squirrel monkey (Saimiri spp. ) I: gross anatomical and histopathological findings in organs at necropsy

BACKGROUND Non-human primates contribute to the spread of the yellow fever virus (YFV) and the establishment of transmission cycles in endemic areas. OBJECTIVE To describe the severe histopathological aspects of YFV infection, 10 squirrel monkeys were infected with YFV and blood, brain, liver, kidney, spleen, heart, lung, lymph node and stomach were collected at 1-7, 10, 20 and 30 days post-infection (dpi). METHODS Histopathological analysis and detection of the genome and viral antigens and neutralising antibodies were performed by RT-PCR, immunohistochemistry and neutralisation test, respectively. FINDINGS Only one animal died from the experimental infection. The genome and viral antigens were detected in all investigated organs (1-30 dpi) and the neutralising antibodies from seven to 30 dpi. The brain contained perivascular haemorrhage (6 dpi); in the liver, midzonal haemorrhage and lytic necrosis (6 dpi) were observed. The kidney had bleeding in the Bowman's capsule and tubular necrosis (6 dpi). Pyknotic lymphocytes were observed in the spleen (1-20 dpi), the lung had haemorrhage (2-6 dpi), in the endocardium it contained nuclear pyknosis and necrosis (2-3 dpi) and the stomach contained blood in the lumen (6 dpi). MAIN FINDINGS Squirrel monkeys reliably reproduced the responses observed in human cases of yellow fever and, therefore, constitute an excellent experimental model for studies on the pathophysiology of the disease.

Epidemia de Febre Amarela no Estado do Rio de Janeiro entre 2017 e 2018: correlação entre padrão espacial dos casos e da cobertura vacinal / Yellow Fever Epidemic in Rio de Janeiro State between 2017 and 2018: correlation between spatial pattern of cases and vaccination coverage

Nos últimos anos observamos a emergência e reemergência de algumas doenças em diferentes países e territórios das Américas, incluindo o Brasil, destacando neste estudo, a Febre Amarela. A epidemia de Febre Amarela silvestre que atingiu a Região Sudeste do Brasil em 2017, e a ocorrência de óbitos de macacos com confirmação laboratorial da infecção pelo vírus da Febre Amarela em outros Estados, trouxeram novamente a discussão da necessidade da adoção de medidas para redução do risco de reurbanização da doença. Este estudo teve como objetivo geral analisar a correlação espacial da cobertura vacinal em relação aos casos de Febre Amarela no Estado do Rio de Janeiro. Metodologia: Estudo do tipo ecológico, descritivo, a partir de dados secundários oriundos dos sistemas de informação ­ Sistema de Informação de Agravos de Notificação/investigação (SINAN) e APIWEB. O cenário do estudo foi o Estado do Rio de Janeiro, localizado na Região Sudeste do Brasil. O período escolhido a ser estudado foram os anos de 2017 e 2018, período em que ocorreu o surto da Febre Amarela no Brasil, sobretudo na região supracitada. Foram incluídos todos os casos notificados/confirmados como Febre Amarela no SINAN além dos dados dos vacinados no APIWEB© e excluídos os campos não preenchidos e ignorados. As variáveis utilizadas foram número de casos confirmados, sexo, faixa etária, raça/cor, escolaridade, evolução do caso, vacinados e dados clínicos. Foram construídos mapas temáticos por municípios para a análise espacial da incidência dos casos de febre amarela e cobertura vacinal da população. Foi calculado o índice de Moran Local e Global uni e bivariado pelo software GeoDA, que assumiu como hipótese nula a ausência de autocorrelação espacial da cobertura vacinal e da incidência de Febre Amarela, com significância de 5%. Aprovado pelo Comitê de Ética e Pesquisa da Universidade do Estado do Rio de Janeiro (UERJ), sob parecer nº 3.450.227, aprovado no dia 11 de julho de 2019. Resultados: a Febre Amarela atingiu mais pessoas do sexo masculino, de cor branca, com mais de 60 anos de idade. Ser vacinado é fator de proteção tanto para hospitalização quanto para óbito dos casos confirmados da doença. Os municípios que mantiveram as maiores coberturas vacinais do Estado no período estudado foram Casimiro de Abreu e Comendador Levy Gasparian e os que tiveram maior incidência foram Rio das Flores e Sumidouro. O município de maior incidência em relação a cobertura vacinal (2008 ­ 2018) foi Duas Barras. Conclusões: Esta pesquisa traz dados importantes que auxiliam na compreensão da difusão espacial da Febre Amarela no Estado do Rio de Janeiro em período de surto, dados estes que podem ser facilmente utilizados como modelo para realização de pesquisas de outras doenças.

In recent years we have observed the emergence and reemergence of some diseases in different countries and territories of the Americas, including Brazil, highlighting in this study, Yellow Fever. The outbreak of yellow fever in the southeastern region of Brazil in 2017, and the occurrence of monkey deaths with laboratory confirmation of yellow fever virus infection in other states, brought again the discussion of the need to adopt measures to reduce risk of disease reurbanization. This study aimed to analyze the spatial correlation of vaccination coverage in relation to cases of yellow fever in the state of Rio de Janeiro. Methodology: Ecological, descriptive study, based on secondary data from information systems - Notification / Investigation Disease Information System (SINAN) and APIWEB. The study scenario was the State of Rio de Janeiro, located in the Southeast Region of Brazil. The chosen period to be studied was 2017 and 2018, the period in which the Yellow Fever outbreak occurred in Brazil, especially in the aforementioned region. All cases reported / confirmed as Yellow Fever in SINAN were included, as well as data from APIWEB © vaccinates, and unfilled and ignored fields were excluded. The variables used were number of confirmed cases, gender, age group, race / color, education, case evolution, vaccinated and clinical data. Thematic maps were constructed by municipalities for the spatial analysis of the incidence of yellow fever cases and population vaccination coverage. The univariate bivariate Local and Global Moran index was calculated using the GeoDA software, which assumed that the absence of spatial autocorrelation of vaccine coverage and the incidence of Yellow Fever, with a significance of 5%, was null. Approved by the Research Ethics Committee of the State University of Rio de Janeiro (UERJ), under Opinion No. 3.450.227, approved on July 11, 2019. Results: Yellow Fever struck more males, white, over 60 years of age. Being vaccinated is a protective factor for both hospitalization and death of confirmed cases of the disease. The municipalities that maintained the highest vaccination coverage in the state during the study period were Casimiro de Abreu and Comendador Levy Gasparian and those with the highest incidence were Rio das Flores and Sumidouro. The municipality with the highest incidence in relation to vaccination coverage (2008 - 2018) was Duas Barras. Conclusions: This research provides important data that help in understanding the spatial diffusion of Yellow Fever in the State of Rio de Janeiro during an outbreak period, which can be easily used as a model for conducting research on other diseases.

Development and Clinical Evaluation of a Rapid Diagnostic Test for Yellow Fever Non-Structural Protein 1

A rapid diagnostic test (RDT) kit was developed to detect non-structural protein 1 (NS1) of yellow fever virus (YFV) using monoclonal antibody. NS1 protein was purified from the cultured YFV and used to immunize mice. Monoclonal antibody to NS1 was selected and conjugated with colloidal gold to produce the YFV NS1 RDT kit. The YFV RDTs were evaluated for sensitivity and specificity using positive and negative samples of monkeys from Brazil and negative human blood samples from Korea. Among monoclonal antibodies, clones 3A11 and 3B7 proved most sensitive, and used for YFV RDT kit. Diagnostic accuracy of YFV RDT was fairly high; Sensitivity was 0.0% and specificity was 100% against Dengue viruses type 2 and 3, Zika, Chikungunya and Mayaro viruses. This YFV RDT kit could be employed as a test of choice for point-of-care diagnosis and large scale surveys of YFV infection under clinical or field conditions in endemic areas and on the globe.

Comparative assessment of the replication efficiency of dengue, yellow fever, and chikungunya arboviruses in some insect and mammalian cell lines

Abstract INTRODUCTION: Insect cell cultures play an essential role in understanding arboviral replication. However, the replicative efficiency of some of these viruses such as dengue (DENV), yellow fever (YFV), and chikungunya (CHIKV) in a new cellular substrate (Lulo) and in the other two recognized cell lines has not been comparatively assessed. METHODS: Vero, C6/36, and Lulo cell lines were infected with DENV, YFV, and CHIKV. The viral progeny was quantified through plaque assays and quantitative reverse transcription-polymerase chain reaction, while for DENV2, the findings were confirmed by immunofluorescence antibody assay. RESULTS: The higher DENV2 titer (from multiplicity of infection 0.001) was obtained on day four post-infection in C6/36 and on day six in Vero cells, while the Lulo cell line was almost impossible to infect under the same conditions. However, C6/36 showed the highest values of viral RNA production compared to Vero cells, while the quantification of the viral RNA in Lulo cells showed high levels of viral genomes, which had no correlation to the infectious viral particles. CONCLUSIONS: C6/36 was the most efficient cell line in the alpha and flavivirus production, followed by Vero cells. Thus, Lulo cells may be a useful substrate to study the mechanisms by which cells evade viral replication.

Development and characterization of a packaging cell line for pseudo-infectious yellow fever virus particle generation

Abstract INTRODUCTION: Pseudo-infectious yellow fever viral particles (YFV-PIVs) have been used to study vaccines and viral packaging. Here, we report the development of a packaging cell line, which expresses the YFV prM/E proteins. METHODS: HEK293 cells were transfected with YFV prM/E and C (84 nt) genes to generate HEK293-YFV-PrM/E-opt. The cells were evaluated for their ability to express the heterologous proteins and to package the replicon repYFV-17D-LucIRES, generating YFV-PIVs. RESULTS: The expression of prM/E proteins was confirmed, and the cell line trans-packaged the replicon for recovery of a reporter for the YFV-PIVs. CONCLUSIONS: HEK293-YFV-prM/E-opt trans-packaging capacity demonstrates its possible biotechnology application.

Single shot of 17D vaccine may not confer life-long protection against yellow fever

The yellow fever (YF) vaccine has been used since the 1930s to prevent YF, which is a severe infectious disease caused by the yellow fever virus (YFV), and mainly transmitted by Culicidae mosquitoes from the genera Aedes and Haemagogus . Until 2013, the World Health Organization (WHO) recommended the administration of a vaccine dose every ten years. A new recommendation of a single vaccine dose to confer life-long protection against YFV infection has since been established. Recent evidence published elsewhere suggests that at least a second dose is needed to fully protect against YF disease. Here, we discuss the feasibility of administering multiple doses, the necessity for a new and modern vaccine, and recommend that the WHO conveys a meeting to discuss YFV vaccination strategies for people living in or travelling to endemic areas.

Metagenomic analysis reveals Hepatitis A virus in suspected yellow fever cases in Brazil

Using a metagenomic approach, we identified hepatitis A virus among cases of acute febrile illnesses that occurred in 2008-2012 in Brazil suspected as yellow fever. These findings reinforce the challenge facing routine clinical diagnosis in complex epidemiological scenarios.

Production and Storage of Virus Simulants

We have examined isolation and identification protocols for three virus simulant candidates to biological warfare agents. MS2 phage, a simulant for yellow fever virus and Hantaan virus, was propagated using as a host an E. coli strain with F pilus. MS2 phage genome was examined by reverse transcription and polymerase chain reaction (RT-PCR). Coat protein of the phage preparation was examined by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and mass spectrometric analysis. Cydia pomonella granulosis virus (CpGV) is a virus simulant candidate to smallpox virus. CpGV was isolated from a commercialized CpGV pellet. In this study, we developed new isolation and identification protocols for CpGV. One disadvantage of using CpGV is that it is not easy to determine viability of the virus. Here, we have included T4 phage as an alternative. We established a high titer production protocol and developed an easy genome identification protocol that does not require purified phage DNA. Stability of these virus preparations was also examined under various storage conditions. When the virus preparations were not subjected to freeze drying, MS2 phage was most stable when it was stored in liquid nitrogen but unstable at 4℃. In contrast, T4 phage was most stable when it was stored at 4℃. CpGV was stable at −20℃ but not at 4℃. Stability during or after freeze drying was also investigated. The result showed that 70~80% MS2 survived the freeze drying process. In contrast, only about 15% of T4 phage survived during the freeze drying. CpGV was found to be degraded during freeze drying.

Estudo de mutações nas proteínas virais E, NS3 e NS4B no genoma do vírus vacinal da Febre Amarela 17D / Study of mutations in the viral proteins E, NS3 and NS4B in the genome of the yellow fever vaccine virus 17D

Resumo: A Febre Amarela (FA) é uma doença infecciosa não contagiosa, causada por um arbovírus e objeto de preocupação sanitária mundial. A principal medida de controle é a vacinação com o vírus atenuado da FA, cepa 17D, que é capaz de induzir resposta imune protetora a longo prazo com administração em dose única. Vírus atenuados são potentes vetores de expressão, pois disseminam o antígeno no hospedeiro e induzem resposta imune protetora, contribuindo para o desenvolvimento de vacinas recombinantes. Diversas estratégias foram desenvolvidas para expressão de genes heterólogos pelo vírus vacinal FA 17D. Entretanto, a inserção induz proliferação viral e imunogenicidade reduzidas. Neste trabalho foi utilizado o vírus vacinal FA 17D recombinante expressando a proteína repórter enhanced green fluorescent protein (EGFP) para avaliar o impacto de mutações específicas que possam modular a replicação viral. Mutações nas proteínas E, NS3 e NS4B foram descritas por aumentarem a proliferação viral em cultura de células e em camundongos, no genoma do vírus da dengue, sorotipos 1 e 2. As mutações em E400 (F→L), E403 (T→I), NS3439 (V→S) e NS4B54 (L→F) foram inseridas no genoma do vírus FA/EGFP, com a finalidade de caracterizar o seu efeito na proliferação viral e na indução de resposta imune humoral. O cDNA do genoma viral FA/EGFP foi utilizado para gerar vírus recombinantes carreando uma, duas ou três mutações. O estudo de proliferação viral foi realizado por cinética de infecção de células das linhagens Vero, Huh7 e C6/36. Os resultados mostram que os vírus da FA recombinantes se proliferam menos que o vírus vacinal FA 17DD. Além disso, a infectividade dos vírus mutantes em células de mamífero é diferente da infectividade em células de mosquitoOs vírus que carreiam as mutações em E400/NS3439 e E400/NS3439/NS4B54 tem a proliferação viral significativamente prejudicada em células de mamífero. Os vírus que carreiam a mutação em E400 apresentaram aumento de proliferação viral em comparação com o vírus FA/EGFP original, em células de mosquito. As diferenças entre os tipos celulares podem ter sido causadas pelas características fisiológicas das células durante a infecção viral e pelas diferenças de propriedades das proteínas virais ocasionadas pela inserção das mutações. Não foi possível recuperar partículas virais infecciosas carreando a mutação em E403. A modelagem molecular das proteínas virais mostrou diferenças discretas de carga, volume de superfície proteica e propriedade físico-química induzidas pelas mutações. Nenhuma das mutações influenciou nas interações intramoleculares. A imunogenicidade foi avaliada por imunização de camundongos das linhagens C57BL/6 e BALB/c com os vírus carreando mutações únicas e os soros foram analisados por PRNT e ELISA para obter os títulos de anticorpos neutralizantes para FA e anticorpos para GFP, respectivamente. Os soros dos camundongos imunizados com os vírus recombinantes apresentam menores títulos de anticorpos neutralizantes em comparação ao grupo imunizado com o vírus vacinal, porém não houveram alterações na indução de anticorpos para GFP. De maneira geral, as mutações em E400 e E403 produziram maiores efeitos sobre a proliferação viral e as mutações NS3439 e NS4B54, na imunogenicidade. (AU)