Serological evidence of orthopoxvirus infection in neotropical primates in Brazil

The genus Orthopoxvirus (OPXV) of the family Poxviridae comprises several viruses that are capable of infecting a wide range of hosts. One of the most widespread OPXVs is the Vaccinia virus (VACV), which circulates in zoonotic cycles in South America, especially in Brazil, infecting domestic and wild animals and humans and causing economic losses as well as impacting public health. Despite this, little is known about the presence and/or exposure of neotropical primates to orthopoxviruses in the country. In this study, we report the results of a search for evidence of OPVX infections in neotropical free-living primates in the state of Minas Gerais, southeast Brazil. The sera or liver tissues of 63 neotropical primates were examined through plaque reduction neutralization tests (PRNT) and real-time PCR. OPXV-specific neutralizing antibodies were detected in two sera (4.5%) from Callithrix penicillata, showing 55% and 85% reduction in plaque counts, evidencing their previous exposure to the virus. Both individuals were collected in urban areas. All real-time PCR assays were negative. This is the first time that evidence of OPXV exposure has been detected in C. penicillata, a species that usually lives at the interface between cities and forests, increasing risks of zoonotic transmissions through spillover/spillback events. In this way, studies on the circulation of OPXV in neotropical free-living primates are necessary, especially now, with the monkeypox virus being detected in new regions of the planet

Desenvolvimento e avaliação de uma plataforma de diagnóstico para meningoencefalites virais por PCR em tempo real

Exportado OPUSMade available in DSpace on 2019-08-11T13:18:30Z (GMT). No. of bitstreams: 1 tese___danilo_bretas_de_oliveira.pdf: 7693858 bytes, checksum: 1fc7bbd0387989e1cb980fcdd91c0e6a (MD5) Previous issue date: 30Infecções no sistema nervoso central (SNC) apresentam distribuição global, acometendo populações em todas as partes do globo. Infecções virais são as principais causas de infecção no SNC em todo o mundo. No Brasil, em média, são notificados 11.500 casos/ano de meningite/meningoencefalite com provável etiologia viral. Entretanto, para a maioria dos casos não há identificação do agente etiológico. Assim, o objetivo deste trabalho foi a utilização da PCR em tempo real (qPCR) para desenvolvimento e avaliação de uma plataforma de diagnóstico para meningoencefalites causadas por vírus. Para isso, foram utilizadas amostras de agentes virais causadores de meningite para obtenção de plasmídeos controle e para padronização da PCR. Foram desenhados iniciadores específicos para detecção de cada vírus separadamente ou grupos virais. Todas as reações desenvolvidas foram padronizadas e apresentaram eficiência satisfatória: ENTV 110%; FLAV 92%%; HHV-1/2 114%%; HHV-3 113%%; SLEV 94%; YFV 96%; JEV 97%; WNV 98%. Todos os testes foram específicos na detecção do alvo e sensíveis, com sensibilidade analítica inferior ou igual a 10 cópias/mL. Os ensaios desenvolvidos apresentaram coeficiente de variação inferior a 5% em testes de reprodutibilidade e de repetibilidade. Além dos ensaios baseados em qPCR foram padronizadas etapas pré-analíticas para aumentar a eficiência de detecção viral. A plataforma de diagnóstico foi avaliada em amostras clínicas de líquido cefalorraquidiano (LCR), sendo capaz de detectar vírus em 62,9% das amostras de pacientes com suspeita de meningoencefalite viral. Foram detectados ENTVs (70%), HHV-3 (13%), DENVs (11%), HSV-1/2 e HHV-5 (2%). O DNA amplificado de amostras de LCR positivas foi sequenciado e os resultados da plataforma qPCR foram confirmados. Também foram detectados vírus em amostras de pacientes com diagnóstico de meningite bacteriana. A plataforma desenvolvida foi eficiente para o diagnóstico de infecções no SNC e apresenta potencial de aplicação no sistema de saúde.Infections on central nervous system (CNS) have a global distribution, affecting people worldwide. Viral infections are the main cause of infection in the CNS. In Brazil, on average, 11,500 cases/year of meningitis/meningoencephalitis with probable viral etiology are reported. However, most cases there are no identification of the etiologic agent. The objective of this work was the use of real time PCR (qPCR) for development and evaluation of a platform of diagnostic for meningoencephalitis caused by viruses. Viruses were used to obtain plasmids for control and standardization of qPCR. Specific primers were designed to detect each virus or viral groups. All reactions were standardized and presented efficiency of: ENTV 110%; FLAV 92%; HHV-1/2 114%; HHV-3 113%; SLEV 94%; YFV 96%; 97% JEV; 98% WNV. All tests were specific in the target detection and analytical sensitivities were equal or less to 10 copies/ml. The developed tests showed coefficient of variation less than 5% in reproducibility and repeatability assays. In addition to qPCR assays pre-analytical steps were standardized to improve viral detection efficiency. Finally, the qPCR platform was applied on clinical samples of cerebrospinal liquid (CSF), and it was able to detect the virus in 62.9% of samples from patients with suspected viral meningoencephalitis. ENTVs were detected (70%), HHV-3 (13%), DENVs (11%), HSV-1/2 and HHV-5 (2%). The amplified DNAs in qPCR positive samples were sequenced and the results were confirmed. Viruses were also detected in samples of patients with bacterial meningitis. So the developed platform showed high efficiency for diagnosis of infections in CNS and has a potential use in health system

A Model to Detect Autochthonous Group 1 and 2 Brazilian Vaccinia virus Coinfections: Development of a qPCR Tool for Diagnosis and Pathogenesis Studies

Vaccinia virus (VACV) is the etiological agent of bovine vaccinia (BV), an emerging zoonosis that has been associated with economic losses and social effects. Despite increasing reports of BV outbreaks in Brazil, little is known about the biological interactions of Brazilian VACV (VACV-BR) isolates during coinfections; furthermore, there are no tools for the diagnosis of these coinfections. In this study, a tool to co-detect two variants of VACV was developed to provide new information regarding the pathogenesis, virulence profile, and viral spread during coinfection with VACV-BR isolates. To test the quantitative polymerase chain reactions (qPCR) tool, groups of BALB/c mice were intranasally monoinfected with Pelotas virus 1—Group II (PV1-GII) and Pelotas virus 2—Group I (PV2-GI), or were coinfected with PV1-GII and PV2-GI. Clinical signs of the mice were evaluated and the viral load in lung and spleen were detected using simultaneous polymerase chain reactions (PCR) targeting the A56R (hemagglutinin) gene of VACV. The results showed that qPCR for the quantification of viral load in coinfection was efficient and highly sensitive. Coinfected mice presented more severe disease and a higher frequency of VACV detection in lung and spleen, when compared to monoinfected groups. This study is the first description of PV1 and PV2 pathogenicity during coinfection in mice, and provides a new method to detect VACV-BR coinfections

Twenty Years after Bovine Vaccinia in Brazil: Where We Are and Where Are We Going?

Orthopoxvirus (OPV) infections have been present in human life for hundreds of years. It is known that Variola virus (VARV) killed over 300 million people in the past; however, it had an end thanks to the physician Edward Jenner (who developed the first vaccine in history) and also thanks to a massive vaccination program in the 20th century all over the world. Although the first vaccine was created using the Cowpox virus (CPXV), it turned out later that the Vaccinia virus was the one used during the vaccination program. VACV is the etiological agent of bovine vaccinia (BV), a zoonotic disease that has emerged in Brazil and South America in the last 20 years. BV has a great impact on local dairy economies and is also a burden to public health. In this review, we described the main events related to VACV and BV emergence in Brazil and South America, the increase of related scientific studies, and the issues that science, human and animal medicine are going to face if we do not be on guard to this virus and its disease

Ocular Vaccinia Infection in Dairy Worker, Brazil

We studied a clinical case of vaccinia virus that caused an ocular manifestation in a dairy worker in Brazil. Biologic and molecular analyses identified a co-infection with 2 isolates from different Brazilian vaccinia virus phylogenetic groups

Infection of the central nervous system with dengue virus 3 genotype I causing neurological manifestations in Brazil

Abstract: A case of dengue virus 3 (DENV-3) genotype I infection with neurological manifestations occurred in Belo Horizonte, Minas Gerais in October 2012. The serotype was detected by PCR, and the genotype was assessed by sequencing and phylogenetic analysis of the C-prM region. The virus causing neurological manifestations clustered with other sequences of DENV-3 genotype I. Because neurological manifestations of DENV are possibly misdiagnosed in Brazil, this study serves as an alert of the importance of DENV diagnoses in CNS infections

Basal Activation of Type I Interferons (Alpha2 and Beta) and 2′5′OAS Genes: Insights into Differential Expression Profiles of Interferon System Components in Systemic Sclerosis

Objective. Systemic sclerosis (SSc) is a complex autoimmune disease in which interferons (IFNs) may play an essential role. We hypothesized that type I and III IFNs may be found in increased levels in patients and be responsible for SSc autoimmune status. Methods. Type I and III IFN and ISG basal expression profiles were measured by qPCR using RNA from PBMCs of patients and controls . Results. Type I IFNs are increased in SSc patients, while no induction of type III IFNs was detected. This induction cannot be related to IRF7, since no upregulation of this gene was seen on patients. Of the ISGs tested, 2′5′OAS levels were increased in patients, while 6–16 and MxA levels were not. Conclusions. While there is no indication of type III IFN induction, increased levels of type I IFNs may lead to abnormal regulation of ISGs that can be responsible for immune system alterations described for SSc