Genetic characterization of the rare Bruconha virus (Bunyavirales: Orthobunyavirus) isolated in Vale do Ribeira (Atlantic Forest biome), Southeastern Brazil

ABSTRACT Brazil is a great source of arbovirus diversity, mainly in the Amazon region. However, other biomes, especially the Atlantic Forest, may also be a hotspot for emerging viruses, including Bunyaviruses (Negarnaviricota: Bunyavirales). For instance, Vale do Ribeira, located in the Southeastern region, has been widely studied for virus surveillance, where Flavivirus, Alphavirus and Bunyaviruses were isolated during the last decades, including Bruconha virus (BRCV), a member of Orthobunyavirus genus Group C, in 1976. Recently, a new isolate of BRCV named Span321532 was obtained from an adult sentinel mouse placed in Iguape city in 2011, and a full-length genome was generated with nucleotide differences ranging between 1.5%, 5.3% and 5% (L, M and S segments, respectively) from the prototype isolated 35 years earlier. In addition, each segment placed BRCV into different clusters, showing the high variety within Bunyavirales. Although no evidence for reassortants was detected, this finding reiterates the need for new surveillance and genomic studies in the area considering the high mutation rates of arbovirus, and also to identify the hosts capable of supporting the continuous circulation of Orthobunyavirus.

Zika virus serological diagnosis: commercial tests and monoclonal antibodies as tools

Zika virus (ZIKV), an emerging arthropod-borne virus (arbovirus) of the Flaviviridae family, is a current issue worldwide, particularly because of the congenital and neurological syndromes associated with infection by this virus. As the initial clinical symptoms of all diseases caused by this group are very similar, clinical diagnosis is difficult. Furthermore, laboratory diagnostic efforts have failed to identify specific and accurate tests for each virus of the Flaviviridae family due to the cross-reactivity of these viruses in serum samples. This situation has resulted in underreporting of the diseases caused by flaviviruses. However, many companies developed commercial diagnostic tests after the recent ZIKV outbreak. Moreover, health regulatory agencies have approved different commercial tests to extend the monitoring of ZIKV infections. Considering that a specific and sensitive diagnostic method for estimating risk and evaluating ZIKV propagation is still needed, this review aims to provide an update of the main commercially approved serological diagnostics test by the US Food and Drug Administration (FDA) and Brazilian National Health Surveillance Agency (ANVISA). Additionally, we present the technologies used for monoclonal antibody production as a tool for the development of diagnostic tests and applications of these antibodies in detecting ZIKV infections worldwide.(AU)

Occurrence of antibodies against Leptospira spp. in donkeys of São Paulo state / Ocorrência de anticorpos contra Leptospira spp. em asininos no estado de São Paulo

Among the diseases that affect equines, bacterial diseases play an important role from a health and economic point of view, especially leptospirosis and brucellosis. The study aimed to provide information on the occurrence of anti-Leptospira spp. and anti-Brucella abortus antibodies in donkeys of São Paulo state. We found a frequency of 62.4% (53/85) antibodies against Leptospira spp. The donkeys were not seropositive for Brucella spp.(AU)

Entre as doenças que acometem os equídeos, as enfermidades bacterianas assumem um papel importante do ponto de vista sanitário e econômico, destacando-se a leptospirose e a brucelose. O estudo teve como objetivo fornecer informações sobre a ocorrência de anticorpos anti-Leptospira spp. e anti-Brucella abortus em jumentos no estado de São Paulo. Estimou-se que 62,4% (53/85) dos animais apresentavam anticorpos anti-Leptospira spp. Os jumentos estudados não foram sororreagentes contra a Brucella spp.(AU)

Simple protocol for population (Sanger) sequencing for Zika virus genomic regions

BACKGROUND A number of Zika virus (ZIKV) sequences were obtained using Next-generation sequencing (NGS), a methodology widely applied in genetic diversity studies and virome discovery. However Sanger method is still a robust, affordable, rapid and specific tool to obtain valuable sequences. OBJECTIVE The aim of this study was to develop a simple and robust Sanger sequencing protocol targeting ZIKV relevant genetic regions, as envelope protein and nonstructural protein 5 (NS5). In addition, phylogenetic analysis of the ZIKV strains obtained using the present protocol and their comparison with previously published NGS sequences were also carried out. METHODS Six Vero cells isolates from serum and one urine sample were available to develop the procedure. Primer sets were designed in order to conduct a nested RT-PCR and a Sanger sequencing protocols. Bayesian analysis was used to infer phylogenetic relationships. FINDINGS Seven complete ZIKV envelope protein (1,571 kb) and six partial NS5 (0,798 Kb) were obtained using the protocol, with no amplification of NS5 gene from urine sample. Two NS5 sequences presented ambiguities at positions 495 and 196. Nucleotide analysis of a Sanger sequence and consensus sequence of previously NGS study revealed 100% identity. ZIKV strains described here clustered within the Asian lineage. MAIN CONCLUSIONS The present study provided a simple and low-cost Sanger protocol to sequence relevant genes of the ZIKV genome. The identity of Sanger generated sequences with published consensus NGS support the use of Sanger method for ZIKV population studies. The regions evaluated were able to provide robust phylogenetic signals and may be used to conduct molecular epidemiological studies and monitor viral evolution.

Outbreak of Yellow Fever among Nonhuman Primates, Espirito Santo, Brazil, 2017

In January 2017, a yellow fever outbreak occurred in Espirito Santo, Brazil, where human immunization coverage is low. Histologic, immunohistologic, and PCR examinations were performed for 22 deceased nonhuman New World primates; typical yellow fever features were found in 21. Diagnosis in nonhuman primates prompted early public health response.

Em janeiro de 2017, ocorreu um surto de febre amarela no Espírito Santo, Brasil, onde a cobertura de imunização humana é baixa. Os exames histológico, imuno-histológico e PCR foram realizados para 22 primatas não humanos do Novo Mundo falecidos; características típicas da febre amarela foram encontradas em 21. O diagnóstico em primatas não humanos levou a uma resposta precoce da saúde pública.

Occurrence of viral diseases in donkeys (Equus asinus) in São Paulo State, Brazil / Ocorrência de enfermidades virais em asininos (Equus asinus) no estado de São Paulo, Brasil

Among the diseases that affect equines, viral diseases play an important role from a health and economic point of view, especially influenza, viral arteritis, herpes infections and vesicular stomatitis. In the Brazilian literature, there is little or no account of the occurrence of infectious diseases in donkeys. Given the importance of donkeys in different activities and the lack of information on infections that may occur in these animals, the aim of this study was to determine the frequency of anti-equine herpesvirus (EHV), anti-equine arteritis virus (EAV), anti-vesicular stomatitis, and anti-equine influenza (H3N8) antibodies in the serum of 85 donkeys bred in some regions of the state of São Paulo. We found the following antibody frequencies: 50.6% (43/85) antibodies against influenza virus subtype H3N8, 47% (40/85) anti-EHV, and 20% (17/85) anti-EAV. The donkeys were not seropositive for vesicular stomatitis. The results suggested that the agents EHV, EAV, and equine influenza subtype H3N8 circulate among donkeys in some regions of the state of São Paulo, Brazil, reinforcing the importance of establishing a routine diagnosis and epidemiological study of this species.(AU)

Dentre as doenças que acometem os equídeos, as enfermidades virais assumem um papel importante do ponto de vista sanitário e econômico, especialmente a influenza, arterite viral, as infecções herpéticas e a estomatite vesicular. Na literatura nacional, existe pouco ou nenhum relato sobre a ocorrência de enfermidades infecciosas nos asininos. Tendo em vista a importância dos asininos para diferentes atividades e a falta de informações sobre as doenças que acometem esses animais, este trabalho teve como objetivo estudar a frequência de anticorpos anti-EHV, antivírus da arterite equina, anti-estomatite vesicular e anti-influenza equina (H3N8) em 85 soros de jumentos criados no estado de São Paulo. Estimou-se que 50,6% apresentavam anticorpos contra o subtipo H3N8 do vírus da influenza; 47% (40/85) apresentavam anticorpos contra o EHV e 20% apresentavam anticorpos contra o vírus da arterite. Os jumentos não foram soro reagentes contra a estomatite vesicular. Os resultados obtidos sugerem que os agentes EHV, vírus da arterite equina e influenza equina subtipo H3N8, circulam entre os jumentos do estado de São Paulo, caracterizando a importância do estabelecimento de uma rotina diagnóstica e estudos epidemiológicos na espécie.(AU)

First complete Genome Sequence of Zika Virus (Flaviviridae, Flavivirus) from an Autochthonous Transmission in Brazil

We report here the genome sequence of Zika virus, strain ZikaSPH2015, containing all structural and nonstructural proteins flanked by the 5' and 3' untranslated region. It was isolated in São Paulo state, Brazil, in 2015, from a patient who received a blood transfusion from an asymptomatic donor at the time of donation.

Inquérito sorológico da infecção por herpesvírus equino no estado de Minas Gerais / Serological survey of equine herpesvirus infection in Minas Gerais state

Os herpesvírus equinos tipo 1 (HVE-1) e 4 (HVE-4) são agentes causadores de diferentes formas de doença em cavalos, das quais as mais comuns são a rinopneumonite, o abortamento, a mortalidade perinatal e a mieloencefalopatia herpética equinas, que causam grandes perdas econômicas. Tem sido descrita mundialmente, havendo poucos estudos no Brasil. O objetivo do presente trabalho foi estudar a ocorrência e a distribuição da infecção por herpesvírus equinos (HVE) em equídeos criados em dez Delegacias Regionais do Estado de Minas Gerais: Almenara, Bambuí, Curvelo, Governador Valadares, Montes Claros, Oliveira, São Gonçalo do Sapucaí, Teófilo Otoni, Unaí e Viçosa. Foi utilizada a técnica de soroneutralização em microplacas com o intuito de detectar anticorpos soro neutralizantes. Das amostras analisadas, 17,6% (145/826) foram soropositivas para o HVE, sendo 18,7% (140/749) cavalos soropositivos, 6,8% (5/73) muares soropositivos e nenhum asinino soropositivo (0/4). Conclui-se que o HVE-1 encontra-se amplamente disseminado no Estado de Minas Gerais, pois todas as regiões estudadas apresentaram animais sororreagentes ao HVE-1. Observou-se maior ocorrência de anticorpos contra o HVE em animais adultos, indicando assim o potencial desses animais como fonte de infecção para os potros.

Equine herpesvirus type 1 (EHV-1) and 4 (EHV-4) are major pathogens affecting horses, and cause respiratory disease, abortion, perinatal mortality and neurological disease, bringing economical losses. This infection has been reported worldwide, but there are only a few studies in Brazil. The present study was undertaken in order to evaluate the occurrence and distribution of equine herpesviruses (EHV) infection in equids from ten regions of Minas Gerais State: Almenara, Bambuí, Curvelo, Governador Valadares, Montes Claros, Oliveira, São Gonçalo do Sapucaí, Teófilo Otoni, Unaí and Viçosa. To detect antibodies against EHV virus neutralization test in microplates was used. We found 17.6% (145/826) positive animals for EHV. 18.7% (140/749) positive horses, 6.8% (5/73) positive mules and none positive (0/4) donkeys. All ten regions studied showed animals reagents to EHV. The results suggest that EHV is widespread in equids of Minas Gerais State. It was observed a higher occurrence of antibodies against EHV in adult animals, indicating the potential of these animals as source of infection for foals.

Ocorrência de leucoencefalomalácia (LEME) em equídeos no estado de São Paulo, Brasil: achados anatomopatológicos / Occurrence of leukoencephalomalacia in equids in São Paulo state, Brazil: anatomopathological findings

O trabalho relata a ocorrência de leucoencefalomalácia em equídeos (LEME) com sintomatologia nervosa e com diagnóstico negativo para raiva, herpesvírus equino e encefalomielite equina durante o período de dois anos, no Estado de São Paulo, Brasil. Foram examinadas 67 amostras de sistema nervoso central e em 10,4% (cinco equinos, um pônei e um asinino) observaram-se lesões macroscópicas de LEME, confirmadas pela análise histopatológica. Os animais acometidos eram cinco machos e duas fêmeas, com idades que variavam de 11 meses a nove anos. Os sete casos ocorreram tanto no inverno como em outras estações do ano. As principais manifestações clínicas relatadas foram incoordenação, ataxia, paralisia dos membros posteriores, profunda depressão, levando ao óbito. Macroscopicamente, observaram-se congestão dos vasos meníngeos, áreas de malácia da substância branca, caracterizadas por coloração amarelada e/ou hemorrágica, com cavitação e amolecimento circundados por hiperemia. As lesões microscópicas observadas em todos os casos eram de necrose de liquefação da substância branca do cérebro, caracterizada por substância eosinofílica amorfa e homogênea, presença de edema axonal e perivascular, hemorragia e vacuolização do neurópilo adjacente e esferoides axonais. Em algumas áreas de malácia havia também células Gitter. Em apenas um animal observou-se manguito perivascular mononuclear. O presente trabalho confirma que o diagnóstico diferencial é importante na distinção da LEME com outras neuropatias encefálicas que acometem equídeos. A ocorrência da LEME relatada neste estudo demonstra que esta enfermidade é importante para a equideocultura do Estado de São Paulo.

This article describes clinical and pathological findings of leukoencephalomalacia in equids with neurological signs which tested negative to rabies, equine herpesvirus and equine encephalomyelitis. This work was carried during the period of two years in São Paulo State, Brazil. A total of 67 brain samples were examined and in 10.4% (five equines, one poney, and one donkey) were observed gross lesions of ELEM, confirmed by histopathological analysis. The animals were five males and two females ranged from 11 months to nine years old. The seven cases happened in all seasons of the year. The most characteristic clinical signs were incoordination, ataxia, paralysis of the hind legs, profound depression and death. Necropsy was performed to collect brain samples for virological and histopathological diagnosis. Gross lesions included congestion of meningeal blood vessels, malacia of the white matter characterized by yellowish depressed areas sometimes hemorrhagic, with cavitations, and softening surrounded by hyperemic area. Microscopically, the lesions were liquefactive necrosis of the white matter brain, characterized by eosinophilic and amorphous material, axonal and perivascular edema, hemorrhage and vacuolization of the neuropil and axonal sferoids. Gitter cells were seen in some areas of malacia. Perivascular mononuclear cuffing was observed in only one case. The present study confirms that differential diagnosis is very important to distinguish equid neuropathies. The occurence of ELEM in the present study shows that the disease is important for the equideoculture in São Paulo State.