Molecular identification of Saint Louis encephalitis virus genotype IV in Colombia

Saint Louis encephalitis virus (SLEV) is a member of the Japanese-encephalitis virus serocomplex of the genus Flavivirus. SLEV is broadly distributed in the Americas and the Caribbean Islands, where it is usually transmitted by mosquitoes of the genus Culex and primarily to birds and mammalian-hosts. Humans are occasionally infected by the virus and are dead-end hosts. SLEV causes encephalitis in temperate regions, while in tropical regions of the Americas, several human cases and a wide biological diversity of SLEV-strains have been reported. The phylogenetic analysis of the envelope (E) protein genes indicated eight-genotypes of SLEV with geographic overlap. The present paper describes the genotyping of two SLEV viruses detected in mosquito-pools collected in northern Colombia (department of Cordoba). We used reverse transcription-polymerase chain reaction to amplify a fragment of theE-gene to confirm the virus identity and completeE-gene sequencing for phylogenetic analysis and genotyping of the two-SLEV viruses found circulating in Córdoba. This is the first report of SLEV genotype IV in Colombia (Córdoba) in mosquitoes from a region of human inhabitation, implicating the risk of human disease due to SLEV infection. Physicians should consider SLEV as a possible aetiology for undiagnosed febrile and neurologic syndromes among their patients who report exposure to mosquito-bites.

Saint louis encpphalitis virus in mato grosso, central-western Brazil / Vírus da encefalite de Saint Louis em Mato Grosso, Centro-Oeste, Brasil

The dengue virus (DENV), which is frequently involved in large epidemics, and the yellow fever virus (YFV), which is responsible for sporadic sylvatic outbreaks, are considered the most important flaviviruses circulating in Brazil. Because of that, laboratorial diagnosis of acute undifferentiated febrile illness during epidemic periods is frequently directed towards these viruses, which may eventually hinder the detection of other circulating flaviviruses, including the Saint Louis encephalitis virus (SLEV), which is widely dispersed across the Americas. The aim of this study was to conduct a molecular investigation of 11 flaviviruses using 604 serum samples obtained from patients during a large dengue fever outbreak in the state of Mato Grosso (MT) between 2011 and 2012. Simultaneously, 3,433 female Culex spp. collected with Nasci aspirators in the city of Cuiabá, MT, in 2013, and allocated to 409 pools containing 1-10 mosquitoes, were also tested by multiplex semi-nested reverse transcription PCR for the same flaviviruses. SLEV was detected in three patients co-infected with DENV-4 from the cities of Cuiabá and Várzea Grande. One of them was a triple co-infection with DENV-1. None of them mentioned recent travel or access to sylvatic/rural regions, indicating that transmission might have occurred within the metropolitan area. Regarding mosquito samples, one pool containing one Culex quinquefasciatus female was positive for SLEV, with a minimum infection rate (MIR) of 0.29 per 1000 specimens of this species. Phylogenetic analysis indicates both human and mosquito SLEV cluster, with isolates from genotype V-A obtained from animals in the Amazon region, in the state of Pará. This is the first report of SLEV molecular identification in MT.

O vírus da dengue (DENV), frequentemente envolvido em epidemias de grande proporção, e o vírus da febre amarela (YFV), responsável por surtos silvestres esporádicos, são considerados os flavivírus circulantes mais importantes no Brasil. Por este motivo, o diagnóstico laboratorial de doença febril aguda indiferenciada durante períodos epidêmicos é frequentemente direcionado para dengue e febre amarela no país, dificultando a detecção de outros arbovírus possivelmente circulantes, incluindo o vírus da encefalite de Saint Louis (SLEV), que é amplamente disperso nas Américas. O objetivo deste estudo foi investigar molecularmente a presença de 11 flavivírus no soro de 604 pacientes durante grande epidemia de dengue no estado de Mato Grosso (MT), Centro-Oeste do Brasil, entre 2011- 2012. Concomitantemente, 3.433 fêmeas de Culex spp. capturadas com aspirador de Nasci na cidade de Cuiabá, MT e alocadas em 409 pools com 1-10 mosquitos em 2013 foram testadas por multiplex seminested RT-PCR para os mesmos flavivírus. O SLEV foi detectado em três pacientes co-infectados com o DENV-4 das cidades de Cuiabá e Várzea Grande, MT. Um dos pacientes apresentava tripla co-infecção com DENV-1. Nenhum paciente referiu histórico recente de viagem ou acesso a áreas rurais/silvestres. Um pool contendo uma fêmea de Culex quinquefasciatus foi positivo para o SLEV, apresentando taxa de infecção mínima (MIR) de 0,29 por 1000 espécimes desta espécie. A análise filogenética indica que ambas as amostras formam um cluster com isolados do genótipo V-A do SLEV obtidos de animais na região amazônica do estado do Pará. Este é o primeiro relato de identificação molecular do SLEV no MT.

Genetic characterization of St. Louis encephalitis virus isolated from human in São Paulo, Brazil

The molecular characterization of SPH253157, a new strain of St. Louis encephalitis virus (SLEV), isolated in 2004 from the first case of human infection recognized in the state of São Paulo, Brazil, is reported. The patient, presenting a febrile illness without neurological involvement, was hospitalized as a probable case of dengue fever. Genomic RNA was isolated from the supernatant of C6/36 cells infected with acute phase-serum specimen of the patient and the envelope gene was amplified by reverse-transcription-polymerase chain reaction. The complete nucleotide sequence of the envelope gene of this isolate was directly sequenced from the amplified products and compared with other Brazilian and American SLEV strains. Phylogenetic analyses were carried out under maximum likelihood criterion with outgroups both included and excluded. Outgroups comprised four flavivirus of the Japanese encephalitis group. Phylogeny also included Bayesian analysis. The results indicated that the new SLEV isolate belongs to lineage III, being closely related to an Argentinean strain recovered from Culex sp. in 1979. It is concluded that there are at least 3 lineages of SLEV in Brazil.

St. Louis encephalitis vírus: first isolation from a human in São Paulo state, Brasil

O presente estudo relata o isolamento do vírus da encefalite São Luis (SLEV) de um caso febril humano suspeito de dengue, em São Pedro, Estado de São Paulo. MAC-ELISA realizado com soros das fases aguda e convalescente foi inconclusivo e anticorpos IgG foram detectados por inibição da hemaglutinação para flavivirus. Imunofluorescência indireta com cultura de células C6/36 inoculadas com soro da fase aguda foi positivo para flavivirus mas negativo quando testado com anticorpos monoclonais para dengue. O RNA extraído de cultura de células infectadas foi amplificado na presença de primers universais para o gênero Flavivirus, deduzidos de uma região da proteína não estrutural 5 e diretamente sequenciado. Os resultados da pesquisa no BLAST indicaram que a seqüência apresenta 93% de similaridade de nucleotídeos com a seqüência de SLEV (cepa MS1.7), confirmado por RT-PCR, realizado com primers específicos para SLEV. O fato de SLEV ter sido identificado como a causa de doença humana indica a necessidade de aprimorar a vigilância a fim de detectar precocemente esse agente no Estado de São Paulo e no Brasil. Esse caso é também um alerta para os profissionais de saúde sobre a necessidade de investigações clínicas e epidemiológicas mais completas sobre doenças febris como no caso relatado. Infecções por SLEV podem não ser reconhecidas ou confundidas com outras causadas por arbovírus como a dengue.