An indirect immunofluorescence assay to detect antibodies against St. Louis Encephalitis virus

An in house indirect immmunofluorescence assay ( IFA ) in relation to neutralization (NT) reference test, was assessed as a fast and cheap method to carry out serological surveys for St. Louis Encephalitis virus (SLE). Sera obtained from 213 blood donors were analyzed by both tests. The prevalence of seropositivity obtained with IFA was lower than (30.98%) that observed on NT (41.78%). The relative specificity rate of IFA was 96.77% whereas its relative sensitivity rate was 69.66%. Kappa index showed a good correlation between both tests. The results indicate that neutralization assay is still the serological test with the highest sensitivity and specificity relative rates for detecting antibodies against SLE virus. Nevertheless, the IFA could be useful as an alternative test in order to learn the circulation of the Flavivirus genus in a certain area.

Age-related seroprevalence study for St. Louis encephalitis in a population from Cordoba, Argentina

A serological screening was performed in 615 individuals aged 0-87 years, living in the city of Cordoba, Argentina to study the relationship between antibody prevalence for the SLE virus and age. A 13.98% prevalence of neutralizing antibodies was obtained and its relation to age was significantly high (p = 0.045). The highest seroprevalence was noted on individuals over 60 years old (>20%), whereas no subject under 10 was seropositive for this virus. Our results confirm that the agent is endemic in this area and neurological pathology studies should be performed on those individuals aged 60 since they represent the most susceptible group to SLE virus.

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

This paper reports the isolation of St. Louis encephalitis virus (SLEV) from a febrile human case suspected to be dengue, in São Pedro, São Paulo State. A MAC-ELISA done on the patient's acute and convalescent sera was inconclusive and hemagglutination inhibition test detected IgG antibody for flaviviruses. An indirect immunofluorescent assay done on the C6/36 cell culture inoculated with the acute serum was positive for flaviviruses but negative when tested with dengue monoclonal antibodies. RNA extracted from the infected cell culture supernatant was amplified by RT-PCR in the presence of NS5 universal flavivirus primers and directly sequenced. Results of BLAST search indicated that this sequence shares 93% nucleotide similarity with the sequence of SLEV (strain-MSI.7), confirmed by RT-PCR performed with SLEV specific primers. Since SLEV was identified as the cause of human disease, it is necessary to improve surveillance in order to achieve early detection of this agent in the state of São Paulo and in Brazil. This finding is also an alert to health professionals about the need for more complete clinical and epidemiological investigations of febrile illnesses as in the reported case. SLEV infections can be unrecognized or confused with other ones caused by an arbovirus, such as dengue.

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.

Seroconversion for west Nile and St. Louis encephalitis viruses among sentinel horses in Colombia

We prospectively sampled flavivirus-naïve horses in northern Colombia to detect West Nile virus (WNV) and St. Louis encephalitis virus (SLEV) seroconversion events, which would indicate the current circulation of these viruses. Overall, 331 (34.1%) of the 971 horses screened were positive for past infection with flaviviruses upon initial sampling in July 2006. During the 12-month study from July 2006-June 2007, 33 WNV seroconversions and 14 SLEV seroconversions were detected, most of which occurred in the department of Bolivar. The seroconversion rates of horses in Bolivar for the period of March-June 2007 reached 12.4% for WNV and 6.7% for SLEV. These results comprise the first serologic evidence of SLEV circulation in Colombia. None of the horses sampled developed symptoms of encephalitis within three years of initial sampling. Using seroconversions in sentinel horses, we demonstrated an active circulation of WNV and SLEV in northern Colombia, particularly in the department of Bolivar. The absence of WNV-attributed equine or human disease in Colombia and elsewhere in the Caribbean Basin remains a topic of debate and speculation.

Serological detection of St. Louis encephalitis virus and West Nile virus in equines from Santa Fe, Argentina

St. Louis encephalitis virus (SLEV) and West Nile virus (WNV) present ecological and antigenic similarities and are responsible for serious human diseases. In addition, WNV is a significant pathogen in terms of equine health. The purpose of our study was to analyse the seroprevalence of SLEV and WNV in equine sera collected in Santa Fe Province, Argentina. The seroprevalence determined using the plaque reduction neutralisation test was 12.2% for SLEV, 16.2% for WNV and 48.6% for a combination of both viruses. These results provide evidence of the co-circulation of SLEV and WNV in equines in Santa Fe.

Estudos sorológicos para pesquisa de anticorpos de arbovírus em população humana da região do Vale do Ribeira: II - inquérito em pacientes do Hospital Regional de Pariquera-Açú, 1980

Foi realizado inquérito sorológico para pesquisa de anticorpos inibidores de hemaglutinaçãc de arbovírus em 516 moradores das zonas urbana e rural da região do Vale do Ribeira, Brasil, área extensamente coberta de florestas onde ocorreu recentemente uma epidemia de encefalite atribuída ao Flavivirus Rocio. Verificou-se que 24,2% destas pessoas tinham anticorpos IH para um ou mais arbovírus (11,2% para Alphavirus; 13,2% para Flavivirus; 4,6% para o Bunyavirus Caraparu e 0,8% para outros arbovírus). Alguns dos investigados, sem antecedente de vacinação contra febre amarela, apresentaram anticorpos neutralizantes para o vírus da encefalite equina do Leste, St. Louis e da febre amarela, os dois últimos ainda não isolados na região. A análise das características dos indivíduos com sorologia positiva sugeria que a transmissão de arboviroses não era fato recente e estava se fazendo em pelo menos 9 municípios da área, não só no ambiente silvestre como fora do mesmo. Os indivíduos de sexo masculino e entre estes os que trabalham em pesca, em geral no período vespertino e noturno, apresentaram maior risco à infecções arbovíricas.

Encefalite na região do Vale do Ribeira, São Paulo, Brasil, no período pós-epidêmico de 1978 a 1983: situação do diagnóstico etiológico e características epidemiológicas

Relatam-se os resultados do levantamento de dados referentes ao diagnóstico etiológico, distribuição e letalidade das encefalites na região do Vale do Ribeira, São Paulo, Brasil, no período de 1978 a 1983, após uma epidemia de encefalite por arbovírus. Verificou-se que não foi possível o diagnóstico etiológico em nenhum dos 80 casos conhecidos. Em apenas 9 doentes (11,25%) foram coletadas duas amostras pares de soro, não se tendo observado conversão sorológica, Em 29 dos 33 pacientes em que se dispunha de pelo menos uma amostra de soro realizou-se teste de inibição de hemaglutinação para os flavivírus Rocio, Ilhéus e St. Louis, observando-se em 8 a presença de títulos > 20 para esses vírus. Testes para os alphavírus VEE, EEE e Mucambo foram efetuados em quatro dos 33 pacientes, com resultados negativos. A distribuição espacial, etária e por sexo dos casos apresentou padrões semelhantes aos do período epidêmico, ao contrário de letalidade que mostrou um aumento significante, de 9,9% para 25,0%. Os autores enfatizam a urgência no incremento da Vigilância epidemiológica das encefalites por arbovírus na região, sugerindo medidas dirigidas ao diagnóstico etiológico.

Studies on mosquitoes (Diptera: Culicidae) and anthropic environment: 10- survey of adult behaviour of Culex nigripalpus and other species of Culex (Culex) in South-Eastern Brazil

A survey of adult behaviour of Culex (Culex) species was carried out from August 1992 through December 1993 in a human modified (anthropic) environment in the Ribeira Valley, S.Paulo State, Brazil. Culex nigripalpus dominated the catches at several sites and it's tendency to increase in the anthropic environment became quite clear. Nevertheless no high level of synanthropy was demonstrated. So it seems that the mosquito may have a restricted role in natural arbovirus cycles. Nonetheless, Cx. nigripalpus must be considered a potential vector of arboviruses, especially St. Louis encephalitis virus outside dwellings.

Inquérito soroepidemiológico para pesquisa de infecções por arbovírus em moradores de reserva ecológica

OBJETIVO: Inquéritos sorológicos têm evidenciado ampla circulação de arbovírus causadores de doença humana na Região do Vale do Ribeira, São Paulo, Brasil. Com o propósito de estabelecer a prevalência de infecções por esses agentes em reserva ecológica, localizada naquela área, pesquisou-se a presença de anticorpos, bem como suas possíveis associações com características individuais e familiares dos investigados. MÉTODOS: Pesquisaram-se anticorpos para os antígenos dos vírus Rocio (ROC), Ilhéus (ILH), encefalite de St. Louis (SLE), encefalites eqüinas do leste (EEE), oeste (WEE) e venezuelana (VEE), em 182 pessoas pertencentes a 58 famílias residentes na Estação Ecológica de Juréia-Itatins, utilizando-se testes de inibição de hemaglutinação e neutralização com redução de placas. Usou-se Mac-Elisa para pesquisar anticorpos IgM para os vírus ROC, ILH e EEE. RESULTADOS: Foi observada a presença de anticorpos para todos os arbovírus testados, com exceção do vírus WEE. A prevalência total de anticorpos foi 26,9% (21,4% para alfavírus e 12,6% para flavivírus). Não foram encontrados anticorpos IgM. Entre as várias características pesquisadas dos indivíduos e de suas famílias, a idade, a ocupação, a naturalidade e o hábito de entrar na mata mostraram-se estatisticamente associados a infecções por arbovírus (p<0,05). CONCLUSÕES: Existe intensa circulação de arbovírus patogênicos, especialmente do alfavírus VEE, na população estudada. Aparentemente, a principal forma de exposição a vetores de arbovírus nessa população é o hábito de entrar na mata. Sugerem-se outras investigações sobre a responsabilidade de aves atuando como amplificadores de vírus dentro dos domicílios.

Epidemiologia das encefalites por arbovírus na amazônia brasileira

Os autores revêem os aspectos ecoepidemiológicos apresentados pelos virus da encefalite de St. Louis (SLE), encefalites equinas Leste (EEE), Oeste (WEE) e Venezuelana [subtipos III, Mucambo (MUC) e IV, Pixuna (PIX)], decorrentes dos estudos realizados em diversas áreas da Região Amazônica brasileira, especialmente ao longo das rodovias e projetos de desenvolvimento. Esses vírus são amplamente distribuídos na Amazônia e pelo menos quatro deles, EEE, WEE, MUC e SLE já demonstraram ser patógenos do homem. O diagnóstico da doença humana foi feito por sorologia, sendo que de MUC e SLE obteve-se também isolamento viral. O vírus PIX, parece ser o menos prevalente e foi isolado em poucas oportunidades. Virtualmente se desconhecem os vetores do PIX e WEE. As aves silvestres constituem os hospedeiros principais de todos esses vírus, exceto do MUC, para o qual constituem os roedores. O quadro clínico apresentado pelos pacientes infectados na Amazônia é discutido, comparando-o ao apresentado em outras áreas, especialmente nos EUA, onde periodicamente SLE, EEE e WEE causam surtos de doença humana. Nenhuma epidemia foi até o presente detectada, embora em 1960 uma epizootia em eqüinos causada pelo EEE tenha sido registrada em Bragança, Pará, onde em um rebanho de 500 animais ocorreu uma letalidade de 5%. Quatro outras pequenas epizootias determinadas pelo SLE ocorreram nas florestas adjacentes a Belém, envolvendo aves silvestres e animais sentinelas.

Surveillance of arbovirus infections in the atlantic forest region, State of São Paulo, Brazil: I. detection of hemagglutination-inhibition antibodies in wild birds between 1978 and 1990

We report data related to arbovirus antibodies detected in wild birds periodically captured from January 1978 to December 1990 in the counties of Salesópolis (Casa Grande Station), Itapetininga and Ribeira Valley, considering the different capture environments. Plasmas were examined using hemagglutination-inhibition (HI) tests. Only monotypic reactions were considered, except for two heterotypic reactions in which a significant difference in titer was observed for a determined virus of the same antigenic group. Among a total of 39,911 birds, 269 birds (0.7%) belonging to 66 species and 22 families were found to have a monotypic reaction for Eastern equine encephalitis (EEE), Venezuelan equine encephalitis (VEE), Western equine encephalitis (WEE), Ilheus (ILH), Rocio (ROC), St. Louis encephalitis (SLE), SP An 71686, or Caraparu (CAR) viruses. Analysis of the data provided information of epidemiologic interest with respect to these agents. Birds with positive serology were distributed among different habitats, with a predominance of unforested habitats. The greatest diversity of positive reactions was observed among species which concentrate in culture fields.

Molecular characterization of two rocio flavivirus strains isolated during the encephalitis epidemic in são paulo state, brazil and the development of a one-step rt-pcr assay for diagnosis

Rocio virus (ROCV) was responsible for an explosive encephalitis epidemic in the 1970s affecting about 1,000 residents of 20 coastland counties in São Paulo State, Brazil. ROCV was first isolated in 1975 from the cerebellum of a fatal human case of encephalitis. Clinical manifestations of the illness are similar to those described for St. Louis encephalitis. ROCV shows intense antigenic cross-reactivity with Japanese encephalitis complex (JEC) viruses, particularly with Ilheus (ILHV), St. Louis encephalitis, Murray Valley and West Nile viruses. In this study, we report a specific RT-PCR assay for ROCV diagnosis and the molecular characterization of the SPAn37630 and SPH37623 strains. Partial nucleotide sequences of NS5 and E genes determined from both strains were used in phylogenetic analysis. The results indicated that these strains are closely related to JEC viruses, but forming a distinct subclade together with ILHV, in accordance with results recently reported by Medeiros et al. (2007).

Production and diagnostic application of recombinant domain III of West Nile envelope protein in Brazil

INTRODUCTION: West Nile virus (WNV) is a flavivirus with a natural cycle involving mosquitoes and birds. Over the last 11 years, WNV has spread throughout the Americas with the imminent risk of its introduction in Brazil. METHODS: Envelope protein domain III of WNV (rDIII) was bacterially expressed and purified. An enzyme-linked immunosorbent assay with WNV rDIII antigen was standardized against mouse immune fluids (MIAFs) of different flavivirus. RESULTS: WNV rDIII reacted strongly with St. Louis encephalitis virus (SLEV) MIAF but not with other flaviviruses. CONCLUSIONS: This antigen may be a potentially useful tool for serologic diagnosis and may contribute in future epidemiological surveillance of WNV infections in Brazil.