Seasonality of viral respiratory infections in Southeast of Brazil: the influence of temperature and air humidity

Viruses are the major cause of lower respiratory tract infections in childhood and the main viruses involved are Human Respiratory Syncytial Virus (HRSV), Human Metapneumovirus (HMPV), Influenzavirus A and B (FLUA and FLUB), Human Parainfluenza Virus 1, 2 and 3 (HPIV1, 2 and 3) and Human Rhinovirus (HRV). The purposes of this study were to detect respiratory viruses in hospitalized children younger than six years and identify the influence of temperature and relative air humidity on the detected viruses. Samples of nasopharyngeal washes were collected from hospitalized children between May/2004 and September/2005. Methods of viral detection were RT-PCR, PCR and HRV amplicons were confirmed by hybridization. Results showed 54% (148/272) of viral positivity. HRSV was detected in 29% (79/272) of the samples; HRV in 23.1% (63/272); HPIV3 in 5.1% (14/272); HMPV in 3.3% (9/272); HPIV1 in 2.9% (8/272); FLUB in 1.4% (4/272), FLUA in 1.1% (3/272), and HPIV2 in 0.3% (1/272). The highest detection rates occurred mainly in the spring 2004 and in the autumn 2005. It was observed that viral respiratory infections tend to increase as the relative air humidity decreases, showing significant association with monthly averages of minimal temperature and minimal relative air humidity. In conclusion, viral respiratory infections vary according to temperature and relative air humidity and viral respiratory infections present major incidences it coldest and driest periods.

Seasonality of viral respiratory infections in Southeast of Brazil: the influence of temperature and air humidity

Viruses are the major cause of lower respiratory tract infections in childhood and the main viruses involved are Human Respiratory Syncytial Virus (HRSV), Human Metapneumovirus (HMPV), Influenzavirus A and B (FLUA and FLUB), Human Parainfluenza Virus 1, 2 and 3 (HPIV1, 2 and 3) and Human Rhinovirus (HRV). The purposes of this study were to detect respiratory viruses in hospitalized children younger than six years and identify the influence of temperature and relative air humidity on the detected viruses. Samples of nasopharyngeal washes were collected from hospitalized children between May/2004 and September/2005. Methods of viral detection were RT-PCR, PCR and HRV amplicons were confirmed by hybridization. Results showed 54% (148/272) of viral positivity. HRSV was detected in 29% (79/272) of the samples; HRV in 23.1% (63/272); HPIV3 in 5.1% (14/272); HMPV in 3.3% (9/272); HPIV1 in 2.9% (8/272); FLUB in 1.4% (4/272), FLUA in 1.1% (3/272), and HPIV2 in 0.3% (1/272). The highest detection rates occurred mainly in the spring 2004 and in the autumn 2005. It was observed that viral respiratory infections tend to increase as the relative air humidity decreases, showing significant association with monthly averages of minimal temperature and minimal relative air humidity. In conclusion, viral respiratory infections vary according to temperature and relative air humidity and viral respiratory infections present major incidences it coldest and driest periods.

Comparison between ectodomain and G2 region of G glycoprotein for genotyping of HRSV

The Human Respiratory Syncytial Virus (HRSV), isolated in 1955, is the main cause of hospitalization of babies and infants with respiratory illness. Several studies have been conducted worldwide aiming the development of a safe and effective vaccine against HRSV. The G2 region of glycoprotein G is used as genotyping default. In the present study, we performed a phylogenetic analysis of G protein and a comparative study between G2 region and ectodomain of attachment glycoprotein. Fifty-three nasal swab samples from children less than 5 years old and presenting symptoms of acute respiratory illness, assisted at the University Hospital (UH) of University of Sao Paulo (USP) in 2004, were submitted to sequencing by PCR and compared with GenBank sequences. We concluded that the G2 region is adequate for HRSV genotyping.

O vírus respiratório sincicial humano (HRSV), isolado em 1955, é a principal causa da hospitalização de bebês e crianças pequenas com sintomas de doença respiratória. No mundo inteiro, vários estudos para o desenvolvimento de uma vacina segura e eficiente contra o HRSV têm tido alta prioridade. A região G2 da glicoproteína G é usada como padrão para genotipagem do HRSV. Neste estudo, foi realizada a análise filogenética da glicoproteína G e o estudo comparativo entre a região G2 e o ectodomínio dessa glicoproteína. Cinquenta e três amostras de swab nasal de crianças com menos de cinco anos de idade, apresentando doença respiratória aguda, atendidas no Hospital Universitário (HU) da Universidade de São Paulo durante o ano de 2004, foram submetidas a sequenciamento por PCR e comparadas com seqüências do GenBank. A região G2 mostrou ser adequada para a genotipagem do HRSV.

Investigation of influenza in migrating birds, the primordial reservoir and transmitters of influenza in Brazil

Birds are the most important reservoirs of the influenza virus. Its maintenance in its natural hosts, including man, allows the influenza virus to reassorts its strains. The recent report of an avian influenza A (H5N1) virus in humans, was in a child with fatal respiratory illness in China, 1997. The current study was conducted to elucidate the transportation of the influenza by birds that migrate, annually, through the both Northern and Southern hemispheres, with special attention paid to the Vireo olivaceus [Juruviara(BR) or Red-eyed vireo(USA)] species, which travels from the USA to Brazil, and vice versa, and the Elaenia mesoleuca [Tuque(BR) or (USA)] species that flies over the entire Southern Hemisphere. There are two species of birds, which breed and migrate in São Paulo State, Brazil, and which were demonstrated to carry Influenza virus, were selected. The viral particles isolated were observed by electron microscopy. The influenza virus was detected by the House Duplex/PCR and Gloria molecular biology tests. The results demonstrated that the Elaenia mesoleuca and Vireo olivaceus bird species are carrying the Influenza virus whilst crossing both the Northern and Southern hemispheres. To understand the role that these migrating birds may play in epidemic influenza, in Brazil, characterization of avian influenza subtypes will be done.

Os mais importantes reservatórios do vírus influenza são os pássaros. A manutenção do vírus influenza em hospedeiros naturais, inclusive o homem, permite que esse vírus realize rearranjos entre as suas cepas. O recente relato de uma cepa influenza aviária A(H5N1), em humanos, se deu em uma criança com doença respiratória fatal, na China em 1977. O presente estudo foi conduzido para elucidar o transporte da influenza por pássaros que migram, anualmente, através de ambos hemisférios o do Norte e do Sul, com especial atenção voltada à espécies Vireo olivaceo [Juruviara(BR) e Red-eyed vireo(USA)] que viaja do USA para o Brasil, e vice-versa, e a espécie Elaenia mesoleuca [Tuque(BR) e (USA)] que voa por todo o Hemisfério Sul. Essas espécies de pássaros, que residem e migram em São Paulo, e que demonstram transportar o vírus influenza, foram selecionadas. As partículas virais isoladas foram observadas por microscópio eletrônico. O vírus influenza foi detectado pelos testes: House Duplex/PCR e Gloria. Os resultados revelam que os pássaros das espécies: Elaenia mesoleuca e Vireo olivaceus são transportes do vírus influenza enquanto cruzam ambos Hemisférios. Para o conhecimento da função que os pássaros migratórios podem desempenhar na epidemia de influenza, no Brasil, caracterização dos subtipos deste vírus estão sendo realizados.

Construction of adenoviral vectors expressing F and G glycoproteins of human respiratory syncytial virus (HRSV)

Human Respiratory Syncytial Virus (HRSV) was first characterized in 1957 and has since been recognized as the most common viral cause of severe respiratory tract infection in young infants worldwide. Despite many years of research there is still no effective treatment or any immediate prospect of a vaccine. The HRSV genome is composed of single stranded negative sense RNA and the virion consists of a nucleocapsid packaged within a lipid envelope. The envelope contains spike-like projections, each being a homo-oligomer of one of three transmembrane viral envelope proteins: the attachment protein G, the fusion protein F involved in viral penetration and the small hydrofobic protein SH. The aim of this work was to construct two recombinant replication-defective adenoviruses carrying separately F and G genes from HRSV. This system was chosen because adenovirus delivers genes into target cells with high efficiency in a variety of cell lines and can be used in vitro and in vivo. In order to obtain the recombinant viruses, we did RT-PCR of RNA extracted from the HRSV A2 strain, the genes F and G were cloned in to pAdeno-X vectors. pAdeno-F and pAdeno-G were transfected in HEK-293 cells for the production of recombinant viruses, that expressed efficiently these two proteins and provide us the means for doing functional assays and immunization tests.

O Vírus Sincicial Respiratório Humano (HRSV) foi isolado e caracterizado pela primeira vez em 1957 e é considerado como o patógeno viral mais freqüente do trato respiratório de bebês e crianças. Apesar de muitos anos de pesquisa, não há ainda um tratamento específico ou uma vacina licenciada. Seu genoma é composto por uma fita simples de RNA polaridade negativa e o vírion consiste em um nucleocapsídeo empacotado por um envelope lipídico. O envelope contém projeções, chamadas espículas, constituídas de homoligômeros de uma das 3 glicoproteínas de membrana: a proteína de ligação G ("attachment"), a proteína de fusão F ("fusion") e a proteína SH ("small hydrofobic"). O objetivo deste trabalho foi construir dois adenovirus recombinantes defectivos em replicação expressando separadamente os genes F e G do HRSV. Este sistema foi escolhido porque os vetores adenovirais possuem a capacidade de inserir genes em uma grande variedade de linhagens celulares in vitro e in vivo. Para obtenção destes vetores adenovirais, um RT-PCR de RNA extraído do protótipo A2 de HRSV foi feito e os genes F e G clonados em vetores pAdeno-X. pAdeno-F e pAdeno-G foram transfectados em células HEK-293 para a produção do vírus recombinante, que expressaram corretamente essas duas proteínas constituem-se ferramentas para imunização e estudos funcionais.

Construction of adenoviral vectors expressing F and G glycoproteins of human respiratory syncytial virus (HRSV)

Human Respiratory Syncytial Virus (HRSV) was first characterized in 1957 and has since been recognized as the most common viral cause of severe respiratory tract infection in young infants worldwide. Despite many years of research there is still no effective treatment or any immediate prospect of a vaccine. The HRSV genome is composed of single stranded negative sense RNA and the virion consists of a nucleocapsid packaged within a lipid envelope. The envelope contains spike-like projections, each being a homo-oligomer of one of three transmembrane viral envelope proteins: the attachment protein G, the fusion protein F involved in viral penetration and the small hydrofobic protein SH. The aim of this work was to construct two recombinant replication-defective adenoviruses carrying separately F and G genes from HRSV. This system was chosen because adenovirus delivers genes into target cells with high efficiency in a variety of cell lines and can be used in vitro and in vivo. In order to obtain the recombinant viruses, we did RT-PCR of RNA extracted from the HRSV A2 strain, the genes F and G were cloned in to pAdeno-X vectors. pAdeno-F and pAdeno-G were transfected in HEK-293 cells for the production of recombinant viruses, that expressed efficiently these two proteins and provide us the means for doing functional assays and immunization tests.

O Vírus Sincicial Respiratório Humano (HRSV) foi isolado e caracterizado pela primeira vez em 1957 e é considerado como o patógeno viral mais freqüente do trato respiratório de bebês e crianças. Apesar de muitos anos de pesquisa, não há ainda um tratamento específico ou uma vacina licenciada. Seu genoma é composto por uma fita simples de RNA polaridade negativa e o vírion consiste em um nucleocapsídeo empacotado por um envelope lipídico. O envelope contém projeções, chamadas espículas, constituídas de homoligômeros de uma das 3 glicoproteínas de membrana: a proteína de ligação G ("attachment"), a proteína de fusão F ("fusion") e a proteína SH ("small hydrofobic"). O objetivo deste trabalho foi construir dois adenovirus recombinantes defectivos em replicação expressando separadamente os genes F e G do HRSV. Este sistema foi escolhido porque os vetores adenovirais possuem a capacidade de inserir genes em uma grande variedade de linhagens celulares in vitro e in vivo. Para obtenção destes vetores adenovirais, um RT-PCR de RNA extraído do protótipo A2 de HRSV foi feito e os genes F e G clonados em vetores pAdeno-X. pAdeno-F e pAdeno-G foram transfectados em células HEK-293 para a produção do vírus recombinante, que expressaram corretamente essas duas proteínas constituem-se ferramentas para imunização e estudos funcionais.

Human respiratory syncytial virus detection in children admitted at a community hospital in Botucatu, SP, Brazil

Human Respiratory Syncytial Virus (HRSV) is recognized as the single most important pathogen responsible for acute viral infection of the lower respiratory tract in young children. In this study, 84 samples from children under two years of age presenting acute respiratory symptoms were obtained from September 2000 to November 2001. Analysis by immunofluorescence and reverse-transcription followed by PCR, revealed that 18% (15/84) of all the samples were positive for HRSV, in which 80% (12/15) of the cases was observed in children under six months of age and the subgroups A and B co-circulated in the studied period. These are the first data obtained for Botucatu city and seasonality is evident by the higher circulation of the virus between May and July.

O Vírus Respiratório Sincicial Humano (VRSH) é descrito como o mais importante patógeno viral causador de doenças respiratórias agudas das vias respiratórias inferiores em crianças. Neste estudo 84 amostras de crianças com idade abaixo dos dois anos apresentando sintomas de doença respiratória aguda, foram obtidas no período de setembro de 2000 a novembro de 2001. Analise por imunofluorescência indireta e transcrição reversa seguida de PCR, revelou que 18% (15/84) das amostras foram positivas, sendo que em 80% (12/15) dos casos a detecção de VRSH foi observada em crianças abaixo dos seis meses, e também que os subgrupos A e B co-circularam. Estes são os primeiros dados obtidos para a cidade de Botucatu, sendo que a sazonalidade mostrou-se evidente pela maior circulação desse vírus entre os meses de maio e julho

Genetic diversity of the VP1/VP2 gene of canine parvovirus type 2b amplified from clinical specimens in Brazil

We evaluated the genetic diversity in the VP1/VP2 gene of CPV type 2b isolates from symptomatic dogs in Brazil. A total of 21 isolates collected from 1990 through 1995 previously typed as CPV2b by PCR assay were studied. Overall we found a high degree of similarity among sequences from different CPV clinical isolates collected. Genetic analysis of this selected region gave no indication of a specific Brazilian parvovirus lineage.

Neste estudo foi avaliada a diversidade genética no gene VP1/VP2 do parvovírus canino tipo 2b a partir de amostras isoladas de cães sintomáticos no Brasil. Foram estudadas 21 amostras coletadas no período de 1990 à 1995, previamente caracterizadas como CPV 2b pela técnica de PCR. Observou-se alto grau de similaridade entre as seqüências estudadas e a análise genética da região selecionada não indicou a presença de uma linhagem brasileira específica.

Time course of virus-like particles (VLPs) double-stranded rna accumulation in toxigenic and non-toxigenic strains of Aspergillus flavus

Two strains of Aspergillus flavus, non-toxigenic NRRL 6550 and toxigenic NRRL 5940, were studied over a period of 44 days, in order to detect the presence of virus-like particles (VLPs) by means of electron microscopy (EM) and nucleic acids electrophoresis. Only the toxigenic strain contained VLPs, presenting three-segmented dsRNA. An increase in VLPs number was observed during the exponential phase of fungal growth, up to day 12 of culture; after this, higher levels of aflatoxin production in toxigenic NRRL 5940 mycelia occurred in parallel with decreased VLPs replication.

Neste trabalho, foram estudadas duas cepas de Aspergillus flavus não toxigênica (NRRL 6550) e toxigênica (NRRL 5940). As cepas foram cultivadas durante 44 dias objetivando a pesquisa de partículas semelhantes a vírus (VLPs), por meio de microscopia eletrônica e pesquisa de ácidos nucléicos, através de eletroforese. Somente as cepas toxigênicas continham "VLPs", as quais apresentaram 3 segmentos de RNA de fita dupla. Um aumento no número de "VLPs" foi observado com 12 dias de cultivo, período correspondente a fase exponencial de crescimento de A. flavus.

Clinical and Laboratory evaluation of measleslike rash in children and young adults

A clinical and laboratory evaluation of 11 children and young adults with measleslike rash was done during the measles outbreak in the Greater São Paulo Metropolitan area at the end of 1996 and spread over the country during 1997. Measles was laboratory confirmed in 07 patients by specific IgM detection in acute serum specimens using an IgM-capture EIA, by specific IgG seroconversion in serum pairs, and by reverse transcription PCR and virus isolation in peripheral blood lymphocytes. Clinical presentations were not always classic; one of the 07 cases had received measles vaccine and corresponded to modified clinical case of measles. The 4 remaining cases were negative for measles and were diagnosed as exanthem subitum (2 cases), scarlet fever and Kawasaki disease. The present study reinforces the view that clinical features alone are not sufficient for establishing an accurate diagnosis in the post-vaccine era, and a surveillance system based on sensitive laboratory results is needed so that it can confirm IgM-negative measles cases.

Apresenta-se um estudo sobre a avaliação clínica e laboratorial de crianças e adultos jovens com exantema semelhante ao sarampo, durante a ocorrência do surto epidêmico de sarampo na região metropolitana da Grande São Paulo no final de 1996 que se alastrou por todo o país durante o ano de 1997. O diagnóstico laboratorial de sarampo foi firmado em 7 dos 11 pacientes examinados, por detecção direta de IgM específico em amostras de soro da fase aguda da doença, por soroconversão de IgG em um par de amostras da fase aguda e convalescente, pelo isolamento do vírus em culturas de linhagem B95a e pela detecção do RNA viral por PCR em linfócitos do sangue periférico e na urina. O quadro clínico apresentado pelos pacientes nem sempre correspondeu à forma clássica do sarampo. Um jovem adulto, que havia sido vacinado contra o sarampo, apresentou um quadro clínico modificado. Os 4 casos restantes foram negativos em todas as provas laboratoriais para o sarampo, tendo diagnóstico final de exantema súbito (2 casos), escarlatina e Doença de Kawasaki. O presente estudo reforça o ponto de vista de que os sinais e sintomas clínicos são geralmente insuficientes para estabelecer um diagnóstico preciso de sarampo na era pós-vacinal, sendo indispensável um sistema de vigilância baseado em resultados laboratoriais que confirmam todos os casos suspeitos, inclusive os casos de sarampo com sorologia IgM-negativa