Systems analysis of subjects acutely infected with the Chikungunya virus.

The largest ever recorded epidemic of the Chikungunya virus (CHIKV) broke out in 2004 and affected four continents. Acute symptomatic infections are typically associated with the onset of fever and often debilitating polyarthralgia/polyarthritis. In this study, a systems biology approach was adopted to analyze the blood transcriptomes of adults acutely infected with the CHIKV. Gene signatures that were associated with viral RNA levels and the onset of symptoms were identified. Among these genes, the putative role of the Eukaryotic Initiation Factor (eIF) family genes and apolipoprotein B mRNA editing catalytic polypeptide-like (APOBEC3A) in the CHIKV replication process were displayed. We further compared these signatures with signatures induced by the Dengue virus infection and rheumatoid arthritis. Finally, we demonstrated that the CHIKV in vitro infection of murine bone marrow-derived macrophages induced IL-1 beta production in a mechanism that is significantly dependent on the inflammasome NLRP3 activation. The observations provided valuable insights into virus-host interactions during the acute phase and can be instrumental in the investigation of new and effective therapeutic interventions.

Sequence analysis of the 2009 pandemic influenza A H1N1 virus haemagglutinin gene from 2009-2010 Brazilian clinical samples.

In this paper, we analysed the haemagglutinin (HA) gene identified by polymerase chain reaction from 90 influenza A H1N1 virus strains that circulated in Brazil from April 2009-June 2010. A World Health Organization sequencing protocol allowed us to identify amino acid mutations in the HA protein at positions S220T (71%), D239G/N/S (20%), Y247H (4.5%), E252K (3.3%), M274V (2.2%), Q310H (26.7%) and E391K (12%). A fatal outcome was associated with the D239G mutation (p < 0.0001). Brazilian HA genetic diversity, in comparison to a reference strain from California, highlights the role of influenza virus surveillance for study of viral evolution, in addition to monitoring the spread of the virus worldwide.

Molecular characterization of dengue viruses type 1 and 2 isolated from a concurrent human infection.

In 2001, an autochthonous case of dual viremia, resulting from naturally acquired dengue virus DEN-1 and DEN-2 infections was detected during the dengue outbreak that occurred in Barretos, a city with about 105,000 inhabitants in the North region of São Paulo State. Serotype identification was based on virus isolation to C6/36 mosquito cells culture and immunofluorescence assays using type-specific monoclonal antibodies. The double infection was also confirmed by reverse transcriptase polymerase chain reaction (RT-PCR). Comparative analysis of the 240-nucleotide sequences of E/NS1 gene junction region between the genome of DEN-1 and DEN-2 isolates of the corresponding reference Nauru and PR 159S1 strains, respectively, showed some nucleotide differences, mainly silent mutations in the third codon position. Results of maximum likelihood phylogenetic analysis of E/NS1 gene sequences indicated that both genotypes of DEN-1 and DEN-2 viruses recovered from double infection in Barretos belonged to genotypes I and III, respectively.

Molecular characterization of influenza B virus outbreak on a cruise ship in Brazil 2012.

In February 2012, an outbreak of respiratory illness occurred on the cruise ship MSC Armonia in Brazil. A 31-year-old female crew member was hospitalized with respiratory failure and subsequently died. To study the etiology of the respiratory illness, tissue taken at necropsy from the deceased woman and respiratory specimens from thirteen passengers and crew members with respiratory symptoms were analyzed. Influenza real-time RT-PCR assays were performed, and the full-length hemagglutinin (HA) gene of influenza-positive samples was sequenced. Influenza B virus was detected in samples from seven of the individuals, suggesting that it was the cause of this respiratory illness outbreak. The sequence analysis of the HA gene indicated that the virus was closely related to the B/Brisbane/60/2008-like virus, Victoria lineage, a virus contained in the 2011-12 influenza vaccine for the Southern Hemisphere. Since the recommended composition of the influenza vaccine for use during the 2013 season changed, an intensive surveillance of viruses circulating worldwide is crucial. Molecular analysis is an important tool to characterize the pathogen responsible for an outbreak such as this. In addition, laboratory disease surveillance contributes to the control measures for vaccine-preventable influenza.

Mumps virus genotypes identified during disease outbreaks in the state of São Paulo, Brazil: 2011-2016 / Genótipos dos vírus da caxumba identificados durantesurtos da doença no estado de São Paulo, Brasil: 2011 - 2016

In São Paulo the mumps virus (MuV) outbreaks have been increasing from In São Paulo the mumps virus (MuV) outbreaks have been increasing from 2011 to nowadays. MuV epidemiological surveillance has been improving by using the polymerase chain reaction in real time (rRT-PCR) in addition to the specific IgM antibody (IgM-Ab) detection; in some cases,genome sequencing studies were performed. Increased virus transmission and recent outbreakshave raised interest on MuV genotyping, as a means to understand the transmission pathways andto identify the vaccine-associated cases. From January 2011 to August 2016, MuV infection was analyzed at Institute Adolfo Lutz. A total of 232 (77.33 %) throat wash samples showed positivity to mumps genome, and 68 (22.66 %) were negative when analyzed by rRT-PCR. Among 15 samples for molecular analysis, 10 serum samples from respective patients were also available for detecting anti-MuV IgM-Ab; and from these, four (40%) samples were seropositive. Vaccination statuswas available only for patients from Cedral and Araraquara. Phylogenetic analysis revealed the circulation of the following mumps virus genotypes in the investigated periods: 2011(M), 2012, and 2013 (K); 2014 (N); 2015 (G, K, and N); 2016 (G). Knowledge on MuV molecular epidemiology in São Paulo-Brazil could contribute to the surveillance and epidemiological program in Brazil, and globally as well.

No estado de São Paulo têm ocorrido surtos de caxumba desde 2011. O diagnóstico laboratorialtem sido realizado no Instituto Adolfo Lutz utilizando-se a técnica de identificação de material genético viral por meio de reação de cadeia de polimerase-em tempo real (rRT-PCR) e peladetecção de anticorpos IgM (Ac-IgM) específicos circulantes. Os recentes surtos de caxumbatêm aumentado o interesse em investigar os genótipos dos vírus prevalentes para identificaros casos associadas à vacina. De janeiro de 2011 a agosto de 2016, 300 amostras de lavadosda orofaringe coletadas de pacientes suspeitos de infecção foram analisadas. O material genéticoviral específico foi detectado em 232 (77,33 %) amostras e 68 (22,66 %) foram negativas.Das 10 amostras analisadas pelo teste sorológico, quatro (40 %) demonstraram positividadepara Ac-IgM específicos anti-vírus da caxumba e seis foram negativas. Somente os municípios Cedral e Araraquara forneceram os dados referentes à vacinação. Análise filogenética mostroua circulação dos seguintes genótipos do vírus da caxumba no período investigado: 2011 (M),2012 e 2013 (K); 2014 (N); 2015 (GKN); 2016 (G). A vigilância virológica é mundialmente imprescindível, para identificar a diversidade e a distribuição dos diferentes genótipos, com vistasà composição de vacinas específicas.

Mumps virus genotypes identified during disease outbreaks in the state of São Paulo, Brazil: 2011 – 2016 / Genótipos dos vírus da caxumba identificados durante surtos da doença no estado de São Paulo, Brasil: 2011 - 2016

In São Paulo the mumps virus (MuV) outbreaks have been increasing from 2011 to nowadays. MuV epidemiological surveillance has been improving by using the polymerase chain reaction in real time (rRT-PCR) in addition to the specific IgM antibody (IgM-Ab) detection; in some cases, genome sequencing studies were performed. Increased virus transmission and recent outbreaks have raised interest on MuV genotyping, as a means to understand the transmission pathways and to identify the vaccine-associated cases. From January 2011 to August 2016, MuV infection was analyzed at Institute Adolfo Lutz. A total of 232 (77.33 %) throat wash samples showed positivity to mumps genome, and 68 (22.66 %) were negative when analyzed by rRT-PCR. Among 15 samples for molecular analysis, 10 serum samples from respective patients were also available for detecting anti-MuV IgM-Ab; and from these, four (40%) samples were seropositive. Vaccination status was available only for patients from Cedral and Araraquara. Phylogenetic analysis revealed the circulation of the following mumps virus genotypes in the investigated periods: 2011(M), 2012, and 2013 (K); 2014 (N); 2015 (G, K, and N); 2016 (G). Knowledge on MuV molecular epidemiology in São Paulo-Brazil could contribute to the surveillance and epidemiological program in Brazil, and globally as well.

No estado de São Paulo têm ocorrido surtos de caxumba desde 2011. O diagnóstico laboratorial tem sido realizado no Instituto Adolfo Lutz utilizando-se a técnica de identificação de material genético viral por meio de reação de cadeia de polimerase-em tempo real (rRT-PCR) e pela detecção de anticorpos IgM (Ac-IgM) específicos circulantes. Os recentes surtos de caxumba têm aumentado o interesse em investigar os genótipos dos vírus prevalentes para identificar os casos associadas à vacina. De janeiro de 2011 a agosto de 2016, 300 amostras de lavados da orofaringe coletadas de pacientes suspeitos de infecção foram analisadas. O material genético viral específico foi detectado em 232 (77,33 %) amostras e 68 (22,66 %) foram negativas. Das 10 amostras analisadas pelo teste sorológico, quatro (40 %) demonstraram positividade para Ac-IgM específicos anti-vírus da caxumba e seis foram negativas. Somente os municípios Cedral e Araraquara forneceram os dados referentes à vacinação. Análise filogenética mostrou a circulação dos seguintes genótipos do vírus da caxumba no período investigado: 2011 (M), 2012 e 2013 (K); 2014 (N); 2015 (GKN); 2016 (G). A vigilância virológica é mundialmente imprescindível, para identificar a diversidade e a distribuição dos diferentes genótipos, com vistas à composição de vacinas específicas.

Molecular charatterization of influenza b virus outbreak on a cruise ship in Brazil 2012 / Caracterização molecular de surto de vírus influenza B em cruzeiro marítimo no Brasil 2012

In February 2012, an outbreak of respiratory illness occurred on the cruise ship MSC Armonia in Brazil. A 31-year-old female crew member was hospitalized with respiratory failure and subsequently died. To study the etiology of the respiratory illness, tissue taken at necropsy from the deceased woman and respiratory specimens from thirteen passengers and crew members with respiratory symptoms were analyzed. Influenza real-time RT-PCR assays were performed, and the full-length hemagglutinin (HA) gene of influenza-positive samples was sequenced. Influenza B virus was detected in samples from seven of the individuals, suggesting that it was the cause of this respiratory illness outbreak. The sequence analysis of the HA gene indicated that the virus was closely related to the B/Brisbane/60/2008-like virus, Victoria lineage, a virus contained in the 2011-12 influenza vaccine for the Southern Hemisphere. Since the recommended composition of the influenza vaccine for use during the 2013 season changed, an intensive surveillance of viruses circulating worldwide is crucial. Molecular analysis is an important tool to characterize the pathogen responsible for an outbreak such as this. In addition, laboratory disease surveillance contributes to the control measures for vaccine-preventable influenza.

Em fevereiro de 2012, durante a temporada de verão no Brasil, um surto de doença respiratória ocorreu no navio de cruzeiro MSC Armonia. Mulher de 31 anos, membro da tripulação, foi internada com insuficiência respiratória e morreu. Com o objetivo de estudar a etiologia da doença foram investigadas necrópsia de tecido do caso fatal e secreções respiratórias de 13 passageiros e membros da tripulação com sintomas respiratórios. O teste de influenza por RT-PCR em tempo real foi realizado e o gene completo da hemaglutinina (HA) das amostras positivas foi sequenciado. O vírus influenza B foi detectado em sete indivíduos, sugerindo-o como a causa do surto de doença respiratória a bordo do navio. A análise da sequência do gene da HA indicou que os vírus estão fortemente relacionados com o vírus B/Brisbane/60/2008, linhagem Victoria, componente da vacina de influenza para 2011-2012 no hemisfério sul. Uma vez que a composição da vacina foi alterada para uso na temporada de 2012-2013, é essencial a vigilância ativa dos vírus circulantes em todo o mundo. A análise molecular é uma ferramenta importante para caracterização do patógeno responsável pelo surto. Além disso, a vigilância de doenças baseada em dados laboratoriais contribui para as medidas de controle da influenza, uma doença imunoprevinível.

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.

Sequence analysis of the 2009 pandemic influenza A H1N1 virus haemagglutinin gene from 2009-2010 Brazilian clinical samples

In this paper, we analysed the haemagglutinin (HA) gene identified by polymerase chain reaction from 90 influenza A H1N1 virus strains that circulated in Brazil from April 2009-June 2010. A World Health Organization sequencing protocol allowed us to identify amino acid mutations in the HA protein at positions S220T (71 percent), D239G/N/S (20 percent), Y247H (4.5 percent), E252K (3.3 percent), M274V (2.2 percent), Q310H (26.7 percent) and E391K (12 percent). A fatal outcome was associated with the D239G mutation (p < 0.0001). Brazilian HA genetic diversity, in comparison to a reference strain from California, highlights the role of influenza virus surveillance for study of viral evolution, in addition to monitoring the spread of the virus worldwide.

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.

First isolation of dengue 3 in Brazil from an imported case

The authors report the isolation of dengue 3 virus for the first time in Brazil. The patient, resident in Limeira-SP, traveled to Nicaragua on May 16th, 1998, where he stayed for two months. Starting on August 14th he had fever, headache, myalgia, arthralgia, retro-orbital pain and diarrhea. He returned to Brazil on August 16th and was hospitalized in the next day. The patient had full recovery and was discharged on August 20th. The virus was isolated in C6/36 cell culture inoculated with serum collected on the 6th day after the onset of the symptoms. The serotype 3 was identified by indirect immunofluorescence assays performed with type-specific monoclonal antibodies. This serotype was further confirmed by polymerase chain reaction analysis. The introduction of a new dengue serotype in a susceptible population is a real threat for the occurrence of severe forms of the disease. The isolation and identification of dengue virus are important in order to monitoring the serotypes circulating in Brazil and to take the measures necessary to prevent and control an epidemic

Molecular characterization of Dengue viruses type 1 and 2 isolated from a concurrent human infection

In 2001, an autochthonous case of dual viremia, resulting from naturally acquired dengue virus DEN-1 and DEN-2 infections was detected during the dengue outbreak that occurred in Barretos, a city with about 105,000 inhabitants in the North region of São Paulo State. Serotype identification was based on virus isolation to C6/36 mosquito cells culture and immunofluorescence assays using type-specific monoclonal antibodies. The double infection was also confirmed by reverse transcriptase polymerase chain reaction (RT-PCR). Comparative analysis of the 240-nucleotide sequences of E/NS1 gene junction region between the genome of DEN-1 and DEN-2 isolates of the corresponding reference Nauru and PR 159S1 strains, respectively, showed some nucleotide differences, mainly silent mutations in the third codon position. Results of maximum likelihood phylogenetic analysis of E/NS1 gene sequences indicated that both genotypes of DEN-1 and DEN-2 viruses recovered from double infection in Barretos belonged to genotypes I and III, respectively

Infecção simultânea por Dengue 1 e 3 em Itapevi, SP, Brasil / Concurrent infection by Dengue 1 and 3 in Itapevi, SP, Brazil

Casos de infecções simultâneas por vírus dengue em um mesmo paciente podem ocorrer em epidemia em que circulam múltiplos sorotipos do vírus. No Brasil esse fato foi relatado pela primeira vez em Miranda, MS em 1996 e em Barretos, SP em 2001. Em ambos foram isolados dengue 1 e 2. Em 2003 foi observado mais um caso, em Itapevi, município de São Paulo. O isolamento dos vírus foi obtido em cultura de células C6/36 e a identificação foi feita por imunofluorescência indireta com anticorpos monoclonais para os quatro sorotipos. Foram isolados vírus dengue tipo 1 e tipo 3, resultado confirmado pela reação de RT/PCR, usando RNA extraído do sobrenadante da cultura de células infectada. Este caso demonstra mais uma vez a importância do isolamento viral na vigilância epidemiológica do dengue, pois ele pode ajudar a determinar a freqüencia das infecções com dois ou mais sorotipos, assim como verificar se essas infecções podem estar associadas a formas mais severas da doença