Serious acute respiratory syndrome: a case series in a municipality region of central Brazil

The influenza B virus is more stable than influenza A, with less antigenic drift and consequent immunologic stability, and does not undergo the process of antigenic shift, its participation in epidemics is minimal, being of lesser academic interest. The aim of this work was to describe the occurrence of a series of SARS cases in a municipality in the Central region of Brazil. This is a case series study with a descriptive and quantitative approach of Serious Acute Respiratory Syndrome (SARS) in institutionalized individuals and a health professional from a long-term institution in the municipality where the study was conducted. The variables studied were: age, comorbidities, vaccination status, date of symptoms onset, symptoms, occurrence of death, information regarding provided care (hospital care, exams, and medications). The study was approved by the Research Ethics Committee of the Federal University of Goiás (UFG), under opinion number 2.167.287. Case 1 was diagnosed with influenza B, treated with antibiotic therapy, with no antiviral drugs administered, and culminated in death. Cases 2 and 3 were confirmed as influenza B, being treated with antiviral drugs and discharged due to full recovery. Case 4 was confirmed as influenza B virus by epidemiological link, treated with antiviral drugs and discharged due to full recovery. An early diagnosis, adequate clinical management, transmissible disease research based on the 11 health promotion steps and actions can promote the reduction of morbimortality by influenza type B

Diagnosis of Hantavirus for genomic detection with phylogenetic study and production of recombinant N protein

Hantaviroses são zoonoses distribuídas mundialmente, associadas a roedores (família Muridae) e transmitidas ao homem pela inalação de partículas virais contidas nas excretas destes roedores. Os Hantavírus são causadores de duas doenças principais: a Febre Hemorrágica com Síndrome Renal, de baixa letalidade, que ocorre na Ásia e Europa e a Síndrome Pulmonar e Cardiovascular por Hantavírus (SPCVH) que ocorre nas Américas, associada a roedores da subfamília Sigmodontinae. Hantavirus são um gênero na família Bunyaviridae. São envelopados, com 80 a l10 nm, possuindo RNA de fita simples e polaridade negativa, divididos em 3 segmentos, o grande (L), o médio (M) e o pequeno (S), que codificam respectivamente a polimerase viral, as glicoproteínas de superfíce (G1 e G2) e a proteína do nucleocapsídeo (N). Vários Hantavírus tem sido identificados em países da América do sul, incluindo o Brasil, onde são conhecidas as espécies, Juquitiba (JUQ), Araraquara (ARA), Castelos dos Sonhos (CAS), Anajatuba (ANJ) ou Rio Mearim (RIME). A SPCVH é problema de saúde pública na região de Ribeirão Preto, Estado de São Paulo, onde já se notificou 37 casos desde 1998, com alta letalidade. O diagnóstico de infecções por Hantavírus no país tem sido feito por ELISA, detectando anticorpos contra a proteína N dos vírus Sin Nombre e Andes. Um método alternativo ao ELISA é a RT-PCR. Como parte deste trabalho, selecionou-se primers de alta homologia entre distintos Hantavírus, para utilização em RT-PCR. Por esta técnica, o gene N de Hantavírus foi detectado em 8 de 9 amostras séricas de pacientes com SPCVH (88,9%) e em todas as 9 amostras por uma nested-PCR (100%). Seqüenciou-se nucleotídios dos amplicons da RT-PCR e da nested-PCR evidenciando alta similaridade destas seqüências com o gene N do Hantavírus ARA (94,8% a 99,1 %). A RT-PCR e a nested-PCR mostraram-se métodos úteis ao diagnóstico de infecções por Hantavirus, prestando-se seus ... produtos a estudos filogenéticos. Também, selecionou-se primers para amplificação por RT-PCR dos genes G1 e G2. De 10 pacientes que tiveram amplicons de G1, G2 e N seqüenciados, fez-se análise filogenética. Para tanto, seqüências foram primeiramente alinhadas pelo programa ClustalW e editadas com o BioEdit e os cladogramas feitos com POY 3.1.1, por otimização direta, utilizando algoritmo de máxima-parsimônia e análise simultânea de N, Gl e G2. A edição de árvores foi feita no programa WinClada e mostrou 3 grupos distintos de vírus da América do Sul. O do vírus Andes (AND), o de outros vírus Argentinos, Pergamino (PRG) , Maciel (MAC), Lechiguanas, Hu39694 e o do vírus Araraquara (ARA), que se agrupou com as seqüências de fragmentos genômicos obtidos dos pacientes. A proximidade filogenética mostra que o Hantavirus causador de SPCVH na região de Ribeirão Preto foi o ARA. Também, seqüenciou-se todo o segmento S do ARA, e com este buscou-se expressar o gene da proteína N. Para tanto, selecionou-se vários primers e com eles, produziu-se amplicons de \'DA ORDEM DE\' 500, \'DA ORDEM DE\' 1400, \'DA ORDEM DE\' 1500 e \'DA ORDEM DE\' 1700 pb, que foram clonados e seqüenciados. O segmento S inteiro de ARA possui 1858 nucleotídios (nt) e a sua região codificadora de N, 1287 nt. Comparou-se filogeneticamente o gene de N com o de outros Hantavírus, tendo este se mostrado-se mais similar com MAC e PERG. Expressou-se o gene de N com novos primers e um amplicon com tamanho esperado foi diretamente clonado no vetor de expressão pET 200D, que transformou colônia de Escherichia coli BL21. Esta bactéria, induzida com IPTG produziu altos teores de uma proteína detectável em WesternBlot por anticorpos de soros de pacientes convalescentes de SPCVH. Avaliou-se o grau de solubilidade e purificou-se esta proteína N por coluna de afinidade Ni-NTA, quantificou-se o produto e utilizou-se o mesmo em ELISA juntamente com controle ... negativo (Escherishia coli, contendo plasmídeo sem gene de N). O ELISA foi testado em 24 amostras de soro utilizando a proteína N recombinante de ARA como antígeno, sendo que 17 destas foi detectado anticorpos IgG contra Hantavírus e pode ser utilizado rotineiramente no diagnóstico da SPCVH.Hantaviruses, family Bunyaviridae, have been implicated as etiologic agents for two acute diseases: The Hemorrhagic Fever with Renal Syndrome (HFRS), occurring in the Europe and Asia and Hantavirus Cardio-Pulmonary Syndrome (HCPS), occurring in the Américas. Both diseases are carried by rodent vectors and they are transmitted to humans by contact or through inhalation of aerosols contaminated with saliva, feces or urine of the infected rodent host. In the case of American hantaviruses which cause HCPS, these hosts belong to the order Rodentia, family Muridae, subfamily Sigmodontinae. The genome of these viruses consist of three single-stranded RNA segments, S, M and L, which encode the nucleocapsid (N), glycoproteins (G1 and G2), and the polymerase protein, respectively. Presently, HCPS is more common in countries of South America than in North America. In Brazil more than 540 cases of HCPS have ever been reported. Among to the total reported cases, 37 cases occurred in the Ribeirão Preto region with a case-fatality rate of 50%. This is an emerging infectious disease with profound impact on public health and therefore it demands studies involving laboratory diagnosis and genomic characterization of strains circulating of hantaviruses. Previous studies have characterized 4 different hantavirus genetic lineages in Brazil, which were named according to the site of detection: Juquitiba (JUQ), Castelo dos Sonhos (CAS), Araraquara (ARA) and Anajatuba (ANJ) or Rio Mearim (RIME). The diagnosis of hantavirus infections in Brazil is done by an enzyme-linked immunosorbent assay based on detection of Immunoglobulin G (IgG) and M (IgM) responses to non-homologous recombinant nucleocapsid (N) proteins of Sin Nombre (SN) and Andes (AND) viruses. An altemative molecular approach used to detect gene regions of hantaviruses would be the RT-PCR and nested-PCR. In this study, in order to determine the genotypes and distribution of hantavirus causing HCPS, selected primers anneling to regions of high similarity were used initially to amplify partial N regions of hantavirus in a RT-PCR. Partial sequences of the N gene of hantavirus were detected in 8 out of 9 serum samples obtained from patients with HCPS (88,9%) and in 9 samples by nested-PCR (100%). Nucleotides sequences of the N genes of hantavirus obtained from all amplicons showed high similarity (94,8% to 99,1%) when compared with Araraquara (ARA) virus partial sequences of the N gene. The RT-PCR and Nested-PCR demonstrated to be a useful tool for the molecular diagnosis of hantavirus infection also becoming possible the molecular analysis of genomic characterization of strains circulating of hantaviruses. Additionaly, for a more refined diagnostic and robust phylogenetic analysis two pairs of primers were selected in high homology regions of G1 and G2 glycoprotein genes after multiple alignment among nucleotide sequences of the M segment of hantavirus. These selected primers were used together with previous selected primers for the N gene. RT-PCR detected in 16 blood serum samples obtained from patients with HCPS the production of three amplicons (N, G1 and G2). Three amplicons of each 10 blood serum samples out of 16 were cloned, sequenced and their obtained sequences compared with different hantaviruses. For phylogenetic analysis, after multiple alignment performed with ClustalW software, the sequences obtained from blood serum samples of patients together sequences of different hantavirus were splited into short fragments with BioEdit software to construct the cladograms by POY 3.1.1 software by optimization direct method (DO), using the principie of maximum parsimony with jackknife support. Four cladograms were constructed for each N, G1 and G2 genes, including one of them by simultaneously analysis (N+ G1 + G2). To visualize the trees, WinClada software was used showing in a similar way in four trees ARA virus agrouped with patients sequences, demonstrating that HCPS cases were caused by ARA. Also, in this study we report the entire nucleotide sequence of the S segment of a Brazilian hantavirus aiming the expression of homologous recombinant antigens with N protein of a local virus as well as the phylogenetic analysis with complete S segment. This sequence was obtained by RT-PCR using the combination of new primers selected from terminal nucleotide sequence after multiple aligment of South American Hantavirus with others previously described for diagnosis based on N gene. The S segment of ARA consists 1858 nucleotide (nt) along with a coding region of 1287 nt, similar to others South American Hantavirus. For expression of N protein, new selected primers flanking the coding region of N gene were used in a RT-PCR. This amplicons was directly cloned in pET 200D expression vector and transformed in Escherichia coli, BL21 to induce the production ofthe N recombinant protein with IPTG. High levei of expression of histidine-tagged fusion N protein with a molecular mass of 52 kDa was determined by sodium dodecyl sulfatepolyacrylamide gel electrophoresis (SDS-PAGE) analysis. The N protein was also recognized by sera of patients that they had had HCPS by Western Blotting assay. By chromatography on nickel-agarose column, the expressed N protein was purified and used together with a negative control (E. coli BL21 containing plasmid without N gene) in a ELISA for detection of IgG. Finally, ELISA were tested for 24 serum samples, showing reaction with 17 HCPS patients serum samples. At last, this assay can be used for studies of seroprevalence and serological diagnosis for HCPS

Assessment of Immunogenicity and Neutralisation Efficacy of Viral-Vectored Vaccines Against Chikungunya Virus

Chikungunya virus (CHIKV) has caused extensive outbreaks in several countries within the Americas, Asia, Oceanic/Pacific Islands, and Europe. In humans, CHIKV infections cause a debilitating disease with acute febrile illness and long-term polyarthralgia. Acute and chronic symptoms impose a major economic burden to health systems and contribute to poverty in affected countries. An efficacious vaccine would be an important step towards decreasing the disease burden caused by CHIKV infection. Despite no licensed vaccine is yet available for CHIKV, there is strong evidence of effective asymptomatic viral clearance due to neutralising antibodies against the viral structural proteins. We have designed viral-vectored vaccines to express the structural proteins of CHIKV, using the replication-deficient chimpanzee adenoviral platform, ChAdOx1. Expression of the CHIKV antigens results in the formation of chikungunya virus-like particles. Our vaccines induce high frequencies of anti-chikungunya specific T-cell responses as well as high titres of anti-CHIKV E2 antibodies with high capacity for in vitro neutralisation. Our results indicate the potential for further clinical development of the ChAdOx1 vaccine platform in CHIKV vaccinology

Modular Approach To Energy Efficient Driver Assistance Incorporating Driver Acceptance

Objective: Hantaviruses are rodent-borne RNA viruses that have caused hantavirus cardiopulmonary syndrome in several Brazilian regions. In the present study, geographical distribution, seroprevalence, natural host range, and phylogenetic relations of rodent-associated hantaviruses collected from seven counties of Southeastern Brazil were evaluated. Methods: ELISA, RT-PCR and phylogenetic analysis were used in this study. Results: Antibodies to hantavirus were detected in Bolomys lasiurus, Akodon sp. and Oligoryzomys sp., performing an overall seroprevalence of 5.17%. All seropositive rodents were associated with grasslands or woods surrounded by sugar cane fields. Phylogenetic analysis of partial S- and M-segment sequences showed that viral sequences isolated from B. lasiurus specimens clustered with Araraquara virus. However, a sequence from Akodon sp. shared 100% similarity with Argentinian/Chilean viruses based on the partial S- segment amino acid sequence. Conclusion: These results indicate that there are associations between rodent reservoirs and hantaviruses in some regions of Southeastern Brazil, and suggest the existence of additional hantavirus genetic diversity and host ecology in these areas. Copyright (C) 2008 S. Karger AG, Base

Hantavirus cardiopulmonary syndrome: a report of two cases

Infection with hantavirus, from the family Bunyaviridae, causes hantavirus cardiopulmonary syndrome (HCPS) in the Americas. This highly lethal anthropozoonosis afflicts preferentially individuals in rural areas and is transmitted by aerosol of excreta from infected wild rodents. The aim of this study is to report the almost simultaneous occurrence of two cases of HCPS in the municipality of Jataí, state of Goiás, Brazil

Role of mixed Th1 and Th2 serum cytokines on pathogenesis and prognosis of hantavirus pulmonary syndrome

The hantavirus pulmonary syndrome (HPS) is an emerging syndrome in the Americas. The disease results from intense immune activation and changes in vascular permeability. The aim of this study was to determine the profile of serum cytokines in HPS patients looking for correlation with the clinical parameters, severity and outcome of illness. Studying 21 HPS patients, we found that IL-6 may have an important role in the pathogenesis of HPS, being associated with fatal outcome. Our results also support a mixed Th1/Th2 immune response during the course of HPS and that the magnitude of Th1 response effector cytokines is correlated to HPS severity. The decreased levels of TGF-beta observed in HPS patients suggest that immunoregulatory activity could be damaged in these patients. (c) 2008 Elsevier Masson SAS. All rights reserved.Fundaqao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP), Brazil[02/14149-1

Evaluation of an Enzyme-Linked Immunosorbent Assay Based on Araraquara Virus Recombinant Nucleocapsid Protein

Laboratory diagnosis of hantavirus cardiopulmonary syndrome (HCPS) in Brazil has been performed mostly by a detection of IgM antibodies to recombinant antigen purified from Sin Nombre virus and Andes Virus (ANDV). Recently, a recombinant nucleocapsid (rN) protein of Argentina virus (ARAV), a Brazilian hantavirus, was Obtained in Escherichia coli. To evaluate ARAV rN as antigen for antibody detection, serum samples from 30 patients front Argentina seropositive for hantavirus were tested. All samples were positive for IgG and IgM by enzyme-linked immunosorbent assay (ELISA) using either ARAV rN or ANDV rN antigens. In Brazil, six of 00 serum samples from patients With suspected HCPS (10%) were positive for IgM by ELISA Using ARAV rN antigen and 7 were positive Using ANDV rN antigen. For results obtained with 90 serum samples analyzed by IgM ELISA with ANDV rN antigen, the sensitivity of the IgM ELISA using ARAV rN antigen was 97.2%,, the specificity was 100%, the positive predictive value was 100% and the negative predictive value was 98.1%. The results show that ARAV rN is a Suitable antigen for diagnosis Of hantavirus infection in Brazil and Argentina

Public Health from UFG; coordinator of the Epidemiological Surveillance Center of the Municipal Health Office in Jataí

abstRaCt Infection with hantavirus, from the family Bunyaviridae, causes hantavirus cardiopulmonary syndrome (HCPS) in the Americas. This highly lethal anthropozoonosis afflicts preferentially individuals in rural areas and is transmitted by aerosol of excreta from infected wild rodents. The aim of this study is to report the almost simultaneous occurrence of two cases of HCPS in the municipality of Jataí, state of Goiás, Brazil

Association of-308G/A polymorphism in the tumor necrosis factor-alpha gene promoter with susceptibility to development of hantavirus cardiopulmonary syndrome in the Ribeiro Preto region, Brazil

Activation of the immune response in hantavirus cardiopulmonary syndrome (HCPS) leads to a high TNF production, probably contributing to the disease. The polymorphic TNF2 allele (TNF -308G/A) has been associated with increased cytokine production. We investigated the association of the TNF2 allele with the outcome of hantavirus infection in Brazilian patients. A total of 122 hantavirus-exposed individuals (26 presenting HCPS and 96 only hantavirus seroconversion) were studied. The TNF2 allele was more frequently found in HCPS patients than in individuals with positive serology for hantavirus but without a history of HCPS illness, suggesting that the TNF2 allele could represent a risk factor for developing HCPS.Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP), Brazil[02/14149-1