Molecular characterization of influenza viruses collected from young children in Uberlandia, Brazil - from 2001 to 2010.

BACKGROUND: Influenza remains a major health problem due to the seasonal epidemics that occur every year caused by the emergence of new influenza virus strains. Hemagglutinin (HA) and neuraminidase (NA) glycoproteins are under selective pressure and subjected to frequent changes by antigenic drift. Therefore, our main objective was to investigate the influenza cases in Uberlândia city, Midwestern Brazil, in order to monitor the appearance of new viral strains, despite the availability of a prophylactic vaccine. METHODS: Nasopharyngeal samples were collected from 605 children less than five years of age presenting with acute respiratory disease and tested by immunofluorescence assay (IFA) for detection of adenovirus, respiratory syncytial virus, parainfluenza virus types 1, 2, and 3 and influenza virus types A and B. A reverse transcription-PCR (RT-PCR) for influenza viruses A and B was carried out to amplify partial segments of the HA and NA genes. The nucleotide sequences were analyzed and compared with sequences of the virus strains of the vaccine available in the same year of sample collection. RESULTS: Forty samples (6.6%) were tested positive for influenza virus by IFA and RT-PCR, with 39 samples containing virus of type A and one of type B. By RT-PCR, the type A viruses were further characterized in subtypes H3N2, H1N2 and H1N1 (41.0%, 17.9%, and 2.6%, respectively). Deduced amino acid sequence analysis of the partial hemagglutinin sequence compared to sequences from vaccine strains, revealed that all strains found in Uberlândia had variations in the antigenic sites. The sequences of the receptor binding sites were preserved, although substitutions with similar amino acids were observed in few cases. The neuraminidase sequences did not show significant changes. All the H3 isolates detected in the 2001-2003 period had drifted from vaccine strain, unlike the isolates of the 2004-2007 period. CONCLUSIONS: These results suggest that the seasonal influenza vaccine effectiveness could be reduced because of A H3N2 variants that circulated in 2001-2003 years. Thus, an early monitoring of variants circulating in the country or in a region may provide important information about the probable efficacy of the vaccine that will be administered in an influenza season.

Respiratory virus infections in hospitalized and non-hospitalized children: determinants of severe course of the disease.

INTRODUCTION: Viral respiratory disease constitutes a great burden worldwide mainly among children. OBJECTIVE: One pursued to compare disease characteristics of children who required hospitalization from those who did not require hospitalization due to a viral respiratory disease. METHODOLOGY: Medical and demographic data were collected through questionnaires and nasopharyngeal aspirates were tested for detection of respiratory disease viruses of in and outpatients up to five years old, presenting acute respiratory infection. RESULTS: Respiratory syncytial virus predominated among hospitalized children while other viruses (Human rhinovirus, Influenza virus, Parainfluenza virus, Adenovirus, and Human metapneumovirus) together predominated among non-hospitalized patients. Although children with underlying risk condition required longer hospitalization, previously healthy children presented severe disease and required hospitalization as well. Also, clinical characteristics were not found that may distinguish RSV infected children who had comorbidities from those previously healthy. CONCLUSIONS: Children who were hospitalized due to respiratory distress had well defined characteristics: early age, respiratory syncytial virus infection, bronchiolitis and presence of comorbidity. Nevertheless, rapid respiratory syncytial virus identification among early age children may be of great value in order to avoid medical misconduct, such as unnecessary antibiotic prescription and preventive health care before an eventual clinical worsening encompassing previous health status.

Human rhinovirus in the lower respiratory tract infections of young children and the possible involvement of a secondary respiratory viral agent.

Human rhinoviruses (HRV) are usually associated with mild respiratory symptoms in children. However, some studies have found that HRV can cause severe disease, especially when the patient is co-infected with a second virus. In this study, 532 nasopharyngeal aspirates (NPAs) were collected over a nine-year period from children at the Clinics Hospital of Uberlândia. The collected NPAs were then tested for HRV RNA using the reverse transcription-polymerase chain reaction. Eighty-three specimens from children diagnosed with lower respiratory tract illness (LRTI) were positive for HRV RNA and were then tested for the presence of eight other respiratory viruses. A second virus was detected in 37.3% (31/83) of the samples. The most frequent clinical diagnosis was bronchiolitis, followed by other LRTI and then pneumonia. The frequency of severe disease in children infected with more than one virus was not significantly different from the frequency of severe disease in children infected with HRV alone. Children infected with both HRV and parainfluenza virus (1.5 m.o.) were significantly younger than those infected by HRV alone (5.0 m.o.) (p = 0.0454). Overall, these results suggest that infection with a second virus does not lead to a higher frequency of severe syndromes in children presenting with LRTI.

Molecular characterization of adenoviruses from children presenting with acute respiratory disease in Uberlândia, Minas Gerais, Brazil, and detection of an isolate genetically related to feline adenovirus.

Human adenoviruses (HAdV) are a major cause of acute respiratory diseases (ARD), gastroenteritis, conjunctivitis and urinary infections. Between November 2000-April 2007, a total of 468 nasopharyngeal aspirate samples were collected from children with ARD at the Clinics Hospital of Uberlândia. These samples were tested by immunofluorescence assay (IFA) and 3% (14/468) tested positive for the presence of HAdV. By performing polymerase chain reaction (PCR) to detect HAdV DNA in samples that tested negative or inconclusive for all viruses identifiable by IFA (respiratory syncytial virus, parainfluenza viruses 1, 2 and 3, influenza viruses A and B and HAdV), as well as negative for rhinoviruses by reverse transcription-PCR, additional 19 cases were detected, for a total of 33 (7.1%) HAdV-positive samples. Nucleotide sequences of 13 HAdV samples were analyzed, revealing that they belonged to species B, C and E. Further analyses showed that species C (HAdV-2) was the most prevalent among the sequenced samples. To our knowledge, this is the first report describing the presence of HAdV-4 in Brazil. We also detected an isolate that was 100% identical to a part of the feline adenovirus hexon gene sequence.

Human rhinovirus and disease severity in children.

OBJECTIVE: To evaluate retrospectively human rhinovirus (HRV) infections in children up to 5 years old and factors involved in disease severity. METHODS: Nasopharyngeal aspirates from 434 children presenting a broad range of respiratory infection symptoms and severity degrees were tested for presence of HRV and 8 other respiratory viruses. Presence of host risk factors was also assessed. RESULTS: HRV was detected in 181 (41.7%) samples, in 107 of them as the only agent and in 74 as coinfections, mostly with respiratory syncytial virus (RSV; 43.2%). Moderate to severe symptoms were observed in 28.9% (31/107) single infections and in 51.3% (38/74) coinfections (P = .004). Multivariate analyses showed association of coinfections with lower respiratory tract symptoms and some parameters of disease severity, such as hospitalization. In coinfections, RSV was the most important virus associated with severe disease. Prematurity, cardiomyopathies, and noninfectious respiratory diseases were comorbidities that also were associated with disease severity (P = .007). CONCLUSIONS: Our study showed that HRV was a common pathogen of respiratory disease in children and was also involved in severe cases, causing symptoms of the lower respiratory tract. Severe disease in HRV infections were caused mainly by presence of RSV in coinfections, prematurity, congenital heart disease, and noninfectious respiratory disease.

Serum mannose-binding lectin levels are linked with respiratory syncytial virus (RSV) disease.

The innate immune response facilitates the quality of the adaptive immune response and is critical to an individual's susceptibility to infection and disease. Mannose-binding lectin (MBL) is a plasma protein with anti-microbial properties that binds a wide range of pathogens to flag them for immune destruction independent of antibodies. In this study, serum MBL levels were measured in 81 children <5 years old experiencing acute respiratory syncytial virus infection and in 40 control children to determine the association with disease severity. Almost 70% of all RSV-infected children had low to intermediate MBL levels (<500 ng/ml) compared to controls, and most of the <6 months old RSV interned patients had low to intermediate levels. No differences were detected in MBL levels between case and control children <1 month old. Analysis of the T-cell compartment in peripheral blood mononuclear cells (PBMC) from acute RSV-infected and control children showed that the percent CD4+ T cells was statistically lower in RSV-infected children > or =6 months old compared to controls, while the percent CD8+ T cells in RSV-infected and control PBMC was generally similar. These results suggest that low serum MBL levels may be a marker of RSV disease severity in children and that MBL may be important in limiting RSV disease pathogenesis.

Human rhinovirus in the lower respiratory tract infections of young children and the possible involvement of a secondary respiratory viral agent

Human rhinoviruses (HRV) are usually associated with mild respiratory symptoms in children. However, some studies have found that HRV can cause severe disease, especially when the patient is co-infected with a second virus. In this study, 532 nasopharyngeal aspirates (NPAs) were collected over a nine-year period from children at the Clinics Hospital of Uberlândia. The collected NPAs were then tested for HRV RNA using the reverse transcription-polymerase chain reaction. Eighty-three specimens from children diagnosed with lower respiratory tract illness (LRTI) were positive for HRV RNA and were then tested for the presence of eight other respiratory viruses. A second virus was detected in 37.3 percent (31/83) of the samples. The most frequent clinical diagnosis was bronchiolitis, followed by other LRTI and then pneumonia. The frequency of severe disease in children infected with more than one virus was not significantly different from the frequency of severe disease in children infected with HRV alone. Children infected with both HRV and parainfluenza virus (1.5 m.o.) were significantly younger than those infected by HRV alone (5.0 m.o.) (p = 0.0454). Overall, these results suggest that infection with a second virus does not lead to a higher frequency of severe syndromes in children presenting with LRTI.

Molecular characterization of adenoviruses from children presenting with acute respiratory disease in Uberlândia, Minas Gerais, Brazil, and detection of an isolate genetically related to feline adenovirus

Human adenoviruses (HAdV) are a major cause of acute respiratory diseases (ARD), gastroenteritis, conjunctivitis and urinary infections. Between November 2000-April 2007, a total of 468 nasopharyngeal aspirate samples were collected from children with ARD at the Clinics Hospital of Uberlândia. These samples were tested by immunofluorescence assay (IFA) and 3 percent (14/468) tested positive for the presence of HAdV. By performing polymerase chain reaction (PCR) to detect HAdV DNA in samples that tested negative or inconclusive for all viruses identifiable by IFA (respiratory syncytial virus, parainfluenza viruses 1, 2 and 3, influenza viruses A and B and HAdV), as well as negative for rhinoviruses by reverse transcription-PCR, additional 19 cases were detected, for a total of 33 (7.1 percent) HAdV-positive samples. Nucleotide sequences of 13 HAdV samples were analyzed, revealing that they belonged to species B, C and E. Further analyses showed that species C (HAdV-2) was the most prevalent among the sequenced samples. To our knowledge, this is the first report describing the presence of HAdV-4 in Brazil. We also detected an isolate that was 100 percent identical to a part of the feline adenovirus hexon gene sequence.

Identification and molecular characterization of the infectious bursal disease virus (IBDV) from an outbreak in a broiler flock in midwestern Brazil

Para identificar e caracterizar o agente causador de um quadro clínico sugestivo de doença de Gumboro (DG) que afetou um plantel de frangos de corte com 34 dias de idade, em Buriti Alegre (estado de Goiás, centro-oeste do Brasil), no ano de 2001, procedeu-se uma combinação de métodos virológicos clássicos e modernos. Análises histopatológicas de bursas revelaram necrose, depleção de folículos linfóides, infiltração de heterófilos, edema e formação de cistos, lesões compatíveis com DG. A inoculação em ovos embrionados de galinhas SPF (specific pathogen-free) de uma suspensão de macerado de amostras de bursas resultou em mortalidade embrionária e lesões macroscópicas compatíveis com aquelas provocadas pelo VDIB. Amostras de bursas foram submetidas à técnica de transcrição reversa-reação em cadeia da polimerase (RT-PCR), utilizando-se oligonucleotídeos específicos para amplificação da região hipervariável do gene da VP2. Essa reação produziu um fragmento do tamanho esperado, que foi digerido pelas enzimas de restrição TaqI, StyI e SspI, mas não foi digerido com SacI. Este padrão foi o mesmo observado com cepas de VDIB hipervirulentas (vvVDIB). Análise da seqüência nucleotídica revelou a presença dos aminoácidos alanina, isoleucina e isoleucina nas posições 222, 256 e 294, respectivamente, característica dessas cepas. Além disso, análise filogenética realizada agrupou a cepa encontrada, denominada de BR-GO, com outras cepas de vvVDIB.

Estudo do comportamento "in vitro"e em animais inoculados, de 29 amostras patogênicas de Staphylococcus aureus reeconhecidas pela fagotipagem / Behaviour \"in vitro\" and \"in vivo\" of strains of Staphylococcus aureus detected by Phagotypage

29 amostras de Staphylococcus aureus, padronizadas e pertencentes aos diversos tipos bacteriológicos: I: 52/52 A-79/80, II: 3A/3C/55/71, III: 42E/47/53/54/75/77/83A/84/85, nåo Classificados: 94/95/96/81, Experimentais: 86/88/89/90/92/DIIHK2, Extra - 42D/187, foram submetidas aos testes clássicos de reconhecimento de patogenicidade. Alguns foram feitos "in vitro" tais como:DNAse, lecitinase, coagulase em lâminas e em tubos, produçåo de hemólise em ágar sangue de carneiros, coelhos e humanos, sensibilidade a 6 agentes antibióticos e fermentaçåo do manitol. Outros testes, "in vivo", foram realizados em camundongos e coelhos de algumas das amostras escolhidas aleatoriamente. Pretendeu-se avaliar o valor prático de cada um dos testes e a sensilidade dos mesmos na detecçåo das amostras patogênicas, comparando-os com os resultados encontrados na literatura pertinente (4,5,10,12). Todas as amostras, com exceçåo da PS 52 demonstraram a atividade de DNAse, embora em diferentes níveis; alguns quase imperceptíveis. Quanto à prova de lictinase foi considerada fortemente positiva e fraca para outras, como por exemplo a PS 52. A coagulase ocorreu em todas as amostras testadas em lâminas e em tubos, também em diferentes níveis, à exceçåo da PS 3A que nåo apresentou coagulase livre. Nem todas as amostras foram hemolíticas nos ágar sangue: humano, de coelho ou de carneiro. As PSs 52 e 47 nåo foram hemolíticas em nehum destes tipos de ágar sangue utilizados. Todas as amostras foram sensíveis à rifamicina e resistentes à penicilina natural. O manitol foi fermentado por todas as amostras, em 36 horas. (4). Apenas a PS 84 foi fatal para o camundongo (injeçåo intraperitoneal de 0,1 ml, aproximadamente 100.000 germes), em 72 horas; e várias provocaram reaçöes inflamatórias localizadas em coelhos (inoculaçöes da mesma suspensåo utilizadas para camundongos), na coxa direita

Prevalência de estreptococos beta-hemolíticos em populaçäo infantil sadia de Goiânia - Goiás - 1989 / Prevalent beta-hemolytic streptococci in normal children in Goiânia - Goiás - 1989

Vinte e três amostras de estreptococos beta-hemolíticos foram isoladas de 279 escolares aparentemente sadios e incluídos na faixa etária de cinco a 10 anos, em Goiânia, Goiás, 1989. Das amostras reconhecidas laboratorialmente, 10 foram provenientes de 143 crianças de escolas particulares do centro da cidade e elevado padräo de vida e 13 de 136 crianças de escolas públicas dos subúrbios, com padräo de vida baixo. As amostras foram identificadas através dos testes bioquímicos e sorológicos. Os estreptococos beta-hemolíticos do grupo A (Streptococus pyogenes) foram prevalentes sobre os outros estreptococos hemolíticos. Os fatores sócio-econômicos, tamanho das famílias e estado nutricional parecem influenciar estes resultados

Soroprevalência de anticorpos para o vírus da rubéola em populaçåo urbana de Goiânia-Go - Brasil / Soroprevalence of antibodies against rubella virus among urban population in Goiânia-Go - Brazil

Análise sorológica visando detecçåo de anticorpos para o vírus da rubéola foi feita em 1025 indivíduos da cidade de Goiânia - Goiás, no período de 1982 a 1990. A faixa etária variou de < 1 mês a 74 anos de idade. Observou-se uma soroprevalência de 86,5 pôr cento através da técnica de Inibiçåo da Hemaglutinaçåo, tomando como padråo de soropositividade o título maior ou igual a 1:16. A soroprevalência aumentou com a idade para indivíduos maiores que 6 anos