Parainfluenza Virus Types 1, 2, and 3 in Pediatric Patients with Acute Respiratory Infections in Beijing During 2004 to 2012 / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Although human parainfluenza virus (HPIV) has been determined as an important viral cause of acute respiratory infections (ARIs) in infants and young children, data on long-term investigation are still lacking to disclose the infection pattern of HPIV in China.</p><p><b>METHODS</b>Nasopharyngeal aspirates were collected from 25,773 hospitalized pediatric patients with ARIs from January 2004 through December 2012 for respiratory virus screen by direct immuno-fluorescence assay.</p><p><b>RESULTS</b>Out of these specimens, 1675 (6.50%, 1675/25,773) showed HPIV positive, including 261 (1.01%, 261/25,773) for HPIV1, 28 (0.11%, 28/25,773) for HPIV2, and 1388 (5.39%, 1388/25,773) for HPIV3, 2 of the samples were positive for both HPIV1 and HPIV3, and 36 were co-detected with other viruses. The positive rates of HPIVs were higher in those younger than 3 years old. HPIV3 was detected from all age groups, predominantly from patients under 3 years of age, and the highest frequency was found in those 6 months to 1-year old (352/4077, 8.63%). HPIV3 was the dominant type in each of the years detected between May and July. HPIV1 showed a peak in every odd year, mainly in August or September. HPIV was detected most frequently from patients with upper respiratory infection (12.49%, 157/1257), followed by bronchitis (11.13%, 176/2479), asthma (9.31%, 43/462), bronchiolitis (5.91%, 150/2536), pneumonia (6.06%, 1034/17,068), and those with underlying diseases (1.0%, 15/1506). HPIV3 is the dominant type in these six disease groups referred above, especially in the asthma group.</p><p><b>CONCLUSIONS</b>HPIV is one of the important viral causes of ARIs in infants and young children in Beijing based on the data from the hospitalized children covering a 9-year term. HPIV3 is the predominant type in all these years and in most of the disease groups. HPIVs with different types show different seasonality.</p>

Epidemiological aspects of influenza A related to climatic conditions during and after a pandemic period in the city of Salvador, northeastern Brazil

During the influenza pandemic of 2009, the A(H1N1)pdm09, A/H3N2 seasonal and influenza B viruses were observed to be co-circulating with other respiratory viruses. To observe the epidemiological pattern of the influenza virus between May 2009-August 2011, 467 nasopharyngeal aspirates were collected from children less than five years of age in the city of Salvador. In addition, data on weather conditions were obtained. Indirect immunofluorescence, real-time transcription reverse polymerase chain reaction (RT-PCR), and sequencing assays were performed for influenza virus detection. Of all 467 samples, 34 (7%) specimens were positive for influenza A and of these, viral characterisation identified Flu A/H3N2 in 25/34 (74%) and A(H1N1)pdm09 in 9/34 (26%). Influenza B accounted for a small proportion (0.8%) and the other respiratory viruses for 27.2% (127/467). No deaths were registered and no pattern of seasonality or expected climatic conditions could be established. These observations are important for predicting the evolution of epidemics and in implementing future anti-pandemic measures.

Establishment of a high-throughput respiratory virus detection technology without RNA purification and reverse transcription / 中国医学科学院学报

<p><b>OBJECTIVE</b>To establish a convenient and high-throughput respiratory virus detection method to facilitate epidemiological viral monitoring.</p><p><b>METHODS</b>We used high-throughput microsphere-based flexible multi-analyte profiling technology (xMAP) coupled with signal amplification molecules to simultaneously detect RNAs of 8 viruses including influenza viruses A and B, parainfluenza viruses type 1, 2 and 3, respiratory syncytial viruses A and B, and metapneumovirus in a 96-well plate format. The sensitivity and specificity of the method for the synthetic viral RNAs were evaluated.</p><p><b>RESULTS</b>There was no cross-reactivity among the 8 respiratory viral target RNAs. The detection limits for the 8 viral in intro-transcribed RNAs ranged from 1204 to 4695 RNA copies.</p><p><b>CONCLUSION</b>We establish a sensitive, specific, convenient, and high-throughput multiplex detection method suitable for detecting multiple respiratory viral RNAs for epidemiological viral monitoring.</p>

Identification of Adenovirus, Influenza Virus, Parainfluenza Virus, and Respiratory Syncytial Virus by Two Kinds of Multiplex Polymerase Chain Reaction (PCR) and a Shell Vial Culture in Pediatric Patients with Viral Pneumonia

PURPOSE: Early identification of causative agents in lower respiratory infection of pediatric patients can reduce morbidity and prevent an overuse of antimicrobials. Two kinds of multiplex polymerase chain reaction (PCR) and a commercial shell vial viral culture were performed to identify causative agents in pediatric patients. MATERIALS AND METHODS: Nasopharyngeal aspirates of 220 children diagnosed with viral pneumonia were obtained. Two kinds of multiplex PCR (Seeplextrade mark RV detection kit, and Labopasstrade mark RV detection kit), and a shell vial culture by R-Mix were performed. RESULTS: Positive samples from 220 total samples by two multiplex PCRs were 52.7% and 46.4%, respectively. We also cultured 103 samples that showed positive results of the adenovirus, influenza virus, parainfluenza virus, and respiratory syncytial virus (RSV) by two multiplex PCR. The RSV was most frequently detected in 53.0% (Seeplex) and 51.7% (Labopass) of patients. The detection rate of adenovirus (AdV) was 10.3% and 12.1%, influenza virus (IFV) A and B was 12.5% and 3.4%, and parainfluenza virus (PIFV) 1, 2, and 3 were 2.9% and 2.6%. Shell vial cultures showed concordant results with each multiplex PCR by 96.1% and 77.7%, respectively. Sequencing results were 90% consistent with multiplex PCR. CONCLUSION: Multiplex PCR showed more positivity than the shell vial culture and it can be an effective primary test. Other complementary efforts such as viral cultures and sequencing analysis could be considered, according to clinical and laboratory conditions.

Parainfluenza virus infections in a tropical city: clinical and epidemiological aspects

Little information on the epidemiology and clinical characteristics of human parainfluenza virus (HPIV) infections, especially in children from tropical countries, has been published. The aim of this study was to determine the frequency of HPIV infections in children attended at a large hospital in Fortaleza in Northeast Brazil, and describe seasonal patterns, clinical and epidemiological characteristics of these infections. From January 2001 to December 2006, a total of 3070 nasopharyngeal aspirates collected from children were screened by indirect immunofluorescence for human parainfluenza viruses 1, 2, and 3 (HPIV-1, 2 and 3) and other respiratory viruses. Viral antigens were identified in 933 samples and HPIV in 117. The frequency of HPIV-3, HPIV-1 and HPIV-2 was of 83.76 percent, 11.96 percent and 4.27 percent, respectively. Only HPIV-3 showed a seasonal occurrence, with most cases observed from September to November, and with an inverse relationship to the rainy season. Most HPIV-3 infections seen in outpatients were diagnosed as upper respiratory tract infections.

Human parainfluenza virus infections in infants and young children with acute respiratory infections in Beijing / 中华儿科杂志

<p><b>OBJECTIVE</b>To understand the impact of human parainfluenza virus (HPIV) on acute respiratory infections in infants and young children in Beijing.</p><p><b>METHODS</b>Multiplex reverse transcription-PCR was used to amplify the hemagglutinin (HA) gene fragment of HPIV from clinical specimens. Primer pairs derived from a conserved region of the HA genes of HPIV were used to develop the multiplex RT-PCR for detecting and typing HPIV. The sensitivity and specificity of the method were determined by using various RNA and DNA viruses as controls. Specimens collected from 3519 children with acute respiratory infections from Aug. 2003 to Apr. 2006 were analyzed for HPIV by the multiplex RT-PCR as well as for other respiratory viruses by virus isolation and/or indirect immunofluorescent assay (IFA). Ten amplicons with expected molecular weight matching different types of HPIV were randomly selected for sequence analysis.</p><p><b>RESULTS</b>Only the cDNA from the isolated strains of HPIV 1 and 3 was positive by the multiplex RT-PCR. Phylogenetic analysis for those 10 amplicons' sequences which belong to HPIV 1 - 4 types respectively as determined by multiplex-PCR indicated that these specimens were truly HPIV positive. These 10 HPIV positive specimens included two specimens of type 4 which was further subtyped as HPIV4A and 4B by sequence analysis. With the multiplex RT-PCR, HPIV were detected in 349 out of 3519 specimens with the positive rate of 9.9% (349/3519), which is higher than 4.8% by the methods of virus isolation and/or IFA. And the HPIV positive rates were high in patients with not only acute upper but also lower respiratory tract infection. No regular seasonality distribution of HPIV infection was found. HPIV 1 and 3 were more common than HPIV 2 and 4.</p><p><b>CONCLUSION</b>With higher sensitivity and specificity than virus isolation and IFA, multiplex RT-PCR is beneficial for the etiologic and epidemiologic studies on HPIV, as well as for HPIV typing. The data from this study indicate that HPIV is one of the important etiological viruses of acute respiratory tract infections in infants and young children in Beijing.</p>

Frequency of viruses associated with acute respiratory infections in children younger than five years of age at a locality of Mexico City

A locality in the district of Tlalpan, Mexico City, was selected in order to identify the viral agents in children younger than 5 years of age with acute respiratory infection (ARI). A total of 300 children were randomly selected and were included in this study for a period of 13 months. During this period nasopharyngeal exudates were collected for the isolation of viral agents. Monoclonal fluorescent antibodies were used for viral identification after cell culture. Viral infection was detected in 65 percent of the specimens. The respiratory syncytial virus (RSV) was the most common virus agent detected. Children required an average of two consultations during the study period. Two high incidence peaks were observed, one during the summer and the other during winter; the most frequent viruses during these seasons were influenza A and RSV, respectively. The largest number of viruses was isolated in the group of children between 1 and 2 years of age and in the group between 4 and 5 years of age. This study demonstrated the presence of ARI and of different viruses in a period of 13 months, as well as the most frequent viruses in children younger than 5 years of age from a community of Mexico City.

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

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 per center (15/84) das amostras foram positivas, sendo que em 80 per center (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.

Isolamento do vírus Parainfluenza bovino tipo 3 no Rio GRande do Sul, Brasil / Isolation of bovine Parainfluenza virus type 3 in Rio Grande do Sul, Brazil

É descrito o isolamento do vírus Parainfluenza bovino tipo 3 (bPI-3) a partir de secreçöes nasais coletadas de um bovino com infecçäo respiratória. A identificaçäo do agente foi realizada através de isolamento em cultivo celular e confirmada por testes de hemaglutinaçäo, inibiçäo da hemaglutinaçäo, hemadsorçäo e imunofluorescência direta. Este é o primeiro registro do isolamento do vírus no Rio Grande do Sul.

Experimental study on effect of recombined decoction on mumps virus / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To evaluate possible inactivating effect of recombined decoction in on mumps virus.</p><p><b>METHODS</b>By adopting tissue cell culturing technology, a group of viruses including the mumps virus, herpes simplex virus (type I, II), rubella virus, cytomegalovirus (CMV), herpes zoster virus, influenza virus, parainfluenza virus, adeno viruses, respiratory syneytial virus (RSV) were cultured. The cells infected with the viruses were treated with the decoction.</p><p><b>RESULTS</b>The decoction showed remarkable inhibitory and killing effects on the mumps virus while had no obvious inhibitory and killing effects on host's cells, herpes simplex virus (type I, II), rubella virus, cytomegalovirus (CMV), herpes zoster virus, influenza virus, parainfluenza virus, adeno viruses, respiratory syneytial virus (RSV).</p><p><b>CONCLUSIONS</b>The decoction had obvious inhibitory and killing effects on mumps virus during single layer cells culture.</p>

Implementación del método rápido de diagnóstico de virus por inmunofluorescencia en niños hospitalizados por infecciones respiratorias agudas / A quick viral diagnostic method by immunofluorescence in acute respiratoty illnes in hospitalized children

Las infecciones respiratorias agudas (IRA) constituyen una causa importante de morbimortalidad en pediatría donde los virus tienen un papel relevante, los cuales pueden ser diagnosticados por la inmunofluorescencia directa IFD con sensibilidad similar al aislamiento en cultivos. El objetivo del estudio fue evaluar la frecuencia de agentes virales asociados a IRA en niños internados por esta causa en diferentes hospitales del país y conocer el aspecto clínico-epidemiológico. Se realizó un estudio de corte transversal donde se analizaron por inmunofluorescencia, aspirados nasofaríngeos, secreciones traqueales, nasales, lavados bronquiales. Se estudiaron 111 muestras de enero a octubre de 1997 de niños de 15 días a 10 años identificándose un agente viral en 59 de los casos. Los agentes hallados fueron Virus Sincicial Respiratorio (VSR) 47 por ciento, el Parainfluenza 1, 2, 3 (5,4 por ciento), el Adenovirus(3,6 por ciento), el Influenza A (5,4 por ciento), Influenza B (1,8 por ciento). Los patrones clínicos fueron Bronquiolitis (49 por ciento), Neumonías (42 por ciento) y Croup (8,5 por ciento). Los VSR se identificaron con mayor frecuencia con pico en agosto; (63 por ciento) en lactantes menores. El virus de influenza A se presentó con temporalidad similar y se relacionó con casos severos en niños mayores de 5 años, en el Chaco. El Parainfluenza se observó en menores de 1 año. El Adenovirus fue más esporádico pero uno de ellos se relacionó a un caso fatal. Se observaron asociaciones de agentes virales. Con esta técnica diagnóstico útil se dispone del primer dato sobre la epidemiología de las afecciones respiratorias agudas de probable etiología viral en la población infantil del país que requirió hospitalización, siendo similar a lo observado en otros países

Cytotoxicity of anthrax lethal factor microinjected into macrophage cells through Sendai virus envelopes.

Lethal toxin (LT) secreted by Bacillus anthracis consists of two proteins, protective antigen (PA) and lethal factor (LF). LT causes lysis of macrophages and derived cell lines at low concentrations. PA binds to the cell surface receptors and mediates translocation of LF into cytosol of mammalian cells. Internalization of LF into cytosol by osmotic lysis of pinocytic vesicles requires high concentration of LF for cell lysis. To examine the possible cell lysis by LF at low concentration, we introduced LF directly into cytosol of J774A.1 cells through reconstituted Sendai virus envelopes. The introduction of LF lysed J774A.1 cells in a concentration dependent manner. Internalization of PA alone through virosome had no toxic effect on J774A.1 cells. In the process of cytotoxicity LF was not cleaved by cellular proteases. Unlike many protein toxins, golgi was not involved in the expression of lethal toxin activity. These results indicate that LF is the toxic component of anthrax lethal toxin and prior proteolytic processing or trafficking through golgi is not required for its activity.

Identificación temprana de virus respiratorios en niños asmáticos / Early identification the viral in reactions in asthmatic children

Se describe el papel de las infecciones virales como parte de los factores ambientales desencadenates de asma en sujetos atópicos, su predominio por grupo de edad, así como algunos factores de riesgo reconocidos para el desarrollo de sibilancias de repetición. Se mencionan algunos mecanismos patogénicos por los cuales los virus promueven la inflamación de la vía aérea y se plantea su identificación temprana por pruebas de laboratorio, para la aplicación de medidas preventivas y tratamiento antiinflamatorio precoz

Inoculación experimental del paramixovirus de ojo azul en ratas de laboratorio (cepa Wistar), vía intramuscular / Experimental inoculation of blue eye paramyxovirus in laboratory rats (Wistar) intramuscularly

Se utilizaron un total de 22 ratas (cepa Wistar) de las cuales 2 animales fueron testigo y 20 ratas fueron inoculadas con 1 ml del paramixovirus del ojo azul (POA) con un título de 10 5.55 DICC/ml, por vía intramuscular. El día 0 se tomaron muestras sangúineas por vía intracardiaca de 2 animales testigos, los cuales se sacrificaron posteriormente. Los días 1, 3, 5, 7, 10, 15, 20, 25, 30, 35 posinoculación (PI) se tomaron muestras de sangre para la detección serológica de anticuerpos contra POA, utilizando la técnica de inhibición de la hemaglutinación (IHA), seroneutralización (SN), para realizar biometrías hemáticas (BH) y la obtención de la capa flogística para aislamiento viral. Además se obtuvieron muestras de órganos (encéfalo, pulmón y tonsila) para intentar el aislamiento en tres líneas celulares (MDBK, PK-15, BT) e inmunofluorescencia en cultivo celular (IFCC); de estas mismas muestras se hicieron estudios histopatológicos (HP). Se recolectaron heces y orina durante los 35 días, para el aislamiento e IFCC. En los resultados de las pruebas serológicas se detectaron anticuerpos a partir del décimo día PI, con títulos de 1:4 hasta 1:256 para SN y de 1:8 hasta 1:64 para IHA. En el aislamiento viral de órganos, capas flogísticas, heces y orina se pudo aislar el virus durante todo el periodo de experimentación en las tres líneas celulares, coincidiendo estos resultados con las IFCC; las BH mostraron variación en los valores de neutrófilos, linfocitos y monocitos. No hubo presencia de lesiones significativas tanto macro como microscópicas

Evaluation of WHO monoclonal antibody kit for diagnosis of acute respiratory viral infections.

In 1990 and 1991, six laboratories located in the WHO Western Pacific Region (WPR) and South East Asian Region (SEAR) were selected, based on their experience in the immunofluorescence antibody technique (IFAT), to participate in the evaluation of a WHO monoclonal antibody (Mab) kit to detect respiratory syncytial (RS) virus, influenza A virus, influenza B virus, parainfluenza virus and adenovirus. Despite differences in the initial standardization procedures, the WHO monoclonal antibodies were found to be of high quality, sensitivity and specificity when tested on clinical specimens. The constant supply of affordable high quality reagents from WHO would enable their use in clinical virological laboratories in the developing countries as well as promote the utilization of IFAT as an adjunct to cell culture isolation in the diagnosis of acute respiratory viral infections.

Comparative evaluation of a simple and sensitive assay for detection of orthomyxo and paramyxoviruses

Studies were done to evaluate comparatively the traditional HA assay and a more recently introduced lectin-neuraminidase (LN) methodologyin search of a simple and sensitive assay for virus detection during laboratorial diagnosis. The results proved the value of LN assay as a sensitive methodologyfor detection of virus particles, presenting results at least equal to those obtained by HA (hemagglutination) assay, with significant values of accumulated frequencies for LN/HA factors (ratios between LN and HA titers) higher than two. The accumulated values of frequencies for LN/HA factors as high as four were very significant, 72.7 (per cent) for influenzavirus and 60.7 (per cent) for Newcastle disease virus (NDV), moreover accumulated frequencies for LN/HA factors even as high as 32 were due to influenzavirus (45.4 per cent) and NDV (7.2 per cent) samples. After the storage period, most of those concentraded samples that even did not present HA titers could be detected through LN assay, demonstrating a lower threshold for virus detection

Detección de antígenos virales en lactantes con infección respiratoria aguda baja / Viral antigens in nasopharingeal exudates from infants with acute lower respiratory tract infections

Con el propósito de medir la frecuencia con que se aíslan determinados antígenos virales, en niños con infecciones respiratorias agudas bajas graves (IRAB), se realizaron ensayos de inmunofluorescencia (IF) para antígenos de virus sincicial respiratorio (VSR), adenovirus (ADV), virus parainfluenza 1, 2 y 3 (VPI), virus influenza a y b (VIa y VIb) en las secreciones nasofaríngeas de 660 niños menores de 24 meses que cumplían con criterios clínicos y radiológicos de IRAB grave comúnmente aceptados. En 322 casos (48,8%) se obtubvieron resultados positivos. Los antígenos más frecuentemente identificados fueron los de VSR (56,2%) y ADV (26,7%). La letalidad en niños con IRAB e IF positiva fue 9,9% y 6,2% en aquellos con IF negativa (p NS). En 24 de 32 (75%) pacientes fallecidos con IF positivas, el antígeno identificado correspondía a ADV, siendo el riesgo de morir en estos casos significativamente mayor que en el de pacientes portadores de otros antígenos o ninguno (Z crítico -1,65. Z calculado -1,178). La apnea fue la principal manifestación clínica en 46 pacientes, todos menores de 3 meses y 19 con reacciones de inmunofluorescencia positivas que, con una sola excepción, correspondían a antígenos de VSR

Infección respiratoria aguda baja; IRAB, en lactantes hospitalizados: aplicación de técnicas rápidas de diagnóstico etiológico / Acute lower respiratory infections; ALRI in hospitalized infants: use of rapid techniques for etiological diagnosis

Las IRAB representan, en el período del lactante, una causa importante de morbilidad, hospitalización y mortalidad, siendo los virus sus principales agentes etiológicos. Por clínica es difícil precisar etiología, siendo tratados con antibióticos. Se estudiaron en 1988, en forma prospectiva, 97 lactantes, menores de 2 años, portadores de IRAB, diagnosticada clínica y radiológicamente, con menos de 6 días de evolución. Se tomó radiografía de tórax y se investigó etiología viral mediante IF y aislamiento. Se comprobó que esta afección predomina en los lactantes menores de 1 año, principalmente durante los meses fríos y que el VRS es el más frecuente (45,5%) en casi todos los meses, con un máximo entre julio y agosto. El rendimiento de la IF es excelente para el VRS, Parainfluenza 3 e Influenza A. Pero el hallazgo de Adenovirus se duplica al agregar el aislamiento (p=<0.05). El costo de la IF es bajo y se fundamentaría en 52,6% de los pacientes la posibilidad de no dejar o de suspender el tratamiento antibiótico. Este hecho significaría: 1) Disminuir los días de hospitalización; 2) Menos agresiones al niño; 3) Ahorro de insumos; 4) Prevenir infecciones intrahospitalarias y 5) Ahorro de trabajo al personal de enfermería