Development and verification of multiplex real-time RT-PCR assay for classing and identification of HPIV1,HPIV2 and HPIV3 / 中国生物制品学杂志

@#ObjectiveTo develop and verify a multiplex real-time RT-PCR assay for simultaneous identification of human parainfluenza virus type 1(HPIV1),type 2(HPIV2)and type 3(HPIV3).MethodsThe whole genome sequences of HPIV1,HPIV2 and HPIV3 were downloaded from the database for alignment analysis,and the conserved regions were selected. Specific primers and probes were designed for the three viruses respectively to develop a multiplex real-time RTPCR assay with human ribonuclease P(RNase P)as theinternal quality control. The method was verified for the sensitivity,specificity and precision and used to detect 192 clinical samples.ResultsAfter optimization,the multiplex real-time RTPCR reaction system was determined to be 30 μL,including 10 × NeoscriptRTase/UNG Multi mix 3 μL,5 × Neoscript RT Premix Multi Buffer 6 μL,upstream and downstream primers of HPIV1,HPIV2 and HPIV3(100 μmol/L)0. 1 μL respectively,HPIV1,HPIV2,HPIV3 probes(100 μmol/L)0. 05 μL respectively,RNase P upstream and downstream primers(50 μmol/L)0. 06 μL respectively,RNase P probe(50 μmol/L)0. 03 μL respectively,template 15 μL,and ddH2O supplemented to 30 μL. The reaction conditions were 50 ℃ 20 min,95 ℃ 3 min and 45 cycles of 95 ℃ 15 s and54 ℃ 30 s. Fluorescence signals were collected during annealing in each cycle. The minimum detection limits of HPIV1,HPIV2 and HPIV3 were all 500 copies/mL by the multiplex real-time RT-PCR assay;The method showed no cross-reaction with influenza A virus,influenza B virus,respiratory syncytial virus andnovel coronaviruses. The coefficients of variation(CVs)in intra-and inter-groups of the recombinant plasmid standard mixture with three different concentrations were all less than 3%. HPIV1,HPIV2 and HPIV3 were detected in 192 clinical samples,and the positive rates were7. 81%,0. 05% and 3. 1%,respectively.ConclusionThe multiplex real-time RT-PCR assay for detection of HPIV1,HPIV2 and HPIV3 developed in this study has good sensitivity,specificity and precision,which has a high clinical application prospect in the field of rapid diagnosis and identification of HPIV.

Characteristics of hemagglutinin-neuraminidase gene of human parainfluenza virus type 3 among patients with acute respiratory tract infection in Henan Province / 中华微生物学和免疫学杂志

Objective:To analyze the molecular characteristics of hemagglutinin-neuraminidase (HN) gene of human parainfluenza virus type 3 (HPIV3) among the cases with acute respiratory tract infection (ARI) in Henan Province.Methods:Nasal/throat swab samples collected from patients with severe acute respiratory tract infection (SARI) in Luohe and patients with influenza-like illness (ILI) in Zhengzhou were used in this study. HPIV nucleic acids in the samples were detected using real-time fluorescent PCR. HPIV3-positive samples were subjected to RT-PCR for the amplification of HN genes and the sequences were analyzed with Sanger method. CExpress and MEGA7.0 software were used for sequences editing, evolution tree construction and gene sequence analysis.Results:A total of 374 throat swab samples collected form ARI cases in Luohe and Zhengzhou were tested and 20 (5.3%) of them were positive for HPIV3. Eighteen HPIV3 HN gene sequences were successfully amplified and all belonged to C3 subgroups, including 16 sequences of C3f genotype and two sequences of C3a genotype. The 18 HN gene sequences shared the homology of 97.6%-100.0% in nucleotide and 99.3%-100.0% in amino acid, but the differences between them and the prototype strain Wash/47885/57 were significant. There were 12 amino acid mutations shared by them, including four function-related mutations (H295Y, I391V, D556N and I53T). There were no significant differences in the nucleotide or amino acid sequences as compared with the epidemic strain of China/BCH4210A/2014.Conclusions:The C3f and C3a branches of HPIV3 were the epidemic genotypes in Henan Province in recent years and a local circulating prevalence might be established. Continuous and in-depth monitoring of HPIV3 C3 subtype would be of great significance for the prevention and control of HPIV3-associated diseases.

Viral spectrum of 36 outbreaks of influenza like-illness in Shandong province / 中华实验和临床病毒学杂志

Objective@#To understand the viral spectrum of influenza-like illness (ILI), Shandong province during 2013-2014.@*Methods@#The data of 36 ILI outbreaks were collected and analyzed. Multiple respiratory pathogens were detected with RT-PCR in pharynx swab specimens.@*Results@#Totally, 35 outbreaks occurred in winter and spring, and dispersed in 12 cities; 27 outbreaks happened in primary and secondary schools. Many of the outbreaks, 17 (47.2%), were caused by influenza virus type B (FluB), followed by FluA H1 and H3, 5 (13.8%) outbreaks, respectively. Mixed infection caused 7 outbreaks. Totally 437 samples were collected, with 235 (53.8%) positive specimens, in which FluB was mostly detected, 142 (32.5%). Except Flu, parainfluenza virus type 2 (PIV2) was the most frequently detected, followed by coronavirus (CoV). The constituent ratio of FluB under 15 years of age was the highest, such as CoV in 25-59 years old group and FluA H1 in others groups.@*Conclusions@#ILI outbreaks occurred mostly in primary and secondary schools. Virus was the main pathogen, with the dominant strains of FluB. However, the dominant strain in different age groups was different.

Parainfluenza virus pneumonia after lung transplantation :a case report and literature review / 中华器官移植杂志

Objective To explore the clinical characteristics ,diagnosis ,treatment and prognosis of parainfluenza virus (PIV) pneumonia after lung transplantation .Methods One case of PIV pneumonia after lung transplantation was retrospectively analyzed . The relevant domestic and foreign cases and literature review were summarized .Results The recipient underwent sequential bilateral lung transplantation for chronic obstructive pulmonary disease ,bullae and respiratory failure .Donor lung was sourced from donation after cardiac death .Routine anti-rejection therapy was prescribed postoperatively .At 14 months ,cough and shortness of breath lead to hospitalization for over 1 month .At 15 months ,sputum/fungal smear and culture showed that nucleic acid of PIV was positive . The definite diagnosis was PIV pneumonia after lung transplantation .After ribavirin antiviral therapy ,tracheal intubation and invasive ventilation ,followed by imipenem plus doxycycline plus anti-infective therapy ,ganciclovir antiviral therapy ,repeated bronchoscopic sputum aspiration and lavage treatment ,the patient's condition deteriorated and died from breathing failure and septic shock at 16 months .Conclusions Preventing PIV infection after lung transplantation is of vital importance .PCR is essential for a rapid detection of virus infection .However ,there is no curative treatment of PIV infection .Specific parainfluenza immunoglobulin and DAS 181 aerosol inhalation may be applied for future treatment of PIV infection in lung transplant recipients .

Human parainfluenza virus surveillance in pediatric patients with lower respiratory tract infections: a special view of parainfluenza type 4 / Vigilância dos vírus parainfluenza humanos em pacientes pediátricos com infecções do trato respiratório inferior: uma visão especial do parainfluenza tipo 4

Abstract Objective: Characterize the role of human parainfluenza virus and its clinical features in Brazilian children under 2 years of age presenting with acute lower respiratory tract infections. Methods: Real-time assays were used to identify strains of human parainfluenza virus and other common respiratory viruses in nasopharyngeal aspirates. One thousand and two children presenting with acute lower respiratory tract illnesses were enrolled from February 2008 to August 2010. Results: One hundred and four (10.4%) patients were human parainfluenza virus positive, of whom 60 (57.7%) were positive for human parainfluenza virus-3, 30 (28.8%) for human parainfluenza virus-4, 12 (11.5%) for human parainfluenza virus-1, and two (1.9%) for human parainfluenza virus-2. Seven (6.7%) patients had more than one strain of human parainfluenza virus detected. The most frequent symptoms were tachypnea and cough, similar to other viral respiratory infections. Clinical manifestations did not differ significantly between human parainfluenza virus-1, -2, -3, and -4 infections. Human parainfluenza virus-1, -3, and -4 were present in the population studied throughout the three years of surveillance, with human parainfluenza virus-3 being the predominant type identified in the first two years. Conclusion: Human parainfluenza viruses contribute substantially to pediatric acute respiratory illness (ARI) in Brazil, with nearly 30% of this contribution attributable to human parainfluenza virus-4.

Resumo Objetivo: Caracterizar o papel do VPH-4 e suas características clínicas em crianças brasileiras com menos de dois anos de idade com infecções agudas do trato respiratório inferior. Métodos: Ensaios em tempo real foram utilizados para identificar tipos de VPH e outros vírus respiratórios comuns em aspirados nasofaríngeos. Mil e duas crianças com doença aguda do trato respiratório inferior foram inscritas para participar de fevereiro de 2008 a agosto de 2010. Resultados: 104 (10,4%) pacientes eram VPH positivos, dos quais 60 (57,7%) eram positivos para VPH-3, 30 (28,8%) para VPH-4, 12 (11,5%) para VPH-1 e dois (1,9%) para VPH-2. Sete (6,7%) pacientes apresentaram mais de um tipo de VPH detectado. Os sintomas mais frequentes foram tosse e taquipneia, semelhantes a outras infecções respiratórias virais. As manifestações clínicas não diferiram de forma significativa entre as infecções por VPH-1, -2, -3 e -4. Os VPH-1, -3 e -4 estavam presentes na população estudada ao longo dos três anos de vigilância, e o VPH-3 foi o tipo predominante identificado nos primeiros dois anos. Conclusão: Os VPHs contribuem substancialmente para a DRA pediátrica no Brasil com quase 30% dessa contribuição atribuível ao VPH-4.

Transferência de imunidade passiva (TIP) e dinâmica de anticorpos específicos em bezerros naturalmente expostos para as viroses respiratórias / Passive immune transfer (PIT) and dynamics of specific antibodies in calves naturally exposed to the respiratory viruses

Esta pesquisa avaliou a TIP e a dinâmica de anticorpos (ACs) específicos em bezerros naturalmente expostos aos agentes causadores da doença respiratória bovina (DRB). Foram selecionados 19 bezerros Holandeses alimentados com colostro proveniente de doadoras vacinadas para DRB. Amostras de soro foram obtidas antes e após a ingestão do colostro (48h) para a soroneutralização (SN). Os valores médios (log2) detectados após colostragem foram de 11,5±1,6 (BVDV), 8,8±1,3 (BoHV-1), 5,5±1,6 (BRSV) e 8,4±1,5 (BPIV-3). Cinco bezerros foram criados do nascimento aos 240 dias de vida, observando-se decréscimo nos títulos de ACs para BVDV, BoHV-1 e BPIV-3 ao longo do tempo (P≤0,001). As taxas de infecções detectadas entre o D14 e o D240 foram de 40% (2/5), 20% (1/5), 80% (4/5), e 60% (3/5), respectivamente, para BVDV, BoHV-1, BRSV e BPIV-3. A maioria dos bezerros manifestou broncopneumonia após as infecções virais. Os bezerros apresentaram ACs para todas as viroses às 48 horas de vida, porém os títulos adquiridos para o BRSV foram baixos. A susceptibilidade para as infecções variou de acordo com os níveis e a duração dos títulos de ACs maternos.(AU)

This research evaluated the PIT and the dynamics of specific antibody (Ab) for calves naturally exposed to the viral agents involved in Bovine Respiratory Disease (BRD). Nineteen Holstein calves fed colostrum from vaccinated donors for DRB. Serum samples were obtained before and after colostrum intake (48h) for serum neutralization (SN). Mean values (log2) detected after colostrum feeding were 11.5±1.6 (BVDV), 8.8 ±1.3 (BoHV-1) 5.5±1.6 (BRSV) and 8.4±1.5 (BPIV-3). Five calves were raised from birth to 240 days of life and presented a decrease in Ab titers for BVDV, BoHV-1 and BPIV-3 over time (P≤ 0.001). Infection rates from D14 to D240 were of 40% (2/5), 20% (1/5), 80% (4/5) and 60% (3/5), respectively for BVDV, BoHV-1, BRSV and BPIV-3. Most of the calves presented bronchopneumonia after seroconversion to the virus. Calves presented Ab for all viruses at 48 hours of life, however BRSV Ab titer were low. Levels and persistence of maternal antibody titers determined the susceptibility to viral infections.(AU)

Innate Immune Response to Viral Infections in Primary Bronchial Epithelial Cells is Modified by the Atopic Status of Asthmatic Patients

PURPOSE: In order to gain an insight into determinants of reported variability in immune responses to respiratory viruses in human bronchial epithelial cells (HBECs) from asthmatics, the responses of HBEC to viral infections were evaluated in HBECs from phenotypically heterogeneous groups of asthmatics and in healthy controls. METHODS: HBECs were obtained during bronchoscopy from 10 patients with asthma (6 atopic and 4 non-atopic) and from healthy controls (n=9) and grown as undifferentiated cultures. HBECs were infected with parainfluenza virus (PIV)-3 (MOI 0.1) and rhinovirus (RV)-1B (MOI 0.1), or treated with medium alone. The cell supernatants were harvested at 8, 24, and 48 hours. IFN-α, CXCL10 (IP-10), and RANTES (CCL5) were analyzed by using Cytometric Bead Array (CBA), and interferon (IFN)-β and IFN-λ1 by ELISA. Gene expression of IFNs, chemokines, and IFN-regulatory factors (IRF-3 and IRF-7) was determined by using quantitative PCR. RESULTS: PIV3 and RV1B infections increased IFN-λ1 mRNA expression in HBECs from asthmatics and healthy controls to a similar extent, and virus-induced IFN-λ1 expression correlated positively with IRF-7 expression. Following PIV3 infection, IP-10 protein release and mRNA expression were significantly higher in asthmatics compared to healthy controls (median 36.03-fold). No differences in the release or expression of RANTES, IFN-λ1 protein and mRNA, or IFN-α and IFN-β mRNA between asthmatics and healthy controls were observed. However, when asthmatics were divided according to their atopic status, HBECs from atopic asthmatics (n=6) generated significantly more IFN-λ1 protein and demonstrated higher IFN-α, IFN-β, and IRF-7 mRNA expressions in response to PIV3 compared to non-atopic asthmatics (n=4) and healthy controls (n=9). In response to RV1B infection, IFN-β mRNA expression was lower (12.39-fold at 24 hours and 19.37-fold at 48 hours) in non-atopic asthmatics compared to atopic asthmatics. CONCLUSIONS: The immune response of HBECs to virus infections may not be deficient in asthmatics, but seems to be modified by atopic status.

Epidemiology and Clinical Characteristics of Parainfluenza Virus Type 4 in Korean Children: a Single Center Study, 2015–2017

PURPOSE: We aimed to identify the epidemiology and the clinical characteristics of human parainfluenza virus type 4 (HPIV-4) infection compared to HPIVs 1–3 infections in Korean children. METHODS: We reviewed medical records of children with HPIV infection who visited Seoul National University Children's Hospital from 2015 to 2017. Detection of respiratory viruses was performed using real time-polymerase chain reaction (rt-PCR), which could differentiate HPIVs 1–4. Diagnosis was classified as a febrile illness, upper respiratory tract infection (URI), croup, bronchiolitis, or pneumonia. The epidemiology, demographic features, and clinical characteristics among HPIV types were compared. The clinical data were analyzed only for the previously healthy children. RESULTS: Of the 472 children diagnosed with HPIV infection, 108 (22.9%) were previously healthy: 24 (22.2%), 19 (17.6%), 39 (36.1%), and 26 (24.1%) in HPIV types 1, 2, 3, and 4, respectively. The median age of children with HPIV-4 infection was 11 (0–195) months: the proportion of children aged 0.05). CONCLUSIONS: We observed seasonal peak of HPIV-4 infection in 2015 and 2017. HPIV-4 was a common respiratory pathogen causing lower respiratory tract infection in hospitalized children.

Research progress in interaction mechanism of enveloped glycoproteins of human parainfluenza virus type 3 / 中华实验和临床病毒学杂志

Human parainfluenza viruses (hPIVs), a series of single-stranded RNA viruses of Paramyxoviridae, are the main pathogen of respiratory tract infection. hPIV3 is the main cause of lower respiratory tract infection leading to bronchiolitis and pneumonias in young children under the age of six months, and it is the second major pathogen only next to respiratory syncytial virus (RSV). In this paper, we mainly discuss two kinds of virulence-related surface glycoprotein of hPIV3: hemagglutinin-neuraminidase (HN) protein and fusion protein (F) by briefly introducing the protein structure and physiological functions of HN and F. According to the latest research progress, we focus on the models which have been postulated to explain how F and HN work in concert to bring about membrane fusion.

Lower Respiratory Tract Diseases Caused by Common Respiratory Viruses among Stem Cell Transplantation Recipients: A Single Center Experience in Korea

PURPOSE: To describe the incidence, clinical courses, and risk factors for mortality of lower respiratory tract diseases (LRDs) caused by common respiratory viruses (CRVs) in stem cell transplantation (SCT) recipients. MATERIALS AND METHODS: We retrospectively reviewed the medical records of 1038 patients who received SCT between January 2007 and August 2011 at a single center in Korea. RESULTS: Seventy-one CRV-LRDs were identified in 67 (6.5%) patients. The human parainfluenza virus (HPIV) was the most common causative pathogen of CRV-LRDs at 100 days [cumulative incidence estimate, 23.5%; 95% confidence interval (CI), 3.3–43.7] and 1 year (cumulative incidence estimate, 69.2%; 95% CI, 45.9–92.5) following SCT. The 30-day overall mortality rates due to influenza-LRDs, respiratory syncytial virus-LRDs, HPIV-LRDs, and human rhinovirus-LRDs were 35.7, 25.8, 31.6, and 42.8%, respectively. Co-pathogens in respiratory specimens were detected in 23 (33.8%) patients. The overall mortality at day 30 after CRV-LRD diagnosis was 32.8% (22/67). High-dose steroid usage (p=0.025), a severe state of immunodeficiency (p=0.033), and lymphopenia (p=0.006) were significantly associated with death within 30 days following CRV-LRD diagnosis in a univariate analysis. Multivariate logistic regression analysis revealed that high-dose steroid usage [odds ratio (OR), 4.05; 95% CI, 1.12–14.61; p=0.033] and lymphopenia (OR, 6.57; 95% CI, 1.80–24.03; p=0.004) were independent risk factors for mortality within 30 days of CRV-LRDs. CONCLUSION: CRV-LRDs among SCT recipients showed substantially high morbidity and mortality rates. Therefore, the implement of an active diagnostic approaches for CRV infections is required for SCT recipients with respiratory symptoms, especially those receiving high-dose steroids or with lymphopenia.

Epidemiological and clinical characteristics of respiratory human parainfluenza virus infection in children / 中华实用儿科临床杂志

Objective To study the epidemiological and clinical features of human parainfluenza virus (HPIV) infection in children in Suzhou,and to provide the evidence-based foundation for early warning,diagnosis and treatment of respiratory infection in children.Methods The sputum specimens and medical history were obtained from children with acute respiratory tract infection hospitalized at the Childen's Hospital Affiliated to Soochow University from January 2006 to December 2015.Seven kinds of common respiratory viruses including respiratory syncytial virus,influenza virus A,influenza virus B,HPIV Ⅰ,HPIV Ⅱ,HPⅣV Ⅲ and adenovirus were detected by using the direct im-munofluorescence.Mycoplasma pneumoniae(MP),chlamydia pneumoniae,human bocavirns (hBoV) were detected by using fluorescence quantitative PCR.Rhinovirus and human metapneumovirus were detected by using reverse transcription-PCR.Sputum was cultured for bacteria.Results In 21 769 cases,the detection rate of HPIV positive was 3.21％ (829 cases),among which,HPIV Ⅰ,HPIV Ⅱ,HPIV Ⅲ were respectively detected in 113 cases (0.52％),16 cases (0.07 ％) and 700 cases (3.21％),respectively.There were 378 cases of simple infection and 428 cases of mixed infection,and the mixed infection was very common in Streptococcus pneumoniae,Haemophilus influenza,MP and hBoV.There was a difference in HPIV infection among genders,and the detection rate of the boys was higher than that of girls[4.14％ (563/13 591 cases) vs.3.25％ (266/8 178 cases),x2 =11.036,P =0.001].In the 28 d-1 year old and ＞ 1-3 year old group,the detection rate of HPIV was higher[4.71％ (494/10 476 cases) and 4.21％ (244/5 793 cases),respectively].In spring and summer,there was a higher detection rate of HPIV infection.The clinical manifestations with simple infection of HPIV Ⅰ and HPIV Ⅲ were cough,fever and wheezing.The rate of fever and shortness of breath in those of HPIV Ⅰ was 71.74％ (33/46 cases),10.87％ (5/46 cases),and that in HPIV Ⅲ was 40.12％ (134/334 cases),2.10％ (7/334 cases),HPIV Ⅰ infection was more likely to cause fever and shortness of breath than those of HPIV Ⅲ,there were significant differences (x2 =16.410,P ＜ 0.001;x2 =10.177,P =0.001).Pneumonia had the highest detection rate of viral infection.Conclusions HPIV Ⅲ is the leading pathogen among the types of HPIV in the hospitalized children in Suzhou area.Among the subtypes of HPIV,the peak of HPIV infection occurs in spring and summer.The children less than 3 years old are the most susceptible to parainfluenza virus,and the HPIV detection rate is gradually declines with age.

Detección de virus parainfluenza en pacientes inmunocomprometidos e inmunocompetentes atendidos en un hospital terciario de la Ciudad de São Paulo, Brasil / Detection of parainfluenza virus in immunocompromised and immunocompetent patients in a tertiary hospital in Sao Paulo, Brazil

Parainfluenza virus infections (PIV) were evaluated in patients with mild and severe infections through real time PCR. One thousand and sixty-seven samples were collected from subjects as follows: 233 adult renal transplanted outpatients, 129 children with congenital heart disease, 381 with adult hematopoietic stem cell patients and 324 hospitalized patients suspected of influenza A (H1N1) pdm09 infection. PIV was detected in 74 (6.9%) samples. VPI-3 was the most frequent (60.8%) and a higher risk was observed for older adults (p = 0.018) and for those who were hematopoietic stem cell transplanted. Further studies are needed to understand the VPI role in patients' at risk for developing serious illness.

Se evaluó la infección por virus parainfluenza (VPI) en pacientes con infecciones leves y graves mediante RPC en tiempo real. Se analizó un total de 1.067 muestras: 233 provenían de pacientes ambulatorios adultos receptores de trasplantes renales, 129 de niños con cardiopatía congénita, 381 de pacientes receptores de trasplantes de precursores hematopoyéticos adultos y 324 de pacientes hospitalizados con sospecha de influenza A (H1N1) pdm09. Se detectó VPI en 74 muestras (6,9%). Siendo VPI-3 el virus más frecuente (60,8%), se observó un mayor riesgo para los adultos mayores (p = 0,018) y para aquellos que fueron receptores de precursores hematopoyéticos. Son necesarios estudios adicionales para entender el papel del VPI en pacientes de riesgo para desarrollar enfermedad grave.

Aspectos clínicos e epidemiológicos de pneumonias infantis associadas aos quatro tipos de vírus para influenza em Fortaleza-CE

As pneumonias são problemas de saúde publica mundial, especialmente em crianças menores que cinco anos de idade. Os vírus para influenza (VPI-1, 2 e 3) são agentes frequentes de pneumonia, pouco se conhecendo sobre a participação do VPI-4devido a dificuldades do seu isolamento em cultura de células, a ausência de antígenos específicos para este vírus nos painéis de rotina de detecção dos vírus respiratórios,além de serem relacionados apenas a casos de infecções respiratórias leves. O objetivo do presente estudo é descrever o perfil epidemiológico e clínico das pneumonias causadas pelos quatro tipos de VPI na população de estudo, no período de janeiro de2013 a dezembro de 2014. Para tanto, aspirados nas ofaríngeos de 542 crianças de até cinco anos atendidas no Hospital Infantil Albert Sabin (HIAS), que receberam o diagnóstico de pneumonia, foram submetidos à RT-PCR para a detecção dos VPI-1, 2 e3 e 4. Estas amostras tinham sido analisadas anteriormente por imunofluores cência indireta para vírus sincicial respiratório (VSR), influenza (A e B), adenovírus e VPI (1,2 e 3). Os VPI foram detectados em 165 casos, seguido de VSR (136), adenovírus (34) e influenza (30). As características clínicas e epidemiológicas de pneumonias pelos VPI foram analisadas em 104 amostras que apresentaram infecção isolada por um dos quatro tipos de VPI. Os VPI mais frequentemente detectados, em ordem decrescente, foram os tipos VPI-3 (64,42%), VPI-4 (19,23%), VPI-1(14,42%) e VPI-2 (1,92%). O VPI-4 foi omais associado a co-infecções. O VPI-4 foi o único VPI cuja circulação esteve associada à estação chuvosa dos dois anos de estudo (p<0,0001). O VPI-3 e o VPI-1apresentaram pico de circulação associado à estação seca. Os VPI são agentes frequentes de pneumonias em crianças menores que cinco anos na cidade de Fortaleza.

Pneumonia are public health problems world wide, especially in childrenyounger than five years old. Human parainfluenza virus (HPIV-1, 2 and 3) are commonagents of pneumonia, little was know about the involvement of HPIV-4 due todifficulties of isolation in cell culture, the absence of antigens specific for this virus inpanels routine detection of respiratory viruses, and are associated only with cases ofmild respiratory infections. The aim of this study is to describe the clinical andepidemiological profile of pneumonia caused by four types of HPIV in the studypopulation, from January 2013 to December 2014. To this end, nasopharyngeal aspiratesof 542 children under five treated at Hospital Infantil Albert Sabin (HIAS), who werediagnosed with pneumonia, were subjected to RT-PCR for the detection of HPIV-1, 2, 3and 4. These samples had been previously analyzed by indirect immunofluorescence forrespiratory syncytial virus (RSV), influenza (A and B), adenovirus, and HPIV (1, 2 and3). The HPIV were detected in 165 cases, followed by RSV (136), adenovirus (34) andinfluenza (30). Clinical and epidemiological characteristics of pneumonia by HPIVwere analyzed in 104 samples with isolated infection with one of four types of HPIV.The HPIV most frequently detected, in descending order, were the HPIV-3 types(64.42%), HPIV-4 (19.23%), HPIV-1 (14.42%) and HPIV-2 (1.92 %). The HPIV-4 wasthe most associated with co-infection. The HPIV-4 was the only HPIV whosecirculation was associated with the rainy season of two years of study (p <0.0001). The HPIV-3 and HPIV-1 had a circulation peak associated with the dry season. The HPIVare frequent agents of pneumonia in children younger than five years in the city of Fortaleza.

The epidemiology, clinical features, phylogenetics analysis of human parainfluenza virus / 国际儿科学杂志

Human parainfluenza virus (HPIV)is one of the most common pathogens of community acquired respiratory infection.The epidemiology and clinical features vary in different subtypes.HPIV also affect other systems except respiratory system.The sequences of nucleotide and amino acid showed high identity among the strains in the same type,while remaining some variation.Most research shows the variation associated with time and district.Hemagglutinin-neuraminidase(HN) protein shows most polymorphic among all the proteins of HPIV.Some amino acid variation among the active site of HN protein will affect its activity.The influence of many gene mutations,which lead to amino acid variation,and affect the activity of HN protein remains uncertain.Further study on the epidemiology,clinical features,phylogenetics analysis of HPIV plays an important role in the prevention and control of its prevalence.

Rapid culture and identification of human parainfluenza viruses / 医学研究生学报

Objective Parainfluenza virus is an important pathogen of lower respiratory tract infections in infants and young children.This study was to search for a method for rapid culture and identification of human parainfluenza viruses from nasal swabs. Methods Nasal swab specimens were collected from 0-5 years old children with acute respiratory tract infection.The specimens were inoculated onto 96 plates with prefabricated LLC-MK2 cells and then centrifuged for 1 hour at 3000 r/min and also inoculated using the traditional culture method, followed by addition of virus mainte-nance medium containing 4 μg/mL TPCK trypsin.The cytopathic effect was observed daily, and hemagglutination and blood absorption tests were done at 2, 5, and 8 days after inoculation.In case of posi-tive result of either test, the specimen was subjected to immunofluo-rescence staining. Results Six strains of parainfluenza virus were isolated from the 83 nasal swab specimens, with a positive rate of 7.2%.There was a significant difference in the rate of separation be-tween the rapid and traditional culture methods after 2 days of culturing (7.2%vs 0%, P<0.05).The infected cells produced a cy-topathic effect that characterized by syncytium and crush formation.Hemagglutination and blood adsorption tests were positive at 4℃and negative at the room temperature.Immunofluorescence staining exhibited specific apple green fluorescence. Conclusion The method for rapid culture and identification of human parainfluenza viruses in nasal swab specimens was successfully established, which can be used to obtain and identify parainfluenza viruses with virulence and biological activity in 2 days.

Isolation & molecular characterization of human parainfluenza virus in Chennai, India.

Background & objectives: Human parainfluenza virus (HPIV) accounts for a significant proportion of lower respiratory tract infections in children as well as adults. This study was done to detect the presence of different subtypes of HPIV from patients having influenza like illness (ILI). Methods: Throat and nasal swabs from 232 patients with ILI who were negative for influenza viruses were tested by multiplex reverse transcription polymerase chain reaction(mRT-PCR) for the detection of human parainfluenza virus. All samples were inoculated in rhesus monkey kidney (LLC-MK2) cell line. Results: Of the 232 samples, 26(11.2%) were positive by mRT-PCR and nine (34.6%) showed cytopathic effect with syncytium formation for HPIV and all were HPIV-3 serotype, other serotypes like 1,2,4 were negative. The HPIV-3 strains (HN gene) were sequenced and analysed. Two novel mutations were identified at amino acid residues 295 and 297. Interpretation & conclusions: The mRT-PCR assay offers a rapid, sensitive and accurate diagnostic method for detection of HPIV which enables early detection and control. In our study there was a predominance of HPIV among 1-5 yr age group and the school going age group was less affected. Further studies need to be done to characterize HPIV isolated from different parts of the country.

Encefalomielitis aguda diseminada grave: Comunicación de un caso asociado a infección por virus parainfluenza 3 / Severe acute disseminated encephalomyelitis associated with parainfluenza 3 infection: Case report

Disseminated encephalomyelitis (ADEM) is an infrequent condition with considerable morbidity and mortality in adult patients. It requires a high level of suspicion and diagnosis emerges by gathering clinical information, laboratory exams and images studies. ADEM is related to an immunological phenomena occurring after a bacterial/viral infection or recent vaccination. Glucocorticoids are the first line treatment, reserving immunoglobulins and plasmapheresis to refractory cases. We report a male patient aged 25, with ADEM associated to parainfluenza 3 virus respiratory infection that required mechanical ventilation and that had a complete recovery only after plasmapheresis.

La encefalomielitis aguda diseminada es una enfermedad infrecuente pero de elevada morbi-mortalidad en pacientes adultos. Demanda una sospecha y diagnóstico precoz que requiere el concurso de información clínica, pruebas de laboratorio y estudio de imágenes. De sustrato inmunológico, se puede relacionar a una infección viral, bacteriana o inmunización reciente. Los glucocorticoides son el tratamiento de elección, mientras que la inmunoglobulina intravenosa y la plasmaféresis se reservan para casos refractarios. Se presenta el caso de una encefalomielitis aguda diseminada grave, en un paciente de sexo masculino de 25 años, asociado a una infección respiratoria por virus parainfluenza 3. Requirió conexión a ventilación mecánica y tuvo una respuesta completa con plasmaféresis.

Serosurveillance and establishment of a reverse transcription-polymerase chain reaction assay for bovine parainfluenza virus type 5 / 대한수의학회지

Bovine parainfluenza virus type 5 (bPIV5) was isolated from cattle with downer cow syndrome in 2012, and included both respiratory and neurotropic pathogens from a variety of animals. In the current study, we conducted serosurveillance using sera obtained from seven Korean farms and optimized a reverse transcription-polymerase chain reaction (RT-PCR) assay to detect bPIV5. The overall seropositive rate for Korean cattle was 21.4% (163/760). A farm located near the city of Milyang in Gyeoungnam province had a markedly elevated seropositive rate for bPIV5 compared to that of the other six farms. The regional seropositive rates were 4.2% (8/192) for Haman, 19.5% (18/55) for Hwasung, 73.9% (65/88) for Milyang, 26.0% (50/192) for Namwon, 1.0% (1/96) for Uljin, 13.5% (13/96) for Yeongju, and 32.7% (8/41) for Yongin. The sensitivity and specificity of three RT-PCR primer sets used to amplify the conserved fusion gene of bPIV5 were also evaluated. An RT-PCR assay using the bPIVFR3 primer set was 10-fold more sensitive than the assays using the two other primer sets and did not result in non-specific amplification. These results demonstrated that the bPIFR3 primer set can be used to detect bPIV5.

Analysis on the detection situation of respiratory viruses in 399 children with acute respiratory tract infection / 国际检验医学杂志

Objective To investigate the epidemiological feature of respiratory viruses in children with respiratory tract infection and to provide evidences for diagnosis and rational use of drugs .Methods Nasopharyngeal secretion were collected from 399 chil‐dren with acute respiratory tract infection ,and 7 respiratory viruses ,including respiratory syncytial virus (RSV) ,adenovirus(ADV) , influenza virus A (FA ) ,influenza virus B (FB) ,parainfluenza virus Ⅰ(PIVⅠ) ,parainfluenza virus Ⅱ (PIVⅡ )and parainfluenza vi‐rus Ⅲ(PIVⅢ) ,were detected by using direct immunofluorescence assay .The clinical epidemiological characteristics were analyzed by age group ,virus distribution and seasons .Results Among 399 children ,142 cases were positive for 7 viruses ,which included 40 cases of RSV infection(28 .2% ) ,26 cases of ADV infection (18 .3% ) ,43 cases of FA infection (30 .3% ) ,15 cases of FB infection (10 .6% ) ,5 cases of PIVⅠ infection(3 .5% ) ,4 cases of PIVⅡ infection(2 .8% ) and 9 cases of PIVⅢ infection(6 .3% ) .The total positive rate was 35 .6% .The number of infected infants of 3 year group were 54 cases(39 .1 .0% ) .In 4 seasons ,the positive rates were 32 .1% (spring) ,26 .9% (summer) ,29 .3% (autumn) ,45 .0% (winter)respectively .The positive rate in winter was the highest .Conclusion FA and RSV is the major virus in children with respiratory tract infection .FA infection rate in infants(>3 years old )is the highest ,and FA is most prevalent in winter .RSV infection rate in infants (< 1 years old )is the highest ,and RSV is most prevalent in spring .

Detection of virus from throat swabs of patients with respiratory tract in-fection / 中国感染控制杂志

Objective To investigate the detection of respiratory tract virus from throat swabs of patients with lower respiratory tract infection(LRTI)in a hospital,so as to provide reference for clinical diagnosis and treatment. Methods Throat swabs of hospitalized children with LRTI between May 2103 and April 2014 were detected by di-rect immunofluorescence assay,infection/carriage of respiratory syncytial virus(RSV),adenovirus(ADV),influen-za virus A(IV-A),IV-B,parainfluenza virus 1(PIV-1),PIV-2,and PIV-3 in children were surveyed.Results Of 8 425 throat swab specimens of hospitalized patients,1 879 were detected virus,the total detection rate was 22.30%,detection rate of RSV was the highest (11 .31 %),followed by PIV-3(4.37%).There was no significant difference in the total detection rate of respiratory tract virus between children of different genders (P =0.051).Of all age groups,detection rate of RSV in infants and young children was the highest,the detection rates of 7 kinds of virus from preschool children were approximate,detection rate of ADV from primary school children was the high-est (37.50%).In spring and winter,children mainly infected with RSA,accounting for 62.82% and 69.88% re-spectively;in summer,PIV-3 was the predominant virus,accounting for 41 .47%,in autumn,there was little difference in the percentage of each virus.Conclusion The detection rate of 7 kinds of virus isolated from throat swabs of hospitalized children with LRTI is related to age and season;the detection rate of 7 kinds of virus from in-fants and young children is high,RSA is the main virus.