Laboratory diagnosis of respiratory viral infection has traditionally been based upon virus isolation and/or viral antigen identification. Recently, more sensitive and specific nucleic acid detection methods by reverse transcription- polymerase chain reaction (RT-PCR) have been developed, however, conventional RT-PCR can identify only a single suspected virus. To identify the causative agents which belong to Paramyxoviridae of respiratory virus infections, we have developed a single-tube multiplex RT-PCR using four primer sets which can amplify respiratory syncytial virus and parainfluenza virus type 1, 2 and 3 simultaneously. Assay sensitivity of single-tube multiplex RT-PCR allowed a detection in the range of 3~500 TCID50 and there were no cross amplification among other respiratory viral agents based on the test using reference virus stocks. The single-tube multiplex RT-PCR was able to directly detect viruses in respiratory specimens, with virus being detected 11 of 80 samples as compared to 9 of 80 samples detected by indirect immunofluorescence or antigen detection following shell vial culture. This result suggests that the single-tube multiplex RT-PCR can be established as a more sensitive and rapid diagnostic application than shell vial assay for the detection of respiratory infection of Paramyxoviridae.
Clinical Laboratory Techniques ; Fluorescent Antibody Technique, Indirect ; Humans* ; Parainfluenza Virus 1, Human* ; Paramyxoviridae ; Paramyxoviridae Infections* ; Polymerase Chain Reaction* ; Respiratory Syncytial Virus, Human* ; Respiratory Syncytial Viruses ; Reverse Transcription*

Human parainfluenza viruses (HPIVs) is one of the main causes of acute respiratory tract infections in children. HPIVs have been grouped into four serotypes (HPIV1~HPIV4) according to serological and genetic variation. Different serotypes of HPIVs have diverse clinical disease spectrum, epidemic characteristics and disease burden. Based on the nucleotide variation in structural protein genes, HPIVs can be further divided into distinct genotypes and subtypes with diverse temporal and spatial distribution features. The standard molecular typing methods are helpful to clarify the gene evolution and transmission patterns of HPIVs in the process of population transmission. However, the development of molecular epidemiology of HPIVs has been hindered by the lack of a standardized molecular typing method worldwide. Therefore, this study reviewed the viral characteristics, genome structure, existing genotyping methods and evolution of HPIVs, and screened the reference strains for molecular typing, so as to improve the understanding of gene characteristics and molecular typing of HPIVs, and provide an important scientific basis for the monitoring and research of molecular epidemiology of HPIVs in China.
Child ; Humans ; Molecular Typing ; Parainfluenza Virus 1, Human/genetics* ; Parainfluenza Virus 2, Human/genetics* ; Parainfluenza Virus 3, Human/genetics* ; Paramyxoviridae Infections/epidemiology* ; Respiratory Tract Infections/epidemiology*

PURPOSE: This study aimed to investigate the association between respiratory virus infection and pneumococcal colonization in children. METHODS: From May 2009 to June 2010, nasopharyngeal (NP) aspirates were obtained from patients under 18 years old who visited Seoul National University Children's Hospital for respiratory symptoms. NP samples were used to detect respiratory viruses (influenza virus A and B, parainfluenza virus 1, 2 and 3, respiratory syncytial virus A and B, adenovirus, rhinovirus A/B, human metapneumovirus, human coronavirus 229E/NL63 and OC43/HKU1) by RT-PCR and pneumococcus by culture. RESULTS: Median age of the patients was 27 months old. A total of 1,367 NP aspirates were tested for respiratory viruses and pneumococcus. Pneumococcus was isolated from 228 (16.7%) of samples and respiratory viruses were detected from 731 (53.5%). Common viruses were rhinovirus (18.4%), respiratory syncytial virus (RSV) A (10.6%), adenovirus (6.9%), influenza virus A (6.8%). Pneumococcal isolation rate was significantly higher in the cases of positive virus detection than negative detection [21.3% (156/731) vs. 11.3% (72/636), P<0.001]. For individual viruses, pneumococcal isolation rate was positively associated with detection of influenza virus A [24.7% (23/93) vs 16.1% (205/1274), P=0.001], RSV A [28.3% (41/145) vs 15.3% (187/1222), P=0.001], RSV B [31.3% (10/32) vs 16.3% (218/1335), P=0.042], rhinovirus A/B [22.6% (57/252) vs 15.3% (171/1115), P=0.010]. CONCLUSION: The study revealed that pneumococcal isolation from NP aspirates is related with respiratory virus detection. The result of this study could be used to investigate how respiratory viruses and pneumococcus cause clinical diseases.
Adenoviridae ; Child* ; Colon* ; Coronavirus ; Humans ; Metapneumovirus ; Orthomyxoviridae ; Paramyxoviridae Infections ; Respiratory Syncytial Viruses ; Rhinovirus ; Seoul ; Streptococcus pneumoniae

PURPOSE: Acute lower respiratory tract infections (ALRI) in children are mostly caused by viruses. This study aimed to define the causative viruses, and clinical manifestations during 4 years (2002-2006) and to determine seasonal occurrence of viral ALRI in Korean children by using our cumulative 10 year data (1996-2006). METHODS: A total of 3,854 hospitalized patients due to viral ALRI at Samsung Medical Center, from October 2002 to July 2006, were analyzed. Nasopharyngeal aspirate was obtained for virus cultures. Respiratory viruses were identified using indirect immunofluorescent staining. RESULTS: Viral agents were isolated in 9.8% (378 cases). The common identified pathogens were parainfluenza virus type 3 (32.3%), respiratory syncytial virus (RSV) (29.6%) and adenovirus (14.0%). The clinical patterns of viral ALRI were pneumonia (73.0%), bronchiolitis (20.9%), croup (3.7%) and tracheobronchitis (2.4%). The occurrence of viral ALRI was highest under 2 years of age. Pneumonia developed mostly by parainfluenza virus type 3 and RSV. The most frequent cause of bronchiolitis was RSV. Croup was frequently caused by parainfluenza virus. During the past 10 years, infections with parainfluenza virus type 3 and RSV epidemically occurred, whereas adenovirus was isolated throughout the study period. CONCLUSION: We could look through the etiological aspect of ALRI among pediatric inpatients during 10 years in Seoul by adding this 4 year data to the former 6 years data in our hospital. Our results may contribute to prevention and control of viral respiratory tract infections.
Adenoviridae ; Bronchiolitis ; Child ; Child, Hospitalized ; Croup ; Epidemiologic Studies ; Humans ; Inpatients ; Korea ; Parainfluenza Virus 3, Human ; Paramyxoviridae Infections ; Pneumonia ; Respiratory Syncytial Viruses ; Respiratory System ; Respiratory Tract Infections ; Seasons ; Viruses

Apoptosis, as a part of the natural defense mechanisms that protect against viral infection, plays a vital role in the pathogenic mechanisms. It also plays a key role in the pathogenesis of diseases including many viral diseases. Mechanisms of virus-induced apoptosis are not completely understood because of the complexity of the underlying biochemical cascades and all of the participating host factors. Mumps virus belongs to the genus Rubulavirus in the family Paramyxoviridae. It contains single stranded RNA genome with negative polarity. It was observed that mumps virus induced apoptosis in VeroE6 cells, and adsorption and penetration of mumps virus to cell membrane alone were not sufficient for the induction of cell death. When mumps virus was superinfected onto nucleocapsid protein (NP) expressing VeroE6 cells, cell viability and facterial titer were maintained until 13 and 12 day, respectively. The levels of p53, Bax, and Bcl-2 were increased in NP-expressing VeroE6 cells, and the increase in Bax, and Bcl-2 was outstanding. It was observed that NP protein did not directly affect the efficiency of the infection of mumps virus in NP-expressing VeroE6 cells. The levels of p53, and Bax were decreased in both mock-infected VeroE6 cells and NP-expressing VeroE6 cells infected with mumps virus. However, the Bcl-2 level was little affected by the virus infection.
Adsorption ; Apoptosis* ; Cell Death ; Cell Membrane ; Cell Survival ; Defense Mechanisms ; Genome ; Humans ; Mumps virus* ; Mumps* ; Nucleocapsid Proteins* ; Nucleocapsid* ; Paramyxoviridae ; RNA ; Rubulavirus ; Virus Diseases

This study was performed to investigate the etiologic agents, age distribution, clinical manifestations and seasonal occurrence of acute viral lower respiratory tract infections in children. We confirmed viral etiologies using nasopharyngeal aspirates in 237 patients of the ages of 15 years or younger who were hospitalized for acute lower respiratory tract infection (ALRI) from March 1996 to February 1998 at Samsung Seoul Hospital, Seoul, Korea. The overall isolation rate was 22.1%. The viral pathogens identified were adenovirus (12.7%), influenza virus type A (21.1%), -type B (13.9%), parainfluenza virus type 1 (13.5%), -type 2 (1.3%), -type 3 (16.0%) and respiratory syncytial virus (21.5%). The occurrence of ALRIs was highest in the first year of life, although parainfluenza virus type 1 infection occurred predominantly in the second year of life and influenza virus caused illnesses in all age groups. The specific viruses are frequently associated with specific clinical syndromes of ALRI. The respiratory agents and associated syndromes frequently have characteristic seasonal patterns. This study will help us to estimate the etiologic agents of ALRI, and establish a program for the prevention and treatment. An annual nationwide survey is necessary to understand the viral epidemiology associated with respiratory illnesses in Korea.
Acute Disease ; Adenoviridae Infections/epidemiology ; Adolescence ; Age Distribution ; Animal ; Bronchitis/virology ; Bronchitis/epidemiology ; Cell Line ; Child ; Child, Hospitalized/statistics & numerical data ; Child, Preschool ; Croup/epidemiology ; Female ; Human ; Infant ; Influenza/epidemiology ; Influenza A Virus, Human ; Influenza B Virus ; Kidney/cytology ; Korea/epidemiology ; Liver/cytology ; Male ; Parainfluenza Virus 1, Human ; Parainfluenza Virus 2, Human ; Parainfluenza Virus 3, Human ; Paramyxovirus Infections/epidemiology ; Pneumonia, Viral/virology* ; Pneumonia, Viral/epidemiology* ; Respiratory Syncytial Virus Infections/epidemiology ; Respiratory Syncytial Viruses ; Respiratory Tract Infections/virology* ; Respiratory Tract Infections/epidemiology* ; Seasons

Four viruses showing cytopathic effects in MDBK cells were isolated from brains of cattle showing downer cattle syndrome in 2012. The isolates were confirmed to belong to the genus Rubulavirus of the subfamily Paramyxovirinae. Isolate QIA-B1201 had the ability to hemagglutinate red blood cells from several species of animals and was capable of adsorbing guinea pig erythrocytes on the surface of infected Vero cells. Nucleotide sequence analysis showed that two isolates (QIA-B1201 and QIA-B1204) had high similarity with other human and animal PIV5 isolates ranging from 98.1 to 99.8%. The highest sequence similarity of the two isolates corresponded to strain KNU-11 (99.8% at the nucleotide and amino acid level) isolated from suckling piglets in Korea in 2012. To evaluate the virulence of strain QIA-B1201, we inoculated bPIV5 into 5 week-old mice via both the intraperitoneal and intracranial route. Body weight was not significantly altered in mice inoculated with QIA-B1201. In this study, we isolated and characterized novel bPIV5s from brain samples showing downer cattle syndrome, but were not able to elucidate the pathogenicity of the bPIV5s in mice.
Animals ; Base Sequence ; Body Weight ; Brain ; Cattle* ; Erythrocytes ; Guinea Pigs ; Humans ; Incidence* ; Korea ; Mice ; Parainfluenza Virus 5 ; Paramyxoviridae Infections* ; Paramyxovirinae ; Rubulavirus ; Vero Cells ; Virulence

PURPOSE: This study was performed to investigate the epidemiologic and clinical features of respiratory viruses in children with acute lower respiratory tract infection (ALRTI) in Busan and Gyeongsangnam-do, Korea. METHODS: From May 2010 to April 2011, we tested nasopharyngeal aspiration specimens in 1,520 hospitalized children with ALRTI with multiplex real time-polymerase chain reaction (RT-PCR) to identify 7 kinds of common pathogens (adenovirus [ADV], influenza virus type A [influ A], influenza virus type B [influ B], human metapneumovirus [hMPV], parainfluenza virus [PIV], human rhinovirus [hRV], respiratory syncytial virus [RSV]). We analyzed positive rates and clinical features by retrospective review of the chart. RESULTS: Virus agents were isolated from 72.5% of cases. The identified pathogens were RSV, 35.5%; hRV, 25.6%; PIV, 13.8%; ADV, 12.8%; hMPV, 7.1%; influ A, 5.0%; and influ B, 0.3%. The major period of viral ALRTI was the first year of life. Clinical diagnoses of viral ALRTI were pneumonia, 52.3%; bronchiolitis, 21.2%; tracheobronchitis, 1.0%; croup, 10.8%; and asthma, 8.8%. The most frequent case of pneumonia and bronchiolitis was RSV. Croup was frequently caused by PIV. The number of hMPV infections peaked between April and June and were primarily caused due to pneumonia. CONCLUSION: Although this study was confined to one year, this study described the features of ALRTI associated with 7 respiratory viruses in children in Busan and Gyeongsangnam-do, Korea. Additional investigations are required to define the role of respiratory viruses in children with ALRTI in this area.
Asthma ; Bronchiolitis ; Child ; Child, Hospitalized ; Croup ; Humans ; Korea ; Metapneumovirus ; Orthomyxoviridae ; Paramyxoviridae Infections ; Pneumonia ; Respiratory Syncytial Viruses ; Respiratory System ; Respiratory Tract Infections ; Retrospective Studies ; Rhinovirus ; Viruses

PURPOSE: In this study, the clinical and epidemiological characteristics of patients admitted for viral croup were analyzed to evaluate disease severity based on the organism that caused the infection. METHODS: We retrospectively reviewed the medical records of 302 patients who were admitted to the Department of Pediatrics at the Wonju Severance Hospital between May 2013 and December 2016 for viral croup. Patients who showed positive results on multiplex polymerase chain reaction were subsequently diagnosed with respiratory virus infection. The Westley scoring system was used to evaluate the severity of viral croup. RESULTS: Of the 302 patients, 149 were admitted due to severe viral croup, including 88 boys and 61 girls, with a boy-to-girl ratio of 1.44:1. About 110 cases of parainfluenza virus infection have been reported, which accounted for almost half of the total cases. The other identified viruses included influenza virus, human rhinovirus, and respiratory syncytial virus. Analysis of the association between severe viral croup and causative pathogen revealed that only parainfluenza type 2 virus showed a significantly high risk. Parainfluenza type 2 virus did not show an age-based difference in frequency but showed relatively a higher frequency of infections during the summer and fall. CONCLUSIONS: In this study, parainfluenza virus type 2 was the only virus associated with severe viral croup. To facilitate proper preventive management, treatment, and prognosis evaluation of viral croup, prospective and multicenter studies should assess the additional variables and the severity of the virus. Additionally, further studies should be conducted to assess age-dependent influences, as well as the regional and seasonal incidence of viral infection.
Child ; Child, Hospitalized ; Croup ; Epidemiology ; Female ; Gangwon-do ; Humans ; Incidence ; Medical Records ; Multiplex Polymerase Chain Reaction ; Orthomyxoviridae ; Parainfluenza Virus 2, Human ; Paramyxoviridae Infections ; Pediatrics ; Prognosis ; Prospective Studies ; Respiratory Syncytial Viruses ; Retrospective Studies ; Rhinovirus ; Seasons ; Severity of Illness Index

PURPOSE: Despite the availability of molecular methods, identification of the causative virus in children with acute respiratory infections (ARIs) has proven difficult as the same viruses are often detected in asymptomatic children. METHODS: Multiplex reverse transcription polymerase chain reaction assays were performed to detect 15 common respiratory viruses in children under 15 years of age who were hospitalized with ARI between January 2013 and December 2015. Viral epidemiology and clinical profiles of single virus infections were evaluated. RESULTS: Of 3,505 patients, viruses were identified in 2,424 (69.1%), with the assay revealing a single virus in 1,747 cases (49.8%). While major pathogens in single virus-positive cases differed according to age, human rhinovirus (hRV) was common in patients of all ages. Respiratory syncytial virus (RSV), influenza virus (IF), and human metapneumovirus (hMPV) were found to be seasonal pathogens, appearing from fall through winter and spring, whereas hRV and adenovirus (AdV) were detected in every season. Patients with ARIs caused by RSV and hRV were frequently afebrile and more commonly had wheezing compared with patients with other viral ARIs. Neutrophil-dominant inflammation was observed in ARIs caused by IF, AdV, and hRV, whereas lymphocyte-dominant inflammation was observed with RSV A, parainfluenza virus, and hMPV. Monocytosis was common with RSV and AdV, whereas eosinophilia was observed with hRV. CONCLUSION: In combination with viral identification, recognition of virus-specific clinical and laboratory patterns will expand our understanding of the epidemiology of viral ARIs and help us to establish more efficient therapeutic and preventive strategies.
Adenoviridae ; Child ; Child, Hospitalized* ; Eosinophilia ; Epidemiology ; Humans ; Inflammation ; Metapneumovirus ; Orthomyxoviridae ; Paramyxoviridae Infections ; Polymerase Chain Reaction ; Respiratory Sounds ; Respiratory Syncytial Viruses ; Respiratory Tract Infections ; Reverse Transcription ; Rhinovirus ; Seasons