OBJECTIVEHuman parainfluenza virus (HPIV) types 1, 2 and 3 are major viral pathogens responsible for upper and lower respiratory tract infections. In this study, a real-time RT-PCR was developed using multiplex primers-probe (HPIV-1, 2, 3) for the simultaneous detection of both HPIV1, HPIV2 and HPIV3 genomes.

METHODSOptimal primers and probes were designed using specialized software. The conditions for multiplex real-time RT-PCR had been optimized. The synthesis of RNA standards of HPIV1, 2, 3 were used a T7 RNA polymerase. Check the specificity sensitivities and stability of one step RT-PCR assay.

RESULTSObtained in a 10-fold dilution series assay demonstrate a high sensitivity of the assay with a lowest detection limit of 10 copies for HPIV1, 100 copies for HPIV2 and 100 copies for HPIV3.

CONCLUSIONThe assays demonstrates an improved sensitivity and scope of detecting HPIV1, 2, 3 viruses relative to routine antigen detection assays while the quantitative utility may facilitate investigation of the pre-diagnosis and respiratory virus pathogenesis.

Humans ; Oligonucleotides ; genetics ; Parainfluenza Virus 1, Human ; isolation & purification ; Parainfluenza Virus 2, Human ; isolation & purification ; Parainfluenza Virus 3, Human ; isolation & purification ; Real-Time Polymerase Chain Reaction ; methods

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*

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

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

BACKGROUNDAlthough 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.

METHODSNasopharyngeal 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.

RESULTSOut 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.

CONCLUSIONSHPIV 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.

Beijing ; epidemiology ; Child ; Child, Preschool ; Female ; Humans ; Infant ; Male ; Parainfluenza Virus 1, Human ; pathogenicity ; Parainfluenza Virus 3, Human ; pathogenicity ; Respirovirus ; pathogenicity ; Respirovirus Infections ; diagnosis ; virology

Using atomic force microscope (AFM), we investigated the images of Pars influenza virus (PIV) under different treatment conditions and observed the different appearances of the virus and its ultra-microstructure from the exterior to the interior. From the 2D images under transmission electron microscope (TEM), we could see that the surfaces of PIV particles exhibited spherical and band-shaped 'tufts'; from the 3D images under AFM, we could further observe the whole spherical virus particles and their detailed surfaces, which exhibited round and band-shaped 'tufts'. Comparing the images under TEM with those under AFM, we found that the latter could reveal the surface topograph and ultramicrostructure of viruses more truly than did the former. The samples of viruses were treated by Tritonx-100, the lipid envelopes of virions were partly or completely resolved, and then most of their capsids were exposed. We could observe the different appearances of the virions under AFM, the lipid envelopes of which were gradually removed. The samples of viruses were also treated by SDS, and the RNA was released from the virions. From the AFM images, we could see the structure of the RNA. It was thus clear that AFM could be used to investigate the different appearances and ultramicrostructure of viruses rapidly and efficiently.
Microscopy, Atomic Force ; Parainfluenza Virus 1, Human ; ultrastructure ; Parainfluenza Virus 2, Human ; ultrastructure

BACKGROUND: Survivors of childhood cancers are recommended to receive revaccinations after chemotherapy, although the universally recommended vaccination schedule for such children has not been established. We evaluated immune response following post-chemotherapy vaccinations in childhood cancer survivors.METHODS: The study included 59 patients who survived at least 5 years after completion of chemotherapy without evidence of recurrence. The patients received hepatitis-B virus (HBV) and measles, mumps, and rubella (MMR) vaccines 1 year after finishing chemotherapy according to our institutional protocol. Immune response to HBV and MMR vaccines was measured and seropositivity and factors hindering immune response to HBV and MMR vaccines were analyzed.RESULTS: The seropositivity for HBV was 88%; with a higher rate in patients with non-hematologic malignancies (100%, 18/18) than those with hematologic malignancies (78.3%, 18/23) (P=0.05) and reciprocally associated with the duration of chemotherapy (P=0.0043). The seropositivity for MMR viruses was 61%, 37% and 83% respectively, showing significantly lower response to mumps and was not different between hematologic malignancy group and non-hematologic malignancy group. Unlike HBV, the duration of chemotherapy did not affect seropositivity for MMR viruses. Ten children who failed to be immune to any of the MMR viruses received booster vaccination which resulted in seropositivity of 60% (3/5), 56% (4/9), 100% (2/2) respectively.CONCLUSION: Longer duration of chemotherapy and underlying hematologic malignancies were adversely associated with achieving immune response to HBV vaccine, but not to MMR vaccine. Our results also underline the need for booster vaccinations in non-responders to vaccinations post-chemotherapy.
Appointments and Schedules ; Child ; Drug Therapy ; Hematologic Neoplasms ; Hepatitis B virus ; Humans ; Immunization, Secondary ; Measles ; Measles-Mumps-Rubella Vaccine ; Mumps ; Recurrence ; Rubella ; Survivors ; Vaccination ; Vaccines

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

INTRODUCTIONMeasles, mumps and rubella (MMR) are viral infections causing significant mortality and morbidity for which effective and safe vaccines are available. The safety, reactogenicity and immunogenicity of a combined MMR vaccine when administered to healthy Singaporean children were evaluated in this study.

MATERIALS AND METHODSA total of 150 children aged 12 to 18 months were vaccinated in this open, single-group, single-centre study [209762/147]. Solicited local and general symptoms reported within 4 days of vaccination and fever, parotid/salivary gland swelling and signs of meningism in the 43 days following vaccination were recorded using diary cards. Serious adverse events occurring during the study period were monitored. Immunogenicity was assessed at 42 days post-vaccination.

RESULTSRedness (8.7%) and pain (7.2%) at injection site were the most commonly reported solicited local symptoms during the 4-day follow-up period after vaccination. Percentage of subjects reporting drowsiness, irritability and loss of appetite during the 4-day follow-up after vaccination was 7.2%, 8% and 7.2%, respectively. None of the solicited symptoms reported during the 4-day follow-up period was of grade "3" intensity. Fever (42.8%) was the most commonly reported solicited general symptom, with 5.1% of the children reporting fever >39.0 degrees C (axillary). No serious adverse events considered to be related to vaccination were reported. Seroconversion rates were 100% for measles and rubella antibodies and 98.1% for mumps antibodies.

CONCLUSIONSGlaxoSmithKline Biologicals' MMR vaccine was shown to be well tolerated and highly immunogenic when used in Singaporean children 12 to 18 months of age.

Female ; Health Status ; Humans ; Infant ; Infant Welfare ; Male ; Measles ; prevention & control ; Measles-Mumps-Rubella Vaccine ; adverse effects ; immunology ; Mumps ; prevention & control ; Mumps virus ; Prospective Studies ; Rubella ; prevention & control ; Singapore

Objective: To determine IgG antibody levels of measles, rubella, mumps in healthy population in Shanghai from 2010 to 2020 and analyze the trend of antibody changes in different age groups. Methods: 10 828 healthy people without measles, rubella and mumps in Shanghai were included in the study from 2010 to 2020. Serum samples were collected from 12 age groups, and the serum IgG antibody of measles, rubella and mumps were detected by ELISA. The difference of antibody positive rates and antibody levels were analyzed. Results: The median age M (Q₁, Q₃) of 10 828 objects were 8 years old (9 months old, 20 years old). Males accounted for 48.34% (5 234/10 828) and females accounted for 50.92% (5 514/10 828). Unknown gender information accounted for 0.74% (80/10 828), and 27.03% (2 927/10 828) of participants had unknown MMR immunization history. The total positive rates of measles, rubella and mumps IgG antibody were 76.78%, 64.46% and 64.29% and their GMCs were 541.45 mIU/ml, 31.76 IU/ml and 133.73 U/ml respectively. There were significant differences in serum IgG antibody GMC of measles, rubella and mumps in each year (F_measles=180.74, P<0.001; F_rubella=189.95, P<0.001; F_mumps=122.40, P<0.001). The positive rate of measles antibody was higher than that of rubella and mumps, and the difference was statistically significant (χ²=518.09, P<0.001). Conclusion: The level of measles IgG antibody in healthy people in Shanghai is higher, while the level of rubella and mumps IgG antibody is slightly lower.
Adult ; Antibodies, Viral ; Child ; China/epidemiology* ; Female ; Humans ; Immunoglobulin G ; Infant ; Male ; Measles/prevention & control* ; Measles-Mumps-Rubella Vaccine ; Mumps/prevention & control* ; Mumps virus ; Rubella/prevention & control* ; Young Adult