BACKGROUNDHuman metapneumovirus (hMPV) has recently been recognized as a notable respiratory pathogen in children. However, no isolation processes and only a limited understanding of hMPV epidemiology present in Chinese children are documented by far.

METHODSNasopharyngeal aspirates from 86 hospitalized children with acute respiratory tract infections from December 2004 to July 2005 were inoculated onto Vero-E6 cells for hMPV isolation. Total RNA was extracted from infected cells with a cytopathic effect and then subjected to a RT-PCR amplification of the N and F genes of the hMPV. Nucleotide sequences of amplified F gene products were examined using a variation analysis with the MegAlign program and the phylogenetic tree construction using the neighbor-joining algorithm with a Phylip package.

RESULTSSix strains of hMPV were isolated from the samples during winter, spring and summer. The most common symptoms were coughing and cyanosis, and the diagnoses were bronchiolitis, bronchopneumonia, infantile asthma, or upper respiratory tract infection. All isolates were in the A2 genetic sublineage and shared a high percentage of homology with the F gene in the nucleotide (99.8%-100%) and amino acid (99.3%-100%) sequences.

CONCLUSIONSThis report indicates that hMPV is an important viral agent for acute respiratory tract infections present in Chongqing, China. Knowledge of phylogeny and genes will benefit the studies on the treatment and prophylaxis of hMPV infection.

China ; Humans ; Infant ; Infant, Newborn ; Metapneumovirus ; classification ; genetics ; isolation & purification ; Paramyxoviridae Infections ; virology ; Phylogeny ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo understand the genotypes of human metapneumovirus (hMPV) and the genetic character of hMPV attachment protein G sequence in Hunan, China.

METHODS232 nasopharyngeal aspirates (NPA) samples from hospitalized children with acute respiratory infections were collected from Hunan, China in 2005. HMPV was detected. The full length of G glycoprotein genes were amplified and sequenced. Bioinformatics soft-wares were employed to analyze the sequences.

RESULTS17/232 (7.3%) were showed hMPV positive. And co-infection rate with other viruses is 35%. The diagnoses of these hMPV positive cases are pneumonia, bronchiolitis and bronchopneumonia. Phylogenetic analysis for G genes from 13 hMPVs revealed the existence of four major subgroups: A1, A2, B1, B2 in Hunan, China in 2005. There are four types of sequence lengths of hMPV G glycoprotein, which are 711, 675, 660, 696nt. It is different in potential N-linked glycosylation sites and number of cysteine residues among these hMPVs of Hunan, China and Beijing, China. Also it is different from those in Japan and North America.

CONCLUSIONThe investigation of hMPV from Hunan, China in 2005 revealed the high speed of genetic variation and the marked character of geographic epidemic differences.

Amino Acid Sequence ; Child ; China ; epidemiology ; Genotype ; Glycoproteins ; classification ; genetics ; Humans ; Metapneumovirus ; classification ; genetics ; isolation & purification ; Molecular Sequence Data ; Phylogeny ; RNA, Viral ; genetics ; Respiratory Syncytial Virus Infections ; epidemiology ; virology ; Sequence Homology, Amino Acid ; Viral Proteins ; classification ; genetics

BACKGROUND: Direct antigen test (DAT) and culture are primary tests to diagnose infections by respiratory viruses, but are mainly available for the traditional viral pathogens such as respiratory syncytial virus (RSV), influenza virus, parainfluenza virus (PIV), and adenovirus in clinical laboratories. The objective of this study was to evaluate a multiplex reverse transcriptase-PCR method using Seeplex(TM) RV Detection kit (Seegene, Korea) for the detection of rhinovirus, coronavirus, and human metapneumovirus (hMPV). METHODS: From January to May 2007, nasopharyngeal aspirates (NPAs) from pediatric patients negative for culture and DAT of traditional viral pathogens were tested with Seeplex(TM). All the amplicons were directly sequenced and homology of the sequences was searched in the National Center for Biotechnology Information (NCBI) database. Patients' medical records were reviewed for clinical and demographic features. RESULTS: Forty-seven (26.4%) of 178 NPAs were positive: 18 rhinovirus, 15 hMPV, 4 RSV A, 3 coronavirus OC43, 3 influenza virus A, 2 adenovirus, 1 coronavirus NL63, and 1 RSV B. Based on maximum identity, each of the sequences indicating rhinovirus, hMPV, and coronavirus OC43 matched to the corresponding viruses with homology of 94-98%, 96-99%, and 98-100%, respectively. Seeplex(TM) positive patients were 0-11 yr old with a male:female ratio of 1.5:1. Clinical diagnoses included 9 pneumonia, 6 bronchiolitis, 2 cold, 1 asthma exacerbation for rhinovirus; 10 pneumonia, 4 bronchiolitis, and 1 clinical sepsis for hPMV; and 1 pneumonia, 2 croup, and 1 cold for coronavirus. CONCLUSIONS: Multiplex reverse transcriptase-PCR method using Seeplex(TM) RV Detection kit is a reliable test to detect rhinovirus, hMPV, and coronavirus. It may improve the diagnostic sensitivity for RSV, influenza virus, PIV, and adenovirus.
Adolescent ; Child ; Child, Preschool ; Coronavirus/classification/*isolation & purification ; Coronavirus 229E, Human/classification/genetics/isolation & purification ; Coronavirus OC43, Human/classification/genetics/isolation & purification ; Female ; Humans ; Infant ; Infant, Newborn ; Male ; Metapneumovirus/classification/genetics/*isolation & purification ; Phylogeny ; Reagent Kits, Diagnostic ; Respiratory Tract Infections/*diagnosis/virology ; Reverse Transcriptase Polymerase Chain Reaction/*methods ; Rhinovirus/classification/genetics/*isolation & purification ; Sequence Analysis, DNA

OBJECTIVETo investigate the molecular epideiological and clinical feature of human metapneumovirus in children with acute respiratory tract infection in Nanjing city, China.

METHODNasopharyngeal aspirates and nasopharyngeal swab were taken from 642 outpatients or hospitalized pediatric patients with acute at the Children Hospital of Nanjing, Jiangsu Province, China, between August 2009 and July 2010. Respiratory speciments were tested for the M gene of hMPV by reverse-transcription polymerase chain reaction (RT-PCR). All RT-PCR positive products were sequenced and phlogenetic analysis was conducted.

RESULThMPV was detected in 35 (5.5%) of the 642 children. Phylogenetic analysis revealed that 51.4% of the hMPV were B1, 31.4% were A2b. The peak of the positive rate was in April. The majority of the hMPV-positive patients(71.4%) were 0-1 years old. Of the 35 hMPV-positive patients, 15 (42.8%) were co-infected with other respiratory viruses, and human rhinovirus (HRV) were the most common additional respiratory virus. The most common clinical diagnosis was pneumonia (48.6%).

CONCLUSIONHuman metapneumovirus is an important pathogen of acute respiratory tract infection in children in Nanjing city. The subtype B1 was the predominating lineage in 2009-2010 in Nanjing city. No significant differences were found for clinical characteristics between genotype A and genotype B human metapneumovirus infection in children in Nanjing.

Acute Disease ; Child, Preschool ; China ; epidemiology ; Female ; Humans ; Infant ; Male ; Metapneumovirus ; classification ; genetics ; isolation & purification ; Phylogeny ; Respiratory Tract Infections ; epidemiology ; virology ; Reverse Transcriptase Polymerase Chain Reaction