Objective To elucidate the prevalence and clinical characteristics of human metapneumovirus(hMPV)in hospital- ized children with respiratory infection.Methods A total of 452 hospitalized children with lower respiratory tract infection were observed from Aug 2004 to Jan 2005.Respiratory tract aspirates were collected from all patients within 48 hours after admis sion.The specimens were routinely tested for respiratory syncytial virus,influenza virus A and B,parainfluenza virus 1 to 3 and adenovirus by direct fluorescent assay(DFA).The 245 specimens negative by DFA were tested for hMPV by RT-PCR. PCR products of hMPV M gene from some patients were randomly selected for sequencing analysis.Results hMPV was identi- fied in 59(24.1%)of the 245 specimens tested,hMPV infection alone accounted for 13.1% of the infections in the 452 chil- dren under study,The prevalence of hMPV was higher than other respiratory viruses in winter.The mean age of hMPV-infec- ted children(n=59)was 27.7 months.There was no significant difference between age groups in terms of the prevalence of hMPV(P＞0.05).There were no statistically significant difference in demographics and clinical symptoms between hMPV in- fection and other common respiratory virus infection.Genotyping for the hMPV M gene from 23 Shanghai patients showed two distinct hMPV genotypes.Sequence analysis of these hMPV M genes showed 82.8%-100% homology to the registered se- quence in GenBank.There was no significant difference in clinical characteristics between the 2 genotypes.Conclusions hMPV plays an important pathogenic role in lower respiratory tract infection of children,hMPV prevailed in the winter of 2004.Clini- cally,hMPV infection can not be discriminated from the infection of other respiratory viruses.Clinical manifestation is similar between the two hMPV genotypes.

OBJECTIVE: To study the forensic application of Goldeneye DNA ID 26Y Kit in the She nationality. METHODS: Through capillary electrophoresis, the genotype of 26 Y-STR loci were analyzed in 53 unrelated male individuals from Fujian She nationality. The population genetics parameters such as allele frequency and haplotype diversity were calculated. The comparisons among the She nationality and the other nationalities were analyzed. RESULTS: A total of 126 alleles were observed on the 26 Y-STR loci of 53 unrelated male individuals. The allele frequencies and GD value ranged from 0.010 1 to 0.886 8 and 0.211 2 to 0.846 2, respectively. The GD value was greater than 0.5 in the 19 loci. A total of 47 haplotypes were observed. Based on R(ST), multidimensional scaling plot indicated that the genetic relationship among Fujian She nationality and Minnan Han nationality was closest, followed by Southern China Han nationality and Northern China nationality. CONCLUSION Goldeneye™ DNA ID 26Y Kit including 26 Y-STR loci has good polymorphism in the She nationality. As an additional system, it has forensic application value in some special cases.
Asian People/genetics* ; China ; Chromosomes, Human, Y/genetics* ; Ethnicity/genetics* ; Forensic Genetics ; Genetic Markers ; Genetics, Population ; Humans ; Polymerase Chain Reaction ; Polymorphism, Genetic ; Population Groups

Objective To find out the importance of human bocavirus (HBoV) as an infectious agent for population in Beijing, China. Seroprevalence study was conducted by using expressed recombinant major capsid VP2 protein as an antigen.Methods Serum specimens collected from infants and children who visited the Children's Hospital Affiliated to the Capital Institute of Pediatrics for health check-up and adults visiting the Xuanwu Hospital, Beijing for diseases other than respiratory infections from April 1996 to March 1997 were used for the investigation. The major capsid protein VP2 from HBoV was expressed in E. coli strain BL21 (DE3) with the transformed PET30b vector inserted with full-length VP2 gene of HBoV and the specific antigenicity of this expressed protein was validated by previous study. Western blotting was used to detect specific IgG antibody against HBoV in collected serum specimens diluted to 1:200. Mock expressed protein was E. coli cells strain BL21 (DE3) with the transformed PET30b vector without insert. Anti-His monoclonal antibody and rabbit anti-HBoV VP2 polypeptides hyper-immune serum were used as positive control for antibody detection.Results Out of 677 serum specimens tested, 400 (59.1% ) were positive for HBoV by Western blotting. About 45.3% (34/75) of the newborns under 1 month of age had anti-HBoV antibodies, and antibody positive rates were decreased in age groups of 1 and 2 months (41.4% and 31.3%, respectively) then increased in the following ages from 6 months to 7 years old ( from 45.6% to 69.7% ). The antibody positive rates were maintained at a relatively constant level ( about 70% ) in the age groups from 7 years to 40 years of age and became lower ( 61.8% - 62. 8% ) in those over 50 years.Conclusions The high seroprevalence of antibody against recombinant HBoV VP2 protein and early age antibody acquisition indicate that HBoV has been circulating in population of Beijing, China as early as in 1996 and most of children had been exposed to HBoV by the age of 7 years. Infants under the age of 6 months were susceptible to this virus.

OBJECTIVETo establish the triple-transgenic mouse model and study their biological characteristics by molecular biology, behavior and pathology.

METHODSHybrid the Tau and amyloid precursor protein (APP)/presenilins (PS1) transgenic mouse, the genotype of offspring mice were identified by PCR. Transcribed target genes were detected by RT-PCR. The protein expression of exogenous genes was detected by Western-blot. The pathological change of neurofibrillary tangles and senile plaque were observed by Bielschowsky silver staining and ABC immunohistochemical method. The changes time of learning and memory were observed by Morris water maze.

RESULTSAPP, PS1 and Tau genes were transcript in Tau/APP/PS1 mice. In 6 to 8 months old Tau/APP/PS1 mice, the neurofibrillary tangles and senile plaque could be found in cortex and hippocampus. In 6 months old Tau/APP/PS1 mice, the learning and memory abilities were worse.

CONCLUSIONWith the behavior change and pathological changes in Tau and beta-amyloid protein (AP), the Tau/APP/PS1 triple-transgenic mice can be used as a further study animal model of AD's pathogenesis and the target of drug treatment.

Alzheimer Disease ; pathology ; Amyloid beta-Protein Precursor ; genetics ; Animals ; Brain ; pathology ; Disease Models, Animal ; Learning ; Male ; Memory ; Mice ; Mice, Transgenic ; Neurofibrillary Tangles ; pathology ; Plaque, Amyloid ; pathology ; Presenilin-1 ; genetics ; tau Proteins ; genetics

objective To understand whether human metapneumovirus(hMPV)is one of the pathogens leading to the children's respiratory infections in Urumqi.Methods A total number of 209 samples were collected in the People's General Hospital of Xinjiang Uygur Autonomous Region from November 2006 to April 2007 with Some from the hospitalized children.while the others from outpatient clinic.Specimens included nasopharyngeal aspirates(NPA)and swabs were analyzed.Samples were all tested hMPV M gene by RT-PCR while the two positive PCR amplicons were sequenced and compared with other hMPV in GenBank by Blast and DNAstar.Results Of all the 209 samples.two positive ones were tested.The identities between them were 83.8%.Results from Phylogenetic analysis showed that they might belong to two different clusters.Conclusion hMPV was one of the pathogens leading to the children's respiratory tract infections in Urumqi.with two different hMPV groups existed in the same season.

Objective The objective of this study was to develop a rapid,sensitive and specific method for identifying and typing for adenovirus from clinical specimens and to learn about the viruses identified in Beijing on the molecular bases.Methods Primers were designed using hexon gene of adenovirus as target.One primer pair was designed as universal primers for amplifying a 1278 bp gene fragment located at the hcxon gene of adenovirus types 3,7,11 and 21.Four primer pairs located within the region of this 1278 bp fragment were designed specifically for amplifying adenovirusea types 3,7,II and 21.Which were used for a multiplex nest-PCR in a single tube.The products from this multiplex nest-PCR were 502 bp(for type 3),311 bp(for type 7),880 bp(for type 11)and 237 bp(for type 21)，respectively.The type of the adenovirus being tested could be determined after agarose eleetrophoresis analysis of the PCR products．Sequencing was performed for part of the Hexon genes at the 5'end from types 3．7 and 11 strains isolated from clinical specimens and the sequences were compared with corresponding genes published in GenBank．Results PCR products with predicted sizes were visualized in the agarose gel for prototype strains of adenovirus types 3,7,11 and 21,but not for other respiratory viruses,indicating that the technique is specific for typing without cross reaction with other viruses.Out of 61 clinicaI specimens which had been proved to be adenovirus positive by tissue culture and/or immunofluerescence assay,37 were found as adenovirus type 3(37161,60.73％),17 8S adenovirus type 7 (17/61,27.9％),3 Was adenovirus type 11(3161,4.9％).1 Was positive for both type 3 and 7(1／61,1.6％),suggesting that the patient Was co-infected with type 3 and 7 adenoviruses．No adenovirns type 21 was detected.Out of the 61 positive specimens,three showed positive on both tissue culture and immunofluerescence but could not be identified under the methods we used,suggesting that these 3 strains (4.9％)were with the types other than types 3,7,11 and 21．Data from sequence analysis indicated that adeno~ruses types of 3.7 and 11 in this study shared high homology with corresponding types of the strains published in GenBank.Three of the type 3 adenovirus in this study shared highest homology with the adenovirus type 3 identified in Guangzhou,China in 2005.Three of the type 7 adenovirus shared highest homology with the adenovirus type 7 identified in Japan in 1998 and 3 of type 11 adenovirus shared highest homology with the adenovirus type 11 identified in Japan in 2004.Comparing with types 3 and 7.The type 11 in this study showed highest diversity with the corresponding type in GenBank,indicated by the dispersing of the varied amino acids within the region of HVRl and HVR, of the Hexon genes.Conclusion This multiplex nest-PCR method had the advantages of rapid,sensitive and specific and could be used for identilying types of adenoviruses in clinical specimens.Although adenovirus types 3.7 and 1l from Beijing strains shared high homology with the corresponding genes in GenBank,some variances were noticed,especially in type 11 strains.

OBJECTIVEAn outbreak of acute respiratory infections in children occurred in Beijing from November to December, 2002. To investigate the etiological agents of affected children who were in day care centers and primary schools.

METHODSThroat swab specimens were collected from one primary school children with acute respiratory infections visiting one outpatient department. After centrifuging, supernatant from the specimens were inoculated into MDCK and Hep-2 cells for virus isolation and pallets for viral antigen detection and using indirect immunofluorescent assay on common respiratory viruses. Nested polymerase chain reaction was used at the same time for detection of respiratory viruses and Mycoplasma pneumoniae (Mp).

RESULTSA total number of 80 specimens were collected during the outbreak. Among them influenza B virus were detected from 18 specimens, with a positive rate of 22.5% (18/80) while Mp were detected from 13 specimens, with a positive rate of 16.3% (13/80). Influenza A3 were also detected from 2 patients (2.5%, 2/80). However, influenza A1, RSV, adenovirus and parainfluenza viruses were not found from these specimens. Influenza B virus and Mp were detected simultaneity in two specimens and influenza A3 virus and Mp were detected in one specimen.

CONCLUSIONThe outbreak of acute respiratory infection in children during the period of investigation was caused by both influenza B virus and Mp.

Adolescent ; Child ; Child, Preschool ; China ; epidemiology ; Disease Outbreaks ; Female ; Humans ; Infant ; Influenza B virus ; isolation & purification ; Influenza, Human ; epidemiology ; Male ; Mycoplasma Infections ; epidemiology ; Mycoplasma pneumoniae ; isolation & purification ; Respiratory Tract Infections ; epidemiology ; microbiology

The aim of this study was to obtain the capsid protein VP2 of human bocavirus (HBoV) identified in Beijing recently and construct virus-like particles (VLPs) in insect cells for further study of this virus. The full-length VP2 gene of HBoV from BJ3722 was inserted into the baculovirus expression transfer vector (pFastBac 1) to obtain the recombinant Bacmid, and generation of recombinant baculoviruses was followed by transfection of the recombinant Bacmid into insect cells. Then the recombinant VP2 protein was recognized by SDS-PAGE using Coomassie-blue staining and Western blot using hyper-immune serum against VP2 of HBoV from rabbit. The recombinant baculoviruses were harvested and amplified to gain large amounts of viruses with high titers to infect insect cells at a multiplicity of infection (MOI) of 0. 5. After 7-10 days or 4-5 days of the infection, the supernatants of culture or the cell lysates treated with lysing solution were harvested, and ultracentrifuged twice through 40% sucrose cushion to obtain purified VLPs, which were followed by Western blot and IFA for VLPs' composition and specificity analysis, by electron microscopy for VLPs' morphologic structure. The recombinant VP2 protein with molecular weight of approximately 61 kD expressed in recombinant baculoviruses was recognized by SDS-PAGE using Coomassie-blue staining and Western blot. The presence of VP2 on VLPs was demonstrated by Western blot and IFA from samples collected during the purification of VLPs from the supernatants of culture or the cell lysates, and the expression of VP2 in insect cells led to the formation of VLPs which formed the typical icosahedral appearance of parvoviruses with a diameter of approximately 20 nm. In conclusion, the recombinant baculoviruses were constructed, the HBoV VP2 protein was expressed in insect cells with high specific antigenicity and VLPs was formed successfully.
Animals ; Blotting, Western ; Capsid Proteins ; genetics ; metabolism ; Cell Line ; Electrophoresis, Polyacrylamide Gel ; Fluorescent Antibody Technique, Indirect ; Human bocavirus ; genetics ; metabolism ; Microscopy, Electron, Transmission ; Polymerase Chain Reaction ; Spodoptera ; Virion ; genetics ; metabolism ; ultrastructure

The aim of this study was to characterize the N and E protein encoding genes of a new human coronavirus (HCoV-NL63) which was identified from one of the clinical specimens (BJ8081) collected from a 12 years-old patient with acute respiratory infection in Beijing. The complete N and E gene sequences of HCoV-NL63 were amplified from clinical sample by RT-PCR, then were cloned into the pCF-T and pUCm-T vectors respectively and sequenced. The complete sequences of N and E genes were submitted to GenBank by Sequin and compared with N and E genes of prototype HCoV-NL63 and the other coronaviruses published in GenBank. The secondary structure and the characteristics of sample BJ8081 N and E proteins were predicted by bioinformatics. It was indicated that the N and E genes amplified from sample BJ8081 were 1134 bp and 234 bp in length and the predicted proteins including 377 amino acids and 77 amino acids, respectively. The data suggested that the region of amino acids 78-85 within N protein probably was the conserved region for all coronaviruses identified so far including HCoV-NL63. The region of amino acids 15-37 for E protein was probably the transmembrane domain. In conclusion, the recombinant plasmids pCF-T-8081 N and pUCm-T-8081 E were successfully constructed and sequenced, and the data predicted by bioinformatics are helpful for the further analysis of HCoV-NL63.
Amino Acid Sequence ; Child ; China ; Computational Biology ; methods ; Coronavirus ; classification ; genetics ; metabolism ; Coronavirus Infections ; virology ; Humans ; Molecular Sequence Data ; Nucleocapsid Proteins ; chemistry ; genetics ; metabolism ; Phylogeny ; Protein Structure, Secondary ; Sequence Analysis, DNA ; Sequence Homology, Amino Acid ; Viral Envelope Proteins ; chemistry ; genetics ; metabolism

BACKGROUNDHuman bocavirus (HBoV) is a newly identified human parvovirus that was originally detected in the respiratory secretions of children with respiratory infections. This study aimed to learn about the importance of HBoV infections by revealing the prevalence of serum antibodies against HBoV in Beijing population.

METHODSTwo batches of serum specimens collected in different periods were tested by Western blotting for specific IgG against HBoV using recombinant VP2 as antigen.

RESULTSOut of 677 serum specimens collected during April 1996 to March 1997, 400 (59.1%) were positive and antibody positive rate for another batch of 141 serum specimens collected in August, 2005 from adults aged from 20 years to over 60 years was 78.7% (111/141). Comparison of the sero-prevalence profiles for serum specimens collected during 1996 - 1997 to those collected in 2005 indicated that the antibody positive rate for specimens collected in 2005 was higher than that of the corresponding age groups collected during 1996 - 1997.

CONCLUSIONSThe data suggest that HBoV has been circulating in Beijing population for at least over 10 years, and most of children had been exposed to HBoV by age of 7 years. Higher HBoV antibody positive rate shown in the serum specimens collected in 2005 suggested that infections by HBoV have been increased in Beijing population in recent years.

Adult ; Antibodies, Viral ; blood ; Blotting, Western ; Bocavirus ; pathogenicity ; China ; epidemiology ; Humans ; Immunoglobulin G ; immunology ; Middle Aged ; Parvoviridae Infections ; blood ; epidemiology ; immunology ; Seroepidemiologic Studies ; Viral Proteins ; immunology ; Young Adult