| Acute respiratory tract infection (ARI) is a leading cause of morbidity and mortality worldwide. With the development of molecular technologies, new viruses have been identitified recently. Especially with the emperging of SARS-CoV, and the swine H1N1, it reveals the importance of the study on new emergeing infectious diseases.Human metapneumovirus is an emerging human respiratory pathogen first discovered in 2001. It is closely related to respiratory syncytial virus (RSV) and included in the Pneumovirinae subfamily of the Paramyxoviridae family. It can cause both upper and lower tract disease, and has been associated with serious illness in the young, among the immunosuppressed, and in the chronically ill. Human metapneumovirus has now been detected worldwide.So far, the characterization of whole genome, and the seroprevalence of HMPV in China remain unclear. In addition, the need for development of rapid test technologies for diagnosis for HMPV make it essential to obtain the linear epitopes knowleges about the HMPV Fusion protein, which is relatively conserved in nucleotide sequence and amino acid sequence and dominant immunogen.To obtain the information above, this study was to conducted on the following areas,A the complete genome characherization of HMPV of Beijing isolatesB seroprevalence of HMPV in Beijing Area.C Linear epitopes for HMPV F proteinIn this study, it was shown that1524 specimens collected in 2 years (from march,2007 to April,2009) were detected by RT-PCR.43 (2.8%) out of 1524 RT-PCR were positive for hMPV, then the genome of metapneumovirus were determined by random PCR and sequenced. Phylogenetic analysis of hMPV isolates showed that the hMPV isolates belonged to hMPV genetic cluster A2. High divergency between G proteins was found, while high identity between F protein sequence was shown.Besides, HMPV N protein was expressed and purified. To define the seroepidemiology of HMPV, an enzyme-linked immunosorbent assay (ELISA) based on expression of the N protein of hMPV (hMPV N) was developed and 1157 serum specimens were tested. The percentages of hMPV seropositive individuals were 66.67%in children<6 months old,58.6% in children 6 to 11 months old,73.4%in children 12 to 23 months old.100%in adult. In contrast, The percentages of RSV seropositive individuals were 71.40%in children<6 months old,84.48%in children 6 to 11 months old,85.8%in children 12 to 23 months old.100%in adult..Finally, we Immunized the BALB/C mice with the recombinant Ad viruses expressing HMPV F/N and the RSV F/N in 293 cell line, we got the antibody specific to HMPV F,N and HRSV F/N respectively. Meawhile, in use of recombinant baculovirus system expressing the HMPV and HRSV F proteins, the crossreaction study between the Fhttp://www.pipi.cn/#tps and N protein of HMPV F/N was undertaken. It showed that it is truely that there are crossreaction between HMPV and HRSV F proteins or HMPV and HRS N proteins.In addition, we showed that the Fusion protein of HMPV is highly immunogen and contains nine linear immunodominant sites as determined by Pepscan analysis with a set of synthetic peptides overlapping almost the F protein sequenc against positive sera from Human.Antisera from immunized with recombinant Adnovirus expressing HRSV F was also tested by Pepscan analysis. It is found that there are some linear peptides overlapping with the HMPV F positive humen sera.To sum up, the study established the foundation for strategy of disease prevention, the development of diagnotic technologies and seroepidemiology for HMPV. |
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