臺灣博碩士論文加值系統

人類間質肺炎病毒(hMPV)是最近發現與呼吸道感有關的重要新興致病原，其主要感染對象為幼童及老年人。我們建立以N重組蛋白為抗原的酵素連結免疫吸收技術(ELISA)系統偵測血清中hMPV的IgG抗體，以研究在台灣地區人類間質性肺炎病毒的流行病學。共偵測台灣地區1120例健康人類血清，包括男性及女性分別為516、604例。結果顯示在台灣地區人類間質性肺炎病毒在每個年齡層的流行率為5.3％(5/94) (0-0.6歲)，5.3％(5/94) (0.6-1歲)，39％(39/100) (1-2歲)，43％(43/100) (2-3歲)，45.1％(43/51) (3-4歲)，53.1％(26/49) (4-5歲)，62％(62/100) (5-6歲)，71.1％(69/97) (6-10歲)，86％(86/100) (10-20歲)，84％(110/131) (20-30歲)，90.6％(96/106) (30-50歲)，91.8％(90/98) (>50歲)。人類間質性肺炎病毒已經在台灣地區流行且第一次感染大都發生在1-2歲，且hMPV的流行率在男性與女性之間沒有差異。我們也偵測13例感染hMPV病患的成對血清，發現急性期及恢復期的血清中確實出現hMPV抗體有77％由陰性變成陽性。由本研究顯示本實驗所建立的N重組蛋白為抗原的酵素連結免疫吸收技術是一種研究人類間質性肺炎病毒流行病學有利的工具，並可應用於臨床診斷。

Human metapneumovirus (hMPV) is a newly identified virus. It causes severe respiratory disease in children and elderly people. To study the seroprevalence of hMPV in Taiwan, we develops the indirect immuno-fluorescence assays based on nucleocapsid (N) recombinant protein for detection of hMPV IgG antibody in human sera. A total of 1120 human sera from health people, including 516 male and 604 female, were examined by ELISA system based on N recombinant protein. The data show that the positive rate of each age group in Taiwan was 5.3％(5/94) (0-0.6 years), 5.3％(5/94) (0.6-1 years), 39％(39/100) (1-2 years), 43％(43/100) (2-3 years), 45.1％(43/51) (3-4 years), 53.1％(26/49) (4-5 years), 62％(62/100) (5-6 years), 71.1％(69/97) (6-10 years), 86％(86/100) (10-20 years), 84％(110/131) (20-30 years), 90.6％(96/106) (30-50 years), 91.8％(90/98) (>50 years). Human metapneumovirus is circulating in Taiwan and the first infection usually occurs in the age group of 1-2 years. The positive rate of hMPV in male is the same with the female. We also detected 13 paired sera from children with hMPV infected, the data indicated that the 10 of 13 (77％) human had a seroconversion. The ELISA based on N recombinant protein is a useful tool for seroepidemiological study of hMPV and can apply to clinical
diagnosis.

指導教授推薦書…………………………………………………………i
口試委員會審定書………………………………………………………ii
誌謝……………………………………………………………………...iii
中文摘要………………………………………………………………...iv
英文摘要………………………………………………………………...v
目錄……………………………………………………………………...vi
圖表目錄………………………………………………………………..vii
第一章 緒論……………………………………………………………1
第一節 hMPV病毒學特性………………………………………..1
第二節 hMPV基因體組成………………………………………..2
第三節 hMPV流行病學…………………………………………..5
第四節 hMPV臨床症狀…………………………………………..5
第五節 偵測血清中hMPV專一性抗體的方法學……...………..7
第二章 研究目的……………………………………………………....8
第三章 實驗設計………………………………………………………9
第四章 實驗方法及材料..……………………………………………11
第五章 實驗結果…………..…………………………………………20
第六章 討論…………………………………………………………..26
參考文獻………………………………………………………………..29
附圖……………………………………………………………………..32
附表……………………………………………………………………..41

1.Hamelin, M.E., Y. Abed, and G. Boivin, Human metapneumovirus: a new player among respiratory viruses. Clin Infect Dis, 2004. 38(7): p. 983-90.
2.VAN DEN HOOGEN, et al., A newly discovered human pneumovirus isolated from youngchildren with respiratory tract disease. NATURE MEDICINE 2001. 7( 6): p. 719-724.
3.van den Hoogen, B.G., et al., Analysis of the genomic sequence of a human metapneumovirus. Virology, 2002. 295(1): p. 119-32.
4.Esper, F., et al., A 1-year experience with human metapneumovirus in children aged <5 years. J Infect Dis, 2004. 189(8): p. 1388-96.
5.Esper, F., et al., Human metapneumovirus infection in the United States: clinical manifestations associated with a newly emerging respiratory infection in children. Pediatrics, 2003. 111(6 Pt 1): p. 1407-10.
6.Peret, T.C., et al., Characterization of human metapneumoviruses isolated from patients in North America. J Infect Dis, 2002. 185(11): p. 1660-3.
7.Boivin, G., et al., Virological features and clinical manifestations associated with human metapneumovirus: a new paramyxovirus responsible for acute respiratory-tract infections in all age groups. J Infect Dis, 2002. 186(9): p. 1330-4.
8.Nissen, M.D., et al., Evidence of human metapneumovirus in Australian children. Med J Aust, 2002. 176(4): p. 188.
9.Peiris, J.S., et al., Children with respiratory disease associated with metapneumovirus in Hong Kong. Emerg Infect Dis, 2003. 9(6): p. 628-33.
10.Ebihara, T., et al., Seroprevalence of human metapneumovirus in Japan. J Med Virol, 2003. 70(2): p. 281-3.
11.Jartti, T., et al., Metapneumovirus and acute wheezing in children. Lancet, 2002. 360(9343): p. 1393-4.
12.Stockton, J., et al., Human metapneumovirus as a cause of community-acquired respiratory illness. Emerg Infect Dis, 2002. 8(9): p. 897-901.
13.Vicente, D., et al., Human metapneumovirus and community-acquired respiratory illness in children. Emerg Infect Dis, 2003. 9(5): p. 602-3.
14.Freymouth, F., et al., Presence of the new human metapneumovirus in French children with bronchiolitis. Pediatr Infect Dis J, 2003. 22(1): p. 92-4.
15.Mejias, A., S. Chavez-Bueno, and O. Ramilo, Human metapneumovirus: a not so new virus. Pediatr Infect Dis J, 2004. 23(1): p. 1-7; quiz 8-10.
16.Biacchesi, S., et al., Genetic diversity between human metapneumovirus subgroups. Virology, 2003. 315(1): p. 1-9.
17.Bastien, N., et al., Sequence analysis of the N, P, M and F genes of Canadian human metapneumovirus strains. Virus Res, 2003. 93(1): p. 51-62.
18.Boivin, G., et al., Global genetic diversity of human metapneumovirus fusion gene. Emerg Infect Dis, 2004. 10(6): p. 1154-7.
19.Ishiguro, N., et al., High genetic diversity of the attachment (G) protein of human metapneumovirus. J Clin Microbiol, 2004. 42(8): p. 3406-14.
20.Andrew, J., J.B. Easton, and D.a.H.F. Rosenberg, Animal Pneumoviruses: Molecular Genetics and Pathogenesis. CLINICAL MICROBIOLOGY REVIEWS, 2004: p. p. 390–412.
21.Skiadopoulos, M.H., et al., The two major human metapneumovirus genetic lineages are highly related antigenically, and the fusion (F) protein is a major contributor to this antigenic relatedness. J Virol, 2004. 78(13): p. 6927-37.
22.Tang, R.S., et al., Effects of human metapneumovirus and respiratory syncytial virus antigen insertion in two 3' proximal genome positions of bovine/human parainfluenza virus type 3 on virus replication and immunogenicity. J Virol, 2003. 77(20): p. 10819-28.
23.van den Hoogen, B.G., D.M. Osterhaus, and R.A. Fouchier, Clinical impact and diagnosis of human metapneumovirus infection. Pediatr Infect Dis J, 2004. 23(1 Suppl): p. S25-32.
24.Maggi, F., et al., Human metapneumovirus associated with respiratory tract infections in a 3-year study of nasal swabs from infants in Italy. J Clin Microbiol, 2003. 41(7): p. 2987-91.
25.Boivin, G., et al., Human metapneumovirus infections in hospitalized children. Emerg Infect Dis, 2003. 9(6): p. 634-40.
26.Wolf, D.G., et al., High seroprevalence of human metapneumovirus among young children in Israel. J Infect Dis, 2003. 188(12): p. 1865-7.
27.Leung, J., et al., Seroepidemiology of human metapneumovirus (hMPV) on the basis of a novel enzyme-linked immunosorbent assay utilizing hMPV fusion protein expressed in recombinant vesicular stomatitis virus. J Clin Microbiol, 2005. 43(3): p. 1213-9.
28.Zhao, L.Q., et al., [Seroprevalence of antibody against human metapneumovirus in Beijing]. Zhonghua Er Ke Za Zhi, 2005. 43(12): p. 904-7.
29.Liu, L., et al., Seroprevalence of human metapneumovirus (hMPV) in the Canadian province of Saskatchewan analyzed by a recombinant nucleocapsid protein-based enzyme-linked immunosorbent assay. J Med Virol, 2007. 79(3): p. 308-13.
30.Hamelin, M.E. and G. Boivin, Development and validation of an enzyme-linked immunosorbent assay for human metapneumovirus serology based on a recombinant viral protein. Clin Diagn Lab Immunol, 2005. 12(2): p. 249-53.
31.Ishiguro, N., et al., Immunofluorescence assay for detection of human metapneumovirus-specific antibodies by use of baculovirus-expressed fusion protein. Clin Diagn Lab Immunol, 2005. 12(1): p. 202-5.
32.Gulati BR, C.K., Seal BS, Goyal SM, Halvorson DA, Njenga MK., Development of a highly sensitive and specific enzyme-linked immunosorbent assay based on recombinant matrix protein for detection of avian pneumovirus antibodies. J Clin Microbiol., 2000 Nov. 38(11): p. 4010-4.
33.Gulati BR, M.S., Patnayak DP, Goyal SM, Kapur V., Detection of antibodies to U.S. isolates of avian pneumovirus by a recombinant nucleocapsid protein-based sandwich enzyme-linked immunosorbent assay. J Clin Microbiol. , 2001 Aug. 39(8): p. 2967-70.
34.Zhang SP, Z.G., Goldman E., Low-usage codons in Escherichia coli, yeast, fruit fly and primates. Gene, 1991 Aug 105(1): p. 61-72.
35.BJ Del Tito, J., JM Ward, J Hodgson, CJ Gershater, H Edwards, LA Wysocki, FA Watson, G Sathe, and JF Kane Effects of a minor isoleucyl tRNA on heterologous protein translation in Escherichia coli. J. Bacteriol., 1995. 177: p. 7086-7091.
36.Kurland, C.a.G., J. , , Errors of heterologous protein expression. . Curr. Opin. Biotechnol., 1996. 7,: p. 489-493.