14
由盥洗台培養出的棘阿米巴原蟲，絕大部分 (79.2％) 屬 T4 基因型；T3 型及 T6 型分別占 12.5％及 8.3％，此次的調查研究沒有分離出此三型以外之基因型。
由先前學者 (Schuster & Visvesvara，2004) 的研究得知，此三型棘阿米巴皆會造成 棘阿米巴角膜炎。因此推測盥洗台上的汙水塵埃是汙染隱形眼鏡盒及保養液甚至
參考文獻
Auran JD, Starr MB, Jakobiec FA, 1987. Acanthamoeba keratitis. A review of the literature. Cornea 6: 2-26.
Awwad ST, Petroll WM, McCulley JP, Cavanagh HD, 2007. Updates in
Acanthamoeba keratitis. Eye Contact Lens 33: 1-8.
Beattie TK, Tomlinson A, McFadyen AK, Seal DV, Grimason AM, 2003.
Enhanced attachment of Acanthamoeba to extended-wear silicone hydrogel contact lenses: a new risk factor for infection? Ophthalmology 110: 765–771.
Bottone EJ, Madayag RM, Qureshi MN, 1992. Acanthamoeba keratitis: synergy between amebic and bacterial cocontaminants in contact lens care systems as a prelude to infection. Journal of Clinical Microbiology 30: 2447-2450.
Booton GC, Kelly DJ, Chu Y-W, Chu YW, Seal DV, Houang E, Lam DSC, Byers TJ, Fuerst PA, 2002. 18S ribosomal DNA typing and tracking of Acanthamoeba
species isolates from corneal scrape specimens, contact lenses, lens cases, and home water supplies of Acanthamoeba keratitis patients in Hong Kong. Journal of Clinical
Microbiology 40: 1621–1625.
Booton GC, Visvesvara GS, Byers TJ, Kelly DJ, Fuerst PA, 2005. Identification and distribution of Acanthamoeba species genotypes associated with nonkeratitis infections. Journal of Clinical Microbiology 43: 1689-93.
16
Boost M, Cho P, Lai S, Sun WM, 2008. Detection of acanthamoeba in tap water and contact lens cases using polymerase chain reaction. Optometry and Vision Science 85: 526-530.
Bovee EC, 1963. Studies concerning the effects of nutrition on morphology of
amoeba. II. Acanthamoeba castellanii (Douglas) on abundant and starvation quantities.
The American Midland Naturalist 69: 173-181.
Bowers B, Korn ED , 1968. The fine structure of Acanthamoeba castellanii. I. The trophozoite. The Journal of Cell Biology 39: 95–111.
Bowers B, Korn ED, 1974. Localization of lipophosphonoglycan on both sides of
Acanthamoeba plasma membrane. The Journal of Cell Biology 62: 533–540.
Chang JCH, Ossoff SF, Lobe DC, Dorfman MH, Dumais CM, Qualls RG, Johnson JD, 1985. UV inactivation of pathogenic and indicator microorganisms. Applied
and Environmental Microbiology 49: 1361-1365.
Costas M, Griffith AJ, 1980. The suitability of starch-gel electrophoresis of esterases and acid-phosphatases for the study of Acanthamoeba taxonomy. Archiv für
Protistenkunde 123: 272-279.
Costas M, Griffith AJ, 1984. The esterases and acid-phosphatases of
Acanthamoeba (Amoebida, Acanthamoebidae) . Protistologica 20: 27-31.
Costas M, Griffith AJ, 1985. Enzyme composition and the taxonomy of
Aanthamoeba. Journal of Protozoology 32: 604-607.
Culbertson CG, 1971. The pathogenicity of soil amoebas. Annual Review of
Microbiology 25: 231-254.
Culbertson CG, Smith JW, Miner JR , 1958. Acanthamoeba observation on animal pathogenicity. Science 127: 1506.
Culbertson CG, Smith JW, Cohen HK ,Miner JR , 1959. Experimental infection of mice and monkeys by Acanthamoeba. The American Journal of Pathology 35:
185–197.
Cursons RT, Brown TJ, Keys EA, Moriarty KM , Till D , 1980. Immunity to pathogenic free-living amoebae: role of humoral antibody. Infection and Immunity 29: 401–407.
Daggett PM, Sawyer TK, Nevad TA, 1982. Distribution and possible interrelationships of pathogenic and nonpathogenic Acanthamoeba from aquatic environments. Microbial Ecology 8: 371-386.
De Jonckheere JF, 1983. Isoenzyme and total protein analysis by agarose isoelectric focusing and taxonomy of the genus Acanthamoeba. Journal of
Protozoology 30: 701-706.
18
Donzis PB, Mondino BJ, Weissman BA, Bruckner D A, 1987. Microbial contamination of contact lens care systems. American Journal of Ophthalmology 104: 325-333.
Felsenstein J , 1985. Confidence limits on phylogenies: An approach using the bootstrap. Evolution 39: 783-791.
Fong CF, Tseng CH, Hu FR, Wang IJ, Chen WL, Hou YC, 2004. Clinical characteristics of microbial keratitis in a university hospital in Taiwan.
American Journal of Ophthalmology 137: 329-36.
Gast RJ, Ledee DR, Fuerst PA, Byers TJ, 1996. Subgenus systematics of
Acanthamoeba: Four nuclear 18S rDNA sequence types. The Journal of Eukaryotic Microbiology 43: 498-504.
Griffin JL, 1972. Temperature tolerance of pathogenic and non – pathogenic free-living amoebas. Science 277: 1174-1179.
Gunderson JH, Sogin ML, 1986. Length variation in eukaryotic rRNA: small subunit rRNA from the protists Acanthamoeba castellanii and Euglena gracilis. Gene 44: 63-70.
Gutell RR, 1994. Collection of small subunit (16 S and 16 S-like) ribosomal RNA structures. Nucleic Acids Research 22: 3502-3507.
Hirst LW, Green WR, Merz W, Kaufmann C, Visvesvara GS, Jensen A, Howard M, 1984 Management of Acanthamoeba keratitis. A case report and review of the
literature. Ophthalmology 91: 1105-11.
Jakovljevich R, Talis B, 1969 Recovery of a hartmanelloid ameba in the purulent discharge from a human ear. The Journal of Protozoology 16(suppl): 36
John T, Deai D, Sahm D, 1989. Adherence of Acanthamoeba castellanii cysts and trophozoites to unworn soft contact lenses . American Journal of Ophthalmology 108: 658-664.
Johnson AM, Fielke R, Christy PE, Robinson B, Baverstock PR,
1990. Small subunit ribosomal RNA evolution in the genus Acanthamoeba. Journal
of General Microbiology 136: 1689-1698.
Jones DB, Visvesvara GS, Rovinson NM, 1975. Acanthamoeba polyphaga keratitis and Acanthamoeba uveitis associated with fatal meningoencephalitis.
Transactions of the Ophthalmological Societies of the United Kingdom 95: 221-232.
Key SN 3rd, Green WR, Willaert E, Stevens AR, Key SN Jr, 1980 Keratitis due to Acanthamoeba castellanii. A clinicopathologic case report. Archives of
Ophthalmology 98: 475-9.
Khan NA , 2003. Pathogenesis of Acanthamoeba infections. Microbial
Pathogenesis 34: 277–285.
20
Khan NA, 2006. Acanthamoeba : biologyand increasing importance inhuman health.
FEMS Microbiology Reviews 30 : 564–595.
Kimura M , 1980. A simple method for estimating evolutionary rate of base substitutions through comparative studies of nucleotide sequences. Journal of
Molecular Evolution 16: 111-120.
Klink F, Alizadeh H, He Y, Mellon JA, Silvany RE, McCulley JP, Niederkorn JY, 1993. The role of contact lenses, trauma, and Langerhans cells in a Chinese hamster model of Acanthamoeba keratitis. Investigative Ophthalmology & Visual Science 34: 1937-44.
Ko LS, 1984. The problem of myopia in Taiwan. Korean Journal of
Ophthalmology 25: 591-604
Larkin DF, 1991. Acanthamoeba keratitis. International Ophthalmology Clinics 31: 163-172.
Larkin DF, Kilvington S, Easty DL, 1990. Contamination of contact lens storage cases by Acanthamoeba and bacteria. The British Journal of Ophthalmology 7:
133–135.
Lindquist TD, Doughman DJ, Rubenstein JB, Moore JW, Campbell RC, 1988 .
Acanthamoeba-contaminated hydrogel contact lenses. Susceptibility to disinfection.
Cornea 7: 300-303.
Lee SM, Choi YJ, Chung DL, 1997. Contamination of Acanthamoeb in contact lens care system. Korean Journal of Ophthalmol 38:53l.
Lee JJ, Hutner SH, Bovee EC, 1985. An illustrated guide to the protozoa.
Society of Protozoologists, Lawrene, Kansas U.S.A
Maghsood AH, Sissons J, Rezaian M, Nolder D, Warhurst D, Khan NA, 2005.
Acanthamoeba genotype T4 from the UK and Iran and isolation of the T2 genotype
from clinical isolates. Journal of Medical Microbiology 54: 755–759.Marciano-Cabral F, Cabral G , 2003. Acanthamoeba spp. as agents of disease in humans. Clinical Microbiology Reviews 16: 273–307.
Martinez AJ, 1985. Free-Living Amebas: Natural History, Prevention, Diagnosis, Pathology, and Treatment of Disease. CRC Press, Boca Raton, Florida.
Martinez AJ, 1987. Clinical manifestation of free-living amebic infection.In:
Rondanelli E.G. (ed). Amphizoic amoebae : human pathology. Piccin, Padua, Italy.
PP. 161-177.
Martinez AJ, 1991. Infections of the central nervous system due to Acanthamoeba.
Reviews of Infectious Diseases 13: S399–S402.
Martinez AJ, Visvesvara GS, 1991. Laboratory diagnosis of pathogenic free-living amoebas: Naegleria, Acanthamoeba, and Leptomyxid. Clinics in Laboratory Medicine
22
11: 861–872.
Martinez AJ & Visvesvara GS, 1997. Free-living, amphizoic and opportunistic amebas. Brain Pathology (Zurich, Switzerland) 7: 583–598.
Melissa KH , Bret SB , Paul TM , Christopher PS, 2003. Identification of a new
Acanthamoeba 18S r RNA gene sequence type, corresponding to the species
Acanthamoeba jacobsi Sawyer, Nerad and Visvesvara,1992 ( Lobosea :
Acanthamoebidae). Acta Protozoologica 42: 325-329.Mergeryan H, 1991. The prevalence of Acanthamoeba in the human environment.
Reviews of Infectious Diseases 13 Suppl 5: S390-1.
Moore MB, McCulley JP, Newton C, Cobo LM, Foulks GN, O'Day DM, Johns KJ, Driebe WT, Wilson LA, Epstein RJ, 1987 Acanthamoeba keratitis. A growing problem in soft and hard contact lens wearers. Ophthalmology 94: 1654-61.
Neefs JM, Van de Peer YD, De Rijk P, Chapelle S, De Wachter R, 1993.
Compilation of small ribosomal subunit RNA structures . Nucleic Acids Research 21: 3025-3049.
Niederkorn JY, Alizadeh H, Leher H, McCulley JP, 1999. The pathogenesis of
Acanthamoeba keratitis. Microbes and infection 1:437–443.
Page FC, 1967a. Taxonomic criteria for limax amoebae, with descriptions of 3 new species of Hartmannella and 3 of Vahlkampfia. The Journal of Protozoology 14:
499–521.
Page FC, 1967b. Re-definition of the genus Acanthamoeba with descriptions of three species. The Journal of Protozoology 14: 709–724.
Page FC, 1988. A New Key to Fresh Water and Soil Amoebae. Freshwater Biological Association Scientific Publications, Cumbria, UK.
Saitou N , Nei M, 1987. The neighbor-joining method: A new method for reconstructing phylogenetic trees. Molecular Biology and Evolution 4:406-425.
Sawyer T, 1971. Acanthamoeba griffini a new species of marine amoeba. The
Journal of Protozoology 18: 650–654.
Schroeder JM, Booton GC, Hay J, Niszl IA, Seal DV, Markus MB, Fuerst PA , Byers TJ, 2001. Use of subgenic 18S ribosomal DNA PCR and sequencing for genus and genotype identification of Acanthamoebae from humans with keratitis and from sewage sludge. Journal of Clinical Microbiology 39: 1903–1911.
Schuster FL, Visvesvara GS, 2004. Free-living amoebae as opportunistic and nonopportunistic pathogens of humans and animals. International Journal for
Parasitology 34: 1001–1027.
24
Seal D, Stapleton F, Dart J, 1992. Possible environmental sources of
Acanthamoeba spp in contact lens wearers. The British Journal of Ophthalmology 76:
424-427.
Singh BN , Das SR, 1970. Studies on pathogenic and nonpathogenic small
free-living amoebae and the bearing of nuclear division on the classification of the order Amoebida. Philosophical Transactions of the Royal Society of London.Series A:
Mathematical and Physical Sciences 259: 435–476.
Stehr-Green JK, Baily TM, Visvesvara GS, 1989. The epidemiology of
Acanthamoeba keratitis in the United States. American Journal of Ophthalmology
107: 331–336.Stothard DR, Schroeder-Diedrich JM, Awwad MH, Gast RJ, Ledee DR,
Rodriguez-Zaragoza S, Dean CL, Fuerst PA, Byers TJ, 1998. The evolutionary history of the genus Acanthamoeba and the identification of eight new 18S rRNA gene sequence types. The Journal of Eukaryotic Microbiology 45: 45-54.
Tamura K, Dudley J, Nei M, Kumar S , 2007. MEGA4: Molecular Evolutionary Genetics Analysis (MEGA) software version 4.0. Molecular Biology and Evolution 10.1093/molbev/msm092.
Torno MS Jr, Babapour R, Gurevitch A, Witt MD, 2000. Cutaneous acanthamoebiasis in AIDS. Journal of the American Academy of Dermatology 42:351–354.
Visvesvara GS , 1991. Classification of Acanthamoeba. Reviews of Infectious
Diseases 13: 369–372.
Visvesvara GS, Balamuth W, 1975. Comparative studies on related free-living and pathogenic amoebae with special reference to Acanthamoeba. The Journal of
Protozoology 22: 245-256.
Visvesvara GS, Martinez AJ, Schuster FL, Leitch GJ, Wallace SV, Sawyer TK Anderson M, 1990. Leptomyxid ameba, a new agent of amebic
meningoencephalitis in humans and animals. Journal of Clinical Microbiology 28:
2750–2756.
Zhang Y, Sun X, Wang Z, Li R, Luo S, Jin X, Deng S, Chen W, 2004.
Identification of 18S Ribosomal DNA Genotype of Acanthamoeba from Patients with Keratitis in North China Invest. Investigative ophthalmology & visual science 45:
1904-1907.
李清真, 1994. 大台北地區公共場所盥洗台伺機性阿米巴—棘阿米巴類之調查 研究。 國立台灣大學醫學院微生物研究所寄生蟲學組碩士論文。
丘子宏、曾錦華、包麗菊、黃宏仁, 1987. 配戴隱形眼鏡之問題分析.中華民國眼 科醫學會會刊。 26：88-91.
26
林思源、周文穎、林隆光、潘玟利、施永豐, 2001. 台灣學生隱形眼鏡配戴之狀 態－十年之變遷。 中華民國眼科醫學會雜誌 43:320-325.
林耕國、侯鈞賀、楊孟玲、李建興, 2005. 隱形眼鏡。 中華民國眼科醫學會雜 誌 44:272-276.
黃國銅, 1994. 針對台北地區土壤及淡水內自由生活性阿米巴之研究。 國立 台灣大學醫學院微生物研究所寄生蟲學組碩士論文。
Acanthamoeba trophozoites form
Acanthamoeba cysts form
圖一. 棘阿米巴之生活史
棘阿米巴生活史中包含兩種形態；活動之營養體 (active trophozoite) 及 呈休眠狀態的囊體，當遇到不適當的環境 (例如食物被吃完時) 營養體便會囊化 (encyst) 而形成雙層含纖維素囊壁的囊體 。
Unf av our abl e cond it ions
Fa vo u ra b le cond it ions
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圖二. 棘阿米巴 Rns 中之十二個高度可變區 (expansion segment) 之分布圖。
此圖為 Gunderson， Sogin 所繪製 Acanthamoeba castellanii 之 18S r RNA 二 級結構圖。圈圈區域代表高度可變區 (Stothard et al., 1998)。
A.
A. ASA.S1 (Acanthamoeba-specfic amplimer S1) region，其大小約為 423~551 bp，位於 18S rRNA 之 936 bp 到 1402 bp 間 (Schroeder et al., 2001)。 B. ASA.S1 region 為 18S r RNA 二級結構中的 stems E23-2 到-30，十二個高度可變區 (expansion segment) 中就佔了三個
(
Gast et al., 1996)。30
圖四
.
棘阿米巴之營養體在 100 倍放大倍率下所觀察到的形態。檢體接種後第一天，可見到檢體中的棘阿米巴分裂增殖，游離的阿米巴向四 周生長。
圖五. 棘阿米巴之囊體在 100 倍放大倍率下所觀察到的形態。
培養後第三天,可見到部分棘阿米巴已變成圓形，進而形成囊體。
32
.圖六. 油鏡下所觀察到棘阿米巴營養體之形態 (相位差顯微鏡)。
將棘阿米巴營養體從培養瓊脂轉置於滴有 Page's ameba saline 的玻片上，待 數分鐘後開始作緩慢的變形運動，此時可見到尖銳透明的棘狀偽足 (N 代表核，
CV 代表伸縮泡) 。
N
CV
圖七. 油鏡下所觀察到棘阿米巴囊體之形態 (光學顯微鏡)。
將棘阿米巴囊體從培養瓊脂轉置於滴有 Page's ameba saline 的玻片上，此時 可見到其雙層囊壁。
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100 bp DNA ATCC Ladder 18 43 80 85 90 100 106 112 2390 2412 30234
圖八. 培養分離的棘阿米巴蟲株之 DNA，以 JDP1/2 作為引子所得 Acanthamoeba-specfic amplimer S1 (ASA.S1) Region 洋菜醣膠電泳圖。在 500 bp 出現 band 者即視為陽性。
500 bp
圖九..Neighbor-joining distance tree based on partial 18S rDNA sequences
二十五株棘阿米巴中，有兩株蟲體其核酸序列與 T6 之核酸序列相似；有三株蟲體 其核酸序列與 T3 之核酸序列相似；其餘二十株蟲體其核酸序列與 T4 之核酸序列 相似。
36
圖十. Parsimony tree based on partial 18SrDNA sequences
二十五株蟲體中，有兩株蟲體其核酸序列與 T6 之核酸序列相似；有三株蟲體其核 酸序列與 T3 之核酸序列相似；其餘二十株蟲體其核酸序列與 T4 之核酸序列相似。
38
T4/7 GG--T-GCGGTCGTCCTTGGCGTC--GG----TTTC---GGCCGG--CGCGGGGGTGGCTTAGCCCG-G 50 GG--T-GCGGTCGTCCTTGGCGTCGGT---TC---GGC--GGCGCGGGGGCGGCTTAGCCCG-G 62 GG--T-GCGGTCGTCCTTGGCGTCGGT----CTTTC---GGGGC-CGGCGCGGGGACGGCTTAGCCCG-G 2720 GG--T-GCGGTCGTCCTTGGCGTCGGT---TCC---TGC-GGGCCCGGGGGCGGGTTACCCCG-G
T4/1 GG--T-GCGGTCGTCCTTGGCGTCTCGG----TCCTTCACGGGGCCGGGGCGCGGGGGCGGCTTAGCCCG-G T4/8 GG--T-GCGGTCGTCCTTGGCGTCTCGG----TCCTTCACGGGGCCGGGGCGCGGGGGTGGCTTAGCCCG-G T4/9 GG--T-GCGGTCGTCCTTGGCGTCTCGG----TTTC---GGCCGGGGTGCGGGGACGGCTTAGCCCG-G T4/5 GG--T-GCGGTCGTCCTTGGCGGTTGGT----CTTCG----AAAGCCAGCGGCGGGGGCGGCTTAGCCCG-G T4/3 GG--T-GCGGTCGTCCTTGGCGGTTGGT----CTTCA----AAAGCCAGCGGCGGGGGCGGCTTAGCCCG-G 22 GG--T-GCGGTCGTCCTTGGCGGTTGGT----CTTCA----AAAGCCAGCGGCGGGGGTGGCTTAGCCCG-G 92 GG--T-GCGGTCGTCCTTGGCGGTTGGT----CTTCA----AAAGCCAGCGGCGGGGGTGGCTTAGCCCG-G 97 GG--T-GCGGTCGTCCTTGGCG-TTGGT----CTTCA----AAAGCCAGCGGCGGGGGTGGCTTAGCCCG-G 151 GG--T-GCGGTCGTCCTTGGCG-TTGGT----CTTCA----AAAGCCAGCG-CGGGGGTGGCTTAGCCCG-G 2628 GG--T-GCGGTCGTCCTTGGCG-TTGGT----CTTCA----AAAGCCAGCG-CGGGGGTGGCTTAGCCCG-G T4/2 GG--T-GCGGTCATCCTTGGCG-TTGGT----CTTCA----AAAGCCAGCG-CGGGGGTGGCTTAGCCCG-G T4/4 GG--T-GCGGCTGTTCTTGGCGTC--GG----TTTC---GGCCGG--CGCGGGGATGGCTTAGCCCG-G 67 GG--C-GCGGTCGTCCTTGGCGTCTGT----CTTTC---GGGGG-CGGCGCGGGGGCGGCTTAGCCCG-G 39 GG--C-GCGGTCGTCCTTGGCGTCTGT----CTTTC---GGGGC-CGGCGCGGGGGCGGCTTAGCCCG-G 100 GG--C-GCGGTCGTCCTTGGCGTCTGT----CTTTC---GGGGC-CGGCGCGGGGGCGGCTTAGCCCG-G 2390 GG--C-GCGGTCGTCCTTGGCGTCGGT----CTTTC---GGGGC-CGGCGCGGGGGCGGCTTAGCCCG-G 2412 GG--C-GCGGTCGTCCTTGGCGTCGGT----CTTTC---GGGGC-CGGCGCGGGGGCGGCTTAGCCCG-G ATCC30234 GG--C-GCGGTCGTCCTTGGCGTCGGT----CCTTCAC---GGGGC-CGGCGCGAGGGCGGCTTAGCCCG-G 71 GG--C-GCGGTCGTCCTTGGCGTCTGTGTCCCTTTCAAC-GGGGGCATGGCGCGAGGGCGGTTTAGCCCG-G 112 GG--C-GCGGTCGTCCTTGGCGTCTGTGTCCCTTTCAAC-GGGGGCATGGCGCGAGGGCGGTTTAGCCCG-G
T4/10 GG--TTGCGGTCGTCCTTGGCGTCTCGG----TTTC---GGCCGGGGCGCGGGGACGGTTTAGCCCG-G T4/6 GG--TTGCGGTCGTCCTTGGCGTCTCGG----TTTC---GGCCGGGGCGCGGGGATGGCTTAGCCCG-G 114 GG--TTGCGGTCGTCCTTGGCGTCTCGG----TTTC---GGCCGGGGCGCGGGGATGGCTTAGCCCG-G 155 GG--TTGCGGTCGTCCTTGGCGTCTCGG----TTTC---GGCCGGGGCGCGGGGATGGCTTAGCCCG-G 2782 GG--TTGCGGTCGTCCTTGGCGTCTCGG----TTTC---GGCCGGGGCGCGGGGATGGCTTAGCCCG-G 3265 GG--TTGCGGTCGTCCTTGGCGTCTCGG----TTTC---GGCCGGGGCGCGGGGATGGCTTAGCCCG-G T3/5 CG-ATTGCGGTCGTCTTTGGTG--TCGT----CCACAGCGATGTGGGCGGCATCGGGATGGCTTAGCTCGCA T3/1 CG-ATTGCGGTCGTCTTTGGTG--TCGC----TCACAAGGC----GGCATC---GGGACGGCTTAGCTCGCA T3/2 CG-ATTGCGGTCGTCTTTGGTG--TCGC----TCACAAGGC----GGCATC---GGGGCGGCTTAGCTCGCA T3/3 CGCATTGCGGTCGTCTTTGGTG--TCGC----TCACAAGGC----GGCACC---GGGGCGGCTTAGCTCGCA T3/4 CGCATTGCGGTCGTCTTTGGTGTGTCGC----TCACAAGGC----GGCATCATCGGGACGGCTTAGCTCGCA 26 CGCATTGCGGTCGTCTTTGGTGTGTCGC----TCACAAGGC----GGCATCATCGGGACGGCTTAGCTCGCA 118 CGCATTGCGGTCGTCTTTGGTGTGTCGC----TCACAAGGC----GGCATCATCGGGACGGCTTAGCTCGCA 2835 CGCATTGCGGTCGTCTTTGGTGTGTCGC----TCACAAGGC----GGCATCATCGGGACGGCTTAGCTCGCA
圖十一. 利用 Clustal W 將所得之 T3 、T4 型棘阿米巴之 ASA.S1 (Acanthamoeba-specfic amplimer S1) region 核酸序列與已知的 DF3
(diagnostic fragment 3) 核酸序列比對。分離出來之二十株 T4 蟲體及三株 T3 蟲體，有四株 T4 蟲體其核酸序列屬於 T4/6 此亞型；有三
株 T3 蟲體其核酸序列屬於 T3/4 此亞型。
表一. 棘阿米巴之基因型與引起人類疾病，如角膜炎及肉芽腫性腦膜炎之相關聯性 (Khan, 2006)。
NA，目前尚未發現其會造成人類的感染。
+，T2 已被證實又可以再細分為 T2a 及 T2b ( Maghsood et al , 2005)。
*，其會造成人類角膜炎及肉芽腫性腦膜炎的感染。
表二. 本研究參考之 Rns 核酸序列及 GeneBank accession numbers (Stothard et al., 1998)。
Sequence type Species classification GenBank accession nuber
T1 Acanthamoeba spp U07400
表三. 本研究參考之DF3核酸序列及Gene Bank accession numbers (Booton et al., 2002)。
Sequence type Gene Bank accession number
T3/1 AF441800 T3/2 AF441814 T3/3 AF441813 T3/4 AF441806 T3/5 AF441805 T4/1 AF441811 T4/2 AF441807 T4/3 AF441794 T4/4 AF441796 T4/5 AF441795 T4/6 AF441801 T4/7 AF441802 T4/8 AF441799 T4/9 AF441803 T4/10 AF441798
表四. 本研究分離所得之棘米阿米巴蟲株之核酸序列比對結果
附錄一.
長戴型隱形眼鏡採檢說明書 Hello!大家好~~
我是台大研究所的學生，因為要研究「隱形眼鏡配帶者其清潔保養液及盥洗 台之棘阿米巴調查研究」，所以需收集以下的檢體。
標示 拋 棄 液 的試管是用來裝已使用過的隱形眼鏡保存液。
標示 生 理 食 鹽 水 的試管是用來裝已開封的生理食鹽水
，量約到 2 的刻度。
標示 隱形眼鏡保存盒擦拭物 的試管請先取試管內的棉棒，用 此棉棒沾取保存盒內部，再放回試管，再將蓋子旋緊即可。
標示 盥洗台擦拭物 的試管請先取試管內的棉棒，用此棉棒沾取水龍頭及其周 圍的水漬、灰塵後再放回試管，再將蓋子旋緊即可。
如果您想知道您的結果，請留下你的 e-mail ，我再將結果 e-mail 給您。
e-mail address:
p.s.有任何收集問題歡迎來信詢問!我的的 e-mail: r94445205@ntu.edu.tw 最後誠摯的謝謝您的幫忙! <此實驗的結果不提供作商業用途!>
附錄二.
拋棄式隱形眼鏡採檢說明書 Hello!大家好~~
我是台大研究所的學生，因為要研究”隱形眼鏡配帶者其清潔保養液及盥洗 台之棘阿米巴調查研究” ，所以需收集以下的檢體。
標示 拋棄式隱形眼鏡 的試管是用來裝已使用過準備丟棄的隱形眼鏡 。 標示 盥洗台擦拭物 的試管請先取試管內的棉棒，用此棉棒沾取水龍頭及其 周圍的水漬,灰塵後再放回試管,再將蓋子旋緊即可。
如果您想知道您的結果，請留下你的 e-mail ，我再將結果 e-mail 給您。
e-mail address:
p.s.有任何收集問題歡迎來信詢問!我的的 e-mail: r94445205@ntu.edu.tw 最後誠摯的謝謝您的幫忙! <此實驗的結果不提供作商業用途!>