臺灣博碩士論文加值系統

三種微孢子蟲分離株分別採集自不同地區之兩不同鱗翅目寄主：緣點紋白蝶(2株：AS-1和AS-2) 及甜菜夜蛾 (1株：SE)。為釐清分離株的分類地位，先進行rDNA基因之定序，紋白蝶微孢子蟲分離株 (AS-1) 之rDNA基因從 5’ 端始依序為SSU rDNA (16S) – ITS – LSU rDNA (23S)。另一紋白蝶微孢子蟲分離株 (AS-2) 及甜菜夜蛾微孢子蟲分離株 (SE) 則為LSU rDNA – ITS – SSU rDNA – IGS – 5S rDNA。根據rDNA序列之親緣關係分析的結果，將紋白蝶之二分離株分別命名為 Vairimorpha sp. AS-1 及 Nosema sp. AS-2；而來自甜菜夜蛾之分離株命名為Nosema sp. SE。 Nosema 屬甜菜夜蛾及紋白蝶微孢子蟲分離株 (Nosema sp. SE和Nosema sp. AS-2) 之整段 rDNA 基因已完成定序， Vairimorpha 屬紋白蝶微孢子蟲分離株 (Vairimorpha sp. AS-1) 之SSU rDNA，ITS 及部分 LSU rDNA 片段亦已完成定序工作。三株分離株的成熟孢子皆呈卵圓形，孢子周圍具折光性，孢子大小以ANOVA分析呈顯著差異。Vairimorpha sp. AS-1之成熟孢子具有單核及雙核兩種形式，而Nosema sp. AS-2及Nosema sp. SE僅具有雙核的孢子，印證了分子標誌之分析結果。根據親緣關係分析及形態觀察，推測 Vairimorpha sp. AS-1應為一新的種類， Nosema sp. SE應屬於含有 Nosema模式種之真微粒子蟲 (true Nosema) 種類，而Nosema sp. AS-2應屬於另一群Nosema 屬種類。

Three microsporidian isolates were collected from two different hosts, Artogeia canidia sordida (2 isolates, AS-1 and AS-2, from different locations in northern Taiwan) and Spodoptera exigua (1 isolate, SE, from middle Taiwan). For clarification of their taxonomic position, the rDNA genes of these isolates were sequenced. The rDNA gene arrangement of AS-1 was SSU rDNA (16S) – ITS – LSU rDNA (23S) from 5’ end while AS-2 and SE was LSU rDNA – ITS – SSU rDNA – IGS – 5S rDNA, which is similar to that of the true Nosema species. According to the phylogenetic analyses based on the gene sequences of rDNA, AS-1, AS-2 and SE was named as Vairimorpha sp. AS-1, Nosema sp. AS-2, and Nosema sp. SE, respectively. The rDNAs of the Nosema isolates were fully sequenced, and most rDNA sequences, including SSU rDNA, ITS and partial LSU rDNA, of the Vairimorpha isolates were carried out. The mature spores were oval and had great refringency. The sizes of the spores were significant difference by ANOVA analysis. The AS-1 contained mononuclear and binuclear spores, wherease only binuclear spores were found in AS-2 and SE isolates. This result confirmed the results of molecular makers. According to the results of the morphology observations and phylogenetic analyses, Vairimorpha sp. AS-1 should be a new species, SE should be grouped in the true Nosema complex while AS-2 belonged to other group of the Nosema species.

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朱耀沂、金小安。1973。臺灣兩種紋白蝶分佈變遷之推測。植物保護學會會刊 15: 93-96。
小山 光男、釜野 靜也。1976。ハズモンヨトウの大量飼養法。 植物防疫30：470-474。
安永 智佐、舟越 正子、河原畑 勇、荒武 義信、岩野 秀俊。1992。シロイチモジヨトウ幼虫から分離された微胞子虫Nosema bombycis (Microsporida: Nosematidae)。應動昆36 : 127-134。
阿部 芳彥、河原畑 勇。1988。モンシロチヨウから分離された微胞子虫類の性狀。日蠶雜59: 288-292。
徐泰浩。1984。斜紋夜盜微孢子蟲病之研究。台灣大學植物病蟲害學系碩士論文。77頁。
Al Fazairy, A. A. 1986. Natural microsporidian infection in laboratory colonies of Spodoptera spp. Insect Sci. Appl. 7 : 701-705.
Asmuth, D. M., P. C. DeGirolami, M. Federman, C. R. Ezratty, D. K. Pleskow, G. Desai, and C. A. Wanke. 1994. Clinical features of microsporidiosis in patients with AIDS. Clin. Infect. Dis. 18: 819-825.
Baker, M. D., C. R. Vossbrinck, J. V. Maddox, and A. H. Undden. 1994. Phylogenetic relationships among Vairimorpha and Nosema species (Microspora) based on ribosomal RNA sequence data. J. Invertebr. Pathol. 64: 100-106.
Bryan, R. T. 1995. Microsporidiosis as an AIDS-related opportunistic infection. Clin. Infect. Dis. 24: S62-S65.
Canning, E. U., J. Lom, and I. Dykova. 1986. The Microsporidia of Vertebrates. Academic Press, London. 289 pp.
Canning, E. U., A. Curry, S. Cheney, N. J. Lafranchi-Tristem, and M. A. Haque. 1999a. Vairimorpha imperfecta n. sp., a microsporidian exhibiting an abortive octosporous sporogony in Plutella xylostella L. (Lepidoptera: Yponomeutidae). Parasitology 119: 273-286.
Caramello, P., G. Mazzucco, M. Romeo, A. Ullio, G. De Rosa, A. Lucchini, B. Forno, T. Brancale, A. Macor, C. Preziosi, and P. Gioannini. 1995. Clinical and microscopical features of small-intestinal microsporidiosis in patients with AIDS. Infection 23: 362-368.
Curry, A., and E. U. Canning. 1993. Human microsporidiosis. J. Infect. 27: 229-236.
Drobniewski, F., P. Kelly, A. Carew, B. Ngwenya, N. Luo, C. Pankhurst, and M. Farthing. 1995. Human microsporidiosis in African AIDS patients with chronic diarrhea. J. Infect. Dis. 171: 515-516.
Eeftinck Schattenkerk, J. K. M. T., and T. Van Gool. 1992. Clinical and microbiological aspects of microsporidiosis. Trop. Geogr. Med. 44: 287.
Gatehouse, H. S., and L. A. Malone. 1998. The ribosomal RNA gene region of Nosema apis (Microspora) : DNA sequence for small subunit rRNA genes and evidence of a large tandem repeat unit size. J. Invertebr. Pathol. 71: 92-105.
Huang, W. F., S. J Tsai, C. F. Lo, Y. Soichi, and C. H. Wang. 2004. The novel organization and complete sequence of the ribosomal RNA gene of Nosema bombycis. Fungal Gene. Bio. 41: 473-481.
Huang, W. F., M. Bocquet, K. C. Lee, I. H. Sing, and J. H. Jiang. 2007. The comparison of rDNA spacer regions of Nosema ceranae isolates from different hosts and locations. J. Invertebr. Pathol. 95: 1-5.
Huelsenbeck, J. P., and F. Ronquist. 2001. MRBAYES: Bayesian inference of phylogenetic trees. Bioinformatics 17: 754-755.
Ironside, J. E. 2007. Multiple losses of sex within a single genus of Microsporidia. BMC Evo. Bio. 7:48
Keeling, P. J., and N. M. Fast. 2002. Microsporidia: biology and evolution of highly reduced intracellular parasites. Annu. Rev. Microbiol. 56: 93-116.
Kellen, W. R., and J. E. Lindergren. 1968. Biology of Nosema plodiae sp. n., a microsporidian pathogen of the Indian-meal moth, Plodia interpunctella (Hubner), (Lepidoptera: Phycitidae). J. Invertebr. Pathol. 11: 104-111.
Kellen, W. R., and J. E. Lindergren. 1969. Hostpathogen relationships in two previously undescribed microsporidia the Indian-meal moth, Plodia interpunctella (Hubner), (Lepidoptera: Phycitidae). J. Invertebr. Pathol. 14: 328-335.
Kramer, J. P. 1965. Nosema necatrix sp. n., and Thelohania diazoma sp. n. microsporidians from the armyworm, Pseudaletia unipuncta (Haworth). J. Invertebr. Pathol. 7: 117-121.
Ku C. T., C. Y. Wang, Y. C. Tsai, C. C. Tzeng and C. H. Wang. 2007. Phylogentic analysis of two putative Nosema isolates from Cruciferous Lepidopteran pests in Taiwan. J. Invertebr. Pathol. 95: 71-76.
Maddox, J. V., M. D. Baker, M. R. Jeffords, M. Kuras, A. Linde, L. F. Solter, M. L. McManus, J. Vavra, and C. R. Vossbrinck. 1999. Nosema portugal, n. sp., isolated from gypsy moths (Lymantria dispar L.) collected in portugal. J. Invertebr. Pathol. 73: 1-14.
Maddox, J. V., and R. K. Sprenkel. 1978. Some enigmatic microsporidia of the genus Nosema. Misc. Publ. Entomol. Soc. America. 11: 65-84.
Malone, A. L. 1984. A comparison of the Development of Vairimorpha plodiae and Vairimorpha necatrix in the Indian Meal Moth, Plodia interpunctella. J. Invertebr. Pathol. 43: 140-149.
Moore, C. B., and W. M. Brooks. 1994. An ultrastructural study of the episporontal inclusions produced during octosporogony by five species/isolates of Vairimorpha (Microspora: Microsporida), J. Invertebr. Pathol. 63: 197-206.
Malone, A. L., and C. A. McIvor. 1996. Use of nucleotide sequence data to identify a microsporidian pathogen of Pieris rapae (Lepidoptera, Pieridae). J. Invertebr. Pathol. 43: 140-149.
Malone, A. L., and P. J. Wigley. 1980. The distribution of Nosema carpocapsae, a protozoan pathogen of the codling moth, Cydia pomonella (Lepidoptera, Tortricidae), in New Zealand. N. Z. Entomol. 2: 152-153.
Patric, J. K., and N. M. Fast. 2002. Microsporidia: biology and evolution of highly reduced intracellular parasites. Annu. Rev. Microbiol. 56: 93-116.
Pilarska, D. K., L. F. Solter, J. V. Maddox, and M. L. McManus. 1998. Microsporidia from gypsy moth (Lymantria dispar L.) populations in central and western Bulgaria. Acta Zool. Bulg. 50: 109-113.
Siegel J. P., L. A. Lacey, and C. R. Vossbrinck. 2001. Impact of a north American isolate of the microsporidium Nosema carpocapsae on a laboratory population of the coding moth, Cydia pomonella. J. Invertebr. Pathol. 78: 244-250.
Sprague, V., J. J. Becnel, and E. I. Hazard. 1992. Taxonomy of phylum Microspora. Crit. Rev. Microbiol. 18: 285-395.
Swofford, D. L. 2003. PAUP*, Phylogenetic analysis using parasimony (* and other methods), Sinauer Associates, Sunderland, MA.
Thompson, J. D., T. J. Gibson, F. Plewniak, F. Jeanmougin, and D. G. Higgins. 1997. The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res. 25: 4876-4882.
Tsai, S. J., C. F. Lo, Y. Soichi, and C. H. Wang. 2003. The characterization of microsporidian isolates (Nosematidae: Nosema) from five important lepidopteran pests in Taiwan. J. Invertebr. Pathol. 83: 51-59.
Tanada Y., and H. K. Kaya. 1993. Protozoan infections: Apicomplexa, Microspora. In: Insect pathology, Academic, San Diego. 414-458
Undeen, A. H., and A. F. Cockburn. 1989. The extraction of DNA from microsporidia spores. J. Invertebr. Pathol. 54: 132-133.
Vossbrink, C. R., M. D. Baker, E. S. Didier, B. A. Debrunner-Vossbrink, and J. A. Shadduck. 1993. Ribosomal DNA sequences of Encephalitozoon hellem and Encephalitozoon cuniculi: species identification and phylogenetic construction. J. Eukaryot. Micorbiol. 40: 354-362.
Vossbrink, C. R., and B. A. Debrunner-Vossbrink. 2005. Molecular phylogeny of the Microsporidia: ecological, ultrastructural and taxonomic considerations. Folia Parasitol. 52: 131-142.
Watanabe, H. 1974. First report of the microsporidian infection of the cabbageworm, Pieris rapae crucivora Boisduval (Lepidoptera: Pieridae) in Japan. Appl. Ent. Zool. 9: 133-142.
Weiss, L. M. 1999. Moleculor biology, molecular phylogeny, and molecular diagnostic approaches to the microsporidia. pp. 85-128. In: Murray Wittner, and L. M. Weiss, eds. The microsporidia and microsporidiosis.