臺灣博碩士論文加值系統

The extremophilic red algae, Cyanidiales, have long attracted many research attentions due to their highly potential biotechnological applications. Although high genetic diversity has been previously revealed, a further systematic taxonomic re-examination with a more comprehensive taxon sampling is still largely unexplored. To this end, I re-examined their taxonomy and species diversity with the integration of morphological, physiological, and multilocus evidence based on 21 strains distributed around the world. Using seven genetic markers from plastid (psaA, psbA, rpoC1, dnaK, and rbcL) and mitochondrion (cox1 and cox2) for phylogenetic analyses, my study revealed at least 15 species in Cyanidiales that has been previously overlooked. Combining morphological (length, width, cell size, and the number of endospore), physiological (the effect of light and exogenous glucose on the growth rate and lipid content), and ecological features (the occurred microhabitats), my study showed that (1) Galdieria maxima should be transferred to the genus Cyanidioschyzon as Cd. maximum (Sentova) Huang et Liu comb. nov.; (2) all commonly used taxonomic characters failed for the species identification without the molecular assistance. With a better understanding of the taxonomic diagnosis in Cyanidiales, this study not only molecularly revealed their hidden cryptic species, but also provides more phenotypic information that further future taxonomic works in Cyanidiales.

Acknowledgment
Table of Contents
Abstract
1. Introduction
2. Materials and Methods
2.1 Species isolation and growth condition
2.2 DNA extraction and polymerase chain reaction
2.3 Phylogenetic analyses
2.4 Morphological observations
2.5 Physiological experiments
2.6 Statistical analysis
3. Results
3.1 Phylogeny of rbcL gene
3.2 Phylogeny of seven concatenated genes and the insertion within rpoC1
3.3 Morphological observation of different species in Cyanidiales
3.4 Growth rate and lipid content of different Cyanidiales species under different physiological conditions
4. Discussion
4.1 Taxonomic transfer of Galdieria maxima to Cyanidioschyzon.
4.2 Phylogenies support high species diversity in Cyanidiales.
4.3 Ecological advantage of mixotrophy and the association between cell size and trophic mode
5. Conclusions
6. Reference
List of Tables and Figures

Adamec, L., 1997. Mineral nutrition of carnivorous plants: A review. Bot. Rev. 63, 273-299.
Azúa-Bustos, A., González-Silva, C., Mancilla, R.A., Salas, L., Palma, R.E., Wynne, J.J., McKay, C.P., Vicuña, R., 2009. Ancient photosynthetic eukaryote biofilms in an Atacama Desert coastal cave. Microb. Ecol. 58, 485-496.
Banito, G.M., Gryganskyi, A.P., Trappe, J.M., Vilgalys, R., 2010. A global meta-analysis of Tuber ITS rDNA sequences: species diversity, host associations and long-distance dispersal. Mol. Ecol. 19, 4994-5008.
Chen, W., Zhang, C., Song, L., Sommerfeld, M., Hu, Q., 2009. A high throughput Nile red method for quantitative measurement of neutral lipids in microalgae. J. Microbiol. Methods 77, 41-47.
Ciniglia, C., Yoon, H.S., Pollio, A., Pinto, G., Bhattacharya, D., 2004. Hidden biodiversity of the extremophilic Cyanidiales red algae. Mol. Ecol. 13, 1827-1838.
Ciniglia, C., Yanh, E.C., Pollio, A., Pinto, G., Iovinella, M., Vitale, L., Yoon, H.S., 2014. Cyanidiophyceae in Iceland: plastid rbcL gene elucidates origin and dispersal of extremophilic Galdieria sulphuraria and G. maxima (Galdieriaceae, Rhodophyta). Phycologia 53, 542-551.
Ciniglia, C., Pinto, G., Pollio, A., 2017. Cyanidium from caves: A reinstatement of Cyanidium chilense Schwabe (Cyanidiophytina, Rhodophyta). Phytotaxa 295, 86.
De Luca, P., Taddei, R., Varano, L., 1978. "Cyanidioschyzon merolae": a new alga of thermal acidic environments. Webbia 33, 37-44.
Edgar, R.C., 2004. MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Res. 32, 1792-1797.
Felsenstein, J., 1985. Confidence Limits on Phylogenies: An Approach Using the Bootstrap. Evolution 39, 783-791.
Geitler, L., 1933. Diagnoses neuer Blaualgen von den Sunda-Insela. Arch. Hydrobiol. Suppl. 12, 622-634.
Gross, W., 1999. Revision of comparative traits for the acido- and thermophilic red algae Cyanidium and Galdieria. J Seckbach, ed, Enigmatic Microorganisms and Life in Extreme Environments. Kluwer Academic Publishers, Dordrecht, The Netherlands, 437–446
Guiry, M.D., Guiry, G.M., 2019. AlgaeBase. World-wide electronic publication, National University of Ireland, Galway. http://www.algaebase.org; searched on 19 July 2019.
Hsieh, C.J., Zhan, S.H., Lin, Y.C., Tang, S.L., Liu, S.L., 2015. Analysis of rbcL sequences reveals the global biodiversity, community structure, and biogeographical pattern of thermoacidophilic red algae (Cyanidiales). J. Phycol. 51, 682-694.
Hsieh, C.J., Zhan, S.H., Liao, C.P., Tang, S.L., Wang, L.C., Liu, S.L., 2018. The effects of contemporary selection and dispersal limitation on the community assembly of acidophilic microalgae. J. Phycol. 54, 720-733.
Iovinella, M., Eren, A., Pinto, G., Pollio, A., Davis, S.J., Cennamo, P., Ciniglia, C., 2018. Cryptic dispersal of Cyanidiophytina (Rhodophyta) in non-acidic environments from Turkey. Extremophiles 22, 713-723.
Merola, A., Castaldo, R., De Luca, P.,Gambardella, R., Musacchio, A., Taddei, R., 1981. Revision of Cyanidium caldarium. Three species of acidophilic algae. Giornale botanico italiano 115, 189-195.
Minoda, A., Sakagami, R., Yagisawa, F., Kuroiwa, T., Tanaka, K., 2004. Improvement of culture conditions and evidence for nuclear transformation by homologous recombination in a red alga, Cyanidioschyzon merolae 10D. Plant Cell Physiol. 45, 667-671.
Moriyama, T., Mori, N., Sato, N., 2015. Activation of oxidative carbon metabolism by nutritional enrichment by photosynthesis and exogenous organic compounds in the red alga Cyanidioschyzon merolae: evidence for heterotrophic growth. Springerplus 4, 559.
Müller, K.M., Oliveira, M.C., Sheath, R.G., Bhattacharya, D., 2001. Ribosomal DNA phylogeny of the Bangiophycidae (Rhodophyta) and the origin of secondary plastids. Am. J. Bot. 88, 1390-1400.
Myers, J., Kratz, W.A., 1955. Relation between pigment content and photosynthetic characteristics in a blue-green algae. J. Gen. Physiol. 20, 11-22.
Nabhan, A.R., Sarkar, I.N., 2012. The impact of taxon sampling on phylogenetic inference: a review of two decades of controversy. Brief. Bioinform. 13, 122-134.
Nilsson, R.H., Kristiansson, E., Ryberg, M., Hallenberg, N., Larsson, K.H., 2008. Intraspecific ITS variability in the kingdom fungi as expressed in the international sequence databases and its implications for molecular species identification. Evol. Bioinform. online 26, 193-201.
Pinto, G., Ciniglia, C., Cascone, C., Pollio, A., 2007. Species Composition of Cyanidiales Assemblages in Pisciarelli (Campi Flegrei, Italy) and Description of Galdieria phlegrea sp. Nov, in: Seckbach, J. (Eds.), Algae and Cyanobacteria in Extreme Environments. Springer, Dortrecht, pp. 487–502.
Pinto, G., Albertano, P., Ciniglia, C., Cozzolino, S., Pollio, A., Yoon, H.S., Bhattacharya, D., 2003. Comparative approaches to the taxonomy of the genus Galdieria merola (Cyanidiales, Rhodophyta). Cryptogamie Algol. 24, 13–32.
Qiu, H., Price, D.C., Weber, A.P.M., Reeb, V., Tang, E.C., Lee, J.M., Kim, S.Y., Yoon, H.S., Bhattacharya, D., 2013. Assessing the bacterial contribution to the plastid proteome. Trends Plant Sci. 18, 680–687.
R Core Team (2018). R: A language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. URL https: //www.R-project.org/.
Schneider, C.A., Rasband, W.S., Eliceiri, K.W., 2012. NIH Image to ImageJ: 25 years of image analysis. Nat. Methods 9, 671-675.
Schönknecht, G., Chen, W.H., Ternes, C.M., Barbier, G.G., Shrestha, R.P., Stanke, M., Bräutigam, A., Baker, B.J., Banfield, J.F., Garavito, R.M., Carr, K., Wilkerson, C., Rensing, S.A., Gagneul, D., Dickenson, N.E., Oesterhelt, C., Lercher, M.J., Weber, A.P., 2013. Gene transfer from bacteria and archaea facilitated evolution of an extremophilic eukaryote. Science 339, 1207-1210.
Selosse, M.A., Charpin, M., Not, F., 2017. Mixotrophy everywhere on land and in water: the grand écart hypothesis. Ecol. Lett. 20, 246-263.
Sentsova, O.J., 1994. The study of Cyanidiophyceae in Russia, in: Seckbach, J. (Eds.), Evolutionary Pathways and Enigmatic Algae: Cyanidium caldarium (Rhodophyta) and Related Cells. Kluwer Academic Publishers, Dordrecht, pp. 167-174.
Skorupa, D.J., Reeb, V., Castenholz, R.W., Bhattacharya, D., McDermott, T.R., 2013. Cyanidiales diversity in Yellowstone National Park. Lett. Appl. Microbiol. 57, 459-466.
Spijkerman, E., Lukas, M., Wacker, A., 2017. Ecophysiological strategies for growth under varying light and organic carbon supply in two species of green microalgae differing in their motility. Phytochemistry 144, 43-51.
Stoecker, D.K., Gustafson, D.E., 2003. Cell-surface proteolytic activity of photosynthetic dinoflagellates. Aquat. Microb. Ecol.. 30, 175-183.
Tamura, K., Stecher, G., Peterson, D., Filipski, A., Kumar, S., 2013. MEGA6: Molecular Evolutionary Genetics Analysis version 6.0. Mol. Biol. Evol. 30, 2725-2729.
Toplin, J.A., Norris, T.B., Lehr, C.R., McDermott, T.R., Castenholz, R.W., 2008. Biogeographic and phylogenetic diversity of thermoacidophilic Cyanidiales in Yellowstone National Park, Japan, and New Zealand. Appl. Environ. Microbiol. 74, 2822-2833.
Wang, X., Zang, R., Yin, Z., Kang, Z., Huang, L., 2014. Delimiting cryptic pathogen species causing apple Valsa canker with multilocus data. Ecol Evol. 4, 1369-1380.
Ward, B.A., Follows, M.J., 2016. Marine mixotrophy increases trophic transfer efficiency, mean organism size, and vertical carbon flux. Proc. Natl. Acad. Sci. U S A. 113, 2958-2963.
Yoon, H.S., Hackett, J.D., Pinto, G., Bhattacharya, D., 2002. The single, ancient origin of chromist plastids. Proc. Natl. Acad. Sci. U S A. 99, 15507-15512.
Yoon, H.S., Hackett, J.D., Ciniglia, C., Pinto, G., Bhattacharya, D., 2004. A molecular timeline for the origin of photosynthetic eukaryotes. Mol. Biol. Evol. 21, 809-818.
Yoon, H.S., Muller, K.M., Sheath, R.G., Ott, F.D., Bhattacharya, D., 2006. Defining the major lineages of red algae (Rhodophyta). J. Phycol. 42,