BIOSYSTEMATIC STUDY OF THE FERN GENUS DIPLAZIUM IN WEST MALESIA
TITIEN NGATINEM PRAPTOSUWIRYO
DEPARTMENT OF BIOLOGY THE GRADUATE SCHOOL BOGOR AGRICULTURAL UNIVERSITY BOGOR 2008
STATEMENT OF RESEARCH ORIGINALITY AND INFORMATION SOURCE This is to verify that my dissertation entitled: Biosystematic Study of the Fern Genus Diplazium in West Malesia is my own work and never been submitted to any institution before. All of the incorporated data and information are valid and stated clearly in the text, and listed in the references.
Bogor, September 2008 Titien Ngatinem Praptosuwiryo NIM P176 00005
ABSTRACT TITIEN NGATINEM PRAPTOSUWIRYO. Biosystematic Study of the Fern Genus Diplazium in West Malesia. (Under the supervision of Prof. Dr. Ir. Edi Guhardja, M.Sc., Prof. Dr. Mien A. Rifai, M.Sc., Prof. Dr. Masahiro Kato, M.Sci., and Dr. Dedy Darnaedi, M.Sc. ) Diplazium is a large genus consisting of about 400 species occur mainly in the tropics, sparingly in the subtropic and only locally extending into temperate. It was estimated that 300 species of the records were occurred in Malesia. Taxonomically, Diplazium is very difficult and quite insufficiently known. Therefore a comprehensive study on Diplazium in West Malesia was conducted by using morphological, ecological, geographic distribution, anatomical, palinological, cytological, as well as DNA analysis to understand the diversity and relationship among species. Based on gross morphological study on 1051 collection number of specimens as well as living collections, it was concluded that West Malesian Diplazium comprises of 69 species with 14 varieties. Thirteen species of them are proposed and described as new species, namely Diplazium asymmetricum, D. batuayauense, D. crameri, D. densisquamatum, D. halimunense, D. loerzingii, D. megasegmentum, D. megasimplicifolium, D. meijeri, D. parallelivenium, D. profluens, D. subalternisegmentum, and D. subvirescens. Two new varieties are poposed, namely D. accedens var. spinosum and D. silvaticum var. pinnae-ellipticum. D. pallidum var. montanum and D. accedens var. ridleyi are proposed as new status. Based on their main habitats, Diplazium can be classified into three major groups, viz. dryland (dominant), riparian and rheophytic species. Species diversity was culminated at 10001500 m above sea level. The individuals with different genetic load in the same species sometimes grow in the different habitat gradients. Based on the range of the geographical distribution, West Malesian species can be divided into three types: (1) very wide species (19 species), (2) Malesian species (27 species), and (3) locally endemic species (23 species). Anatomical study on the transversal section of stipe of 27 species showed that the vascular bundle shape is varying among species. Therefore the leaf-trace shapes are important diagnostic features which support species delimitation in Diplazium. Spore morphology study showed that perine ornamentations support in delimitating species in Diplazium. However the phylogenetic analysis using parsimony revealed that morphological variation of spore is inadequate to depict natural relationship among Diplazium species. Cytological study on 117 collection number from 54 localities included in 31 species found that West Malesian Diplazium has six ploidy levels with x = 41 (diploid, triploid, tetraploid, pentaploid, hexaploid, and octoploid). New cytological information for science on 19 species are recorded. They are D. aequibasale (2n = 164), D. angustipinna (2n = 123). D. asymmetricum (2n = 123), D. batuayauense (2n = 164, 205), D. crenatoserratum (2n = 123, 164), D. halimunense (2n = 123), D. hewittii (2n = 123), D. profluens (2n = 164), D. loerzingii (2n = 82, 123), D. pallidum (2n = 82), D. petiolare (2n = 82), D. porphyrorachis (2n = 164), D. riparium (2n = 82, 123), D. spiniferum (2n = 82), D. subserratum (2n = 82, 123, 164), D. subvirescens (2n = 123), D. tomentosum (2n = 82, 205), D. xiphophyllum (2n = 82, 246), and D. wahauense (2n=164). Phylogenetic analysis on morphological data sets of 69 species using parsimony revealed that the phylogenetic relationship among species in the genus Diplazium was very difficult to explain due to the lack of or weak support Bootstrap value. However the lack of or weak support for a phylogenetic tree does not strictly indicate that the pattern observed is incorrect but it does limit the amount of confidence that can be placed in the relationships between taxa.and the conclusions can be drawn from them. This study showed that some terminal clades formed are consisting of species that presumed to be closely related by formerly authors. DNA analysis resulted new gene rbcL sequences data on 25 species. Gene rbcL sequence is very well in supporting species delimitation and revealing the intraspecific diversity within species of Diplazium. Phylogenetic analysis on 29 species from West Malesia and 9 references species outside Malesia using parsimony revealed that gene rbcL is more informative than morphological data in inferring phylogeny of Diplazium and showed that West Malesian Diplazium is monophyletic. The position of D. porphyrorachis at the basal clade of the morphological tree is supported by the phylogenetic tree generated from molecular data (gene rbcL sequence). This study also showed the congruence between the clade of riparium group drawn by gene rbcL tree and the clade of imparipinnate frond group drawn by morphological tree.
RINGKASAN TITIEN NGATINEM PRAPTOSUWIRYO. Biosystematic Study of the Fern Genus Diplazium in West Malesia. (dibimbing oleh Prof. Dr. Ir. Edi Guhardja, M.Sc., Prof. Dr. Mien A. Rifai, M.Sc., Prof. Dr. Masahiro Kato, M.Sci. dan Dr. Dedy Darnaedi, M.Sc.). Diplazium merupakan marga besar tumbuhan paku yang beranggotakan lebih kurang 400 jenis yang sebagian besar ditemukan di daerah tropis, sedikit di daerah sub tropis dan hanya secara lokal meluas ke daerah beriklim sedang. Dari jumlah tersebut, diperkirakan 300 jenis terdapat di kawasan Malesia.
Marga
tumbuhan paku terestrial ini mempunyai ciri-ciri diagnosa sebagai berikut: Alur tangkai daun dan tulang daun utama terbuka dan alur ini diteruskan sampai tulang anak daun berikutnya; alur daun berbentuk U dengan dasar pipih pada sebagian besar jenis; anak daun basal yang mengarah ke rembang (acroscopic side) seimbang atau lebih kecil, pinggir lembaran daun tidak menulang; sori menggaris, ganda (diplazioid) atau tunggal (asplenoid), yang tunggal membuka ke arah uraturat daun utama atau urat-urat daun pusat dari cuping utama, yang ganda membuka dengan arah bertolak belakang. Secara taksonomi, Diplazium sangat sulit dan kurang dipahami. Tumbuhan muda mungkin saja subur dan sulit untuk dikenali sebagai suatu jenis. Banyak takson memiliki variasi morfologi. Adanya poliploidi, apomiksis dan hibrid dalam marga paku ini menambah sulit dalam membuat pembatasan jenis. Pengelompokan anak marga dari marga ini secara alami belum pernah dilakukan walaupun variasi morfologinya sangat luas. Pembatasan marga Diplazium juga masih meragukan. Di Malesia pada umumnya dan Malesia Barat pada khususnya, penelitian sistematika
Diplazium dengan menggunakan pendekatan biologi secara
menyeluruh belum pernah dilakukan untuk seluruh kawasan. Oleh karena itu penelitian biosistematika Diplazium dengan menggunakan pendekatan morfologi, ekologi, distribusi geografi, anatomi, palinologi, sitologi dan juga analisa DNA dilakukan untuk memahami keanekaragaman jenis dan hubungan kekerabatannya.
Berdasarkan pengamatan morfologi pada 1051 nomor koleksi specimen dan juga koleksi hidup, disimpulkan bahwa Diplazium Malesia Barat terdiri dari 69 jenis dan 14 varitas. Tiga belas jenis diantaranya diusulkan sebagai jenis baru, yaitu
Diplazium
asymmetricum,
D.
batuayauense,
D.
crameri,
D.
densisquamatum, D. halimunense, D. loerzingii, D. megasegmentum, D. megasimplicifolium,
D.
meijeri,
D.
parallelivenium,
D.
profluens,
D.
subalternisegmentum dan D. subvirescens. Dua varitas baru diusulkan, yaitu D. accedens var. spinosum dan D. silvaticum var. pinnae-ellipticum. D. pallidum var. montanum dan D. accedens var. ridleyi diusulkan sebagai status baru. Berdasarkan habitat utamanya, Diplazium dapat dikelompokkan dalam tiga group utama, yaitu jenis lahan kering (dryland species, 64 species), jenis riparian (5 jenis) dan reofit (2 jenis). Keanekaragaman jenis memuncak pada ketinggian 1000-1500 m dpl. Individu-individu dengan tingkat ploidi berbeda pada jenis yang sama seringkali menempati habitat ketinggian.
Berdasarkan jangkauan distribusi
berbeda berdasarkan
geografinya, jenis Diplazium
Malesia Barat dapat dibagi dalam tiga tipe: 1) jenis tersebar luas (19 jenis), 2) jenis Malesia (27 jenis) dan 3) jenis endemik setempat (23 jenis). Penelitian anatomi irisan melintang tangkai daun pada 27 jenis Diplazium memperlihatkan bahwa bentuk pembuluh vaskular bervariasi diantara jenis dan penting untuk menyokong pembatasan jenis. Pengamatan morfologi spora pada 46 nomor koleksi yang tercakup dalam 26 jenis memperlihatkan bahwa hiasan perine menyokong pembatasan jenis Diplazium. Walaupun demikian analisa filogeni dengan menggunakan parsimoni menunjukkan bahwa variasi perine tidak cukup untuk menggambarkan hubungan kekerabatan alami diantara jenis Diplazium. Penelitian sitologi pada 117 nomor koleksi dari 54 lokasi yang mencakup 31 jenis menemukan bahwa Diplazium Malesia Barat mempunyai enam tingkat ploidi dengan jumlah kromosom dasar x = 41 (diploid, triploid, tetraploid, pentaploid, heksaploid dan oktoploid). Informasi sitologi baru bagi dunia ilmu pengetahuan dilaporkan untuk 19 jenis, yaitu D. aequibasale (2n = 164), D. angustipinna (2n = 123). D. asymmetricum (2n = 123), D. batuayauense (2n = 164, 205), D. crenatoserratum (2n = 123, 164), D. halimunense (2n = 123), D. hewittii (2n = 123), D. profluens (2n = 164), D. loerzingii (2n = 82, 123), D.
pallidum (2n = 82), D. petiolare (2n = 82), D. porphyrorachis (2n = 164), D. riparium (2n = 82, 123), D. spiniferum (2n = 82), D. subserratum (2n = 82, 123, 164), D. subvirescens (2n = 123), D. tomentosum (2n = 82, 205), D. xiphophyllum (2n = 82, 246) dan D. wahauense (2n=164). Analisa filogeni pada seri data morfologi dari 69 jenis dengan menggunakan parsimoni menunjukkan bahwa hubungan kekerabatan filogeni diantara jenis Diplazium sangat sulit dijelaskan karena tidak adanya atau lemahnya nilai Bootstrap.
Bagaimanapun,
tidak adanya atau lemahnya
penyokong statistik bagi pohon filogeni tidaklah menandakan bahwa pola-pola yang diamati tidak benar namun ini hanya membatasi tingkat kepercayaan yang dapat ditempatkan pada hubungan kekerabatan diantara takson dan dari nilainilai tersebut kesimpulan dapat ditarik. Beberapa cabang ujung pohon morfologi tersusun dari jenis-jenis yang diduga berkerabat dekat oleh para peneliti sebelumnya. Analisa DNA menghasilkan data baru sekuensi gene rbcL dari 25 jenis Diplazium. Sekuensi gene rbcL sangat bagus untuk menyokong pembatasan jenis, mengungkap keanekaragaman genetik dalam jenis dan juga lebih banyak memberikan informasi untuk menduga filogeni Diplazium dibanding data morfologi. Berdasarkan analisa filogeni pada 29 jenis Malesia Barat dan 9 jenis referensi dari luar Malesia, Diplazium terbukti monofiletik berdasarkan gene rbcL. Kedudukan group porphyrorachis pada cabang pangkal pohon filogeni morfologi disokong oleh pohon filogeni gene rbcL. Cabang group daun menyirip gasal pada pohon morfologi selaras dengan cabang group riparium pada pohon gene rbcL. Penelitian ini tidak mengusulkan suatu kerangka sistematika di dalam marga Diplazium, sebab: (1) Pohon hipotesa filogeni yang dihasilkan dari data morfologi tidak didukung oleh alasan-alasan statistik yang obyektif; (2) Pohon hipotesa filogeni yang dihasilkan dari sekuensi gene rbcL tidak kokoh karena sebagian besar jenis kunci yang diduga dari pohon filogeni morfologi tidak dimasukkan dalam analisa disebabkan ketidaktersediaan sampel material segar pada jenis-jenis tersebut; (3) Analisa filogeni pada seri kombinasi data molekuler dan non molekuler untuk menduga filogeni Diplazium belum dilakukan.
Copyright @ 2008, Bogor Agricultural University All Right Reserved 1. It is prohibited to cite all or part of this dissertation without referring to and mentioning the source. a. Citation only permitted for the sake of education, research, scientific writing, report writing, critical writing or reviewing scientific problems. b. Citation does not inflict the name and honor of Bogor Agricultural University. 2. It is prohibited to republish and reproduce all part of this dissertation without copyright permission from Bogor Agricultural University.
BIOSYSTEMATIC STUDY OF THE FERN GENUS DIPLAZIUM IN WEST MALESIA
TITIEN NGATINEM PRAPTOSUWIRYO
A dissertation submitted to fulfill one of the requirements for the Doctorate Degree at the Study Program of Biology, Graduate School, Bogor Agricultural University
DEPARTMENT OF BIOLOGY THE GRADUATE SCHOOL BOGOR AGRICULTURAL UNIVERSITY BOGOR 2008
Examiner of first examination: Dr. Tatik Chikmawati, M.Si. Department of Biology, Faculty of Mathematic and Natural Sciences, Bogor Agricultural University, Bogor Examiners of second examination: 1. Dr. Sri Sudarmiyati Tjitrosoedirdjo, M.Sc. Department of Biology, Faculty of Mathematics and Natural Sciences, Bogor Agricultural University, Bogor 2. Dr. Rugayah, M.Sc. Herbarium Bogoriense, Botany Division, Research Center for Biology – Indonesian Institute of Sciences, Cibinong Sciences Center, Cibinong
Dissertation Title
: Biosystematic Study of the Fern Genus Diplazium in West Malesia
Name of student
: Titien Ngatinem Praptosuwiryo
Number of student
: P176 00005
Study Program
: Biology
Certified by Supervisor Committee
Prof. Dr. Ir. Edi Guhardja, M.Sc. (Chairman)
Prof. Dr. Mien A. Rifai, M.Sc (Member)
Prof. Dr. Masahiro Kato, M.Sci. (Member)
Dr. Dedy Darnaedi, M.Sc. (Member)
The Biology Study Program
Dr. Dedy Duryadi Solihin, DEA (Head)
Graduate School
Prof. Dr. Ir. Khairil A. Notodiputro, M.S. (Dean)
Examination Date: 25 March 2008
Graduation Date:
PREFACE Research on the West Malesian Diplazium under the tittle Biosystematic Study of the Fern Genus Diplazium in West Malesia was conducted from 20012007. Field study was carried out at 54 localities in the primary and secondary forest of Jawa, Sumatra and Kalimantan. Specimens examination was conducted at Herbarium Bogoriense (BO) and Herbarium of Singapore Botanic Gardens (SING). Study on the anatomy, cytology, and mode of reproduction type were conducted at the Anatomical and Cytological Laboratory of
Herbarium
Bogoriense. Observation on spore morphology by using Scanning Electron Microscope and DNA squencing (gene rbcL squencing) were conducted at the Laboratory of Department of Chemical and Biological Sciences, Faculty of Science, Japan Women s University and the Laboratory of Molecular Systematic of Department of Biological Science, University of Tokyo. A part of this dissertation (Chapter 5) has been published under two different tittles articles: (1) Cytological Study of Some Species of Ferns Genus Diplazium in Java I. that published in Floribunda 2 (5): 128-137 (2004) and (2) Cytological Study of Some Species of Ferns Genus Diplazium in Java II. that published in Floribunda 2 (8): 209-221 (2005). I greatly appreciate to my supervisors: Prof. Dr. Ir. Edi Guhardja, Prof. Dr. Mien A. Rifai, Prof. Dr. Masahiro Kato and Dr. Dedy Darnaedi, for their advice, guidance and encouragements throughout this study. I sincerely thank Dr. Irawati and Dr. Eko Baroto Walujo for their advices and technical support. I am indebted to Prof. Dr. Ryoko Imaichi, Dr. Chie Tsutsumi, Dr. Y. Kita, Dr. Mami Konomi, Dr. Mamiko Sato, and Mie Hashino for their helpful advices and technical supports. I would like express my appreciation to Dr. Chin See Chung and Ms. Serena Lee M. L. for giving me an opportunity to study Diplazium specimens at the Singapore Botanic Gardens Herbarium. I am grateful to Dr. R.J. Johns for fruitful discussion and his guidance when I was studying Diplazium specimens in Singapore Botanic Gardens.
I would like to sincerely thank
Dr. Teguh Triono, Dr. Titik Rugayah, and Dr. Joko Ridho Witono for the fruitful discussion, criticism, and technical supports. I also thank Dr. Sri Sudarmiyati Tjitrosoedirdjo and Dr. Tatik Chikmawati for their suggestions and corrections.
I sincerely thank Dr. Rudy Lukman for his helpful advices and soft literatures. I thank Abdulrokhman Kartonegoro, S.Si. and Dr. Nunik S. Ariyanti for taking some pictures of specimens type and literatures from Leiden. I would like to thank Prof. Dr. Hiroshi Okada and Dr. Hirokazu Tsukaya for involving me in the field study in Java and Central Kalimantan. I also thank to Dra. Esti Munawaroh, Wita Wardani, S.Si., Arief Hidayat, S.Si., Bp. Bambang Purwadi, Bp. Ujang Hapid and Sri Wahyuni, S.P. for sampling and fieldtrips. I am also grateful to Bp. Endjum and Ibu Ahati for maintaining the living collections. I thank the following institute in which I used their facilities:
a)
Herbarium Bogoriense, Botany Division, Research Center for Biology Indonesian Institute of Science (Lembaga Ilmu Pengetahuan Indonesia), b) Center for Plant Conservation, Bogor Botanic Gardens
LIPI, c) Department of
Chemical and Biological Sciences, Faculty of Science, Japan Women s University, d) Department of Biological Science, University of Tokyo, Japan. Finally, my grateful goes to my dear father (Bp. Ng. Praptosuwiryo who has passed away on 25 th February 2006) and mother (Ibu Kardiyem) my sisters and brother (Yu Ni, Dik Tum, and Dimas Bas) for their deep understanding and moral supports. This research was partly supported by Proyek Kompetitif Pengembangan IPTEK LIPI TA 2004 , Prof. Dr. Masahiro Kato (Department of Botany, National Museum of Nature and Science, Amakubo, Tsukuba, Japan), Prof. Dr. Ryoko Imaichi (Department of Chemical and Biological Sciences, Faculty of Science, Japan Women s University, Japan) and The Singapore Botanic Gardens Research Fellowship 2005.
CURRICULUM VITAE
Titien Ngatinem Praptosuwiryo was born on 27 March 1969 in Boyolali, Central Java, the second daughter from four children from father the late Ngadimin Praptosuwiryo and mother Kardiyem. She was graduated from Bogor Agricultural University (Institut Pertanian Bogor) in 1994. In 1994 to 1996, she worked at Herbarium Bogoriense, Puslitbang Biologi LIPI, Bogor as an honorary researcher in ferns. In September 1996, she had an opportunity to continue study at IPB sponsored by Prof. Dr. Masahiro Kato, the University of Tokyo, Japan and admitted to the degree of M.Sc. in August 1999. In 1999 to 2000 she continued working at Herbarium Bogoriense. In September 2000, she had an opportunity to continue study at IPB for her PhD. This study was partly supported by MENRISTEK and LIPI. Since 2001 she has been working at the Center for Plant Conservation-Bogor Botanic Gardens, LIPI, Bogor as a researcher in plant systematic. Her research area is biosystematic study on ferns.
TABLE OF CONTENTS Page LIST OF TABLES
……………………………………………………..
iv
LIST OF FIGURES ………..…………………………………………….
v
LIST OF APPENDIXES …………………………………………………
vi
1 INTRODUCTION Taxonomical Aspects of Diplazium and Its Systematic Problems …………………………………………………............ The Diversity of Diplazium in Malesia .......................................... The Biological Aspects on Systematics Study of Ferns …............ Morphological and Anatomical Evidences in Taxonomy ……………………………………………........ The Constribution of Palynology to Systematics: Spore Morphology Evidence in Pteridophytes ......................... Cytological Evidence in the Revealing Taxonomic Problems on Diplazium and Its Closely Related Genera ........................................................................ The Utility of Molecular Techniques for phylogenetic studies of pteridophytes: Gene rbcl Sequences ....................... Objectives ……………………………………….……………….
1 7 8 8 9
10 11 12
2 DIVERSITY AND ECOLOGY OF DIPLAZIUM Introduction ……………………………………………………… Materials and Methods ……………………………………......... Results and Discussion ……………………………………......... Ecology .………………………………………….................. Rheophytic Diplazium ……………………...................... Riparian Diplazium ………………………....................... Diplazium in Dryland ……..……………........................ Diversity of Diplazium Based on Elevation …............... Conclusions ……………………………………………..............
13 14 14 14 15 16 16 19 22
3 THE DISTRIBUTION OF WEST MALESIAN DIPLAZIUM INSIDE AND OUTSIDE MALESIA Introduction ……………………………………………………… Materials and Methods …………………………………………... Results and Discussion ………………………………………….. Very Wide Distribution Species ………………….................. Malesian Species ………………………………….................
23 24 25 26 26
i
Endemic Species to Island in West Malesia ………................. Conclusions ………………………………………………...........
27 40
4 THE STELAR ANATOMY OF STIPE AND ITS TAXONOMIC SIGNIFICANT IN DIPLAZIUM Introduction …………………………………………………….. Material and Methods …………………………………………... Results and Discussion ………………………………………….
41 42 42
Conclusions ……………………………………………………………
46
5 CYTOLOGICAL AND REPRODUCTIVE STUDIES ON DIPLAZIUM IN WEST MALESIA Introduction …………………………………………………….. Materials and Methods …………………………………………. Results and Discussion …………………………………………. Chromosome Number Variations and Mode Reproduction Types on Diplazium ………………………………………... The Relationship between Ploidy Level and Morphological Variation within Species and Closely Related Species of Diplazium ............................................... Relationship between ploidy level and habitat gradient ....... Correlations between reproductive mode and habitat ........... Conclusions ................................................................................
47 49 50 50
66 71 72 81
6 PHYLOGENETIC STUDIES OF DIPLAZIUM FROM WEST MALESIA: EVIDENCE FROM MORPHOLOGY Introduction …………………………………………………...... Character Selection and Construction …………………………. Character Selection ………………………………………… Character Type …………………………………………….. Character Coding …………………………………………… Character Variation within West Malesian Diplazium ……….. Materials and Methods ……………………………………….... Taxon Sampling ………………………………………….... Character Examination of Diplazium …………………....... Phylogenetic Analysis ……………………………………… Results and Discussion ………………………………………… Conclusions ………………………………………………….....
83 85 85 85 86 87 100 100 100 101 110 117
7 SPECIFIC DELIMITATION AND RELATIONSHIP AMONG SPECIES OF DIPLAZIUM BASED ON SPORE MORPHOLOGY Introduction .................................................................................. Materials and Methods ................................................................. Results and Discussion .................................................................
118 120 122
ii
Spore Characters of Diplazium and Its Use in Supporting Species delimitation and Identification …........... Phylogenetic Analysis ……………………………………... Conclusions .................................................................................
122 137 143
8 MOLECULAR SYSTEMATIC OF DIPLAZIUM FROM WEST MALESIA Introduction …………………………………………………….. Materials and Methods …………………………………………. DNA Analysis ……………………………………………… Phylogenetic analysis ………………………………………. Results and Discussion ………………………………………… Infraspecific Genetic Diversity in Diplazium ……………… Species Delimitation in Diplazium based on Gene rbcL Sequence ……………………………………………... Informative Characters of Gene rbcL Sequences for Inferring Phylogenetic Hypothesis of Diplazium ………. Phylogenetic Analysis ……………………………………… The Monophyly of Diplazium ………………………….. Relationships among species within Diplazium ….…….. Conclusions ……………………………………………………...
146 148 149 161 161 161 163 165 166 166 165 175
9 TAXONOMIC STUDY OF THE FERN GENUS DIPLAZIUM IN WEST MALESIA Introduction …………………………………………………….. Materials and Methods …………………………………………. Taxonomic Treatment …………………………………………..
176 176 177
10 GENERAL DISCUSSION Synthesis ……………………………………………………….. General Discussion …………………………………………….. Systematic Implications for the Genus Diplazium ……………...
283 284 285
11 CONCLUSIONS …………………………………………………...
187
LITERATURES …………………………………………………….......
292
LIST OF TABLES Table 2.1. Classification of Diplazium based on their main habitat ……
18
Table 2.2. Diversity of Diplazium based on elevation ………………….
20
iii
Table 3.1. Distribution of West Malesian Diplazium inside and outside Malesia …………………………………………..….
28
Table 3.2. Endemic Species of Diplazium in West Malesia ……………
39
Table 3.3. Species diversity and endemism of Diplazium in four mainlands of West Malesia ……………………………
40
Table 5.1. Somatic Chromosome Numbers, Ploidy Level and Mode Reproduction Type of Diplazium from West Malesia ...
52
Table 5.2. Polyploid series of Diplazium in West Malesia Based on Present Study .............................................................................
73
Table 6.1. Characters, character states, and coding for 88 characters utilized in construction of morphological data set of Diplazium
102
Table 7.1. Spores Description of Diplazium in West Malesia ................
124
Table 7.2. Characters, character states, and coding for 17 characters utilized in construction of spore morphology dataset of Diplazium …………………………………………
143
Table 7.3. Coding for 17 characters utilized in construction of spore morphology data set of Diplazium ……………………
144
Table 8.1. List of Taxa Used in This Study ……………………………
152
Table 8.2. Primers Used for Amplifying and Sequencing DNA from Diplazium (Hasebe et al, 1994) ……………………………
160
Table 8.3. Infraspecific Genetic Variatons of Diplazium based on Gene rbcL Sequences ………………………………….
162
Table 8.4. Interspecific Genetic Variatons of Diplazium based on Gene rbcL Sequences ………………………………….
164
LIST OF FIGURES Figure 2.1. a-b. Light shade-ferns of Diplazium ……………………….
21
Figure 2.2. Elevational distribution of Diplazium species in West Malesia …………………………………………………….
21
Figure 4.1. Vascular structure of the leaf axis ………………………….
43
Figure 4.2. Leaf-trace shapes in Diplazium ……………………….......
44
iv
Figure 4.3. Leaf-trace shapes in Diplazium. .............................................
45
Figure 5.1. Somatic chromosome of Diplazium. ………………………
74
Figure 5.2. Somatic chromosome of Diplazium ………………………...
75
Figure 5.3. Somatic chromosomes of Diplazium ……...………………..
76
Figure 5.4. Somatic chromosomes of Diplazium ………………………
77
Figure 5.5. Somatic chromosomes of Diplazium cordifolium ……………
78
Figure 5.6. Somatic chromosomes of Diplazium ......................................
79
Figue 6.1. Rhizome appeareance of Diplazium ………………………….
89
Figure 6.2. The variation of scale shapes in Diplazium …………………
91
Figure 6.3. Margin of scales …………………….………………………
92
Figure 6.4. Stipes appearances of Diplazium ………………….……….
93
Figure 6.5. Frond architectures of Diplazium ……………………………
95
Figure 6.6. Venation types of Diplazium ……………………………….
98
Figure 6.7. Sori variation in Diplazium .………………………...….......
99
Figure 6.8. Strict consensus of 8 trees of length 1366 from unweighted morphological dataset comprises 88 morphological caharcters. .......................................................
111
Figure 7. 1. Group I. a and b. D. accedens; c and d. D. bantamense; e. D. lobbianum; f and g. D. pallidum; h-j. D. procumbens; k-l. D. sorzogonense .................................................................
133
Figure 7.2. Group II. a. D. subserratum; b-c. D. vestitum; d-e. D. vestitum var. borneense; Group III. f-g. D. crenatoserratum; h-i. D. prescottianum; Group IV. j-l. D. silvaticum ......................................................
134
Figure 7.3. Group V. a-c D. pallidum; Group VI. d-e. D. cordifolium; g-i. D. tomentosum; j.. D. malaccense; k-l. D. megasegmentum; m -o. D. simplicivenium ………………………………………
135
Figure 7.4. Group VII. a-c. D. profluens; Group VIII. d. D. spiniferum; Group IX. e-f. D. subvirescens ……….....
136
v
Figure 7.5. Tree number 1 of 100 the most parsimonius trees …………
141
Figure 7.6. The strict consensus of 100 the most parsimonious trees …………………………………………..
142
Figure 8.1. Strict consensus of the 200 equally most-parsimonious trees obtained in maximum parsimony analysis of the rbcL sequence data. …………..…………………………………..
169
LIST OF APPENDIXES Appendix 1. Matrix of 88 Morphological Characters for Maximum Parsimony …………………………………………………. Appendix 2. Gene rbcL Sequence Data ………………………………..
309 321
Plate 1. Diplazium asymmetricum Praptosuwiryo …………………….
351
Plate 2. Diplazium batuayauense Praptosuwiryo ……………………...
352
Plate 3. Diplazium crameri Praptosuwiryo ...........................................
353
Plate 4. Diplazium densisquamatum Praptosuwiryo ..............................
354
Plate 5. Diplazium halimunense Praptosuwiryo .....................................
355
Plate 6. Diplazium loerzingii Praptosuwiryo …………………………..
356
Plate 7. Diplazium megasegmentum Praptosuwiryo ……………………
357
Plate 8. Diplazium megasimplicifolium Praptosuwiryo …………………
358
Plate 9. Diplazium meijerii Praptosuwiryo ……………………………..
359
Plate 10. Diplazium parallelivenium Praptosuwiryo ...............................
360
Plate 11. Diplazum profluens Praptosuwiryo ...........................................
361
Plate 12. Diplazium subalternisegmentum Praptosuwiryo .......................
362
Plate 13. Diplazium subvirescens Praptosuwiryo .....................................
363
Plate 14. Diplazium subvirescens Praptosuwiryo ……………………….
364
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