Systematic Studies in Abildgaardieae (Cyperaceae) Kerri Lyn Clarke B.Se.Hons University of New England A thesis submitted for the degree of Doctor of Philosophy of the University of New England October 2005 The University of New England Annidale, NSW 2351, Australia Declaration I certify that the substance of this thesis has not already been submitted for any degree and is not currently being submitted for any other degree of qualification I certify that any help received in preparing this thesis, and all sources used, have been acknowledged in this thesis. Kerri Lyn Clarke ii Prologue Format The fonnat of this thesis follows broadly the Australian Systematic Botany fonnat and recommendations as outlined in the Style Guide of the University of New England (http://www.une.edu.au/tlc/styleguide/). Thesis structure The thesis has been broken down into separate smaller studies to emphasise the Australian taxa and to provide a basis where each chapter may be prepared as papers for publication. ｾｴ ｡ｲ ｰ･ｳ Nomenclature Names for Australian species follow the Australian Plant name index, with the exception of species assigned to Abildgaardia by Goetghebeur (1986) i.e. F. oxystachya, F. macrantha and F. pachyptera, which are included in this thesis under Abildgaardia. All other species names, and authorities, were obtained D'om Index ｋｾ･ｷ ｮｳｩ and the more recent World Checklist of Monocotyledons (2004). Provisional names have been included within the thesis for some species, however, these new names and combinations are not validly published here (Article 29 of the International Code of Botanical Nomenclature). 111 Acknowledgements My sincere thanks are extended to all those who assisted, encouraged and supported me throughout the course of my PhD study. Thanks to my supervisors Assoc. Prof. Jeremy Bruhl (UNE) for his support and patience provided during the finals months of thesis preparation; Adjunct Assoc. Prof. Karen Wilson (Royal Botany Gardens Sydney) for sharing her considerable knowledge of the group, especially regarding TYPE information; and Assoc. Prof. Nallamilli Prakash (UNE, now retired) for significant support and encouragenlent. Sincere thanks to the Botany technicians Doug Clark and Richard Willis, for their friendly assistance and advice; Chris Cooper for digital image help and financial monitoring; and former members of Botany staff Matt Gray, Barb Blenman, Olive Bourke (now retired) and Geraldine Cronin for their assistance while crnployed at Botany and continuing friendship. Patrick Littlefield and Peter Garlick (now retired) provided assistance with scanning electron microscopy (SEM) that is ｨ｣ｕｬｾｮ appreciated. Special thanks to those who aided with accommodation and assistance during field excursions: John Clarkson (MBA) friendly accommodation and assistance at :\1BA; Ian Co\vie (DNA) for his kindness, patience, and expertise in the field at Kakadu National Park; Lana Little, Queensland Parks and Wildlife Service Environmental Protection Agency, Chillagoe, for her good companionship and help in the field. Thanks also to the owners and manager of 'El Questro' resort in the Kimberleys for permission to collect specimens and providing accommodation. IV Many thanks to the following Herbarium directors and curators for providing bench space and to staff for assistance during visits, and providing subsequent large loans: NSW (special thanks to Leonie Purdie for assistance during my ｾ ｓｗ visits, Miguel in the library, and the friendly postgraduates and technicians \\'ho entertained me at lunches), DNA (Clyde Dunlop now retired) BRI (with special thanks to Philip Sharpe), MEL, CANB, PERTH, MBA; also, overseas herbaria KEW', EA (special thanks to Dr Muthama Muasya), PRE, L, GENT, P for providing loan rnaterial. The Directors of Australian National Parks and Wildlife Service and equivalents in N.S.W, N.T., Qld, W.A. provided permission to collect in their service areas. Many thanks to Professor Paul Goetghebeur for his comments on Bulbostylis barbata and Abildgaardia vaginata, and correspondence that provided general information on the range of species in Bulbostylis which ultimately aided in llly drawing the 'sampling' line for the genus in this study. Also, Professor Kare Lye for sharing with me his views on the Australian B. pyriformis and the African B. hispidula group; I included some of the B. hispidula subspecies to assess the limits of B. pyriformis. Thanks and appreciation to fellow postgraduates for comradery and advice where applicable. To my staunch allies and fellow postgrads Kathy Owen, Azadeh Hadaddachi and Mohammad Fatemi, I am indebted for their friendship, support and great advice. Special thanks to Liisa Atherton for her friendship, support and proofreading skills. A big thankyou to friend Sue Williams for providing accommodation during my visits to Sydney and her friendly support. I'd especially like to thank my parents LYll and Gerry Clarke, for their endless encouragement and support, and also to express my appreciation for looking after the v grandkids (during field work, herbaria visits, conference attendence and periodically during thesis construction). Lastly I would like to thank my five children Shannon, Melanie, Candace, Daniel and Poya, who have tolerated many years of my study, accompanied nle on field work and survived great financial hardship and sometimes boredom. Their love and support has been the mainstay of my dedication to completing the doctoral dissertation. My 10 year old son Poya deserves recognition in his own right, as he has lived and breathed this project for as long as he can remember - he will be glad to have a mother again. Provision of a 3 year PhD scholarship as part of a larger ABRS grant obtained by Jeremy Bruhl and Karen Wilson was gratefully received, as was an additional 6 month scholarship provided by UNERS (University of New England Research Scholarship). Financial support from Friends of Botany (1998-2004), Faculty of Sciences Postgraduate Support, UNE (1998-2001), Joyce W. Vickery Scientific Research Fund of the Linnean Society of New South Wales (1998) allowed for SEM, conference attendence in Perth (2000) and additional field work in Queensland (2000). The work in this thesis is entirely my own except where specifically indicated to the contrary. Kerri Clarke vi Table of Contents Declaration Prologue Format Thesis structure NOlnenclature Acknowledgements Table of Contents List o,fTables List o,fFigures Abstract Chapter 1 General introduction Abildgaardieae Lye History of the tribe Abildgaardieae General history of genera of the Abildgaardieae Fimbristylis, Abildgaardia, and Bulbostylis Embryo morphology and Anatomy Gerleral aim l'hesis outline Chapter 2 General materials and methods Plant material Taxa Studied Sampling Phenetic studies Taxa for phenetic analyses Characters for phenetic analyses Analysis preparation Analyses Ordination Classification Network analysis E,valuation Cladistic studies Analysis preparation Ingroup Outgroup C:haracters Polymorphism Leaf blade and culm anatomy Embryo morphology Scanning electron microscopy Inflorescence-synflorescence homology Analyses Evaluation Photomicroscopy Chapter 3 Crosslandia W.Fitzg.: a phenetic and cladistic study , , , , " " " " " i ii ii ii ii iii vi ix x xiii 1 1 1 1 3 3 8 9 9 11 11 11 12 14 14 15 15 17 19 19 21 22 22 24 24 24 24 25 25 26 29 32 34 35 36 37 39 39 Vl1 Introduction Materials and methods ｬｾ｡ｸ Phenetic study Characters Pattern Analyses C:ladistic study Ingroup Embryo morphology and anatomy Inflorescence-synflorescence structure Results I>henetic study Crosslandia, Abildgaardia, Fimbristylis, and Bulbostylis Crosslandia, Fimbristylis spiralis and Abildgaardia vaginata Crosslandia complex C:ladistic analysis Noteworthy characters C)bservations Inflorescence-synflorescence structure Floret sex Basal spikelets Embryo morphology Vegetative anatomy Discussion Nomenclature of Crosslandia '" " " Chapter 4 Abildgaardia Vahl: a phenetic and cladistic study Introduction Materials and Methods C[axa Phenetic Study Pattern Analyses C:ladistic study Ingroup Embryo morphology Anatomy PAUP* Analyses Results J?henetic study Abildgaardia schoenoides group Abildgaardia pachyptera - A. oxystachya group Abildgaardia macrantha group Abildgaardia ovata - F. odontocarpa group C::ladistic Analysis Observations Inflorescence-synflorescence structure Perianth Nut shape and pattern Embryo Anatomy Discussion Nomenclature of Abildgaardia " " " " " " " " " " " " " " " " " 39 42 42 43 43 47 47 47 48 48 50 50 50 57 57 61 68 68 68 75 79 80 80 87 92 94 94 " 94 96 96 99 99 101 101 101 103 103 103 103 109 110 114 114 117 119 119 120 120 126 126 131 135 VIII ｃｨ｡ｰｴ･ｾｲ 5 Phenetic and cladistic analyses of Australian Bulbostylis Kunth 136 136 136 Chapter 6 Testing monophyly of the tribe Abildgaardieae Lye 188 188 Introduction Materials and methods Taxa Phenetic study Pattern analy·ses Cladistic analysis Ingroup Outgroup Characters and homology Embryo morphology Anatomy PAUP* analyses Results '" Pllenetic study Bulbostylis densa group Bulbostylis turbinata-B. sp. aff. burbidgeae group Cladistic analysis Observations Inflorescence-synflorescence structure Amphicarpy Nut sculpturing Embryo morphology Discussion NOlTlenclature ofBulbostylis in Australia Introduction Materials and methods Ingroup Outgroup Characters and homology Embryo morphology and anatomy Analyses Results Cladistic analysis Characters Observations Inflorescence-synflorescence Nut epidermal pattern Embryo Anatomy Discussion General Conclusion Appendices 136 139 139 142 142 146 146 146 146 148 148 148 149 149 155 160 165 167 167 169 169 170 181 186 188 190 190 190 192 192 193 193 193 196 197 197 200 208 212 216 223 236 ix List of Tables Table 1.1 Tribes of the Cyperaceae 2 Table 1.2 Classification of Bulbostylis by Clarke Table 2.1 Ordination stress values " 5 " 21 Table 3.1 Specimens sampled as the focus group in the assessment of the genus Crosslandia '" " '" 44 Table 3.2 Attribute codes and definitions used in the main phenetic analyses for 45 Crosslandia Table 3.3 Taxa included in the cladistic analyses to assess the relationships of Crosslandia setifolia, provisional C. anthelata, Fimbristylis spiralis and Abildgaardia vaginata 49 Table 3.4 Floret sex distribution seen in aerial spikelets for sampled specimens in Crosslandia setifolia and the provisional C. anthelata Table 3.5 Comparison of species to be assigned to Crosslandia "................... 76 93 ".. " Table 4.1 Specimens sampled as the focus group in the assessment of Australian ,. 97 Abildgaardia Table 4.2 Attribute codes and definitions used in the main phenetic analsyses for the Australian Abildgaardia. . .. ... . ... .. ... ... . .. .. .. .. . .. . .. . . .. . . . .. . .. . .. . .. . .. .. ... 100 Table 4.3 Taxa included in the cladistic analyses to assess the relationships of species " 102 in Abildgaardia Table 5.1 Specimens sampled as the focus group in the phenetic assessment of Australian Bulbostylis " ".................. 140 Table 5.2 Attribute codes and definitions used in the main phenetic analyses for Australian Bulbostylis " 143 Table 5.,3 Taxa included in the cladistic analyses to assess the relationships of species in Australian Bulbostylis " ) 47 Table 6" 1 Taxa included in cladistic analysis to assess monophyly of the tribe Abildgaardieae ] 91 x List of Figures Figure 2.1 Schematic representation of the 'type' of variation in the photosynthetic pathway that correlates with the arrangement of tissues within vascular bundles in Cyperaceae 28 (from Soros & Bruhl 2000; Soros & Dengler 2001) Figure 2.2 General en1bryo types of the Cyperaceae (adapted from Haines and Lye 1983 33 after Van der Veken 1965) Figure 3.1 MDS ordination in 2-dimensions (stress = 0.17) 52 53 Figure 3.2 Correlation of attributes with ordination space in figure 3.1 Figure 3.3 WPGMA phenogram ( ｾ］ＭＰＮＱＩ using the Gower metric similarity coefficient showing groups that correspond with the ordination (Figure 3.1) 54 Figure 3.4 MDS ordination in 2-dimensions (stress = 0.11) showing groups formed when Abildgaardia vaginata is included within the Crosslandia complex 58 Figure 3.5 Correlation of characters that fit the ordination space in Figure 3.4 59 using the Gower metric similarity measure showing Figure 3.6 UPGMA phenogram Ｈｾ］ＭＰＮＱＩ four groups 60 Figure 3.7 MDS ordination in 2-dimensions (stress = 0.19) showing three groups: Crosslandia setifolia (G 1), Crosslandia anthelat' (G2), and Fimbristylis spiralis (G3) . ........................................................................................................................................ 62 Figure 3.8 Correlation of attributes with the ordination in Figure 3.7 63 Figure 3.9 Minimum spanning tree (MST) for OTU linkages of Crosslandia setifolia, C. anthelata and Fimbristylis spiralis that correspond to the ordination in Figure 3.4 .. 64 Figure 3.10 UPGMA phenogram Ｈｾ］ＭＰＮＱＩ using the Gower metric similarity measure that 65 corresponds to the ordination in Figure 3.7 Figure 3.11 Cladogram for tree 2 of 4 shortest trees (tree length=725) to assess monophyly for Crosslandia. Crosslandia setifolia, C. anthelata, F. spiralis and A. vaginata form a monophyletic group sister to species of Abildgaardia 67 Figure 3.12 highly reduced anthelodium 69 Figure 3.13. Inflorescence-synflorescence variation observed within Abil((gaardia vaginata. ........................................................................................................................................ 70 71 Figure 3.14 The reduced anthelodium Crosslandia anthelata Figure 3.15 Highly reduced secondary anthelodium Crosslandia anthelata 72 Figure 3.16 A terminal head of spikelets as sessile ramified reduced anthelodia Cros"llandia ｡ｩｬｯｦｾｴ･ｳ 73 Figure 3.17 Lateral head of sessile spikelets in Crosslandia setifolia 74 Figure 3.23 Variation in embryos of Crosslandia setifolia and Abildgaardia vaginata 85 Figure 3.24 Leaf and/or culm transverse sections showing C4 fimbristyloid anatomy.Crosslandia setifolia 86 Figure 4.1 MDS ordination in 2-dimensions for OTUs of Abildgaardia, Crosslandia, Fimbristylis and Bulbostylis 104 Figure 4.2 MDS ordination (stress = 0.13) showing OTUs forming broad species groups 105 within Abildgaardia Figure 4.3. Correlation of attributes with ordination space in Figure 4.2 106 Figure 4.4 Minimum spanning tree (MST) from network analysis that corresponds to the ordination in Figure 4.2 107 Figure 4.5. WPGMA (8=-0.1) phenogram for the full Abildgaardia analyses, using the Gower metric association measure, corresponds to the ordination in Figure 4.2 ......... 108 Figure 4.6 Two dimensional MDS ordination (stress = 0.1) showing OTUs b'Touped as 111 Abildgaardia oxystachya, A. pachyptera and A. sp. aff. pachyptera Figure 4.7 Minimum spanning tree from network analysis that corresponds to the ordination 112 in I<igure 4.6 Figure 4.8 WPGMA (B=-O.I) phenogram, using the Gower metric association measure, that 113 corresponds to the ordination in Figure 4.4 Xl Figure 4.9 MDS ordination in 2-dimensions (stress = 0.17) showing OTUs grouped as Abildgaardia ovata, A. macrantha, F. odontocarpa, and F. sp. aff. odoJltocarpa 115 Figure 4.10 WPGMA (B=-O.l) phenogram, using the Gower metric association measure, that corresponds to the ordination in Figure 4.6 116 Figure 4.11 One of 126 equally most parsimonious trees (TL=987, CI=0.4863, HI=0.5137) . ...................................................................................................................................... 118 Figure 4.12 Variation of the inflorescence-synflorescence structure seen in species of Abi1dgaardia 121 Figure 4.13 Perianth in Abildgaardia 122 Figure 4.14 Scanning electron micrographs (SEM) and light micrographs ＨｌｾＱＩ of nuts in species of Abildgaardia 123 Figure 4.15 Scanning electron micrographs (SEM) of nuts in species of Abildgaardia 124 Figure 4.16 Scanning electron micrographs for Abildgaardia hygrophila and Fimbristylis 125 variegata Figure 4.17 Light micrographs of mostly whole cleared embryos for Abildgaardia-type 127 embryos in species of Abildgaardia Figure 4.18 Light micrographs of whole cleared embryos for some species assigned to Abildgaardia 128 Figure 4.19 Culm and leaf blade transverse sections for two species of Abildgaardia showing the typical outlines, arrangement of sclerenchyma strands per vascular bundle, and ｃｾ fimbristyloid anatomy 129 Figure 4.20 Culm and leaf blade transverse sections for species of Abildgaardia showing the typical outlines, arrangement of sclerenchyma strands per vascular bundle, and C4 fimbristyloid anatomy 130 Figure 5.1. MDS ordination in 2-dimensions (stress = 0.17) from primary phenetic a'1a1ysis (see Chapter 3) highlighting Bulbostylis 150 151 Figure 5.2. MDS ordination for OTUs of Bulbostylis (stress = 0.18) Figure 5.3 Characters that correlate (>80%) to group formation in the ordination shown in Figure 5.1. Figure 5.4. Minimum spanning tree (MST) for OTUs of Bulbostylis corresponding to ordination in Figure 5.2 152 Figure 5.4. Minimum spanning tree (MST) for OTUs of Bulbostylis corresponding to 153 ordination in Figure 5.2 Figure 5.5. WPGMA (/3 = -0.1) phenogram that corresponds with the ordination in Figure 5.2., for all OTUs of Bu1bostylis 154 Fig 5.6 Figure 5.6 MDS ordination in 2 dimensions (stress = 0.18) for the Bulbostylis densa group from the primary Bulbostylis analysis (see Figure 5.2) 156 Figure 5.7 Characters correlated (> 70 %) with the ordination in Figure 5.6 for OTU;; of the Bulbostylis densa group 157 Figure 5.8 Minimum spanning tree (MST) with linkages for the Bulbostylis densa group plotted onto the 2-dimensional ordination in Figure 5.6 158 Figure 5.9 WPGMA (/3 = -0.1) phenogram for the Bulbostylis densa subset (see Figure 5.2 for all species of Bulbostylis) that best correlates with the ordination (Figure 5.(») and Minimum Spanning Tree (MST) (Figure 5.8) 159 Figure 5.10 MDS ordination in 2-dimensions (stress= 0.18) for OTUs of the B. turbinata group from Figure 5.2 161 Figure 5.11 Attributes correlated (>80%) with the ordination in Figure 5.10 f()f OTCs of the Bulbostylis turbinata group 162 Figure 5.12 Minimum spanning tree (MST) with linkages for the Bulbostylis turbinata group 163 plotted onto the 2-dimensional ordination of Figure 5.10 Figure 5.13 WPGMA Ｈ ｾ ］ -0.1) phenogram for the Bulbostylis turbinata subset (see Figure 5.2 for all OTUs of Bulbostylis) that fits the ordination (Figure 5.10) and MST (Figure 5.12) 164 Figure 5.14 Cladogram for tree 1 of 112 shortest trees (tree length = 1490) in the assessment of monophyly for Australian species of Bulbostylis 166 Figure 5.15 Variation of synflorescence structure for some species of ｂｵｬ｢ｯｳｾｹｬｩｳ 167 xii Figure 5.16 Scanning electron micrographs (SEM) showing the variation of nuts for some san1ples of the Bulbostylis densa group 171 Figure 5.17 Scanning electron micrographs (SEM) showing the variation of nuts for samples of the Bulbostylis burbidgeae group 172 Figure 5.18 Scanning electron micrographs (SEM) showing the variation of nuts for samples of the Bulbostylis turbinata group 173 Figure 5.19 Scanning electron micrographs (SEM) showing the variation of nuts for samples fro:m Bulbostylis pyriformis and the B. hispidula complex 174 Figure 5.20 Scanning electron micrographs (SEM) showing the differences between nuts of Bulbostylis barbata and B. sp. aff. barbata 175 Figure 5.21 A. Embryo morphology in Bulbostylis 176 Figure 5.22 Variation in Bulbostylis embryo size, shape, and development of second primordial leaf 177 Figure 5.23 Culm and leaf blade transverse sections in Bulbostylis " 178 Figure 5.24 Culm and leaf blade transverse sections for Bulbostylis 179 Figure 5.25 Culm and leaf blades transverse sections for Bulbostylis 180 Figure 6.1 Cladogran1 for tree 2 of 66 shortest trees (tree length = 1490) for the tribe 194 Abildgaardieae Figure 6.2 Nelmesia melanostachya ISOTYPE showing general habit, including solitary spikes where the lateral spikelet is reduced to a single floret.. 199 Figure 6.3 Scanning electron micrographs (SEM) showing the variation of nut outline and " 202 epidermal sculpturing in some species of Fimbristylis Figure 6.4 Scanning electron micrographs (SEM) showing the variation of nut outline and " 203 epidermal sculpturing in some species of Fimbristylis Figure 6.5 Scanning electron micrographs (SEM) and light micrographs (LM) show-ing the variation of nut outline and epidermal sculpturing in species of ｆｩｭ｢ｲｩｳｾｹｬｩｳ (including 204 Tylocarya) " " Figure 6.6 Scanning electron micrographs (SEM) showing the nut outline and epidermal surface in two species of Nemum " 205 Figure 6.7 Scanning electron micrographs (SEM) showing the variation of nut outline and epidermal sculpturing in species from the provisional tribe Arthrostylideae 206 Figure 6.8 Scanning electron micrographs (SEM) showing the variation in nut outline and epidermal sculpturing for outgroup species of Schoenoplectus and Schoenoplectiella . .............................................................................................................................. , 207 Figure 6.9 Light micrographs of whole cleared embryos showing the variation in shape and size for some species assigned to Fimbristylis, plus schematic embryos for Tylocarya 208 and Nelmesia. Fimbristylis depauperata (K.L. Clarke 305, L. Little) Figure 6.10 Light micrographs of whole cleared embryos for some species from the outgroup used in cladistic analyses: Actinschoenus, Trachystylis and Schoenoplectiella... "...... 211 Figure 6.11 Culm and leaf blade transverse sections for some species from the provisional tribe' Arthrostylideae' selected as outgroup taxa for use in cladistic analyis, showing the typical outlines, arrangement of sclerenchyma strands per vascular bundle, and C 3 anatomy , 214 Figure 6.12 Culm and leaf blade transverse sections for Fimbristylis, Tylocm:ya, and Nemum . ...................................................................................................................................... 215 xiii Abstract The tribe Abildgaardieae Lye is composed of 5-7 genera: Crosslandia, Fimbristylis, Abildgaardia (= Fimbristylis section Abildgaardia), Bulbostylis, Nemum, Nelmesia and Tylocarya (= Fimbristylis nelmesii). There has been little disagreement about the general boundaries of the tribe. However, linlits of the main genera are disputed and unresolved. Some species and generic boundaries of Crosslandia, Abildgaardia and Australian Bulbostylis require assessment across their morphological and geographical range of distribution. The general aim of this thesis is to test monophyly of and within the tribe Abildgaardieae. To address the aim, the limits for Crosslandia, Abildgaardia and Australian Bulbostylis are assessed to determine the species and generic limits on a global level. Data from nl0rphology, vegetative anatomy and embryo morphology were used in phenetic and cladistic analyses. Phenetic analyses of morphological data were used to test and set species limits. Additional characters from morphology, vegetative anatomy and embryo morphology were used in cladistic analyses to test monophyly of the tribe and previous classifications. Representative samples frorn }?imbristylis, Nemurn, and the monotypic genera Nelmesia and Tylocarya (= Fimbristylis nelmesii) were added to those species defined in phenetic analyses. Data were polarised using the outgroup method; with outgroup taxa selected from the provisional Arthrostylideae, Schoenoplectus and Schoenoplectiella. Cladistic analysis revealed that the tribe Abildgaardieae is not monophyletic when Nemum, Nelmesia and Tylocarya are included. Members of' Arthrostylideae' violated monophyly of Abildgaardieae. The limits of Crosslandia and Abildgaardia xiv are revised: Crosslandia 4 spp., all endemic to Australia and Abildgaardia 11 spp., 9 in Australia, 8 endemic. The limits for Australian species of Bulbostylis remain unresolved, with further testing of a wider sample of overseas species needed. A new species of Bulbostylis, B. kakadu ined., is recognised. Species of Nemum are not monophyletic and Nelmesia rendered Abildgaardia non-monophyletic in the full tribal analysis. Tylocarya fonned a clade with Fimbristylis depauperata. Species of Fimbristylis did not fonn a monophyletic group. Suggestions for further work are gIven.