BIOL2023/2923 Botany

Transcription

1 BIOL2023/2923 Botany Manual Part A: Plant Diversity Actinotus helianthi, white flannel flower of the Sydney region. Pracs Weeks Angiosperm Diversity Bryophyte and Lycophytes Diversity Monilophyte and Gymnosperm Diversity Plant Systematics Plants of the Sydney region: Diversity & identification Myrtaceae & Proteaceae Plants of the Sydney region: Diversity & identification Ericaceae & Rutaceae

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3 Week One ANGIOSPERM DIVERSITY General introduction The aim of these practicals and associated audiovisual sessions is to introduce you to the broad diversity of angiosperms and to provide you with training in their identification. To achieve this, you must first become familiar with the descriptive terminology used to communicate about the variation observed in plant morphology. Once you are conversant with botanical descriptive terminology, you should be able to identify any plant in the world, as long as you have a (reliable) taxonomic key. Thus, we will also provide you with training in how to use dichotomous keys as well as open-entry, interactive keys. As with any new language, you must be prepared to practice using botanical terminology. Therefore, throughout this component of the course, we will provide you with the opportunity to practice your newly acquired skills by keying out plant species that we will supply. BIOL2023/2923 Botany 1.1

4 Angiosperm Morphological Terminology In this practical you will be introduced to a range of descriptive terms applied to the vegetative and reproductive morphology of angiosperms. These features, or characters, are used to determine the phylogenetic relationships of angiosperms and to provide a means of identifying unknown species by way of taxonomic keys. It can not be over-emphasized that it is far more efficient for you to learn how to apply morphological terminology correctly than it is to memorise species names. Whilst we have made every effort to provide material that exemplifies a particular character, you should realise that many of the terms should be considered as points along a continuum of morphological variation. You will discover that intermediate conditions exist! Perhaps the best way to conceptualise plant morphology is to think of plants as being modular in their construction (Fig 1). Notice how every module comprises a leaf and its node, an axillary bud and an internode. The branch or stem, therefore, is made up of a number of these modules. Figure 1 Fig 4.2 Judd et al. BIOL2023/2923 Botany 1.2

5 Similarly, inflorescences are shoots that serve for the formation of flowers. Inflorescence characters can be very confusing due to the arbitrary separation of leaf and flower-bearing portions of a shoot. Two fundamentally different types of inflorescence can be recognized (Fig 2). One type is terminated by a flower, whereas the other is not. Thus we have determinate (monotelic) and indeterminate (polytelic) inflorescences. Fig 4.28 Judd et al. Figure 2 BIOL2023/2923 Botany 1.3

6 Flowers also follow a generalized morphological pattern (Fig 3 ) which can be considered as a series of concentric rings whorls of organs. In the case of flowers with both male and female parts, the female reproductive structures (gynoecium) is always surrounded by the male reproductive parts (androecium). The petals (corolla) always surround the androecium and the sepals (calyx) always surround the corolla. Thus, the variation encountered in flowers generally reflects the presence or absence of one of these whorls or the fusion of various components within or between whorls. This consistency of spatial arrangement of organs allows us to formularize the morphology of any flower. Fig 4.16 Judd et al. Figure 3 BIOL2023/2923 Botany 1.4

7 Floral formulas act as an accurate, shorthand means of conveying the form of any flower. It relies on the use of codes arranged in a precise sequence. The following codes each refer to the number of parts and whether they are fused within (are connate) or between (are adnate) whorls. The sequence always follows: 1. Symmetry: irregular or regular.. 2. Number of parts in the calyx (K) 3. Number of parts in the corolla (C) 4. Number of parts in the androecium (A) 5. Number of parts in the gynoecium (G) 6. Whether the gynoecium is: inferior or superior G G If parts are connate (i.e. fused within a whorl) they are surrounded by a circle e.g. If parts are adnate (i.e. fused between whorls) they are linked by a line e.g. Thus a formula of:.. K5, C5, A10, G2 Would describe an irregular flower with 5 free sepals, 5 free petals 10 free stamens and a gynoecium of 2 free, superior carpels. BIOL2023/2923 Botany 1.5

8 Layout and sequence of practical 1 Section 1 A number of stations have been set up throughout the laboratory. Each station illustrates several morphological characters. The stations are grouped into vegetative or reproductive terminology. You can visit the stations in any order. Each station contains a set of tasks for you to complete. Once you have worked through the morphological definitions, you should move onto section 2 of the practical. Section 2 This will involve you in describing a number of rainforest plants using the terminology that you have just defined. You will then identify the rainforest plants you have just described by using an interactive, computerized key. Section 3 You will describe the morphology of a flower using a standardised floral formula. IF YOU ARE UNSURE ABOUT ANY COMPONENT OF THIS PRACTICAL, PLEASE DO NOT HESITATE TO ASK A DEMONSTRATOR. BIOL2023/2923 Botany 1.6

9 Section 1: Vegetative and reproductive morphology In the spaces provided, record your own definitions and/or illustrations of the morphological characters. Habit Vegetative characters Tree. Shrub. Herb. Suffrutescent. Vine. Liana. BIOL2023/2923 Botany 1.7

10 Leaf arrangement Alternate. Opposite. Whorled. Pseudowhorled. Leaf structure Simple (unlobed). Simple (lobed). Compound (pinnate) BIOL2023/2923 Botany 1.8

11 Compound (bipinnate). Compound (palmate) Unifoliolate. Stipules Interpetiolar. Petiolar. Terminal. BIOL2023/2923 Botany 1.9

12 Leaf shape Ovate. Obovate. Elliptic. Oblong. Leaf base Acute. Rounded. Cuneate. Cordate. BIOL2023/2923 Botany 1.10

13 Leaf tip Acute. Acuminate. Mucronate. Emarginate.. BIOL2023/2923 Botany 1.11

14 Leaf margin Entire. Serrate. Dentate Crenate. Domatia Trichomes Simple Branched. BIOL2023/2923 Botany 1.12

15 Reproductive characters Determinate inflorescences Cyme. Raceme. Umbel. Panicle. BIOL2023/2923 Botany 1.13

16 Indeterminate inflorescences Raceme. Umbel. Head (capitulum). Spike. Spadix. BIOL2023/2923 Botany 1.14

17 Carpel fusion (carpels connate) Free carpels (apocarpous). Fused carpels (syncarpous). Carpel fusion (carpels adnate with hypanthium) Superior ovary... Inferior ovary. Section 2 Take one piece of each rainforest plant and fill in the following table. When you have completed the table you will be shown how to use the interactive key to rainforest plants. BIOL2023/2923 Botany 1.15

18 Specimen Habit Leaf arrangement Leaf form Stipules Domatia Leaf aroma Oil dots Leaf margin Leaf length mm Leaf width mm Gland Hairs Petiole length mm Other features Example tree alternate simple absent present present present entire absent absent Leaves soft A B C D Record the family and species name below A. B. C. D. 1.16

19 Section 3 Take a piece of flowering stem with at least two flowers on it. Write a floral formula for the species. We will identify the species in a subsequent practical using the key to angiosperms in the Flora of the Sydney Region. Floral formula: BIOL2023/2923 Botany 1.17

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21 Week Two / Week Three DIVERSITY OF BRYOPHYTES AND TRACHEOPHYTES Contents page Bryophytes (Week 2) Concepts Practical Exercises A. Liverworts B. Hornworts C. Mosses Keywords Review Questions Lycophytes and monilophytes (week 2) Concepts Practical Exercises D. Psilotopsida (Monilophyte) E. Lycophyta (Lycophyte) Keywords Polypodiopsida (Monilophytes) and gymnosperms (Week 3) Concepts Practical Exercises A. Polypodiopsida Keywords Review Questions Gymnosperms (week 3) Concepts B. Cycadophyta C. Ginkophyta D. Pinophyta E. Gnetophyta Keywords Review Questions Recommended books Evert RF and Eichhorn SE Raven: Biology of Plants. 8th Ed. Freeman & Co Publishers. New York. NY. Online material: NOTE: The introductory text in the following sections on bryophytes, ferns and gymnosperms is a brief summary of the basic structural and reproductive features in each group. In the practical exercises you will have the opportunity to study these features in detail. It is best to read the concepts before each practical class, which will help you understand what you are doing and allow you to make rapid progress. BIOL2023/2923 Botany 2.1

22 INTRODUCTION TO BRYOPHYTES, LYCOPHYTES AND MONILOPHYTES Week Two Concepts Land plants. Several lines of evidence indicate that land plants evolved from a charophycean green alga. Like the green algae, land plants have chlorophyll a (primary photosynthetic pigment), chlorophyll b and carotenoids (accessory pigments), starch (carbohydrate reserve) deposited within the chloroplasts, and cellulose (principal component of the cell wall). A number of structural modifications were accomplished during the transmigration from aquatic to terrestrial environment. Unlike the green algae (except Coleochaete), land plants form a phragmoplast and a cell plate during cell division. The phragmoplast consists of a ring of microtubules and actin filaments, oriented perpendicular to the equatorial plane, which expands centrifugally ahead of the cell plate as it grows toward, and eventually fuses with, the parental cell wall. In most algae, in contrast, cell division is accomplished by a phycoplast (microtubules oriented parallel to the new wall, the new wall grows centripetally). All plants have embryos (hence the term embryophytes). The development of the zygotes depends on the protective and nutritional environment of the gametophyte. One of the features that protects the land plants from drying out is a layer(s) of sterile cells around the antheridium (pl. antheridia; male sex organs, produce sperm) and around the archegonium (pl. archegonia; female sex organs, produce eggs and protect the embryo). A sterile jacket layer was also formed around the sporeproducing cells of the sporangium (pl. sporangia). Land plants typically show a heteromorphic alternation of generations. The aerial parts of most plants are covered with a waxy layer, the cuticle (prevents dehydration), which is closely correlated with the presence of stomata (sg. stoma; gas exchange). More advanced features include the evolution of the vascular tissue, vascular cambium, apical and secondary meristems, and leaves. BRYOPHYTES. Bryophytes are the simplest, least specialized, small land plants. They usually inhabit damp environment, although some can tolerate loss of most of their cytoplasmic water and rehydrate when water becomes available again. They have no true xylem and phloem, but they have developed alternative ways of trapping and transporting water. The gametophyte (haploid) is always nutritionally independent whereas the sporophyte (diploid) is permanently attached to, and dependent on, the gametophyte. In other words, the gametophyte is the conspicuous and dominant generation, in contrast to the dominant sporophytes of vascular plants. The gametophyte is usually attached to the substrate by elongate single cells of filaments of cells called rhizoids, which generally serve only to anchor the plants since absorption of water and ions commonly occurs directly through the gametophyte. Archegonia and antheridia both have sterile jacket layers; each archegonium contains a single egg, each antheridium produces numerous, biflagellate sperm. The sperm must swim through water to reach the archegonium. The archegonia are flask-shaped, with a long neck and swollen basal portion, the venter, which encloses a single egg. The central cells of the neck, the neck canal cells, disintegrate when the archegonium is mature, resulting in a fluid-filled tube through which the sperm swim to the egg. After fertilization, the zygote is retained within the archegonium where it develops into an embryo. The cells of the venter divide and enlarge, and the venter keeps pace with the growth of the young sporophyte within the archegonium; the enlarged archegonium is called calyptra. The subsequent details of development differ among the various bryophytes. Traditionally the term "bryophytes" has been used as a collective term for three distinct divisions of these relatively unspecialized plants: A. Liverworts (Hepatics), lacking stomata and specialized conducting cells; B. Hornworts, lack specialized conducting cells but have stomata, C. Mosses (Musci), which have stomata and specialized water-conducting and food-conducting cells in both their gametophytes and their sporophytes. BIOL2023/2923 Botany 2.2

23 Exercise A: LIVERWORTS (allow 45 minutes) The name liverwort (wort=herb) dates from the ninth century when it was thought, because of their livershape, that these plants might be useful in treating diseases of the liver. The simplest liverworts have a flattened, dorsoventral gametophyte called thallus. The most familiar and widespread genus is Marchantia. The thallus, which grows by repeated divisions of a single apical cell, contains pores and air chambers, gemma cups with gemmae (for vegetative reproduction), and elongated, disk- or spokeheaded stalks antheridiophores and archegoniophores. In a large group of "leafy" liverworts, the gametophyte differentiates into primitive leaves that are arranged into two or three rows. The discharge of spores is aided by specialized hygroscopic cells, elaters. A diagram of the life cycle of Marchantia is included on pages 2.6 & 2.7. Examine the colonies of the dichotomously branching gametophytes of Marchantia (demonstration), which is a common liverwort that grows in damp, nutrient-rich environments such as the flower pots in plant nurseries. A1. Are these thalli haploid or diploid? Using a dissecting microscope, examine a small sample of Marchantia and note the dark-green thallus with a midrib-like region, air pores, and gemma cups on the upper surface. A2. What is the function of gemmae? Examine the lower surface and note the many rhizoids, which are tubular structures of one or more cells that anchor the thallus to the substrate. You may also find multicellular scales on the lower surface. A3. [slide B320]. On a prepared slide of TS Marchantia thallus, note the air pores and chambers, chlorenchyma tissue (green), and storage tissue. A4. As shown in the demonstration of reproductive Marchantia, the gametophytes are unisexual, with the gametangia being borne on specialized, erect branches of the male and female plants. The antheridia are on disk-headed branches antheriodiophores, and the archegonia on spoke-headed archegoniophores. A5. [slide B323B]. Examine a prepared slide with LS antheridial head and find the elongate antheridia. Each antheridium consists of a short stalk and a mass of spermatogenous tissue surrounded by a sterile jacket layer that is one cell thick. A6. Are the sperm nuclei produced by meiosis or mitosis? BIOL2023/2923 Botany 2.3

24 A7. Draw and label a diagram of the antheridium In nature, the antheridial disk-head serves as a "splash platform" for raindrops. A8. How does this facilitate fertilization? Explain the journey of the spermatozoid. Examine a prepared slide LS archegonial head [slide B325]. Locate a fully developed archegonium and identify the swollen basal portion, or venter; the egg cell within the venter; and the elongate neck, with its neck canal cells. The archegonium may be surrounded by a collar-shaped flap of tissue. A9. Are the archegonia located on the upper or lower surface of the head? A10. Is the egg produced by meiosis or mitosis? Explain : A11. Draw and label a diagram of the archegonium Using a low-power objective, examine a prepared slide of LS mature sporophyte [slide B328]. Note the sporophyte is attached to the gametophyte by a large foot, which grows through the base of the venter into the storage tissue of the archegonial head. A12. Is the sporophyte haploid or diploid? : A13. What is the function of the foot? A14. Explain whether the sporophyte is a dependent organism and how? BIOL2023/2923 Botany 2.4

25 Examine the same slide under a high-power objective and identify the stalk (seta), capsule (sporangium), numerous spores and elongated elaters, all enclosed within the sterile jacket layer of the capsule. The elaters have hygroscopic wall thickenings that twist with changes in humidity and aid in spore dispersal. The remnants of the old archegonium form a cap, or calyptra. A15. What is the ploidy level of the calyptra? Explain: A16. Draw a diagram of the sporophyte and label. Examine the demonstration of thallose liverworts and leafy liverworts and note the morphological features that identify them. BIOL2023/2923 Botany 2.5

26 Figure 1: Life cycle of Marchantia BIOL2023/2923 Botany 2.6

27 From Raven et al. BIOL2023/2923 Botany 2.7

28 Exercise B: HORNWORTS (allow 15 min) Unlike liverworts, hornworts often have stomata (sg. stoma) on their gametophytes and sporophytes. The sporophyte is a highly specialized, upright structure. It consists of a bulbous foot, which is embedded in the gametophyte, and a cylindrical capsule. In the basal portion of the capsule is a meristem that continues producing new cells for several months. A sporogenous tissue inside the capsule surrounds a central, sterile columella. As the capsule matures beginning from its tip, it begins to dehisce along two lines from the tip toward the base, giving a "horned" appearance of the adult sporophyte. The most familiar genus is Anthoceros. B1. Examine thalloid gametophytes with attached sporophytes. Note the rosette-like shape of the gametophyte with irregular branching and lobed margins, with rhizoids underneath. The thallus may have cavities on the underside, containing dark masses of the blue-green alga Nostoc. These may be visible under the microscope. B2. Can you think why this association occurs?: Examine the demonstration microscope with a small piece of thallus dissected out. Note the number of chloroplasts in a cell and the presence or absence of pyrenoids. B3. What other group of plants contain pyrenoids and similar number of chloroplasts?: Note the mode of dehiscence of the sporangium and look for the thread-like, colourless columella. Tease out a portion of the sporangium in water on a slide and observe under a microscope. B4. Describe the shape of the spores and the pseudo-elaters: B5. Examine the two demonstration slides of LS young sporangium of Anthoceros [slides B300 and B304] showing a foot, meristem tissue, developing sporogenous tissue, and columella. BIOL2023/2923 Botany 2.8

29 Exercise C: MOSSES (allow 1.5 hour) Mosses constitute a diverse group of about 10,000 species, which inhabit an amazingly wide range of environments, from damp, tropical areas to Antarctica, and in deserts and on dry rocks. Moss gametophytes are leafy", with the leaves usually arranged in a spiral. The sporophytes are partly photosynthetic, differentiated into a stalk (seta) and a sporangium (capsule); there are no elaters. The gametophyte of "true mosses" (class Bryidae, eg. Funaria, Dawsonia) first forms as a protonema, which gives rise to a bud with an apical cell, and eventually to the leafy gametophyte. In many mosses, the stems of the gametophytes and sporophytes have a central strand of water-conducting cells (hydroids), which in some genera are surrounded by food-conducting cells (leptoids). The sporangia (capsules) are generally elevated on a stalk (seta) and spore dispersal is aided by a ring of hygroscopic teeth (peristome). Stomata are usually present on moss sporophytes although some consist only of a single, doughnut-shaped guard cell. The sequence of colour photographs on the large poster Life of a Moss shows the salient features of these organisms and their reproductive cycle. The poster is complemented by a set of demonstration slides from which the photographs were originally taken. You may notice that the principal features of the Moss life cycle are similar to those of Marchantia that you have studied earlier. A diagram of a moss life cycle is included on pages 2.12 & C1. Examine the colour photographs on the poster, comparing with the original slides if necessary, and make brief notes about the main features in the space below. BIOL2023/2923 Botany 2.9

30 Some mosses have primitive conducting tissue consisting of hydroids and leptoids. Identify the rhizome and aerial stem of live Dawsonia. C2. Examine a prepared slide of TS aerial stem Dawsonia [slide B384]. Identify the hydroids and leptoids: Examine demonstration photographs of transfer cells in the foot of Dawsonia capsule. These cells have extensive wall ingrowths, which increase the surface area/volume ratio and thus enhance the efficiency of transport of solutes. C3. How would you describe the shape of the ingrowths? : Look for antheridia and archegonia in young, reproductive plants of Funaria, or a similar moss, by dissecting the plant under a microscope (old plants will have only adult sporophytes). C4. (slides B381 & B361B). Examine slides with LS of moss archegonia and antheridia. Draw and label an archegonium and an antheridium. Are they different from those in the liverwort? : Examine a mature sporophyte on a live moss. Note the capsule, calyptra, operculum, and peristome. Breathe on the peristome teeth and with a hand-lens observe their hygroscopic movements. C5. Which way do the peristome teeth move in high humidity? Which way do they move in dry air? : Mosses of diverse morphologies are shown in the demonstration, including specimens with branched gametophytes, elongated capsules, etc. C6. Describe the apparent differences in their gametophytes and sporophytes : BIOL2023/2923 Botany 2.10

31 Peat mosses (sub-class Sphagnidae) The leaves of "peat mosses" (class Sphagnidae, eg. Sphagnum) typically consist of a combination of large, dead cells (hold water), which are surrounded by narrow, green living cells. Peat mosses form extensive peat bogs, covering about 1% of the world's land surface. Examine a live specimen of Sphagnum and note the radial arrangement of leaves on the stem. Mount a few leaves on a slide and examine under low and high power. Note the narrow elongated photosynthetic cells, and large hyaline cells (colourless) with perforations and wall thickenings. C7. Draw and label the two distinct cell types : BIOL2023/2923 Botany 2.11

32 Figure 2: Life cycle of a moss BIOL2023/2923 Botany 2.12

33 From Raven et al. BIOL2023/2923 Botany 2.13

34 KEYWORDS antheridiophore antheridium Anthoceros apical cell archegoniophore archegonium (arche=beginning) calyptra (kalyptra=covering) capsule (capsula=little box) cell plate columella (columna=column) cuticle (cutis=skin) Dawsonia elater (elatus=lift, carry away) embryo embryophyte foot Funaria gemma (gemma=bud) hornwort hydroid leptoid (leptos=slender) liverwort Marchantia moss neck canal operculum (operire=lid) peristome (peri=around) phragmoplast (phragmos=fence) phycoplast pore protonema (protos=first) rhizoid (rhiza=root) seta (seta=bristle) spermatozoid (sperma=seed) Sphagnum spore (sporos=seed) sporangium sporophyte stomata (stoma=mouth) thallus (thallos=branch) transfer cells venter (venter=belly) zygote (zygon=yoke) BIOL2023/2923 Botany 2.14

35 REVIEW QUESTIONS 1. Name the three groups of nonvascular plants. 2. Do the three groups have similar life cycles? What parts of the cycle are different? 3. Which of the three groups exhibit heteromorphic alteration of generations? 4. What is the dominant generation? 5. What are the characters that distinguish nonvascular plants from algae? 6. Which parts of nonvascular plants are haploid? Which are diploid? 7. How do liverworts differ from mosses? How do hornworts differ from both? 8. What are the morphological/anatomical differences between leafy liverworts and mosses? 9. How would you describe the anatomy of a moss leaf? 10. Is there vascular tissue in the moss leaf midrib? 11. Is there vascular tissue in the moss stem or seta? 12. Are male gametes produced by meiosis? Sure? 13. Is the egg cell produced by meiosis? 14. What happens to the neck canal cells during development of the archegonium? BIOL2023/2923 Botany 2.15

36 15. Why are nonvascular plants considered to be embryophytes? 16. Where is the embryo located on the nonvascular plant? 17. Where would you look for transfer cells? 18. Describe how the zygote develops into an embryo. 19. Are the spores formed by meiosis? Sure? 20. How are the spores released from the sporophyte? Any specific mechanisms in the three groups? 21. A spore germinates into a filamentous protonema: Where does the apical cell originate? How? 22. What is a bud? What is a bulbil? 23. What is the function of the capsule? 24. What is the function of the sporophyte foot? 25. How do the nonvascular plants deal with long-distance transport of water, minerals, and sugars? 27. What do you think is the evolutionary advantage of having a phragmoplast instead of a phycoplast? BIOL2023/2923 Botany 2.16

37 DIVERSITY IN TRACHEOPHYTES A number of key advances, besides those accomplished by the bryophytes, have accompanied the early invasion of the land by plants. These include the evolution of the shoot and efficient photosynthetic leaves (acquisition of light energy, acquisition of carbon dioxide), roots (absorption of water and nutrients, anchorage of the plant), and an efficient conducting system consisting of xylem (transport of water and nutrients) and phloem (transport of assimilate), which facilitates long-distance transport. Another important feature was the evolution of protective layers such as the cuticle and spores with protective walls. As early plants became increasingly suited to life on land, their gametophytic generation was progressively reduced in size and became increasingly more protected by, and nutritionally dependent on, the dominant sporophyte. Moreover, the life cycle has acquired the selective advantages of two new principles: a) heterospory, the production of microspores that carry genes over large distances, and megaspores that nourish the young embryo; and b) endosporial development of gametophytes, which provides protection for both male and female gametophytes and is the necessary first step in the evolution of seed plants. Construction of the vascular column. In most ferns and fern allies, the plant body consists entirely of primary tissues, there are no secondary meristems. The vascular tissues exhibit three basic arrangements: 1. Protostele, the most primitive type of stele. It consists of a solid central column of vascular tissue, xylem surrounded by phloem. It was found in extinct vascular plants and it is present in the living Psilotum, Lycopodium, and Selaginella. Additionally, this type of stele is found in the roots of most living seed plants. 2. Siphonostele, a hollow vascular cylinder surrounding a parenchymatous pith. In ferns the phloem may be inside as well as outside the cylinder together with an endodermis on both sides, in which case the stele is called solenostele. Vascular strands (leaf traces) depart from the hollow vascular cylinder and lead to the leaves, leaving behind open spaces (leaf gaps) in the cylinder. 3. Eustele, which consists of a system of discrete vascular strands that surround the pith, the strands being separated from one another by ground tissue. This type is present in almost all living seed plants. Figure 3: Types of stele From Raven et al. (a) A protostele, with diverging appendages, the evolutionary precursors of leaves. (b) A siphonostele with no leaf gaps; the vascular traces leading to the leaves simply diverge from the solid cylinder. this sort of siphonostele is found in Selaginella, amongst other plants. (c) A siphonostele with leaf gaps, commonly found in seedless vascular plants. (d) A eustele. Siphonosteles and eusteles appear to have evolved independently from protosteles. BIOL2023/2923 Botany 2.17

38 Origin of leaves. Leaves originated in more than one way. The following are the two most widely accepted theories. 1. Microphylls evolved as superficial lateral outgrowths (enations) of the stem (see diagram below). An outgrowth may or may not have a leaf trace, but if present the leaf trace does not create a leaf gap. Microphylls are associated with protosteles and are characteristic of the lycophytes and Psilotum (monilophytes). 2. Megaphylls, in contrast, are leaves with complex venation and the leaf traces are associated with leaf gaps. They are associated with siphonosteles and eusteles. According to the telome theory, megaphylls evolved by a sequence of modifications of a branch system, namely: a) "overtopping", whereby an equal dichotomous branching becomes overtopped by one dominant branch (the main axis) and the remaining branches become lateral; b) "planation", or flattening of all the lateral branches into one plane; and c) "webbing", whereby a thin sheet of photosynthetic tissue develops in the spaces between the neighboring branches. Two additional processes, "recurvation" and "coalescence", could explain the evolution of fertile leafy structures (sporophylls). Figure 4: Origin of leaves From Raven et al. BIOL2023/2923 Botany 2.18

39 Reproduction. The life cycles of seedless vascular plants are essentially similar to one another: an alternation of heteromorphic generations in which the sporophyte is dominant and free-living. Generally, the gametophytes are reduced in size and dependent on food derived from the sporophyte. An important advance, relative to bryophytes, was the evolution of heterosporous plants (e.g. Selaginella, Isoetes, Marsillea). These produce microspores and megaspores, which germinate and give rise to male gametophytes and female gametophytes (produce antheridia and archegonia). All seedless vascular plants have motile sperm which require water to swim to the archegonium. Diversity. Vascular plants go back at least 420 million years. The earliest ones, now extinct, belonged to the division Rhyniophyta which had simple, dichotomously branching axes without roots or leaves. Extinct Lycophyta, Equisetopisida and fern-like monilophytes were key representatives of the "Coal age plants" MYA, a period of great accumulation of fossil-fuel. The seedless vascular plants that live today may be classified in four divisions: PSILOTOPSIDA (only two living genera, Psilotum and Tmesipteris), which lack roots and usually also lack leaves. LYCOPHYTA (Lycopodium, homosporous; microphylls associated with protosteles. Selaginella, Isoetes, heterosporous), characterized by EQUISETOPSIDA (mostly extinct, only one living genus, Equisetum, homosporous), reduced scale-like leaves, eustele. POLYPODIOPSIDA (a very large group of "TRUE FERNS", eg. Polystichum, Pteridium, mostly homosporous but some are heterosporous), have megaphylls and siphonosteles. Exercise D: Psilotopsida (allow 20 min) As the name indicates (psilos=bare), the genus Psilotum is unique among living vascular plants in that it lacks both roots and leaves. The plant is a common greenhouse weed. The sporophyte consists of a dichotomously branching aerial portion with small scale-like outgrowths. Its vascular tissue, protostele, is the simplest of steles. Another genus, Tmesipteris, is endemic to Australia, New Caledonia, New Zealand and other regions of the South Pacific. Both genera are very simple plants that resemble the Rhyniophytes in their basic structure. Examine the live plant of Psilotum (demonstration). Note the dichotomous branching of the aerial stem, absence of leaves and roots, and large sporangia borne in united groups of three in the axils of scalelike outgrowths. The spores give rise to subterranean, bisexual gametophytes with antheridia and archegonia. As in all ferns and allies, the flagellated sperm require water to swim to the egg. The zygote develops into a simple embryo. Examine a prepared slide of TS Psilotum stem (slide B396) Locate the position of the protostele, phloem, xylem, and protoxylem. Compare with the adjacent 3D-diagram of a protostele. D1. Draw a diagram of the transverse section of Psilotum stem and label. Can you see any leaf gaps? BIOL2023/2923 Botany 2.19

40 Exercise E: Lycophyta (allow 30 minutes) The Lycophyta sporophytes are differentiated into distinct microphylls (one vein, one leaf trace), stem, and root. They usually have a protostele. All living species of Lycopodium (about 200) are small herbs with prostrate rhizomes and true roots, and with short upright branches. Microphylls are spirally arranged on the stem. All Lycopodium species are homosporous. Selaginella plants (about 700 species), which are smaller and more delicate, represent the more derived condition by being heterosporous. The megagametophytes develop inside the megaspore wall. Some species of Selaginella behave as "resurrection" plants, which curl up, turn brown and appear dead upon drying, but uncurl and turn green again when moistened. As a guide for the following exercises, a diagram summarizing the life cycle of Selaginella is attached at the end of this section (pp & 2.23). Examine the live plants of Lycopodium and Selaginella (demonstration). Note differences in the shape of microphylls and their arrangement on the stem. E1. How would you tell the two species apart? Can you see any cones? : As the name homosporous suggests, Lycopodium produces meiospores of only one size. Live sporophylls are pale green and are closely compacted into an elongated cone, or strobilus (strobilos=fircone). Each sporangium originates from the cells near the base of the leaf. Numerous spore mother cells (sporocytes) divide meiotically to form tetrads of spores. E2. Examine a prepared slide of LS strobilus of Lycopodium [slide B458]. Note that all sporangia and the spores they contain are identical in size. After the spores have been liberated from the sporangium, they may remain dormant for days or years but eventually germinate to produce small, inconspicuous gametophytes. Antheridia and archegonia are borne on the same gametophyte, usually intermingled but sometimes segregated into different areas. After fertilization, the zygote divides transversely to the long axis of the archegonium. One daughter cell forms a suspensor, while the other divides to form foot, stem, and root primordia. The rest of the life cycle is similar to that in Polypodiopsida in week 3. Traditionally the Lycophyta have served to demonstrate the evolution from homospory to heterospory. The heterosporous Selaginella produces meiospores of two different sizes: the larger one, the megaspore, which originates from megasporangia that are borne on megasporophylls, develops into the female gametophyte;the smaller one, the microspore, which originates from microsporangia that are borne on microsporophylls, develops into the male gametophyte. All sporophylls are compacted into strobili. In some species, the lower sporophylls of the cone may bear megasporangia while the upper ones bear microsporangia, although the spatial arrangement varies among species. The development of both male and female gametophytes is endosporous, that is, it occurs within the confines of the spore wall (see diagram below). E3. Examine a prepared slide of LS strobilus of Selaginella. [slide B465A]. Note the different size of microsporangia and megasporangia, as well as different size of misrospores (30-80 µm diameter) and megaspores ( µm diameter). E4. Draw a diagram of the heterosporous cone and label BIOL2023/2923 Botany 2.20

41 The microspore nucleus divides repeatedly, giving rise to a jacket one cell thick that surrounds the central spermatogenous cells. Each spermatogenous cell differentiates into a disc-shaped spermatozoid with two anterior flagella. At maturity the jacket cells break down, releasing the free-swimming spermatozoids. Figure 5: Development of heterosporous gametophytes E5. What evolutionary advantages can be attributed to heterospory? What are the advantages of endosporous development? KEYWORDS annulus (ring) antheridium apical cell archegonium cuticle (cutis=skin) dependent gametophyte dictyostele dominant sporophyte eusporangium eustele extant species Equisetum frond heterosporous plant homosporous plant lamina (thin plate) leaf leaf gap leaf trace leptosporangium (lepto=thin/fine/slender) Lycopodium megaphyll (megas=large/great) megaspore microphyll microspore petiole (petiol=leafstalk) phloem Polystichum Pteridium (ptero=wing/feather) prothallus (protos=first) protostele Psilotum (psilos=bare) pteridophyte rhizome (rhiza=root) root Selaginella shoot siphonostele (siphon=pipe/tube) sorus spores stele (stele=standing pillar) tapetum (tapet=carpet/rug) thallus (thallos=young shoot) tracheid (tracheia=windpipe/artery) tree-fern xylem vessel (vas=duct) BIOL2023/2923 Botany 2.21

42 Figure 6: Life cycle of Selaginella BIOL2023/2923 Botany 2.22

43 From Raven et al. BIOL2023/2923 Botany 2.23

44 BIOL2023/2923 Botany 2.24

45 Polypodiopsida (monilophytes) and Gymnosperms Week Three The living plants in the Polypodiopsida number about 11,000 species, the largest group of plants other than flowering plants. They exhibit great morphological diversity including very small plants; a climbing fern Lygodium with a long, twining rachis (extension of petiole) that may grow 30 m long; or Cyathea, a large tree-fern growing 25 m tall and 30 cm in diameter (all primary tissues) with leaves 5 m long. Some monilophytes (i.e. Psilotopsida, Marattiopsida, and Polypodiopsida) may be informally classified according to the method of development of their sporangia which can be either: a) Eusporangiate, represented by Ophioglossales (Psilotopsida) and Marattiopsida; or b) Leptosporangiate, which include the Polypodiopsida. Most Polypodiopsida are homosporous, with at least 10,500 species representing nearly all familiar ferns. However, Salviniales (also Polypodiopsida) are heterosporous and are known as the water ferns. The distinction is of fundamental importance for understanding reproduction in vascular plants. The origin of an eusporangium (eu=well) is multicellular, involving series of divisions in the surface cell layer that generate several outer layers and numerous spores. Leptosporangium (leptos=slender), in contrast, originates from a single initial cell, which first produces a stalk and then a capsule, and gives rise to a relatively small number of spores. In either case, the spores are enclosed within nutritive cell layers, tapetum (tapet=carpet/rug). Cellular details of the origin of eusporangia and leptosporangia are well illustrated in Raven et al. 1992, p.343. The following practical exercises deal with the morphology and anatomy of ferns in the order Polypodiales. The sequence of colour photographs on the large poster Life of a Fern emphasizes the key features of ferns and their life cycle. Details of the morphology, anatomy, and reproduction will be examined in the rest of the exercise. As a guide, a diagram of a homosporous fern life cycle is included at the end of this section (see Figure 5, pp 3.8 & 3.9). The simplest way of dealing with this section is to study the poster first, then do all the practical exercises, and finally go back to the poster and take notes where necessary. Exercise A: (Allow 1.5 hrs) A1. Study the poster series Life of a Fern (and associated demonstration slides 1-14). It is preferable to take your own notes rather than copying all the details. Structure of the gametophyte. To study the real organism, it seems logical to start from the earliest primordial stage. After the haploid spore has landed on a moist, hospitable ground, it absorbs water, swells, its protective wall cracks open, and the cell starts expanding and dividing. All this activity is microscopic, of course. Examine the microscope demonstration of germinating fern spores. Note the emergence of the prothallus with rhizoids. Small prothalli may be filamentous; larger ones will have commenced a twodimensional growth; a mature prothallus is the heart-shaped gametophyte. A2. What type of nuclear division produces the multicellular prothallus? : Examine live fern gametophytes growing in a dish (demonstration) and showing various stages of development. The youngest ones may not have reached the reproductive stage yet; others may be reproductive, having gametangia (archegonia and antheridia) on the lower surface; and large, mature gametophytes may already have a young sporophyte growing alongside them. Collect a small sample of reproductive gametophytes, place them lower surface uppermost on a slide, and examine under your dissecting microscope. Note the position of the single apical cell in the cleft of the heart-shaped gametophyte, rhizoids originating from the central cushion, and the relatively thin wings. A3. Draw the gametophyte and label BIOL2023/2923 Botany 3.1

46 Unlike the bryophytes, the fern gametangia occur on the same gametophyte, although they usually are spatially separated. Typically, the archegonia are located closer to the cleft of the heart-shaped gametophyte. On your specimen, locate the protruding archegonial necks (fertilized archegonia are brown). Antheridia, which are more globose and usually develop before the archegonia (temporal separation ensures outcrossing), are located among the rhizoids. Compare your specimen with the prepared slides of antheridia [slide B405] and archegonia [slide 407] in your slide trays. A4. Complete your drawing above in [A3] by indicating the relative positions of antheridia and archegonia on the gametophyte. If you have successfully located a gametophyte in the reproductive stage, it should have some antheridia with mature spermatozoids. Place the gametophyte on a slide, mount in a drop of water under a coverslip, tap gently with a lead pencil, and examine under 10x and 40x objectives. Mature antheridia undergo a hypotonic shock, which releases the motile spermatozoids into the surrounding liquid medium. A5. Can you see them swim? How would you describe the swimming motion? : Anatomical details of the archegonium are best seen on prepared sections. Examine the slide of LS fern archegonium [slide LS 409] in your slide tray under a HP objective. Identify the venter, egg, neck, and neck canal cells. A6. Draw the archegonium and label. How does it resemble the moss archegonium you saw last week? Following fertilization, the zygote begins to develop into an embryo, which then grows to become a young sporophyte. Examine the demonstration slide [B415] showing a young sporophyte still attached to the parental gametophyte. Note the primary root of the sporophyte, the region of foot (attachment to the gametophyte), and the first leaf (megaphyll) with a dichotomous venation. A7. Does the young sporophyte have a chance to survive out in the bush on its own? How soon? By what means? : Structure of the sporophyte. Despite the tender age and harsh environment, many young sporophytes "make it" and grow into robust, mature plants. Examine a mature sporophyte of live Polystichum with sori (demonstration plant). Identify the underground stem (rhizome) with adventitious roots, young leaves (fiddleheads) near the apex of the rhizome, and large, finely divided mature leaves (fronds). The frond petiole is usually covered in brown scales, and the compound, bipinnate lamina is subdivided into leaflets (pinnae) and further into secondary leaflets (pinnules). A8. Draw a diagram of the sporophyte and label Look for clusters (sori) of sporangia on the lower (abaxial) surface of the leaflets on large fronds in various stages of maturation. Note the variation in size and colour of the sori. Immature sori are very small and light-coloured, while the mature ones are distinct and brown. Break of a leaflet with mediumsized sori, and another leaflet with large, brown sori, and examine them both under your dissecting microscope. Note the spatial relationship of the sori to the veins. BIOL2023/2923 Botany 3.2

47 A9. Are the sori associated with the veins or do they occur along the leaf margins? (on your drawing (A8) of the sporophyte, indicate the position of sori and veins). Continue dissecting your specimens under the microscope. A developed sorus is protected by an umbrella-like, papery cover (indusium). Use a needle to remove the indusium from the sorus to expose the individual, stalked sporangia. Continue searching until you find a sorus with dark-brown, mature sporangia. If you bring your desk-lamp close to the exposed sorus, the sporangia will soon begin to dry out and crack open to discharge the spores. A10. How would you describe mechanistically the process you see? Scrape mature sporangia from an intact sorus onto a slide and mount in a drop of glycerol under a coverslip. Observe under 10x or 40x objectives. Identify the following structures: a) annulus, a peripheral row of cells with thickened inner walls; b) stomium (a "mouth"), thin-walled cells between the end of the annulus and the attachment of the stalk (this is where the sporangium will crack open); c) sporangium side walls; and d) spores, located inside the sporangium. As the glycerol dehydrates the cells of the annulus, the annulus straightens and the resulting tension opens the sporangium at the mouth, releasing the spores. When observed under 40x objective, the individual bean-shaped spores may be surrounded by a bulging "perine" coat or, if exposed, the spores may show a "scar" or "hilum" (the result of attachment of spores in the tetrad). A11. Draw the sporangium under HP and label all the details. a) Explain the mechanism of spore dispersal in this organism. b) In what way is this mechanism similar to that in the bryophytes? c) What type of nuclear division gives rise to the spores Figure 1: Diagram of sporangium BIOL2023/2923 Botany 3.3

48 Examine a prepared slide with TS sorus of Polystichum [TS indusium B432A, TS sporangia B420, fern sporangia B431] in your slide tray. Note the sorus, indusium, and sporangia. Compare with your own preparation above. A12. How would you compare these details with your preparation of the fresh material? Anatomy of the stele. A classical example of protostele was shown for Psilotum in last week s practical. Generally, the Pterophyta utilize a rich variety of siphonosteles. These apparently evolved from the protostele independently from eustele, which is characteristic of all seed plants. On the other hand, some Pterophyta such as Pteridium have evolved xylem elements that resemble the vessels of the seed plants Figure 2: Diagram of a siphonostele Examine a prepared slide of TS rhizome Gleichenia [slide 18] in your slide tray, which shows a siphonostele that has leaf traces but no leaf gaps. It may help if you first look at the slide under the dissecting microscope to see the proportions of the individual tissues, and then use the compound microscope to study the cellular details. Compare the structure with the diagram above. A13. Draw a half-diagram of the Gleichenia siphonostele and identify leaf traces. Examine a prepared slide of TS Pteridium rhizome [slide B436] in your slide tray that shows a dictyostele, which is a variation of siphonostele that has leaf gaps; also, they overlap, that is, two or more gaps appear in any transverse section. Compare with the adjacent diagram. Note the following features: a) Xylem, arranged in two concentric rings interrupted by leaf gaps, with extensive patches of sclerenchyma between the two rings. You may also see leaf traces and traces that depart into adventitious roots. The xylem consists of tracheids, parenchyma, and protoxylem (small tracheids at the periphery) b) Phloem (lacks companion cells) with adjacent pericycle and endodermis, which form a continuous layer around each segment of the xylem. BIOL2023/2923 Botany 3.4

49 Figure 3: Diagram of a dictyostele A14. Draw a map diagram of one half of the section of the Pteridium dictyostele. Can you find cambium? : To complement the view seen in the transverse sections, examine a prepared slide with longitudinal section of Pteridium stele LS Pteridium rhizome [slide 20] in your slide tray. Identify tracheids with unique wall thickenings; primitive vessels with scalariform thickenings and perforations in the end wall; and phloem elements with sieve areas. Finally, examine the slide of macerated rhizome [slide B2400] in your slide tray to see the details of the wall architecture in the individual cell types. Apical meristems. Theoretically, the evolution of specialized conducting cells allows more extensive division of labor among the various tissues of the plant. Downward phloem transport permits roots to extend deep into the dark soil, and ylem conducts water upward to the elongating shoot. Absorption of nutrient solutions by roots, and a steady import of carbohydrate from the photosynthetic megaphyll, allow the development of tissues that perform other specific tasks, such as that of the apical meristems. Most ferns, however, still depend on the activities of simple apical cells similar to those seen in the bryophytes, and unlike the more complex meristems found in the seed plants. Examine the demonstration slide of the shoot apex of Davallia. [slide 4 on demo bench]. Note a single apical cell. With luck, you may be able to find a procambial strand, which is the earliest visible sign of developing vascular tissue. A15. Draw the apical cell with the adjacent, derived, cells on either side. Indicate the location of the procambial strand if you have found it Figure 4: Shoot apical region of a fern BIOL2023/2923 Botany 3.5

50 Examine the demonstration slide of LS Davallia root apex [slide 5 on demo bench]. Note the single triangular apical cell, which cuts off new cells from all three faces. A16. Draw the apical cell. How does it differ from the apical cell in the shoot? : KEYWORDS annulus (ring) antheridium apical cell archegonium cuticle (cutis=skin) dependent gametophyte dictyostele dominant sporophyte eusporangium eustele extant species Equisetum frond heterosporous plant homosporous plant lamina (thin plate) leaf leaf gap leaf trace leptosporangium (lepto=thin/fine/slender) Lycopodium megaphyll (megas=large/great) megaspore microphyll microspore petiole (petiol=leafstalk) phloem Polystichum Pteridium (ptero=wing/feather) prothallus (protos=first) protostele Psilotum (psilos=bare) pteridophyte rhizome (rhiza=root) root Selaginella shoot siphonostele (siphon=pipe/tube) sorus spores stele (stele=standing pillar) tapetum (tapet=carpet/rug) thallus (thallos=young shoot) tracheid (tracheia=windpipe/artery) tree-fern xylem vessel (vas=duct) BIOL2023/2923 Botany 3.6

51 REVIEW QUESTIONS 1. What makes the ferns and fern allies more advanced than the bryophytes? 2. What do the ferns and fern allies lack relative to gymnosperms? 3. What benefits can be attributed to the evolution of vascular tissue? 4. What would be the selective advantage of a tall, upright habit? 5. What is the selective advantage of a water-proofing cuticle? 6. Would it make sense to develop stomata before developing an impermeable cuticle? 7. What is the function of roots, relative to rhizoids? 8. Why should vascular tissue precede the evolution of roots and active apical meristems? 9. How could lignified cells be useful in plants? 10. How would you describe the main differences between protostele, siphonostele, and eustele? 11. Would siphonostele and eustele be superior to protostele? 12. Do any living plants have a protostele? 14. Could a protostele form a leaf trace? 15. Do angiosperms have a protostele? In roots? In stems? 16. Do tree-ferns have secondary xylem? 17. What types of cells constitute the xylem in pteridophytes? 18. It could be argued that three types of leaves occur in the plant kingdom. What are they? 19. How would you tell if a small leaf was a microphyll rather than a megaphyll? 20. What is the differences between enation and telome? 21. What is the significance of the microphyll- and megaphyll-lines of evolution? 22. Why is Psilotum regarded as a living fossil? 23. What was the problem with the cambium in ancient lycophytes? 24. What has been the most significant change in gametophyte morphology since 420 MYA? 25. How would you compare the evolutionary potential of haploid and diploid organism? 26. What are the reproductive constraints in ferns and their allies, relative to gymnosperms? 27. What is the significance of heterospory? 28. What is the meaning of "primitive" and "advanced" in describing plant features? 29. Why are the terms "original" (or "relictual") and "derived" more accurate? 30. Why do we study pteriodophytes? BIOL2023/2923 Botany 3.7

52 Figure 5: Life cycle of the fern Polypodium BIOL2023/2923 Botany 3.8

53 From Raven et al. BIOL2023/2923 Botany 3.9

54 GYMNOSPERMS One of the most important innovations to arise during the evolution of vascular plants was the seed and the pollen grain. The seed provides protection for the enclosed embryo, and the stored food supports the germination and establishment of the seedling, thus giving the seed plants a great selective advantage. Another crucial evolutionary advance compared with ferns was the bifacial vascular cambium. This is a secondary meristem, which produces secondary xylem (in gymnosperms mostly tracheids with circular bordered pits); secondary phloem (sieve cells, but no companion cells in gymnosperms); and a secondary meristem, cork cambium. All seed plants have megaphylls. Similarly to ferns, gymnosperms are characterized by an alternation of heteromorphic generations, with large, sometimes gigantic, independent sporophytes and greatly reduced gametophytes. The female gametophyte and origin of the naked seed. Gymnosperms are plants with "naked" seeds (gymnos=naked). Their ovules are covered with integument(s) but they remain exposed on the surface of the scale-like sporophyll, not enclosed within an ovary or a carpel. Formation of the (naked) seed requires five conditions: 1. Heterospory, that is, formation of distinct micro- and mega-spores. 2. Retention of only a single megaspore after meiosis. 3. Endosporous development of the embryo (young sporophyte), that is, the embryo develops within the megagametophyte, which is multicellular and provides the stored food. The mature megagametophyte may contain several archegonia. 4. Formation of an integument as an outgrowth of the sporophyll surface that encloses the ovule. 5. After fertilization, the ovule develops into a seed. The process is accompanied by conversion of megasporangial tissue into a nucellus that surrounds the ovule, and by conversion of the integument(s) into seed coat that encloses the complete package. Pollen and development of the male gametophyte. The male gametophyte develops as pollen grain. Unlike fertilization in mosses and ferns, the male gametes of gymnosperms travel by a combination of two unique means of transport. First, pollination, which is accomplished by various transport mechanisms of the pollen grain from the microsporangium to the megasporangium. This is followed by the formation of a pollen tube (male gametophyte), in which two male gametes (sperm) arise directly from the spermatogenous cell (antheridia are lacking in all seed plants). The pollen tube carries the two male gametes to the archegonium. Subsequently, one sperm of the pollen tube unites with the egg inside the archegonium, while the second sperm has no apparent function in gymnosperms. Thus, free water is no longer necessary for the sperm to reach the egg. Except for cycads and Gingko, which have flagellated sperm, the sperm of seed plants are non-motile. Structural diversity in gymnosperms. Living gymnosperms may be classified into four divisions, although they all have similar life cycles. CYCADOPHYTA (cycads, eg. Cycas, Zamia, Macrozamia) GINKGOPHYTA (Ginkgo) PINOPHYTA (very large group of conifers: eg. Araucaria, Pinus, Callitris) GNETOPHYTA (Gnetum, Ephedra, Welwitschia) BIOL2023/2923 Botany 3.10

55 Exercise B: Cycadophyta (allow 15 min) Observe various Cycadophyta in the lab. Living cycads consist of 11 genera and about 140 species. The present-day cycads are descendants of a cycad flora that was quite extensive during the age of the giant dinosaurs. Today's cycads are only a small remnant of a previously widespread group, being largely confined to isolated areas in the tropics and subtropics. The stems of cycads contain a vascular cambium and undergo secondary growth. A unique feature is a gigantic apical meristem, the biggest in the plant kingdom. Also, the motile male gamete, that measures up to 300 µm in diameter and is the largest found in the plant kingdom, is propelled by the flexing movements of about 40,000 flagella! Macrozamia is a native cycad of Australia. Its life cycle is similar to that of pine (a diagram of Pinus life cycle will be available in the laboratory or in the lecture). Below is a summary of the gametophyte development and fertilization in Macrozamia, accompanied by a set of diagrams on the opposite page. Additional details will be given in lectures. Development of male gametophyte ( Figures on next page ) 1. Unlike conifers, cycads are dioecious, that is, male and female cones are borne on separate plants. The microsporophylls bear clusters of microsporangia on their lower surface (Fig. 1), each containing numerous microspores (=pollen grains) (Fig. 2 ; TS microsporangium). 2. The pollen grain, which is carried by wind, is caught on the micropylar end of the ovule and then lodged in a pollen chamber above the nucellus (Fig. 8). 3. During germination, the pollen grain undergoes endosporic development including several consecutive divisions (Fig. 3, 4). The mature gametophyte consists of a tube cell, a sterile cell, a prothallial cell, and a spermatogenous cell that eventually forms two motile spermatozoids (Fig. 5). 4. Numerous pollen tubes grow into the nucellus, although most of them abort. The floor of the pollen chamber eventually disintegrates, thus facilitating entry into the archegonial chamber (Fig. 6). Development of female gametophyte 5. Two large ovules, each enclosed in an integument, are borne on the base of the megasporophyll (=ovulate scale) (Fig. 7). Development of the female gametophyte is also endosporic. In each ovule, meiotic divisions of a mother cell give rise to several cells, of which only one develops into a megaspore while the others abort. 6. The female gametophyte (=endosperm) is massive and parenchymatous (Fig. 8). It bears up to five archegonia near the archegonial chamber next to the pollen chamber (Fig. 9). Each archegonium contains an egg, a ventral canal cell, and two neck cells (Fig. 8a-c). Fertilization 7. The pollen tube bursts open and releases a pair of multiflagellate spermatozoids (Fig. 6). Fertilization, fusion of the spermatozoid with the egg, depends on a liquid drop secreted from the tube in which the spermatozoids swim. 8. Several archegonia are fertilized, although only one embryo normally develops into a mature seed. A pro-embryo is formed first: its basal part is meristematic and differentiates into the embryo; the apical part differentiates into a suspensor, which pushes the embryo deeper into the female gametophyte (details not shown). 9. After fertilization, the ovule enlarges many times its original size. The integument differentiates into an outer, fleshy layer and an inner stony layer (Fig. 10). Mature seeds are released from the disintegrating female cone, and germination occurs about 16 months from fertilization. BIOL2023/2923 Botany 3.11

56 BIOL2023/2923 Botany 3.12

57 There are two demonstration microscope slides of Macrozamia, showing important stages of the reproductive cycle. Examine demonstration slide LS ovule with mature archegonium [slide B469] on demo bench, and locate the following: pollen chamber, archegonial chamber, archegonia, and megasporangium/nucellus. B1. Draw a diagram of the ovule. Label which components are diploid & which are haploid. Examine demonstration slide LS pollen tubes in nucellus [slide 3] on demo bench showing the antherozoid nuclei and archegonial necks. B2. Identify all of the above features. Exercise C: Ginkgophyta (observe live plant in lab) Similarly to the cycads, the Ginkgophyta once had many genera and species. Today the only surviving relic is a single species, Ginkgo biloba, the maidenhair tree. It has been saved from extinction by the priests of ancient China and Japan who cultivated the trees in their temple gardens. The maidenhair tree is planted extensively for decoration now. The tree is deciduous, dioecious, with short "spur" shoots with microsporangiate cones produced by the males, and "long" shoots with ovuliferous structures produced by the females. BIOL2023/2923 Botany 3.13

58 Exercise D: Pinophyta (allow 1 hour) (observe live plant & demonstration materials) From the four divisions, the conifers are the largest and most widespread group, with about 500 species dominating vast areas of the northern cool temperate forests and representing about one third of all earth forest. The tallest vascular plant, the redwood (Sequoia), which grows up to 117 m tall and over 11 m in diameter, is a conifer. Other conifer genera include Abies (fir), Picea (spruce), Pinus (pine), and Araucaria (Norfolk Island pine), which is restricted to the Southern Hemisphere. This division also includes Wollemia nobilis (the Wollemi pine) that belongs to the family Araucariaceae. Morphological studies of wood anatomy DNA analysis suggest that the Wollemi is a new genus, falling between Agathis and Araucaria. There are various examples of gymnosperms and cones in the laboratory for you to examine. Also, there is a large, colour poster of Life of a Pine on display. The pictures and accompanying text describe the basic morphological and anatomical features of the organism and its life cycle. It is a simple way of learning the important principles of a gymnosperm life cycle. Below are specific tasks and questions for you to answer, as well as extra space for any additional notes you may wish to take. Examine the details of the structure of the male cone. D1. How would you describe the composition of the pine wood? D2. What is a "spur" shoot? Needle leaves? D3. Would you find both male and female cones on the same branch? D4. What are the sacs on the lower side of the microsporophylls? D5. Are the ovules located on the upper or the lower surface of the ovulate scales? D6. What are the components of the ovule before fertilization? D7. What is the nucellus? BIOL2023/2923 Botany 3.14

59 D8. Describe the pathway of the pollen tube during fertilization. D9. What is suspensor? D10. How does the mature seed travel? Exercise E: Gnetophyta (allow 5 min) (photos in lab) This small division consists of three distinct genera: Gnetum, which are lianas that climb to the tops of tall trees in tropical forest. Ephedra, is a profusely branched shrub with inconspicuous, scale-like leaves, which grows in desert areas. Welwitschia, which grows in southwestern African desert, is probably the most bizarre plant of all. Most of the plant is buried in sandy soil. The exposed part consists of a massive, woody, concave disk that produces only two strap-shaped leaves. Because the Gnetophyta have many angiosperm-like features, such as the presence of vessels in xylem, or double fertilization in Ephedra, they have been considered as evolutionary connecting links and the closest living relatives of the angiosperms. Most botanists today, however, regard them as specialized end-points of gymnosperm evolution. BIOL2023/2923 Botany 3.15

60 KEYWORDS Araucaria bifacial vascular cambium cambium (cambium=change) cone (konos=cone) conifer Pinophyta cork cambium Cycadophyta endospory (endon=within/inside) gamete (gametes=spouse) gametophyte generative cell/nucleus Gingko Gnetophyta gymnosperm (gymnos=naked) heteromorphic heterospory integument (inter=between) long shoot Macrozamia megaspore megaspore wall megagametophyte megasporophyll microspore microgametophyte microsporophyll nucellus ovule (ovum=egg) ovuliferous scale Pinus pollen chamber pollen grain pollen tube secondary meristem secondary phloem secondary xylem seed seed coat Sequoia short shoot sieve cell sperm (sperma=seed) spermatogenous cell sporophyte (sporos=seed) suspensor sterile cell tube cell/nucleus tracheid (trachia=windpipe) vessel (vas=vessel) xylem REVIEW QUESTIONS 1. Many conifers have only small scale- or needle-like leaves. Are these microphylls? Why? 2. How long can a gymnosperm needle live? 3. What types of stele do you find in gymnosperms? 4. What is the estimated age of the oldest living conifer? 5. What is cambium? A "bifacial" cambium? BIOL2023/2923 Botany 3.16

61 6. What is the difference between primary and secondary meristems? 7. What are the cell types in conifer xylem? 8. What is wood? 9. What are the components of conifer phloem? 10. What is bark? And its origin? 11. What are the structural components of a cone? 12. Are conifer cones similar to those of lycophytes? How do they differ? 13. What are the differences between conifer and cycad cones? 14. How can you tell the difference between microsporophyll and megasporophyll? 15. What is a pollen sac? Pollen grain? Pollen tube? 16. Which gymnosperms have pollen? Pollen tubes? 17. What is the function of the pollen tube? 18. Which gymnosperms have a motile sperm? Which do not? 19. What are the advantages of retaining the megaspore by the sporophyte? BIOL2023/2923 Botany 3.17

62 20. What parts of the seed provide nutrition to the embryo? 21. What does it mean "three generations under one roof"? 22. How would you define the ovule? 23. Who would you define the seed? 24. What is the meaning of "naked" seed? 25. What is the origin of the seed coat? 26. What is the origin of the integument? 27. How would you describe the evolutionary origin of integuments? 28. What is the origin of nucellus? 29. What is the function of the nucellus? 30. What is the largest living creature on Earth? BIOL2023/2923 Botany 3.18

63 Week Four SYSTEMATICS Aims In today s practical you will develop hypotheses about the evolutionary relationships of Eucalyptus and Angophora, by carrying out a phylogenetic analysis of selected species. Throughout the exercises you should be developing a working knowledge of the vocabulary of systematics. Objectives After completing this practical and the associated reading you should be able to: 1. do an analysis of characters to resolve relationships amongst Eucalyptus and Angophora. 2. devise a phylogenetic classification for Eucalyptus and Angophora. 3. interpret a cladogram. 4. correctly use the vocabulary of systematics (e.g. apomorphy, synapomorphy, plesiomorphy, ancestral, derived, character, character state, cladogram). 5. construct a phylogeny for a group of organisms of your choice. Introduction There is a wonderful diversity of life, both living and extinct. For biologists to easily communicate with each other about these organisms they need to be classified into groups. Ideally, the classification should be based on the evolutionary history of life, such that it predicts properties of newly discovered or poorly known organisms. Classification, however, is only one aspect of the much larger field of systematics. Systematics is an attempt to understand the evolutionary interrelationships of living things and systematists try to interpret the way in which life has diversified and changed over time. The aims of systematics can be summarised as: To discover ALL of the branches of the evolutionary tree of life. To document ALL changes that have taken place during the evolution of those braches To describe ALL of the species (the tips of the branches). Systematics then, is the study of the pattern of relationships amongst taxa (groups of organisms). Taxonomy is a subsidiary discipline of systematics and involves the theory and practice of describing and naming taxa and is currently based on a system developed by Linnaeus in the 18th century. Contemporary taxonomists attempt to make their classifications natural by making them congruent with hypotheses on the evolutionary relationships of the taxa they are concerned with. BIOL2023/2923 Botany 4.1

64 PRINCIPLES OF CLADISTICS Cladistics has been defined as the concepts and methods employed for the determination of the branching patterns of evolution (Stuessy 1990). The results of a cladistic analysis are represented by a branching diagram a cladogram - which, in turn, economically summarises a number of evolutionary trees or hypotheses. Optimally, cladograms are bifurcating networks which define nested sets of taxa on the basis of shared, derived characters. Furthermore, nested sets of taxa are, ideally, monophyletic; that is, they share a common ancestor. Thus, a cladogram may represent the phylogenetic relationships of the group under study. If a classification is based upon this branching pattern, the classification may, also, be thought to represent the phylogeny of the group, and, hence be a natural classification. Procedure: Cladograms can be represented as nested sets of taxa, with each set being defined by shared derived characters. For example the relationships illustrated as a nested set below can be represented by the following data set. A simple example of a set of characters and their states is outlined in the Table 1. 1, 2 4, 5 3 We will make the following assumptions about the data in Table 1: Each taxon is a species (they could equally be genera or above) of angiosperm. The five species form a monophyletic group (i.e. they share a common ancestor). Character state 0 = ancestral 1=derived. BIOL2023/2923 Botany 4.2

65 Characters Species A B C D E F G H I outgroup Table 1. A data matrix recording the states of nine characters (A-I) for five species (1-5) and an outgroup. Thus, by reference to Table 1 we can see that the species-group 1, 2, 3, 4 and 5 is defined by character A, the species-group 1 and 2 is defined by character B, the species-group 3, 4, and 5 is defined by character C (as well as by lacking character B) and species-group 4 and 5 is defined by character D. Characters E - I are unique to the species which possess them, therefore, these characters do not define relationships between species, they only define the species themselves. So, to construct a cladogram we first look for groups of taxa defined by shared characters, then, within those groups, identify sub-groups based on other shared characters. Things are not, however, that simple, for we must also decide whether the shared characters are derived or ancestral. In other words, we must be able to identify if an evolutionary change occurred along the branch that leads to each group of species. How can we distinguish ancestral from derived characters? We need to polarise the characters into ancestral and apomorphic derived states. Then we need to see if the derived states are shared by any of the species-groups. This is most commonly achieved by the use of an outgroup. BIOL2023/2923 Botany 4.3

66 The Outgroup Method Outgroups are chosen on the basis that they are assumed to be closely related to, but not members of, the ingroup. In the example we are using, species 1 to 5 are the ingroup, these might be species of angiosperms. The outgroup might be selected from within the gymnosperms. Characters are then selected (as in Table 1 above) and are polarised by with reference to the outgroup. Any character state that is present in both the ingroup and the outgroup is assumed to have evolved before these two groups diverged it is considered to be ancestral. Any character state that is found only within the ingroup is assumed to have evolved within the ingroup it is considered to be derived. Thus, we can simply and explicitly distinguish between ancestral characters states and derived characters states. The term plesiomorphic refers to ancestral characters states whereas the term apomorphic refers to derived character states. If an apomorphic state is shared by two or more species it is called a synapomorphy state (i.e. a shared, derived state). Conversely, if an ancestral state is shared by two or more species it is called a symplesiomorphic state (i.e. a shared ancestral state). Having identified shared, derived characters states we can now draw a tree which the direction of change of the characters states and, at the same time, depicts hypothetical evolutionary relationships amongst the species (Fig. 1) E F G H I D B C A Figure 1. A cladogram based on the data in Table 1 upon which the characters have been I mapped. I = shared derived characters (synapomorphies). II = unique characters (autapomorphies). NB: The outgroup is not shown in this figure. Having drawn a cladogram (Fig. 1), we can map the characters that define speciesgroups (i.e. synapomorphies) onto the appropriate branches of the cladogram. The theoretical position of characters unique to the terminal taxa (i.e. autapomorphies) can also indicated; note that these characters do not define relationships. BIOL2023/2923 Botany 4.4

67 Once the synapomorphies are mapped on, we can easily determine the phylogenetic length of our cladogram by counting the total number of evolutionary steps (characters state changes) required to give the particular cladogram branching pattern (usually referred to as a topology). In Figure 1 we can see that the length of the tree is 9 steps (i.e. 4 synapomorphies + 5 autapomorphies). We could, also, say that only four characters were informative (the four synapomorphies). Glossary of terms for Systematics Cladistics is rich in terminology, the definitions of which (and their philosophical implications) are the subject of some debate. The following is a selection of the most commonly encountered terms and their definitions: Apomorphic: An apomorphic character state is one that is not present in an outgroup but is present in one or more members of the ingroup. An apomorphic character is a derived characteristic. Autapomorphies (adj. autapomorphic) designates derived characters, or states, possessed by only one taxon in a set. Branch: a line that connects a clade or terminal taxon (a species for example) with a branch point (or node). (Also called an edge or internode) Character: any feature or attribute of an organism that can be used for identification, taxonomy, classification, or study of evolution. Characters can be drawn from many aspects of the biology of an organism, e.g. internal and external morphology, chromosomes, behaviour, DNA sequences, and many, many more. Characters consist of two or more states. Character state: The particular state that a character shows for a given taxon (e.g. present or absent) Clade: see monophyly. Homology: a term used for structures that share an evolutionary history. When we make observations on organisms we assume that similar structures are homologous. Characters and states are, therefore, homology hypotheses. Convergence: the development of similar characters or states in different lineages without a common ancestor. Homoplasy: defines resemblance not due to inheritance from common ancestry, this includes parallelism and convergence. Ingroup: The ingroup is the group of organisms that is of interest to the investigator and is the group that is examined with reference to the outgroup. Internode: a line connecting two nodes in a cladogram (see branch). Monophyletic: A group that includes an ancestor and all its descendents. Such groupings are referred to as clades. Natural taxon: a group (a clade) of organisms that is derived from a single ancestor (i.e. is monophyletic) and contains all of the descendants of that ancestor. BIOL2023/2923 Botany 4.5

68 Node: A branching point in a cladogram. Outgroup: any group used in the analysis that is not included in the ingroup. The outgroup is used as a reference point for examination of the ingroup. The outgroup dictates the ancestral character states. Parallelism: the development of similar characters or states separately in two or more lineages of common ancestry and on the basis of, or channelled by, characteristics of ancestry. Paraphyletic: A group that does not contain all descendants of a common ancestor. Parsimony: That the simplest sufficient hypothesis is the one that is to be accepted, even though others are possible. (Adjective: parsimonious) Phylogenetic tree: a branching structure that is believed to reflect evolutionary history. Phylogeny: the hypothetical evolutionary (historical) relationships among taxa; a phylogeny is a hypothesis about the evolutionary relationships amongst taxa. Plesiomorphic: a state is one that is present in the ancestor of a species and also its descendants. Therefore, a plesiomorphic state is an ancestral character. Plesiomorphic states do not tell us anything about evolution of the ingroup because the character has not changed from ancestor to descendant. (Noun: plesiomorphy) Plesiomorphic (or plesiomorphous) designates ancestral characters or states. Polyphyletic: Polyphyletic groups are artificial because the common ancestor of the group is placed in another group (e.g. taxa are placed in a group that does not share a common ancestor with other members). Root: the internode at the bottom of the cladogram and represents the outer limits of the phylogenetic hypothesis. Some approaches to phylogenetic reconstruction involve unrooted cladograms. Phylogenetic trees are often rooted by means of an outgroup. Sister group: any group (taxon or clade) that is phylogenetically most closely related to any other clade or taxon. It is helpful to use a sister group as the outgroup. Synapomorphy: A synapomorphy is an apomorphic character that is shared with another taxon. The identification of synapomorphy characters is the basis of phylogenetic inference. Symplesiomorphies (adj. symplesiomorphic): designates ancestral characters, or states, shared by two or more taxa. BIOL2023/2923 Botany 4.6

69 Biological nomenclature Our current system of nomenclature is based on a system invented by Linnaeus in the 18th century (the Linnean system). The scientific name of a species includes the name of the genus to which the species belongs (the generic name) and the species epithet. Together these are known as the species name. In our case, Homo is the name of the genus to which the species sapiens has been assigned. The first letter of the genus name is always capitalised; the first letter of the species epithet is always in lower case; and both names are italicised (when typed) or underlined (when hand-written). Each taxon has a name and a rank. The rank is a level in the hierarchical classification. There are rules designed to ensure that every taxon has a single, unambiguous name. Similarly, each name, at each rank from family and above has a particular ending that identifies the name as belonging to that rank the rules of nomenclature can be found in International Code for Botanical Nomenclature. Kingdom Phylum Class Order Family Genus Species Plantae Magnoliophyta (ophyta) Magnoliopsida (opsida) Proteales (ales) Proteaceae (aceae) Telopea Telopea speciosissima Figure 2. The hierarchical relationship between ranks for Telopea speciosissima (waratah). NB the endings for ranks of family, and above, are given parenthetically. BIOL2023/2923 Botany 4.7

70 Exercise 1: Phylogenetic relationships of Eucalyptus and Angophora In this exercise you will explore the pattern of relationships amongst Eucalyptus and Angophora. The principles used in this exercise can be applied to any organisms. STEP 1. COMPLETING THE MATRIX The first step is simply to record the character states for each character in the attached data matrix. We recommend that you examine more than one specimen or consult the posters as well as the specimens. Enter the species name and subgenus name of each specimen in the table. We can indicate evolutionary change by comparing character states in the ingroup (i.e. Eucalyptus and Angophora) with those in an outgroup (i.e. Arillastrum gummiferum). The character states that occur in both groups are deemed to have evolved before the separation of these two groups (i.e. are ancestral and coded 0); and those that are different in the ingroup and the outgroup are deemed to have evolved within the ingroup after their separation from the outgroup (i.e. are derived and coded 1). Arillastrum gummiferum, a species endemic to New Caledonia, is proposed here as the outgroup for this analysis. We could have also used the following species as outgroups: Arillastrum papuana, Allosyncarpia ternata, Stockwellia quadrifida and Eucalyptopsis alauda. Taxa NB. The infrageneric classification follows Pryor and Johnson A B Arillastrum gummiferum (outgroup) Angophora hispida Angophora floribunda G H Eucalyptus sideroxylon Subgen Symphyomyrtus Sect. Adnataria Eucalyptus paniculata Subgen. Symphyomyrtus Sect. Adnataria C Eucalyptus eximia Subgen. Corymbia Sect. Ochraria I Eucalyptus capitellata Subgen. Eucalyptus Sect. Renantheria D Eucalyptus gummifera Subgen. Corymbia Sect. Rufaria J Eucalyptus piperita Subgen. Eucalyptus Sect. Renantheria E Eucalyptus saligna Subgen. Symphyomyrtus Sect. Transversaria F Eucalyptus punctata Subgen. Symphyomyrtus Sect. Transversaria BIOL2023/2923 Botany 4.8

71 Characters NB. We have provided you with illustrations of ArilIlastrum gummiferum (Fig. 4). 1. Adult leaves: relative number of lateral veins: 0. few lateral veins 1. many lateral veins 2. Locule number: 0. 2 locules 1. >2 locules 3. Adult leaves: arrangement: 0. opposite 1. alternate 4. Perianth: whether calyptrate: 0. not calyptrate 1. calyptrate 5. Bud scar: 0. absent 1. present 6. Petals: whether absent: 0. present 1. suppressed 7. Inflorescence structure: 0. compound 1. simple 8. Anther shape: 0. oblong (cuboid) 1. reniform (kidney-shaped) 9. Anthers dehiscence: 0. opening by parallel slits (remaining separate) 1. not opening by parallel slits (confluent) 10. Cotyledon shape: 0. kidney-shaped (reniform) 1. not kidney-shaped 11. Anthers whether versatile: 0. versatile (pivoting) 1. not versatile (rigid) 12. Inflorescence position: 0. axillary 1. terminal 13. Bristle glands: 0. absent 1. present 14. Paratracheal parenchyma: 0. scanty 1. confluent 15. Large oil ducts in the pith: whether present: 0. absent 1. present 16. Fruit shape: 0. not urn-shaped 1. urn-shaped 17. Fruit: whether ribbed: 0. not strongly, longitudinally ribbed 1. strongly longitudinally ribbed BIOL2023/2923 Botany 4.9

72 Figure Bud showing scar left by calycine operculum (E. cinerea). 2. LS of bud showing inflexed stamens (E. sideroxylon). 3. LS of bud (E. gummifera) showing calycine operculum (S) and Coraline operculum (P). 4. TS of flower showing the 5-loculate gynoecium (E. saligna). BIOL2023/2923 Botany 4.10

73 Figure 4. Arillastrum gummiferum. 1. Buds. 2. LS flower (note outer staminodes). 3. Petal. 4. Stamen (anterior view). 5. Stamen (posterior view). 6. Leaf. 7. TS gynoecium. 8. Cotyledon. 9. Flower. og - oil gland, hr hypanthial rime, is inner stamens (after J. Dawson, 1970). BIOL2023/2923 Botany 4.11

74 Figure 5. The floral morphology of Eucalyptus. Taken from Figs. 81, 82 & 84 (Clarke and Lee, 1987). BIOL2023/2923 Botany 4.12