Asteraceae

605

Inflorescence and floral ontogeny in Osteospermum ecklonis (Asteraceae)

Botany Downloaded from www.nrcresearchpress.com by CAUL on 09/17/11 For personal use only.

Mohammad Reza Dadpour, Somayeh Naghiloo, and Gholamreza Gohari

Abstract: Development of the capitulum inflorescence with different types of florets in Asteraceae is an interesting issue in the field of plant evolution and development. In this study, ontogeny of the inflorescence and florets of Osteospermum ecklonis (DC.) Norl., an ornamental and evergreen subshrub, was investigated using epi-illumination light microscopy. The initiation and subsequent development of florets on the highly convex inflorescence apex occurred acropetally, except for the ray florets, which showed a lag in initiation. Organogenesis in disc florets started with unidirectional initiation of corolla lobes from the adaxial side and then proceeded by simultaneous appearance of five stamen and finally two median carpel primordia. Significant developmental features included the lack of pappus differentiation, formation of nonsyngenesious stamens, and formation of the ovule-less ovary. Ray florets showed significant differences from disc florets as reflected by the zygomorphic shape of floral apex and shift of floral merosithy from pentamery to tetramery. Also, expansion of corolla lobes to form the ligule and the formation of staminodia were observed. It is hypothesized that the actinomorphic pentamerous disc florets are most primitive among the family from which the tetramerous ray florets are derived. Accordingly, ray florets evolved from disc florets under long-term selective pressure and play a crucial role in enhancing reproductive success. Key words: Asteraceae, Osteospermum ecklonis, inflorescence and floral ontogeny, epi-illumination technique. Résumé : Chez les Asteraceae, le développement d’inflorescences en capitule formées divers types de fleurons constitue un sujet intéressant dans le champ évo-dévo. Les auteurs ont étudié l’ontogénie de l’inflorescence et des fleurons chez l’Osteospermum ecklonis (DC.) Norl., un sous arbuste ornemental décidu, en utilisant la microscopie photonique en épi-illumination. L’initiation et le développement subséquent des fleurons sur l’apex fortement convexe de l’inflorescence s’effectuent de façon acropétale, sauf pour les fleurs ligulées montrant un délai d’initiation. L’organogenèse chez les fleurs tubulées commence avec une initiation unidirectionnelle des lobes de la corolle à partir du côté adaxial et procède ensuite avec l’apparition simultanée de cinq étamines et finalement de deux primordiums de carpelles médians. Les caractères ontogéniques significatifs incluent l’absence de différenciation d’aigrettes, la formation d’étamines syngénésiques, et la formation d’ovaire sans ovule. Les ligules montrent des différences significatives par rapport aux fleurs tubulées, reflétées par la forme zygomorphe de l’apex floral et une réduction du nombre de verticilles de cinq à quatre. De plus, on observe l’expansion des lobes de la corolle pour former la ligule et la formation de staminodes. On propose l’hypothèse que les fleurs tubulées pentamères actinomorphes seraient plus primitives dans la famille dont les fleurs ligulées tétramères sont dérivées. Conséquemment, les fleurs ligulées auraient évolué de fleurs tubulées sous une pression sélective à long terme, et joueraient un rôle crucial pour augmenter le succès de la reproduction. Mots‐clés : Asteraceae, Osteospermum ecklonisi, inflorescence et ontogénie florale, technique d’épi-illumination. [Traduit par la Rédaction]

Introduction The Asteraceae, comprising about 1600 genera and 25 000 species, is one of the largest families of flowering plants (Anderberg et al. 2007). The family has a worldwide distribution and is mainly found in the tropical and subtropical regions of North America, the Andes, eastern Brazil, southern Africa, the Mediterranean region, central Asia, and southwestern China. This family contains several economically important species used as a source of food (e.g., Cynara scolymus, or artichoke, and Helianthus annuus, or sunflower; Dempewolf et al. 2008). Furthermore, the Asteraceae includes some fa-

miliar ornamental plants such as Zinnia, Dahlia, and Chrysanthemum. The inflorescences and flower types of the Asteraceae present distinct features rarely found in other angiosperm families. Inflorescences are composed of morphologically different types of flowers tightly packed into a flower head (capitulum) that overtly resembles a single flower. The capitulum is surrounded by one or several layers of involucral bracts resembling the calyx of other flowers. The epigynous, bisexual, or unisexual flowers have a calyx known as the pappus, which is sometimes modified as awns, scales, or capillary bristles. The sympetalous corolla is classified as

Received 2 March 2011. Accepted 4 July 2011. Published at www.nrcresearchpress.com/cjb on 6 September 2011. M.R. Dadpour and G. Gohari. Department of Horticultural Science, Faculty of Agriculture, University of Tabriz, Tabriz, Iran. S. Naghiloo. Department of Plant Biology, Faculty of Natural Sciences, University of Tabriz, Tabriz, Iran. Corresponding author: Mohammad Reza Dadpour (e-mail: [email protected]). Botany 89: 605–614 (2011)

doi:10.1139/B11-052

Published by NRC Research Press


606

three structural types: (i) bilabiate (zygomorphic corolla with a short tube having upper and lower lips); (ii) disk (actinomorphic corolla with short to elongate tube bearing generally 5 teeth-like or elongate lobes); and (iii) ray or ligulate (zygomorphic corolla with generally short tube having elongate, flat extension with 3–5 apical teeth). Heads of many asteraceaen taxa have a mixture of central actinomorphic disk flowers and peripheral ray flowers. The alternipetalous, usually syngenesious stamens (with anthers connate but filaments distinct) form a tube surrounding the syncarpous gynoecium (Anderberg et al. 2007). Owing to these specific features, the large family Asteraceae has historically presented intriguing problems for ontogenetic studies. Even though the Asteraceae has been the object of numerous physiological studies (Maksymowych 1990), ontogenetic investigations have received little attention. The most complete developmental study on reproductive structures of Asteraceae is that of Harris (1995), who assembled a thorough investigation of early inflorescence and floral ontogeny within the family. Although this work presented some new prospects about the inflorescence and flower development in the Asteraceae, important developmental aspects, namely the nature and development of the pappus and corolla, as well as the evolution of different floret types, have yet remained ambiguous and herald a renaissance in the study of floral and inflorescence ontogeny in the Asteraceae. In the present study, we included a complete analysis of the inflorescence and flower ontogeny of Osteospermum ecklonis (DC.) Norl. as a representative member of the Asteraceae. Osteospermum, an ornamental genus native to South Africa, comprises evergreen subshrubs or herbaceous species that have been developed into successful pot and garden plants. The genus belongs to the Calenduleae, one of the small tribes of the Asteraceae that is taxonomically fairly well understood at the species level (Norlindh 1977; Nordenstam 1996, 2006; Nordenstam et al. 2006). Osteospermum is the most problematic genus of this tribe. In spite of recent taxonomic rearrangements in the genera, it remains heterogeneous, and future changes in generic taxonomy are inevitable (Barker et al. 2009). Regarding the importance of developmental data in the burgeoning field of phylogenetic analysis, ontogenetic information resulting from this study would be applicable either by adding new characters for comparison or by testing other characters as a consequence of evaluating the homology of the various character states. The aims of this study were (i) to explain the complete floral and inflorescence ontogenesis in the representative member of Asteraceae, O. ecklonis; (ii) to investigate the ontogenetic differences and similarities between ray and disc florets; and (iii) to compare the resulting ontogeny with the developmental data found for other Asteraceae. The current study is part of an ongoing research program that will continue to provide insight into several fundamental questions about floral and inflorescence evolution in the Asteraceae.

Materials and methods Source of materials Inflorescences of O. ecklonis at different developmental stages were collected periodically (from 1 June 2009 until 30 December 2009) from the northwest region of Iran (Tab-

Botany, Vol. 89, 2011

riz, altitudes above 2200 m). The specimens were determined and voucher specimens deposited at the herbarium of Tabriz University (Tabriz, Iran). The materials were fixed in formalin : glacial acetic acid : 50% ethanol (5:5:90 by volume) for at least 24 h. The specimens were then rinsed and dehydrated in 70% and 95% ethanol, respectively. Afterwards, they were stained with alcohol-soluble nigrosin (0.6% nigrosin in 95% ethanol) for at least 2 days according to the method reported by Charlton et al. (1989) with some modifications. The samples were washed in 95% ethanol before morphological microscopic examination. Epi-illumination light microscopy Ontogeny of the inflorescence and floral organogenesis in O. ecklonis was studied using a digital version of epi-illumination light microscopy (Dadpour et al. 2008). The prepared samples were dissected under a SMZ1500 stereomicroscope (Nikon, Tokyo, Japan) and viewed by a Nikon E600 reflected light microscope in dark field mode while the samples were submerged in absolute ethanol. To display all details of specimen in an image, depth of focus was improved automatically using Z-stack acquisition and composition (Dadpour et al. 2008). For this purpose, an image series (Z-stack) of each specimen was acquired by means of a DXM1200F high-resolution digital camera (Nikon) and then processed according to methodology introduced by Peighambardoust et al. (2010). Final digital images were then trimmed and saved as TIF files in grayscale mode (8 bits, 256 levels) with a resolution of 2560 × 2048 pixels.

Results Organography This herbaceous shrub, or subshrub, produces 90–120 flowers on radiate capitula surrounded by 1–3 layers of involucral bracts. The inflorescence has a typical daisy shape represented by tubular central disc florets and a ring of ligulated ray florets (Fig. 1). The female ray florets are white with contrasting blue spots on the base of the corolla’s adaxial surface (arrow). These florets have a markedly forked style and 4 staminodes at the base of the style. The disc florets are pseudobisexual and may vary in color from metallic blue to yellow. Disc florets have 5 stamens, corresponding to the number of corolla lobes, with free filaments and anthers, unlike the syngenesious stamens of most Asteraceae. The style in the disc florets is minutely bifid. The species produces acutely 3-angled achenes with very narrow wings. Inflorescence The shoot apex of O. ecklonis, which was hemispheric at the vegetative state (Fig. 2A), then broadened to attain a conical shape as it differentiated into the inflorescence apex (Fig. 2B). After helical inception of a set of involucral bracts, onset of capitulum development was marked by the formation of the first disc floret primordia in the axil of the smallest and innermost phyllaries (Fig. 2C). Then the initiation of disc florets continued acropetally in orderly parastichies until occupation of the entire inflorescence apex (Figs. 2D, 2E). Primordia were arranged along 13 dextrorse and 21 sinistrorse parastichies. Ray floret primordia were somewhat delayed in their initiation until almost three layers of disc Published by NRC Research Press

Dadpour et al.


Fig. 1. The inflorescence of Osteospermum ecklonis showing the peripheral ray and central disc florets. Arrow indicates blue spots at the corolla base of a ray floret.

607

cells along the rim resulted in early differentiation of style and stigma (Figs. 3H, 3I). Organ development in disc florets By mid-development, the corolla lobes grew over the stamen primordia and then arched inward to cover the interior of the flower (Figs. 4A, 4B). At this time, the pappus developed as small encircling epidermal outgrowths at the top of the ovary. When the disc florets were ca. 0.9 mm long, multicellular trichomes differentiated on the base of the corolla tube (Fig. 4C). At maturity, the disc floret corolla tube was continuous for two-thirds of its length (Fig. 4J), the petal lobes recurving outward at anthesis. Differentiation of the stamen primordia began with distal broadening (Fig. 4D) that yielded the basifixed large anther with the short filament (Fig. 4E). Development of a median abaxial groove and lateral grooves delimited the microsporangia (Fig. 4F). Simultaneously, the anther orientation changed to the dorsifixed state. In mature flowers, short, laciniate acute appendages were formed in apical and basal portions of the anther (arrowheads, Fig. 4I). The carpel grew upward as a straight structure and developed into an inferior ovary, a slightly bilobed style, and a round stigma (Figs. 4G, 4H). At this stage, no sign of ovule formation was observed in the single locule of the ovary.

florets emerged (Fig. 2G). One to two ray floret primordia arose at the base of each parastichy formed by the disc floret primordia, resulting in a mean of 13 rays per capitulum (Figs. 2H, 2I). The lowermost disc floret primordia were the first to begin organogenesis, which then proceeded acropetally on the head (Fig. 2E). Shortly thereafter, ray floret primordia also started organogenesis (Fig. 2H). By the time the inflorescence diameter reached 2.2 mm, both ray and disc florets had completed organogenesis (Figs. 2F, 2I). The receptacle was yet conical at this stage, and the conical shape was retained through late development when the florets were undergoing final corolla expansion prior to anthesis. Organogenesis in disc florets Each floret apex was flattened at first and sank slightly as the corolla ring meristem became distinguishable (Fig. 3A). After formation of this ring meristem, the corolla lobe primordia arose unidirectionally, with the adaxial lobes usually appearing first (Fig. 3B). Next to be initiated were the two lateral corolla lobe primordia (Fig. 3C), followed by the abaxial primordium (Fig. 3D). Concurrently with the formation of the lateral corolla lobes, the five stamen primordia emerged simultaneously in the alternipetalous position (Fig. 3C). Although a surrounding rim became evident at the base of the corolla soon after the inception of the stamen primordia (Fig. 4A), pappus lobes were not produced (Figs. 4B, 4C). Finally, two carpel primordia were initiated at the median position (Figs. 3H, 3I). At the same time that stamen primordia were enlarging (Figs. 3E, 3F), the first sign of carpel initiation appeared. At this time, a central hollow surrounded by a ring of cells was observed at the base of the floral cavity that led to formation of the inferior ovary (Fig. 3G). Subsequent enlargement of

Organogenesis in ray florets The ray florets began organogenesis shortly after neighboring disc floret primordia. The apex was deepened, and a meristematic ring initiated that then developed into the corolla lobes (Fig. 5A). The lateral corolla lobe primordia were the first to appear (Fig. 5B) followed by the abaxial and adaxial lobes (Figs. 5C, 5D). Soon after that the corolla lobes began to expand and arch inward (Fig. 5E), four stamen primordia originated synchronously in the next floral whorl, alternate to the petals (Fig. 5G). At about the same time as the appearance of the stamen primordia, two carpels were initiated (Figs. 5G, 5H). Shortly thereafter the pappus ring was formed (Fig. 6B), encircling the ray floret at the base of the corolla. However, no individual pappus primordia differentiated from this ring during floral development (Fig. 6C). Organ development in ray florets After differentiation of the corolla lobes, they rapidly incurved and grew to cover the inner parts of the floret (Figs. 5E, 6A). Although at first four clear petal lobes developed from the corolla ring meristem, the adaxial lobe was immediately suppressed (Fig. 5E) so that the ligule of the ray floret was formed by the zonal growth of the three remaining lobes (Fig. 5F). By late development, numerous multicellular trichomes emerged from the epidermal cells of the corolla base (Figs. 6C, 6I, 6J). The lateral margins of the corolla tube remained incurved until just before anthesis (Fig. 6J). The stamen primordia enlarged somewhat (Figs. 5H, 5I), but soon ceased development, and the elongating style overtook them in size (Figs. 6D, 6E). In mature ray florets, they were visible as small relictual staminodia at the base of the style (Figs. 6F, 6G). Published by NRC Research Press

608



Fig. 2. Inflorescence initiation and development. (A) Polar view of vegetative apices producing helically arranged leaf primordia. (B) Conversion to the reproductive phase with broadening and concaveness of apical meristem. (C) Initiation of involucral bract primordia and floral primordia along the periphery of inflorescence meristem. (D) Initiation of florets in an acropetal sequence. (E) Polar view of an inflorescence showing completion of floral initiation. Florets begin organogenesis. (F) A mature capitulum with all flowers having completed organogenesis. (G–I) Side view of capitulum in different stage of development showing initiation and subsequent development of ray florets (arrowheads) at the base of each of the disc floret parastichies. F, floral primordia; B, involucral bracts; IM, inflorescence meristem; LP, leaf primordia; SAM, shoot apical meristem.

The carpel primordia enlarged and then underwent differentiation into inferior ovary and markedly bifurcated style (Fig. 6E). The stigma appeared as two stigmatic lines on the adaxial surface of style branches. At mid-development, the formation of an ovule primordium began within the inferior ovary (Fig. 6H).

Discussion Inflorescence The diversification of Asteraceae contributed to their achieving the status of being the largest family of flowering

plants, partly owing to their possession of the capitulum inflorescence. Putative advantages of a capitulum inflorescence are greater attraction to insect pollinators, better protection for the seeds, and a potential capacity for producing more recombinants (Stebbins 1967). On the basis of floret type, different groups of capitula could be recognized (Bremer 1994). In O. ecklonis, a heterogamous radiant capitulum was found with actinomorphic disc florets and few zygomorphic ray florets. Regarding the absence of a strictly terminal floret or other structure in the head (Lawalrée 1948), it is widely accepted that the capitular inflorescence derives from the condensation and thickening of the axis of an indeterminate Published by NRC Research Press

Dadpour et al.

609


Fig. 3. Organogenesis in disc florets. Abaxial side is at the left in all figures. The corolla has been removed in F–I to show the developing stamens and carpels. (A) Formation of the corolla ring meristem. (B) Initiation of first corolla lobes (P1) on the adaxial side. (C) Inception of lateral corolla lobes (P2) and appearance of five stamen primordia on the concave floral apex alternating with the corolla lobes. (D) Formation of abaxial corolla lobe. (E) Polar view showing enlargement and arching of corolla lobes. (F) Polar view of stamen primordia after removal of petals. (G) The first sign of initiation of inferior ovary with the formation of a hollow at the base of floral concavity. (H, I) Enlargement of carpel primordia to form style and stigma. A, stamen; P, petal; C, carpel.

raceme or spike inflorescence (Burtt 1977; Jeffrey 1977; Weberling 1989). Throughout the Asteraceae, it has been judged that initiation of flowers begins on the periphery of the inflorescence meristem and proceeds acropetally, resulting in a single developmental shift between ray identity and disc identity for floral meristems (Cronquist 1955). However, this assumption has been refuted by recent developmental studies. Departure from the strictly acropetal plan were found in species with heteroga-

mous (radiate) capitula, in which the initiation and development of ray florets is delayed with respect to at least the first series of disc floret primordia (Harris et al. 1991; Harris 1995). In fact, in heterogamous capitula, the order is often mixed, being partly acropetal (regarding disc florets) and partly basipetal (regarding ray florets). Similar delay in development of ray florets is also observed in heterogamous capitula of O. ecklonis. In this species, floret primordia were arranged along 13 dextrorse and 21 sinistrorse parastichies. It Published by NRC Research Press

610



Fig. 4. Organ development in disc florets. (A) Appearance of the pappus ring meristem below the corolla (arrowheads). (B) Lateral view of flower with corolla lobes growing and covering the inner organs. (C) Trichomes begin to form at the base of the corolla tube. (D) Lateral view of stamens showing their apical expansion. (E) Differentiation of stamens to distinct anther and filament. (F) A lateral view of flower showing the free anthers and filaments. (G) Lateral view of carpel inception. (H) A flower at mid-development showing the inferior ovary, style, and stigma. No ovule has formed in the ovary. (I, J) Elongation of stamens and corolla shortly before anthesis. Stamen appendages and multicellular trichomes at the base of corolla tube are distinct (arrowheads). A, stamen; C, carpel; O, ovary; P, petal; Sl, style; St, stigma.


Dadpour et al.

611


Fig. 5. Organogenesis in ray florets. Abaxial side is at the base in all figures. The corolla has been removed in G–I to reveal the developing stamens and carpels. (A) Concaveness of floral apex and formation of corolla ring meristem. (B) First corolla lobes appearing in lateral position. (C, D) Initiation of abaxial and adaxial corolla lobes. (E) Polar view of incurved corolla lobes showing reduction of adaxial corolla lobe. (F) Expansion of three remaining corolla lobes to form the ligule. Stomata are forming on the corolla lobes (arrowheads). (G) Polar view showing initiation of stamens alternating with petals and carpels in median position. (H, I) Enlargement of carpels in polar view. A, stamen; P, petal; C, carpel.

is suggested that this Fibonacci pattern in the asteraceaen inflorescence optimizes access to moisture, rainfall, and sunlight. Comparison of ontogeny with the other Asteraceae Despite major structural differences between the two types of floret, the order of floral whorl initiation was not associated with flower type. In both disc and ray florets, the order of whorl initiation was corolla, androecium, and gynoecium, respectively. Although a ring meristem of pappus initiated concurrently with the gynoecium, no pappus lobes differentiated

from this rim. The pappus is an important characteristic structural feature of the Asteraceae. This structure aids in dispersal or defense against herbivory (Stuessy and Garver 1996). Three basic states regarding pappus initiation have been reported in the Asteraceae: complete absence of a pappus; development of five normal pappus lobes; and initiation of two or numerous pappus lobes (Harris 1995). For a long time, the morphological nature of the pappus has remained ambiguous. There have been two theories of the morphological nature of the pappus. In the phyllome theory, the pappus has been considered a Published by NRC Research Press

612



Fig. 6. Organ development in ray florets. (A, B) Pappus ring (arrowheads) initiating on the flank of the flower primordium at the base of corolla. (C) Corolla lobes are growing and forming trichomes at the base (arrowhead). (D, E) Lateral view of flowers showing the continued elongation of the carpels. Stamen primordia (indicated with an asterisk) have nearly stopped their growth. (F) Lateral view of a mature flower in which stamen primordia are becoming clearly staminodial, enlarging distally only slightly. (G) Higher magnification of staminodia in (F) at the base of style. (H) Lateral view of a mature flower showing the formation of the basal ovule primordium in the single locule. The two style branches are distinct. (I, J) Side view of abaxial and adaxial portions of corolla prior to anthesis showing ligule, which curves inward. Multicellular trichomes are abundant at the base of both abaxial and adaxial surface of corolla (at arrowheads). C, carpel; L, ligule; Ov, ovule; P, petal; Sl, style; St, stigma.

more or less modified calyx, while according to another theory, the pappus has the nature of trichomes (Small 1919; Tiagi and Taimni 1963). Based on recent anatomical and morphological analysis, it has been shown that the pappus is a calyx that has undergone modifications in several ways. It is suggested that the pappus in the ancestral Asteraceae consisted of five sepal-like members. Two major lines of specialization can be established in this family from a primitive type of fivelobed pappus. In one line of evolution there has occurred an amplification of the number of whorls and number of members in the whorls of the pappus. In agreement with this hy-

pothesis, in the species with an amplified number of pappus members (i.e., Tragopogon portensis), calyx inception begins with the five original pappus primordia, which then connect basally by a rim, and more pappus primordia are formed centrifugally on this rim (Sattler 1973). In the other line of evolution there has been reduction in the number of pappus members from five to a total loss of pappus along with its vascular supply. Accordingly, the situation observed in O. ecklonis is derived from the primary five-lobed pappus by the suppression of pappus members and therefore could be regarded as an advanced feature. Published by NRC Research Press


Dadpour et al.

During the differentiation of the corolla lobes from a ring meristem, unidirectional initiation from the adaxial side was observed, contrasting with the reported sequential order in Osteospermum fructicosum (Harris 1995). In most Asteraceae, the corolla is initiated as a ring meristem (meristematic rim or ring wall) that develops before the synchronous initiation of the corolla-lobe primordia upon it (Leins and Erbar 2000; Lacroix et al. 2007), although in the non-asteroid tribes an irregular successional development of the corolla-lobe primordia has been reported in the development of bilabiate and ligulate corollas (Harris 1995). The members of the stamen whorl appeared more or less simultaneously in O. ecklonis in contrast to the sequential initiation of stamens that was observed in O. fructicosum (Harris 1995). In other Asteraceae, a spiral or unidirectional sequence of stamen initiation has been observed in addition to a simultaneous order of initiation (Harris 1995). Initiation of the gynoecium, as in most angiosperms with inferior ovaries, showed an appendicularepigynous ground plan in which a distinctive concavity appears in the center of the floral apex during the inception of the perianth (Kaplan 1967; Leins 1972; Magin 1977). However, in O. ecklonis, epigyny is the result of the formation of a secondary hollow in the depth of the floral cavity, which then was followed by the inception of stylar primordia on the edge of the hollow, while based on the appendicularepigynous plan, the inferior position of the ovary is due to deepness of the concavity and initiation of carpels below the stamen primordia. The gynoecium in all members of the Asteraceae consists of two styles and stigmas, whereas there is only one locule, ovule, and placenta. Therefore, the ovary is basically pseudomonomerous, and the two styles are interpreted as ancestral vestiges of a two-carpellate pistil that became evolutionarily reduced to a single ovuled and loculed structure (Simpson 2006). During organ development, significant characteristics of the asteraceaen flower have been found in O. ecklonis. However, contrary to observations for most other Asteraceae, there is no evidence of postgenital fusion of anthers. As a diagnostic character of asteracean florets, anthers are connivent around the style, and this arrangement typically functions as a presentation mechanism for animal pollination. Comparison between ray and disc florets Ontogenetic analysis of ray and disc florets confirms the existence of major structural differences between the two types of floret in terms of both floral merosity and development of the corolla, androecium, and gynoecium. There were four corolla lobes and stamen primordia in ray florets versus five in disc florets. Anatomical studies in some members of the Asteraceae (Actinomeris aquarrosa and Bidens biternata) demonstrated that tetramerous florets in the Asteraceae are derived from pentamerous florets by cohesion or reduction of alternipetalous strands (Singh and Agarwal 2001). Based on this issue, the actinomorphic pentamerous disc florets are the most primitive among the family from which the ray, ligulate, and neutral florets (often with tetramerous ground plan) are derived. There are marked differences between the two types of floret in corolla development. In disc florets, zonal growth between corolla lobes produced a corolla tube with five equal

613

lobes at the end. In spite of initiation of four corolla lobes in ray florets, the adaxial lobe was suppressed soon after, and zonal growth of the three remaining lobes resulted in the formation of the ligule. Multicellular uniseriate trichomes on the corolla of mature ray florets were much more pronounced than those of disc florets. These trichomes probably aid in fruit dispersion by the wind, in the absence of a pappus. Ray florets also exhibited the formation of stomata on the proximal side of the corolla that were not observed in disc florets. It has been recently suggested that stomata on petal surfaces have an important physiological role in petal movement (Azad et al. 2007). Whereas normal development of stamens occurred in disc florets, ray florets displayed intriguing variations related to stamen development. All four stamen primordia cease their growth after some enlargement and are noticeable as relictual staminodal structures in adult stages. Such a phenomenon was reported in some related species such as Chrysanthemoides monilifera (Calenduleae: Asteroideae) and Dresslerothamus spp. (Senecioneae: Asteroideae). In some members of Asteraceae only two stamen primordia are initiated, as in Tithonia aotundifolia (Heliantheae: Asteroideae), and these are aborted and resorbed. Some species, such as Rudbeckia laciniata (Heliantheae: Asteroideae), show no evidence whatsoever of stamen initiation (Harris 1995). Based on the hypothesis that was proposed by Tucker (1988), organ loss in one organ whorl tends to disrupt the next successive whorl, whereas such a tendency does not exist in the case of organ reduction. Our previous work in the flower of Xeranthemum, in which stamen primordia disappeared after initiation, showed that the development of carpels was atypical and no ovule was formed by the ovary. However, in O. ecklonis the stamens remain as staminodes, and the subsequent ontogenetic cascade proceeded normally with the initiation of the two carpels that produced an ovule. It seems that differences between ray and disc floret development are related to their location on the capitulum. Accordingly, it is suggested that ray florets evolved from disc florets under long-term selective pressure and play a crucial role in enhancing reproductive success. Perspective to future systematic analysis There is ample evidence of considerable variation within the genus Osteospermum. On the basis of both internal transcribed spacer and cpDNA gene trees as well as the combined analysis, Osteospermum is paraphyletic, with many placements that are not well supported (Barker et al. 2009). Ontogenetic studies could be helpful in the solution of the taxonomic problem of the genus owing to the addition of new characters for phylogenetic analysis or the testing of other characters by assessing the homology of the various character states. Unfortunately, few ontogenetic studies have been done in this genus, like for other Asteraceae. However, our results revealed that there are differences in the order of organ initiation between our studied species (O. ecklonis) and a previously reported one (O. fructicosum, Harris 1995). It is suggested that this character could be helpful for the taxonomic comparison of the species within the genus. For this purpose, it is necessary to investigate floral ontogeny in representative species of all sections to broaden the data set on Published by NRC Research Press

614

the basis of which a detailed phylogenetic analysis would be possible.

Acknowledgements This work was funded in part by grants from Research Affairs of the University of Tabriz, whose support is appreciated. We would like to thank the anonymous reviewers and the editor for their extensive and valuable comments and corrections.


References Azad, A.K., Sawa, Y., Ishikawa, T., and Shibata, H. 2007. Temperature-dependent stomatal movement in tulip petals controls water transpiration during flower opening and closing. Ann. Appl. Biol. 150(1): 81–87. doi:10.1111/j.1744-7348.2006.00111.x. Barker, N.P., Howis, S., Nordenstam, B., Källersjö, M., Eldenäs, P., Griffioen, C., and Linder, H.P. 2009. Nuclear and chloroplast DNA-based phylogenies of Chrysanthemoides Tourn. ex Medik. (Calenduleae; Asteraceae) reveal extensive incongruence and generic paraphyly, but support the recognition of infraspecific taxa in C. monilifera. S. Afr. J. Bot. 75(3): 560–572. doi:10.1016/j. sajb.2009.05.006. Bremer, K. 1994. Asteraceae: cladistics and classification. Timber Press, Portland. Burtt, B.L. 1977. Aspects of diversification in the capitulum. In The biology and chemistry of the Compositae. Edited by V.H. Heywood, J.B. Harborne, and B.L. Turner. Academic Press, London. pp. 41–59. Charlton, W.A., Macdonald, A.D., Posluszny, U., and Wilkins, C.P. 1989. Additions to the technique of epi-illumination light microscopy for the study of floral and vegetative apices. Can. J. Bot. 67(6): 1739–1743. doi:10.1139/b89-220. Cronquist, A. 1955. Phylogeny and taxonomy of the Compositae. Am. Midl. Nat. 53(2): 478–511. doi:10.2307/2422084. Dadpour, M.R., Grigorian, W., Nazemieh, A., and Valizadeh, M. 2008. Application of epi-illumination light microscopy for study of floral ontogeny in fruit trees. Int. J. Bot. 4(1): 49–55. doi:10.3923/ ijb.2008.49.55. Dempewolf, H.L., Rieseberg, H., and Cronk, Q.C. 2008. Crop domestication in the Compositae: a family-wide trait assessment. Genet. Resour. Crop Evol. 55(8): 1141–1157. doi:10.1007/ s10722-008-9315-0. Harris, E.M. 1995. Inflorescence and floral ontogeny in Asteraceae: a synthesis of historical and current concepts. Bot. Rev. 61(2–3): 93–278. doi:10.1007/BF02887192. Harris, E.M., Tucker, S.C., and Urbatsch, L.E. 1991. Floral initiation and early development in Erigeron philadelphicus L. (Asteraceae: Astereae). Am. J. Bot. 78(1): 108–121. doi:10.2307/2445234. Jeffrey, C. 1977. Corolla form in Compositae — some evolutionary and taxonomic speculations. In The biology and chemistry of the Compositae. Edited by V.H. Heywood, J.B. Harborne, and B.L. Turner. Academic Press, London. pp. 111–118. Kadereit, J.W. and Jeffrey, C. (Editors) 2007. The families and genera of vascular plants. Vol. VIII. Flowering plants, eudicots, asterales. Springer, Berlin. Kaplan, D.R. 1967. Floral morphology, organogenesis and interpretation of the inferior ovary in Downingia bacigalupii. Am. J. Bot. 54(10): 1274–1290. doi:10.2307/2440367.

Botany, Vol. 89, 2011 Lacroix, C.R., Steeves, R., and Kemp, J.F. 2007. Floral development, fruit set, and dispersal of the Gulf of St. Lawrence Aster (Symphyotrichum laurentianum) (Fernald) Nesom. Can. J. Bot. 85(3): 331–341. doi:10.1139/B07-011. Lawalrée, A. 1948. Histogénèse florale et végétative chez quelques Composées. Cellule, 52: 214–294. Leins, P. 1972. Das Karpell im ober- und unterstandigen Gynoeceum. Ber. Dtsch. Bot. Ges. 85: 291–294. [In German.] Leins, P., and Erbar, C. 2000. Die frühesten Entwicklungsstadien der Blüten bei den Asteraceae. Bot. Jahrb. Syst. 122: 503–515. [In German.] Magin, N. 1977. Das Gynoeceum der Apiaceae: Model und Ontogenie. Ber. Dtsch. Bot. Ges. 90: 53–66. [In German.] Maksymowych, R. 1990. Analysis of growth and development of Xanthium. Cambridge University Press, Cambridge. Nordenstam, B. 1996. Recent revision of Senecioneae and Calenduleae systematics. In Compositae: Systematics. Proceedings of the International Compositae Conference, Kew. Edited by D.J.N. Hind and H.J. Beentje. Royal Botanic Gardens, Kew, UK pp. 591–596. Nordenstam, B. 2006. Generic revisions in the tribe Calenduleae (Compositae). Comp. Newsl. 44: 38–49. Nordenstam, B., Källersjö, M., and Eldenäs, P. 2006. Nephrotheca, a new monotypic genus of the Compositae-Calenduleae from the southwestern Cape Province. Comp. Newsl. 44: 32–37. Norlindh, T. 1977. Calenduleae — systematic review. In The biology and chemistry of the Compositae. Edited by V.H. Heywood, J.B. Harborne, and B.L. Turner. Academic Press, London. pp. 961–988. Peighambardoust, S.H., Dadpour, M.R., and Dokouhaki, M. 2010. Application of epifluorescence light microscopy (EFLM) to study the microstructure of wheat dough: a comparison with confocal scanning laser microscopy (CSLM) technique. J. Cereal Sci. 51(1): 21–27. doi:10.1016/j.jcs.2009.09.002. Sattler, R. 1973. Organogenesis of flowers: a photographic text-atlas. University of Toronto Press, Toronto, Ont. pp. 160–168. Simpson, M.G. 2006. Plant systematics. Elsevier/Academic Press, New York. Singh, B.P., and Agarwal, A. 2001. Floral anatomy of Actinomeris squarrosa and Bidens biternata (Asteraceae). Acta Bot. Hung. 43(3): 423–432. doi:10.1556/ABot.43.2001.3-4.15. Small, J. 1919. The origin and development of the Compositae. New Phytol. 18(5-6): 129–176. doi:10.1111/j.1469-8137.1919.tb07297.x. Stebbins, G.L. 1967. Adaptive radiation and trends of evolution in higher plants. In Evolutionary biology. Edited by M.K. Dobzhansky and W.C. Hecht. Appleton-Century-Crofts, New York. pp. 101–142. Stuessy, T.F., and Garver, D. 1996. The defensive role of pappus in heads of Compositae. In Compositae: biology and utilization. Proceedings of the International Compositae Conference, Kew, 1994. Edited by P.D.S. Caligari and D.J.N. Hind. Royal Botanic Gardens, Kew. pp. 81–91. Tiagi, B., and Taimni, S. 1963. Floral morphology and embryology of Vernonia cinerescens Schut., and V. cinerea less. Agra Univ. Jour. Res. Sci. 12: 123–138. Tucker, S.C. 1988. Heteromorphic flower development in Neptunia pubescens, a mimosoid legume. Am. J. Bot. 75(2): 205–224. doi:10.2307/2443887. Weberling, F. 1989. Morphology of flowers and inflorescences. Cambridge University Press, New York.
