Research Article |
Corresponding author: Alexander Vrijdaghs ( alexander.vrijdaghs@kuleuven.be ) Academic editor: Brecht Verstraete
© 2022 Alexander Vrijdaghs, Petra De Block, Karen L. G. De Toni, Erik Smets, Elmar Robbrecht.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Vrijdaghs A, De Block P, De Toni KLG, Smets E, Robbrecht E (2022) Floral ontogeny links Dialypetalanthus (Condamineeae) with the floral developmental morphology of other Rubiaceae. Plant Ecology and Evolution 155(3): 394-403. https://doi.org/10.5091/plecevo.84606
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Background – Vegetative and fruit characters of the Amazonian genus Dialypetalanthus point to a position in Rubiaceae. However, its floral morphology is so deviant that the genus was often placed in a family of its own. Even relationships outside Gentianales were postulated. Current molecular phylogenetic studies firmly show that Dialypetalanthus belongs to Rubiaceae.
Aims – This study aims to understand the idiosyncratic floral morphology in Dialypetalanthus and to compare it with the floral development in two other Condamineeae genera as well as in other Rubiaceae for which ontogenetic data are available.
Material and methods – SEM and LM based floral ontogeny in Dialypetalanthus fuscescens, Mussaendopsis beccariana, and Pogonopus exsertus.
Results and main conclusions – Flowers in Dialypetalanthus develop a stamen-corolla-calyx tube, which can be considered as a floral morphological link between the genus and the other Rubiaceae. The polyandrous androecium originates from an annular intercalary meristem at the adaxial side of the stamen-corolla-calyx tube.
Dialypetalanthus, floral cup, floral development, Mussaendopsis, Pogonopus, SEM, stamen-corolla tube, stamen-corolla-calyx tube
Upon its recognition by
Flowers of Rubiaceae usually are sympetalous, actinomorphic and generally consist of a tetra- or pentamerous calyx and corolla, both often consisting of a tubular base with distal lobes, an androecium with a number of (often epipetalous) stamens usually equal to the number of calyx and corolla lobes. The gynoecium consists of an inferior, bilocular ovary, each locule with one to many unitegmic ovules, and a single style with two stigmas; on the top of the ovary, a gynoecial annular nectary usually surrounds the base of the style (
In contrast to the confusing flower morphology, which was inconclusive as to the placement of the genus, all molecular phylogenetic studies showed that Dialypetalanthus belongs to Rubiaceae, Cinchonoideae sensu lato (
The floral development of only few species of Rubiaceae has been studied until now (see Table
Available floral developmental studies in Rubiaceae. Arranged according to the classification in two subfamilies (
CINCHONOIDEAE | ||
Coffeeae | Coffea L. |
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Octotropideae | Canephora Juss. |
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RUBIOIDEAE | ||
Paederieae | Paederia L. |
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Knoxieae | Pentas Benth. |
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Sacosperma G.Taylor |
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Spermacoceae | Mitrasacmopsis Jovet |
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Pentodon Hochst. |
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Spermacoce L. |
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Anthospermeae-Anthosperminae | Galopina Thunb. |
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Rubieae-Theligoninae | Theligonum L. |
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Rubieae-Galiinae | Asperula L. |
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Asperula tinctoria L. |
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Crucianella Boiss. |
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Cruciata Opiz |
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Cruciata glabra (L.) Ehrend. |
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Cruciata laevipes Opiz. |
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Galium L. |
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Galium verum L. |
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Phuopsis stylosa (Trin.) G.Nicholson |
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Sherardia L |
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Rubieae-Rubiinae | Rubia L. |
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Rubia tinctorum L. |
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According to
Collections were made in ethanol 70%. Partial inflorescences and floral buds of Dialypetalanthus fuscescens at all developmental stages were sampled by Karen De Toni from a tree in the Rio de Janeiro Botanic Garden, Brazil, and are preserved at Meise Botanic Garden, Belgium. Samples of Mussaendopsis beccariana Baill. came from the alcohol collection of Meise Botanical Garden and Pogonopus exsertus (Oerst.) Oerst. was collected from the living collection of Meise Botanic Garden. For voucher data, see Table
Species | Collector and year | Origin | Preserved at |
Dialypetalanthus fuscescens Kuhlm. | Karen De Toni 2017 | Rio de Janeiro Botanical Garden, Brazil | RBv 7855, RB 462363 |
Mussaendopsis beccariana Baill. | Christian Puff, Anton Igersheim, and Gustavo Martinelli 1990 | Sungai Liang Arboretum, Brunei | BR 900885/1/1 |
Pogonopus exsertus (Oerst.) Oerst. | Petra De Block 2020 | Meise Botanic Garden, Belgium | BR 20180855-05 |
For SEM, in preparation to critical point drying, the dissected material was gradually brought from ethanol 70% to a 1:1 mixture of ethanol 70% and dimethoxymethane (DMM) and subsequently to pure DMM. Next, the material was critical point dried by gradually replacing the DMM by liquid CO2 with the aid of a Balzers CPD 030 critical point dryer. The dried samples were mounted on aluminium stubs using carbon adhesive tape and sputter coated with gold with a Balzers SCD 020 sputter coater. SEM images were obtained with a JEOL JSM5800-LV scanning electron microscope at the laboratory of Plant Conservation and Population Biology, KU Leuven, Belgium.
For LM, the samples were gradually dehydrated through an ethanol series and subsequently embedded in KULZER’s Technovit 7100 (based on HEMA, hydroxyethyl-meth-acrylate). Seven μm thick sections were obtained with the help of a rotary microtome Leica RM2135 with disposable blades (Leica DB80). Subsequently, the sections were stained in a 0.1% toluidine blue in aq. dest. solution. Observations were done using an Olympus BX51 microscope equipped with a Color View Soft Imaging System camera at Meise Botanic Garden.
Inflorescence axes show decussate branching, the lateral axes occurring two-by-two, subtended by pairs of opposite bracts. Each pair of bracts and lateral axes is at 90° with respect to the preceding pair. This pattern is continued by the inflorescence unit/flower subtending bracts, also occurring in pairs that are perpendicular to each other (Fig.
Macrograph (A) and SEM images (B–F) of inflorescence (A–B) and successive stages of the early floral development (C–F) in Dialypetalanthus fuscescens. A and B: lateral view. C and E: apical view. D and F: lateral-apical view. A. Inflorescence axis with flowers in bud and at anthesis with calyx lobes, corolla lobes, androecium, and style indicated (resp. light green, red, yellow, and purple arrows). Pedicel of each flower with two opposite, possibly slightly displaced with respect to each other, bracteoles (encircled in dark green). Encircled in white the distal part of the main inflorescence axis and lower, an inflorescence unit. Framed in purple the inferior, bilocular ovary. B. Cymosely branched inflorescence unit with a terminal flower (F); scar of one of its bracteoles (Bo) subtending an inflorescence unit consisting of a developing lateral flower with two bracteoles (dark green arrows), a hairy calyx (light green arrows) and inner flower (blue) with centrally the appearing style (purple arrowhead) surrounded by stamens (yellow arrowhead). C. Flower primordium and two bracteoles. D. First two sepals appearing, opposite to each other and at 90° with respect to the pair of bracteoles. E. Four sepals, two by two, surrounding a flat central floral apex. The most recent pair of sepals opposite the bracteoles. F. Appearance of a first pair of petals, at 90° with respect of the latest developed pair of sepals. The floral axis (red arrow), situated in the centre of a cavity formed by the upwards growth of the bases of the surrounding floral parts. Colour code: blue, developing inner flower–pollen grains; green, calyx/bracteoles; purple, gynoecium; red, corolla; yellow, androecium. Symbols: B, bract; Bo, bracteole; ca, calyx (lobe); cl, colleter; co, corolla (lobe); F, flower.
SEM images of successive stages of the floral development in Dialypetalanthus fuscescens. A–H: apical view, I: lateral view. A. Floral apex (white arrow) becomes a central cavity, surrounded by a second pair of opposite petals, now called corolla lobes at 90° with respect to the previous pair. Sepals are from now onwards called calyx lobes. B. Appearance of a row of stamen primordia (one indicated by yellow arrowhead) on the distal part of the apical central cavity (white arrow). C. Centripetally of the developing stamens, a second ‘whorl’ of stamen primordia appears. Floral apex differentiates into two bulges surrounding a central split (purple arrow). D. The central bulges are raised from a common base (purple arrow), consisting of a single style. E. Below the single style, one out of two locules filled up with a U-shaped placenta (purple arrow). F. Developing style with two stigmas (purple arrowheads). Surrounding the base of the style, a scar of removed stamens shows a common androecial base (yellow arrow). G–H. Variable number of stamens (16–17). Single style/stigmas (purple arrowheads) protruding above developing stamens. I. Stamens with a common base (yellow arrow). Colour code: purple, gynoecium; red, corolla; yellow, androecium. Symbols: a, anther; ca, calyx (lobe); f, filament; cl, colleter; co, corolla (lobe); s, stamen; st, style.
SEM images of successive developmental stages of the gynoecium in Dialypetalanthus fuscescens. All images are lateral views. A. Opened locule with U-shaped early placenta with multiple ovule primordia. B–E. Idem, ovules develop top to bottom. B–C. Side view on U-shaped placenta. B. Developing single style starting to protrude above the stamens, its base surrounded by a whorl of trichomes. D. Adaxial view on placenta with distally semi-mature ovules. E. Adaxial view on placenta with unitegmic mature ovules (purple arrowheads). F. Semi-mature single style and stigmas (purple arrowheads). Colour code: purple, gynoecium; yellow, androecium. Symbols: a, anther; f, filament; pl, placenta; s, stamen; sp, septum; st, style; purple arrow, placenta; white arrow, whorl of hairs at style base.
LM images of transverse (A, C, D) and longitudinal sections (B, E, F) through developing flowers of Dialypetalanthus fuscescens. A. Through pedicel at height of bracteoles. B. Through early floral developmental stage. C. Through successive floral developmental stage with central depression (encircled in white). D. Detail of central depression with early stamens (encircled in yellow). E–F. Through developing flower, with central depression and apical cavity (encircled in purple) with developing stamens (yellow arrows) surrounded by two originating carpellary bulges (purple arrowheads). Colour code: purple, gynoecium; yellow, androecium. Symbols: Bo, bracteole; ca, calyx (lobe); co, corolla (lobe); Pc, pedicel; red asterisk, floral apex.
In a first stage of the floral development, a calyx with abaxially large trichomes and five petals develop, the latter united at the base (Fig.
In (semi-)mature flowers, a calyx is present consisting of a short calyx tube and five calyx lobes (Fig.
SEM images of stages of the floral development in Pogonopus exsertus. A–L: lateral views. A. Longitudinally opened developing flower. B. Tetrasporangiate introrse anther on short (invisible) filament. C. Adaxial view of part of tubular corolla; encircled in red, the haired rims of adherent corolla lobes. Proximally, a ‘whorl’ of hairs between stamen-corolla tube and corolla tube sensu stricto. D–E. Scar of filament at adhesion point of an epipetalous stamen at the tubular corolla. Below the scar, the stamen-corolla tube grows faster than the corolla tube sensu stricto above the scar. E. Longitudinal section of developing flower. An annular nectary (purple arrows) developing at the base of a single style with two stigma branches. F–G. Adaxial view of part of a semi-mature tubular corolla with two out of five epipetalous stamens. F. Proximal part. G. Distal part; the corolla lobes (one indicated by red triangle) with hairy margins. H–L. Development of the gynoecium. H. Developing single style with two style branches. I. Same stage, longitudinally opened flower with opened locule with U-shaped placenta. J–K. Successive stage with developing annular nectary surrounding the style base. Below the nectary, at right hand side, developing ovules. K. Idem from more apical view. Developing ovules encircled in purple. L. Developing ovules, detail of unitegmic ovule in frame. Obviously unitegmic ones indicated by purple arrowheads. Colour code: purple, gynoecium; red, corolla; yellow, androecium. Symbols: a, anther; ca, calyx (lobe); co, corolla (lobe); f, filament (scar of); ne, nectary; o, ovule; pl, placenta; s, stamen; sg, stigma; st, style; red double arrow, corolla tube sensu stricto; red-yellow double arrow, stamen-corolla tube.
SEM images of stages of the floral development in Mussaendopsis beccariana. A–F: lateral views. A–B. Semi-mature flower with part of corolla removed. Calyx consisting of a calyx tube and calyx lobes. Corolla consisting of free petals embedded in the hypanthium (red arrow). Five stamens with tetrasporangiate dorsifixed and introrse anthers. Conspicuous annular nectary surrounding the style base (scar of removed style indicated by purple arrow). C. Longitudinally opened semi-mature flower with encircled in purple a placenta with multiple ovules filling up one out of two locules. Centrally in the flower, scar of removed style indicated by purple arrow. D–E. Mature stamen with stretched filament, longitudinally opened tetrasporangiate dorsifixed and curved anther. E. Detail of anther. F. Detail of style and two stigmas. Colour code: green, calyx; purple, gynoecium; red, corolla; yellow, androecium. Symbols: a, anther; ca, calyx (lobe); f, filament; ne, nectary; pc, pedicel; pe, petal; sg, stigma; st, style.
The inflorescence is essentially decussate and dichasial, with successive pairs of opposite bracts, each subtending a lateral inflorescence axis (Figs
Flowers in D. fuscescens at first view show a quite idiosyncratic development, compared to other Rubiaceae, as already mentioned by
Theoretical outline of the inflorescence (A) and floral development (B, after
Moreover, during floral development, the initially slightly convex floral apex (Fig.
During the development of the perianth lobes, individual stamen primordia appear in a more or less centripetal order at the adaxial side of the central floral cup (Figs
The polyandry exhibited by D. fuscescens can theoretically find its origin in either ‘dédoublement’ of initially individual stamen primordia or polygenesis (
Only after the appearance of most stamens, the development of the gynoecium starts, similar as in other Rubiaceae (
The floral morphology in M. beccariana differs from that in most Rubiaceae studied by the absence of a tubular corolla and epipetaly. However, in contrast to the flowers of Dialypetalanthus, the flowers of M. beccariana have floral features that are considered to be common in Rubiaceae, such as a calyx tube, the same number of stamens as corolla lobes (no polyandry) and the presence of an annular nectary surrounding the base of the single style.
The development of the flower in P. exsertus concurs with the floral development as described by
According to
Following the hypothesis of
The early development of a stamen-corolla-calyx tube sensu
The adaxial part of the central floral cup in flowers of Dialypetalanthus consists of an annular androecial meristem or primary androecium primordium, from which multiple individual stamen primordia originate in a centripetal succession, eventually resulting in two ‘whorls’ of in total more or less 10 stamens. Hence, polyandry and no epipetaly sensu stricto, though the position of the stamens on a stamen-corolla-calyx tube can be considered as another character state of epipetaly (see also Table
Floral developmental characters in the three species studied based on the developmental hypothesis of
Characters | Dialypetalanthus fuscescens | Mussaendopsis beccariana | Pogonopus exsertus | |
ca | free sepals (1) or floral cup* + lobes (0) | 0 | 0 | 0 |
co | stamen-corolla tube | 0 | 0 | 1 |
corolla tube sensu stricto | 0 | 0 | 1 | |
stamen-corolla-calyx tube | 1 | 0 | 0 | |
petal fusion | 0 | 0 | 0 | |
an | free stamens (1) or epipetaly (0)** | 1 | 1 | 0 |
stamens on: stamen-corolla tube (1) or stamen-corolla-calyx tube (0) | 0 | 1 | – | |
4–5 stamens (1) or polyandry (0) | 0 | 1 | 1 | |
gy | inferior (1) or (half)-superior (0) | 1 | 1 | 1 |
bilocular (1) or other (0) | 1 | 1 | 1 | |
single style/ 2 stigmatic branches (1) or two styles (0) | 1 | 1 | 1 | |
ne | gynoecial annular nectary | 0 | 1 | 1 |
The inferior gynoecium in flowers of Dialypetalanthus develops like in all other Rubiaceae studied, but no gynoecial nectary surrounding the base of the single style is developed.
The inflorescence in Dialypetalanthus is characterised by a pairwise pattern of appearance of all lateral parts, including these of the perianth, whereby each pair of leaf-like structures is positioned at 90° with respect to the preceding (or following) one (decussate arrangement).
Mussaendopsis beccariana has a corolla consisting of five free petals and an androecium consisting of five free stamens. Hence, no polyandry occurs, neither does epipetaly. In contrast, the floral ontogeny in P. exsertus concurs with that in all other flowers studied in Rubiaceae.
We are now aware of floral ontogenetically well-documented cases of a switch in the development of floral whorls from a limited number of individual primordia to a primary annular meristem/primordium from which many individual parts originate secondarily.
We thank Iris Van der Beeten (Meise Botanic Garden) for technical assistance with the LM images. We thank the anonymous reviewers for their valuable comments.