Int. J. Plant Sci. 165(5):723738. 2004. 2004 by The University of Chicago. All rights reserved. 1058-5893/2004/16505-0003$15.

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FLORAL MORPHOLOGY AND DEVELOPMENT IN ARAGOA (PLANTAGINACEAE) AND RELATED MEMBERS OF THE ORDER LAMIALES
M. A. Bello,* P. J. Rudall,y F. Gonzalez,1; * and J. L. Fernandez-Alonso*
*Facultad de Ciencias, Universidad Nacional de Colombia, Ap. Ae. 7495 Bogota; and yJodrell Laboratory, Royal Botanic Gardens, Kew, Richmond, Surrey TW9 3AB, United Kingdom

Inorescence and oral morphology and development were investigated in Aragoa (Plantaginaceae) and related genera. Each inorescence of Aragoa is a reduced, axillary raceme, on which the actinomorphic oral apices generally arise successively. The inorescences of Aragoa and Plantago are polytelic and lateral. The ve sepals emerge from the abaxial to the adaxial side of the oral apex, but at maturity, the calyx is actinomorphic. The four stamens arise simultaneously and before emergence of the petals. The four petals emerge unidirectionally united, but the corolla becomes actinomorphic. Aestivation is cochlear ascendent. The two united carpels initiate simultaneously. The abaxial-adaxial inception of the calyx and corolla during early oral development in genera such as Aragoa, Digitalis, Plantago, and Veronica may indicate that the zygomorphic condition is ancestral in those genera. The tetramerous corolla, which is actinomorphic during middle and late development, and the presence of four stamens are possible synapomophies of the clade (Aragoa Plantago). Pentamery of the calyx and corolla appears to be plesiomorphic in the broader AragoaAngelonia clade. Characters related to development and morphology of inorescences and owers of Aragoa are essentially similar to those found in Plantago, which is consistent with the molecular-based sister group relationship between these genera. Keywords: Antirrhinum, Aragoa, oral development, oral morphology, Heliohebe, Lamiales, Plantago, Plantaginaceae, Scrophulariaceae, Veronica.

Introduction
Aragoa Kunth, with 19 species, three subspecies, and four probable hybrids (Fernandez-Alonso 1995), is endemic to the paramos of Colombia and Venezuela, between 2500 and 4000 m altitude (Pennell 1937; Ferna ndez-Alonso 1991, 1993, 1995; Nilsson and Hong 1993). The genus was originally described as Bignoniaceis afnia by von Humboldt et al. (1819), but its afnities have been in dispute since then (for review, see Fernandez Alonso 1995; Bello et al. 2002). Before recent molecular analyses, most authors included Aragoa within Scrophulariaceae sensu lato (e.g., Endlicher 18361840; Wettstein 1895; Fernandez-Alonso 1995), closely related to genera such as Hebe Comm. ex Juss. and Veronica L. in tribe Veroniceae (Bentham 1846, 1876; Hallier 1903; Pennell 1937; Yamazaki 1957; Thieret 1967; Nilsson and Hong 1993). However, Aragoa is atypical among Scrophulariaceae sensu lato because all species are shrubs or small trees and have actinomorphic corollas. These characters states led Pennell (1937) to regard Aragoa as primitive Veroniceae. Other characters of Aragoa, such as the helicoid arrangement of the leaves, the presence of four equal stamens, and the morphology of pollen and seed (Thieret 1967; Nilsson and Hong 1993), are not shared with all other members of the former
1

Author for correspondence; e-mail fagonzalezg@unal.edu.co.

Manuscript received September 2003; revised manuscript received March 2004.

tribe Veroniceae. Nilsson and Hong (1993) separated the genus into its own tribe (Aragoeae) based on pollen morphology but did not clarify its closest relationships within Scrophulariaceae. Fernandez-Alonso (1995) observed some morphologi cal similarities with Capraria L., Chionohebe Briggs & Ehrend., Detzneria Schltr. ex Diels., and Hebe and Scoparia L. but did not clearly propose a close relationship with any of them. Several authors have used oral characters to discuss the systematic relationships of Aragoa. For example, Don (1835) segregated the genus into its own family Aragoaceae based on the convolute aestivation of the corolla, later described by Bentham (1846) as imbricate. Others (Meisner 18301843; Endlicher 18361840; Bentham 1846; Rouy 1909) placed the genus within tribe Veroniceae, primarily on the basis of characters such as its racemose inorescence, ve-lobed calyx, the imbricate aestivation of the four-lobed corolla with funnelrotate shape, and the tetramerous androecium with conuent thecae. Although oral morphology has provided key characters in assessments of the systematic relationships of the genus, a detailed study of its oral development and morphology is lacking, preventing a comparison with related genera that have already been investigated (see, e.g., Payer 1857; Chatin 1873; Saunders 1933; Srinivasan 1940; Yamazaki 1957; Schrock and Palser 1967; Singh 1979; Singh and Jain 1979; Nishino 1983; Awasthi et al. 1984; Leins and Erbar 1988; Endress 1992, 1994, 1998; Wunderlin 1992; Kampny et al. 1993; Kampny and Dengler 1997; Prakash and Singh 1997). Recent molecular and phytochemical data for Aragoa support a previously unsuspected sister group relationship with
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INTERNATIONAL JOURNAL OF PLANT SCIENCES ohebe raoulli were collected from the living collections at the Royal Botanic Gardens, Kew. Various developmental stages covering the complete oral ontogeny of four species (two of them from three different populations) and one putative hybrid of Aragoa were examined under the LM and SEM (vouchers cited in table 1). Samples were xed in FAA and then transferred to 70% ethanol. The samples were then dissected in 90% ethanol under a Wild Heerburg M8 dissecting microscope. The material was then submitted to a dehydration series of 90%, 95%, and 100% ethanol, 1 h in each concentration. The samples used for SEM were critical point dried in a Balzers 030 drier, coated with gold in an Emitech Sputter coater K550, and examined and photographed on a Cambridge Instruments Stereoscan 240 at 15 kV. For light microscopy, the samples were transferred from 100% ethanol through an ethanol-Histoclear series (90 : 10, 70 : 30, 50 : 50, 30 : 70, 10 : 90, 0 : 100) for 18 h. Floral buds were embedded in paraplast for serial sectioning using a rotary microtome (Reichert-Jung 2040). Sections were stained with safranin and Alcian Blue and mounted in DPX resin.

Plantaginaceae, which resulted in the inclusion of Aragoa within Plantaginaceae (Bello et al. 2002; Rnsted et al. 2002; Jensen et al. 2003), with Veronica as its immediate sister group. The aims of this article are to investigate the oral development and morphology of Aragoa and to compare these results with those of genera that are currently included in the clade recognized by Bello et al. (2002), hereafter called the Aragoa-Angelonia clade (Veronicaceae sensu Olmstead et al. 2001). The hierarchical structure of this clade is as follows: ((((((((Aragoa Plantago) Veronica) Hemiphragma) Digitalis) Globularia) (Callitriche Hippuris) Antirrhinum) ((Chelone Collinsia) Tetranema) and other genera of its sister clade (((Amphianthus Gratiola) Bacopa) Angelonia). Heliohebe Garn.-Jones, a member of the Hebe complex sensu Garnock-Jones (1993), and Littorella, a basal genus in the phylogeny of Plantaginaceae s.l. (Rnsted et al. 2002; Hoggard et al. 2003), are here treated as distinct genera in the discussion of oral characters. The inclusion of Aragoa in a clade with Antirrhinum, Plantago, and Veronica makes examination of its oral morphology and ontogeny particularly relevant because recent developmental-genetic work on oral symmetry in Antirrhinum has demonstrated that zygomorphy relies on differential expression of two closely related adaxial identity genes CYCLOIDEA (CYC) and DICHOTOMA (DICH) in the young oral meristem (Coen and Nugent 1994; Coen 1996; Luo et al. 1996, 1999). This discovery has stimulated a renaissance in studies of oral symmetry patterns (e.g., Neal et al. 1998; Endress 1999; Rudall and Bateman 2003). Plantago is often cited as an example of a taxon with actinomorphic owers that is phylogenetically nested within a clade with taxa that produce predominantly zygomorphic owers (Coen and Nugent 1994; Donoghue et al. 1998; Endress 2003). Because owers of Aragoa are also regarded as actinomorphic, such different patterns of symmetry in related taxa would merit more rigorous examination in this clade.

Results Organography of Aragoa

The owers of Aragoa are bisexual (g. 1). The ve sepals are ovate, oblong, or lanceolate; equally sized (ranging from 2.8 to 8 mm long), (sub)coriaceous, frequently with long hairs on the margin and short hairs along the dorsal side. By preanthesis, the calyx shows a cochlear-ascendent aestivation (gs. 2, 3N). The corolla is tetramerous, actinomorphic, rotate (e.g., in Aragoa cundinamarcensis Fernandez-Alonso) to subrotate (e.g., in Aragoa abietina H.B.K., Aragoa cleei Fernandez Alonso, and Aragoa cupressina H.B.K.) or tubular-campanulate (e.g., Aragoa perez-arbelaeziana Romero). The corolla is generally white, except in the yellow-owered A. perez-arbelaeziana). The tube is 1.517 mm long, and the corolla lobes are obovate or spathulate, 3 to 9 3 2:5 to 7 mm (g. 1). The marginal indumentum of the petals is different in the species studied. In A. cleei and A. cupressina H.B.K., it is formed by dense liform hairs (g. 4C); in Aragoa x funzana Fernandez-Alonso,

Material and Methods

Floral buds of Aragoa and Plantago were collected in the eld at several stages of development, whereas those of Heli-

Table 1
Species and Material Examined Species Aragoa abietina Kunth Aragoa cleei Fernandez-Alonso Aragoa cundinamarcensis Fernandez-Alonso Aragoa cupressina Kunth Aragoa x funzana Fernandez-Alonso Heliohebe raoulli (Hook.f.) Garn.-Jones Plantago major L. Voucher COL: Bello 118, 119, 120 COL: Bello 108 COL: Bello 107 COL: Bello 128, 129, 130 COL: Bello 113, 115 HK: 1996-23; CHAD 574 COL: Bello 247 Locality Colombia: Cundinamarca, Parque Nacional Natural Chingaza Colombia: Cundinamarca, Villapinzon, Paramo de Gachaneque Colombia: Cundinamarca, Villapinzon, Paramo de Gachaneque Colombia: Cundinamarca, Paramo de Monserrate Colombia: Cundinamarca, Villapinzon, Paramo de Gachaneque Royal Botanic Gardens, Kew Colombia: Bogota, campus of the National University

Note. COL specimen supported by a voucher in the Colombian National Herbarium, Bogota. HK material collected from the Living Collections, Royal Botanic Gardens, Kew.

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Fig. 1 Flowering shoots of Aragoa

the hairs are scarce, whereas petal margins of A. abietina are glabrous. The corolla aestivation is cochlear ascendent (g. 4A, 4B; g. 5H). The four epipetalous stamens alternate with the corolla lobes (g. 4G) and have liform, glabrous, or distally pubescent laments (g. 5E, 5G) and dithecal, dorsixed, short (0.71.8 mm long) anthers that are introrse and open by longitudinal slits (g. 4G). The gynoecium is bottle shaped, bilocular, and basally surrounded by a hypogynous, ringlike nectariferous disc (g. 4F, 4G; g. 5C, 5J). Placentation is axile, and there are four to eight ovules per locule (gs. 4F, 5J). The style is cylindrical, up to 20 mm long, reaching the same level of the anthers by anthesis (gs. 1B, 4G). The stigma is slightly bilobed, capitate, dry, and papillate (g. 4I).

Floral Development of Aragoa

Inorescences. The inorescences of Aragoa are reduced axillary racemes, which are often congested in adjacent nodes of the distal or middle zones of the branches. The racemes and internodes are both extremely short, resulting in a oral arrangement that appears whorled. Several oral pseudowhorls can develop along a single branch, but they are separated by a long vegetative zone. The youngest racemes are closest to the branch apex, which suggests an acropetal sequence of development (g. 1A). Each raceme is subtended by a leaf and consists of two or three owers (gs. 1A, 2). If the raceme has three owers (e.g., in A. cundinamarcensis, A. cupressina, and A. x funzana), two of them are lateral, and one occupies the median position (g. 2). Each ower is subtended by a single leaike bract. This bract is perpendicularly oriented with respect to the subtending leaf in the lateral owers and opposite to it in the median ower. Within each inorescence, oral apices generally arise successively, but two can occasionally initiate almost simultaneously (g. 3A, 3B). Simultaneous emergence of three oral apices within the same inorescence was not observed. A dense indumentum of white hairs surrounds the buds, especially during early stages of development (g. 3A). Calyx. The oral apex is initially radially symmetrical (g. 3A). Calyx development commences with the simulta-

neous initiation of the two abaxial-lateral sepals, followed by initiation of the two adaxial-lateral ones. The gap between the latter two is soon lled by the fth (adaxial) sepal primordium (g. 3B3D). Thus, the emergence of the sepals is unidirectional, from the abaxial to the adaxial side of the oral apex, indicating a zygomorphic initiation of the calyx. The abaxial-lateral sepals are slightly larger than the adaxiallateral ones, whereas the adaxial sepal is the smallest (g. 3E, 3N); this difference remains evident until preanthesis. The formation of the calyx tube is slightly asymmetric because intercalary growth between the abaxial-lateral sepals occurs earlier than that between the lateral and the adaxial sepals (g. 3F). By preanthesis, the calyx becomes actinomorphic and has cochlear-ascendent aestivation. Sepals have a dense covering of hairs along the margins. Androecium and corolla. The four stamens primordia arise simultaneously opposite to the abaxial-lateral and the lateral sepals, before the emergence of the alternating petal primordia (g. 3G). There is no evidence of a fth stamen opposite the adaxial sepal. Initiation of the four petal primordia is unidirectional because the abaxial and the lateral petal primordia emerge simultaneously alternating with the sepals, and the adaxial initiates later opposite the adaxial sepal (g. 3H). Then, the emergence of the petals results in initial bilateral symmetry for the corolla. Just after organogenesis, the petal primordia are separated from each other by the developing stamens (g. 3I3K). Late sympetaly occurs by growth subjacent to the petals and their sinuses. The stamens in these sinuses thus fuse to the corolla (g. 3I, 3K, 3L). At this time, differentiation of lament and anther becomes apparent, and the corolla is slightly zygomorphic (g. 3L, 3M, 3O). Before anthesis, corolla aestivation becomes cochlear ascendent as the lateral and abaxial lobes enlarge over the adaxial one (g. 4A4C). Corolla aestivation in the lateral owers within an inorescence forms mirror images (cf. g. 4A, 4B). By anthesis, the corolla becomes actinomorphic after the

Fig. 2 Inorescence diagram of Aragoa with the two lateral owers already developed and a third oral apex in median position. Subtending leaf and bracts are in black; sepals are hatched; petals are in white; circle with dot represents shoot axis; circle with cross represents inorescence axis.

BELLO ET AL.FLORAL MORPHOLOGY OF ARAGOA enlargement of petals and the tube, and the four stamens are fully developed and alternate with the corolla lobes (g. 1). Gynoecium. After petal initiation, a gynoecium primordium in the center of the young ower is apparent (g. 3H). Shortly after that, the congenitally fused carpels appear as a ringlike primordium, without protruding median parts (i.e., symplicate carpels); the septum develops later (g. 3H, 3I). The carpels remain united up to the stigmatic region, where there is a narrow furrow radially oriented with respect to the axis of the inorescence (g. 3J, 3O). A narrow style with a central transmitting tract (g. 5E, 5F, 5H, 5J) is formed after elongation of the upper part of the pistil.

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(e.g., A. cupressina). In contrast to the thick cuticle present in mature sepals, petals lack cuticle, at least in some species (g. 5). The inner epidermis of the petal consists of a single layer of domed (papillate) cells; a dense covering of liform hairs is also present at the level of stamen insertion (g. 4G). Stomata were observed on petal surfaces in all species examined. The petal mesophyll consists of a homogeneous parenchyma without intercellular spaces (g. 5E, 5G, 5H). The epidermis of the anthers is formed by a thin layer, and the endothecium consists of regularly shaped cells with thickened walls near the dehiscence area. The tapetum is parietal.

Floral Development of Heliohebe Floral Vasculature and Anatomy of Aragoa

Serial cross sections of Aragoa owers (g. 5) show that the vascular traces along the pedicel form a continuous stele. At the level of calyx insertion, the median vascular trace of each sepal departs directly from the central stele and remains unbranched (g. 5A5E). Additionally, ve commissural bundles depart from the stele, and each divides into two traces at the base of the sepals, forming the two lateral traces of adjacent sepals (g. 5A). As a result, each sepal contains three main vascular bundles. At the level of insertion of the corolla, eight vascular traces depart from the central stele, four of which supply the corolla and the alternating four supply the stamens (g. 5B 5E). At this level, the adaxial petal trace divides into three traces; the median of these three traces enters the center of the adaxial corolla lobe, and the two lateral traces irrigate its anks. The two median carpellary veins are also evident at the level of corolla insertion (g. 5B, 5C). A short distance above this level, each of the two remaining petal traces also splits into three traces. These traces essentially have the same branching pattern as the adaxial trace. The central traces of each petal further divides into ve to eight traces in the distal half of each corolla lobe. In contrast, stamen bundles remain unbranched (g. 5D, 5E). The remaining central vascular traces supply the gynoecium. Each of the two carpels is irrigated by a median and two lateral bundles. The ovules are directly supplied by the marginal traces. There are two traces along the style (g. 5F). Glandular trichomes consisting of a stalk of two to three cells and a head formed by six cells are scattered on both the outer and the inner corolla surfaces (g. 4D, 4E, 4H); they are restricted to the distal part of the tube in some species Inorescence. Inorescences of Heliohebe raoulii are terminal racemes, in which owers develop spirally and acropetally (g. 6A). A single bract covers the corresponding oral bud by the time of emergence of the abaxial-lateral sepals. Calyx. The oral apex is zygomorphic (g. 6A). The two abaxial-lateral sepal primordia initiate rst. They emerge simultaneously. The two adaxial-lateral sepals arise much later (g. 6B, 6C); they also emerge simultaneously and remain smaller than the abaxial-lateral sepals throughout development (g. 6C6F). The whole calyx is therefore zygomorphic. There is no evidence of a fth sepal primordium. The fusion between the abaxial-lateral sepals is greater than that between the adaxial-lateral ones (g. 6D6F). The anks of the abaxial-lateral sepals partially overlap the adaxial-lateral sepals (g. 6C6F). Marginal trichomes become evident on the abaxial-lateral sepals during late development. Androecium and corolla. During development, these oral whorls are highly zygomorphic in H. raoulii. The two stamen primordia initiate simultaneously after the emergence of adaxial-lateral sepals (g. 6C), opposite them. There is no evidence of additional stamen primordia. Later, three petal primordia simultaneously emerge on the abaxial side of the oral bud; the abaxial petal primordium alternates with the two abaxial-lateral sepals and the two lateral petal primordia opposite them (g. 6D). Petal growth is delayed because lobes are still included in the young ower even during late development (g. 6E, 6F). By preanthesis, the corolla aestivation is imbricate because the two lateral petals are external to the medial petals (not shown). Gynoecium. The gynoecium primordium in the center of the young ower appears to initiate simultaneously with petal initiation. Shortly after that, the congenitally fused

Fig. 3 Early and midoral development of Aragoa. A, Aragoa cundinamarcensis. Two oral apices. B, Aragoa x funzana. Upper view of inorescence apex (arrowhead points to primordium of adaxial sepal). C, Aragoa cleei. Floral bud with the abaxial-lateral sepals initiating rst. D, Aragoa cundinamarcensis. Unidirectional development of sepals. E, Aragoa cleei. Calyx before stamen emergence. F, Aragoa cundinamarcensis. Lateral view of inorescence. G, Aragoa cupressina. Stamen emergence. H, Aragoa cundinamarcensis. Upper view of oral bud (sepals removed except the adaxial one); arrowhead indicates the site where adaxial petal will develop. I, Aragoa cundinamarcensis. Gynoecium initiation. J, Aragoa x funzana. Upper view of apex inorescence (sepals removed); the arrowhead shows the intercarpellary furrow. K, Aragoa cundinamarcensis. Lateral view of oral bud (sepals removed). L, Aragoa abietina. Lateral view of oral bud (sepals removed; adaxial petal on the right). M, Aragoa cleei. Floral bud with margins of petals fusing (arrowhead; sepals removed; adaxial petal on the lower left). N, Aragoa x funzana. Cochlear aestivation of the calyx. O, Aragoa cundinamarcensis. Actinomorphic oral bud (sepals removed). a floral apex; b bract; c gynoecium; ea abaxial stamen; ep adaxial stamen; pa abaxial petal; pl lateral petal; s1 abaxial-lateral sepal; s2 adaxial-lateral sepal; s3 adaxial sepal; tc corolla tube; x inflorescence axis. Scale bars 100 mm in A, B, D, E, H, I; 50 mm in C, G; 200 mm in F, J, K, MO; 500 mm in L.

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Fig. 4 Late oral development of Aragoa. A, B, Aragoa abietina. Upper view of mirror image of oral buds (sepals removed). C, Aragoa cleei. Upper view of cochlear-ascendent corolla. D, E, Aragoa cleei. Lateral view of the corolla; capitate hairs arrowed. F, Aragoa x funzana. Placental septum basally surrounded by a ringlike disk (ovary walls removed). G, Aragoa cleei. Inner view of a mature ower. H, Aragoa x funzana. Capitate hair. I, Aragoa cleei. Papillary stigma surface. an anther; f filament; n nectarial disk; o ovule; pa abaxial petal; pl lateral petal; pp adaxial petal; sp placental septum; tc corolla tube; v ovary. Scale bars 500 mm in A, C, E, F; 1 mm in B, D; 2 mm in G; 20 mm in H; 100 mm in I.

carpels appear as a ringlike primordium (i.e., symplicate carpels; g. 6D). Later development of the pistil was not followed in this species. Mature owers show a bottle-shaped gynoecium basally surrounded by a ringlike nectariferous disc with glandular-ciliate border (not shown). The stigma is four lobed, narrowly capitate (not shown).

Floral Development of Plantago

Inorescence. The developing spikes in Plantago show congested oral buds that develop acropetally (g. 7A). Each

oral bud is subtended by a bract that covers it even before organogenesis. Calyx. Floral apices of Plantago are initially radially symmetrical until sepal emergence, which occurs unidirectionally; the initiation of the two abaxial-lateral sepal primordia is followed by the two adaxial-lateral ones (g. 7A, 7B). There is no evidence of the emergence of a fth adaxial primordium. Each calyx lobe soon becomes concave (g. 7B); then, the calyx tube begins to be formed. The calyx tube remains short (g. 7E). The abaxial-lateral calyx lobes are slighly larger than the adaxial-lateral ones (g. 7C7E),

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Fig. 5 LMs of ower sections. AE, Aragoa cupressina, series of transverse sections (TS) through a single ower. F, Aragoa cupressina, TS style. G, H, Aragoa abietina, TS ower bud at preanthesis. J, Aragoa abietina, longitudinal sectional of ower bud at preanthesis. n nectariferous disc; tt stylar transmitting tissue. Scale bar 100 mm.

showing a zygomorphic pattern. Pluricellular trichomes begin to develop on the outer surface (g. 7D). Androecium and corolla. The four stamens emerge simultaneously shortly before petal initiation (g. 7A). Stamen primordia are opposite the sepals. The emergence of the petal

primordia is unidirectional, from the abaxial to the adaxial side (g. 7B). However, the adaxial petal becomes slightly larger than the others by mid-development (g. 7C). There is no evidence that the adaxial petal is formed by the fusion of two petal primordia. The corolla lobes emerge independently

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INTERNATIONAL JOURNAL OF PLANT SCIENCES and polytelic. By contrast, Veronica and Hebe have predominantly polytelic inorescences (cf. g. 9 in Weberling and Troll 1998), but they are often terminal. Flowers have been described as solitary in Angelonia Bonpl., Antirrhinum L., Aragoa, and Collinsia Nutt. (Meisner 18301843; Bentham 1846, 1876; Lemee 1929). How ever, they are primarily arranged in polytelic inorescences, either as lateral (in Aragoa) or terminal racemes (in Angelonia, Antirrhinum, and Collinsia; see Wettstein 1895, their g. 24). Optimization of polytelic versus monotelic inorescences in the Aragoa-Angelonia clade (not shown in g. 8) indicates that polytelic inorescences could be plesiomorphic. However, inorescence architecture is unknown in genera such as Amphianthus and Hemiphragma, which makes interpretation of this character difcult in this clade. Inorescences were described as monotelic in Tetranema Benth. ex Lindl. by Weber (1972), which would be autapomorphic for it. Lateral, polytelic inorescences seem to be plesiomorphic in two subclades: (((((Aragoa Plantago) Veronica) Hemiphragma) Digitalis) Globularia) and ((Chelone Collinsia) Tetranema). However, in these subclades, terminal polytelic inorescences appear in Collinsia, Digitalis, and some species of Veronica. Littorella and Heliohebe (not sampled in Bello et al. 2002) have lateral and terminal polytelic inorescences, respectively. In the remaining members of the Aragoa-Angelonia clade, solitary owers appear independently at least twice in the clade Callitriche Hippuris, in Gratiola, and in Bacopa. Furthermore, terminal polytelic inorescences also appear independently in Angelonia and Antirrhinum. Inorescence architecture is unknown in Amphianthus and Hemiphragma.

Fig. 6 Floral development of Heliohebe raoulii. A, Upper view of inorescence apex with four oral buds. B, Frontal view of oral bud with two abaxial-lateral sepals already initiated. C, Floral bud with the two abaxial-lateral and two adaxial-lateral sepals; the two adaxial-lateral stamens start initiation. D, Floral bud with the four petals (arrowheads) initiated; gynoecium becomes evident. E, F, Midoral bud with strongly zygomorphic calyx. a floral apex; b bract; c fused carpels; e stamen; p petal; s1 abaxiallateral sepal; s2 adaxial-lateral sepal. Scale bars 100 mm in A, DF; 50 mm in B, C.

Calyx
In some groups of plants, the symmetry of the oral apex determines symmetry during early stages of oral organogenesis (Tucker 1984). This seems to occur in Digitalis and Heliohebe, which have zygomorphic oral apices before and after organogenesis (Singh 1979; Wunderlin 1992; g. 6), and in Collinsia, which has radial oral apices and a simultaneous initiation of the sepals (Schrock and Palser 1967). However, the oral apices of Aragoa and Plantago are initially actinomorphic and become zygomorphic because of the abaxialadaxial emergence of the sepals (g. 3A3E; table 3). A similar switch in symmetry of the oral apex has been observed in Angelonia, Antirrhinum, and Veronica (Srinivasan 1940; Singh and Jain 1979; Kampny et al. 1993) (table 2), although in Antirrhinum, zygomorphy is mainly expressed in the corolla and androecium (Endress 1999). During early and middevelopment in Aragoa, Digitalis, Heliohebe, Littorella, Plantago, and Veronica, the abaxial-lateral sepals are larger than either the adaxial sepal (if present) or the adaxial-lateral sepals (Payer 1857; Singh 1979; Kampny et al. 1993; Prakash and Singh 1997; g. 6B6D; table 3). Similar development is observed in the perianth (treated as calyx in Cronquist 1981; Takhtajan 1997) of Hippuris vulgaris (Leins and Erbar 1988). A switch in symmetry during oral ontogeny was considered unusual by Tucker (1984) and Canne-Hilliker (1987) but has since been reported in a number of plants (Armstrong

(g. 7B) and become interconnected by interprimordial growth with the alterning stamens, showing late sympetaly. By mid-development (g. 7C7F), the corolla is actinomorphic, and the stamens are already differentiated in anther and lament (g. 7F). Petal aestivation is imbricate because the abaxial petal is partially covered by the lateral and the adaxial ones (g. 7G). Gynoecium. Gynoecium development and gross shape of the mature pistil in Plantago major is essentially the same to that described in Aragoa (g. 7A7E).

Discussion Inorescence
Our results conrm Fernandez-Alonsos (1995) report that the inorescences of Aragoa are lateral, extremely condensed racemes (g. 1). Plantago often has lateral spikes (see Weberling and Troll 1998 for a detailed description of inorescences in Plantaginaceae). Therefore, inorescences of Aragoa and Plantago are similar in that they are both lateral

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Fig. 7 Floral development of Plantago major. A, Upper view of an inorescence; arrowhead on the left oral bud shows an abaxial-lateral stamen primordium. B, Floral bud with stamen and petal primordia; the adaxial petal primordia are not yet initiated. C, Midoral development. D, E, Floral buds showing the zygomorphic calyx. F, G, Buds at preanthesis (sepals partially removed). a floral apex; b bract; c carpel; ea abaxial-lateral stamen; ep adaxial-lateral stamen; pa abaxial petal; pl lateral petal; pp adaxial petal; s1 abaxial-lateral sepal; s2 adaxial-lateral sepal. Scale bars 100 mm in A, DG; 10 mm in B, C.

and Douglas 1989; Endress 1999). Symmetry during early ontogeny is expressed by several traits as the order of initiation and the phyllotaxis of the oral organs, whereas in later stages, it is expressed by differential growth of the organs (Endress 1999). Similarly, it has been proposed that oral symmetry is affected during early development by the position of the oral buds in the inorescence; for example, in some plants with actinomorphic owers, initial development is strongly zygomorphic, especially in spikes or racemes (Endress 1999). Armstrong and Douglas (1989) suggested that zygomorphy produced only by early ontogeny would repre-

sent primitive zygomorphy in Scrophulariaceae. This could also be applied to the unidirectional emergence of the sepals of Aragoa and related genera such as Plantago, which would represent ancestral zygomorphy of the calyx (g. 9). The unidirectional development of the calyx is present independently in other groups of Scrophulariaceae s.l. such as Calceolaria L. (Ritterbusch 1976) and Lophospermum D. Don (Asarina Miller; Payer 1857). Synsepaly and pentamery are often correlated with zygomorphy of the calyx (Endress 1987). However, in Aragoa, in spite of early zygomorphy shown by the calyx, it becomes

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INTERNATIONAL JOURNAL OF PLANT SCIENCES count that the pentamerous condition of the calyx could be plesiomorphic in the Aragoa-Angelonia clade (g. 8; see also Reeves and Olmstead 1998), the presence of four sepals in the latter four genera indicates a suppression of the adaxial sepal (table 3). This type of suppression in an ontogenetic sequence is considered as phase specic; i.e., it is restricted to one whorl only and does not alter subsequent ontogenetic stages (Hufford 1996). Therefore, calyx variation in members of the Aragoa-Angelonia clade includes pentamery (i.e., as in Antirrhinum, Aragoa, Collinsia, Digitalis, Hemiphragma, Globularia), tetramery (as in some species of Plantago and Veronica), trimery (as in some species of Plantago), and complete suppression or modication (as in Callitriche and Hippuris, respectively). Kampny and Dengler (1997) suggested that suppression of the adaxial sepal could be a synapomorphy of Veronica and allies, Pseudolysimachion (Koch) Opiz and Hebe group, with reversals to pentamery in several species of Veronica, in Chionohebe B. Briggs & Ehrend., Paedorota L. and Paedorotella (Wulff) Kem.-Nat. However, in the Aragoa-Angelonia clade, the presence of four sepals appears independently in Plantago and Veronica and could be homoplastic in these genera (g. 8).

Androecium
In most angiosperms, including Euasteridae, the sequence of formation of the oral whorls (calyx, corolla, androecium, and gynoecium) is centripetal (Endress 1994). However, in Aragoa, the corolla-androecium sequence of initiation is clearly inverted (g. 3G; table 3). This change appears independently at least three times in the Aragoa-Angelonia clade: (1) in (Veronica (Plantago Aragoa)) (Kampny et al. 1993; Kampny and Dengler 1997; Prakash and Singh 1997); (2) in Collinsia (cf. Schrock and Palser 1967); and (3) in Angelonia and Bacopa (cf. Srinivasan 1940; Safeeulla and Govindu 1950). The absence of comparative studies on Amphianthus, Chelone, Globularia, Gratiola, Hemiphragma, and Tetranema makes an interpretation of this character in the AragoaAngelonia clade difcult. Possibly this condition has evolved several times in various groups of Lamiales, although it could be considered as a secondary synapomorphy of the clade ((Aragoa Plantago) Veronica). The emergence of the stamens before the petals has also been observed in other genera such as Besseya (Hufford 1995; Kampny and Dengler 1997), Calceolaria (Ritterbusch 1976), Phlox (Nishino 1983), Scrophularia marylandica (Schertz 1919), Synthyris (Hufford 1995; Kampny and Dengler 1997), Vadellia (Krishna Iyengar 1940), Plumbaginaceae (Prakash and Singh 1997), and Primulaceae (Duchartre 1844; Schertz 1919; Sattler 1962). Prakash and Singh (1997) suggested that in Plantago, this character could indicate a close relationship with Plumbaginaceae rather than with Scrophulariaceae s.l. However, the results of this study and other analyses of oral development in Scrophulariaceae s.l. allow interpretation of independent evolution of this character in different groups of Asteridae. Judd et al. (1999) suggested that the development of the androecium prior to the corolla could be a synapomorphy of Plantaginaceae; however, they included genera such as Digitalis in this family. In Digitalis, the emergence of the stamen primordia alternates with that of the petal primordia (Singh

Fig. 8 Distribution of oral characters in the Aragoa-Angelonia clade, based on the topology achieved in the molecular analysis of Bello et al. (2002). Numbers above lines indicate number of stamens primordia that initiate in bud. Despite the presence of ve petals in some species of Plantago and Veronica, presence of four petals is optimized as a synapomorphy of (((Aragoa Plantago) Veronica) Hemiphragma), based on species-level phylogenies of Veronica and allies (Albach and Chase 2001) and Plantaginaceae (Rahn 1996; Rnsted et al. 2002), in which the basalmost taxa sampled have tetramerous corollas. This suggests that the pentamerous condition could be derived independently in Plantago and Veronica.

actinomorphic before anthesis; the calyx lobes become similar in size and shape, possibly because there is not a mechanical constriction on their edges as they have cochlear aestivation. This also occurs in Angelonia, Digitalis, and some species of Agalinis and Antirrhinum (Wettstein 1895, p. 89; Kampny and Canne-Hilliker 1987; Thompson 1988, pp. 6266; table 2). However, in Heliohebe, Plantago, and Veronica, genera that primarily have four sepals, the calyx can remain zygomorphic in mature owers (g. 6F; table 2). Possibly a tetramerous calyx, with two abaxial-lateral and two adaxial-lateral sepals, allows a permanent zygomorphic symmetry of the calyx during oral development, although Littorella and some species of Plantago and Veronica, with four sepals, exhibit a radial symmetrical calyx (g. 147H in Harms and Reiche 1895). Moreover, Endress (2003) suggested that in genera such as Plantago and Callitriche, merosity reduction of the perianth is related to a change from monosymmetry to polysymmetry. In Aragoa and some species of Veronica, the adaxial sepal is smaller than the other sepals, whereas in Heliohebe, Littorella, Plantago, and other species of Veronica, it is completely suppressed (g. 6; g. 7A7E; tables 2, 3). Taking into ac-

Table 2
Main Floral Features of Aragoa and Genera Included in the Aragoa-Angelonia Clade
Floral apex symmetry ? Radial Radial Sepal emergence ? Abaxialadaxial Abaxialadaxial Abaxialadaxial ? ? ? Simultaneous Abaxialadaxial ? ? Abaxialadaxial ? Abaxialadaxial Abaxialadaxial Abaxialadaxial ? Abaxialadaxial Sepal primordia 5 5 5 Final symmetry of calyx ? Radial Bilateral radial Radial ? ... ? Radial Radial ? Radial Bilateral ? Radial Radial Radial, bilateral ? Radial, bilateral Stamen emergence ? Abaxialadaxial Abaxialadaxial Simultaneous ? ? ? Simultaneous Abaxialadaxial Abaxialadaxial ? Simultaneous ? ... ? Simultaneous ? Simultaneous Vestigial stamens None None Adaxial Fertile stamens 2 4 4 Petal emergence ? ? Simultaneous? Abaxialadaxial ? ... ? ? Abaxialadaxial ? ? Abaxialadaxial ? ... ? Abaxialadaxial ? Abaxialadaxial Petal primordia 4 5 5 Final symmetry of corolla Bilateral Bilateral Bilateral Corolla aestivation Antirrhinoid Antirrhinoid Antirrhinoid

Genera Amphianthus Angelonia Antirrhinum

Inorescences Polytelic? Polytelic terminal Polytelic terminal Polytelic lateral Solitary owers Solitary owers Polytelic lateral Polytelic terminal Polytelic terminal Polytelic lateral Solitary owers Polytelic terminal Solitary owers? Solitary owers Polytelic lateral Polytelic lateral Monotelic lateral Polytelic lateral and terminal

Sympetaly ? ? Late

References 1, 3, 4, 6, 15 1, 3, 4, 15, 35 1, 2, 3, 4, 7, 8, 13, 15, 24, 25, 29, 31, 32, 35 1, 3, 6, 12, 15, 38 1, 3, 4, 15, 34 1, 3, 4, 15, 17, 32 1, 3, 4, 15, 18, 32 1, 3, 4, 15, 27, 32 1, 3, 4, 15, 28, 32, 35, 36 3, 4, 15, 30 1, 3, 4, 15, 35 38 1, 3, 4, 6, 15 5, 30 6, 10, 11, 12, 15, 23, 26 1, 6, 10, 11, 16, 19, 20, 21, 22, 23, 30, 32, 38 1, 3, 4, 15, 33, 35 1, 3, 6, 12, 14, 15, 16, 26, 32, 34, 37

Aragoa Bacopa Callitriche Chelone Collinsia Digitalis Globularia Gratiola Heliohebe Hemiphragma Hippuris Littorella Plantago

Radial ? ? ? Radial Radial or bilateral ? ? Bilateral ? ? ? Radial

5 5 Absent 5 5 5 5 5 4 5 ? 4 4

None None None Adaxial Adaxial Adaxial None Two abaxial None None None None None

4 4 12 4 4 4 4 2 2 4 1 4 (13) 4

4 5 ... 5 5 5 5 5 4 5 ... 4 4

Late ? ... ? ? Late ? ? Late ? ... ? Late

Radial Bilateral ... Bilateral Bilateral Bilateral Bilateral Bilateral Radial Radial ... Radial Radial, bilateral Bilateral Radial, bilateral

Rhinanthoid Antirrhinoid ... Antirrhinoid Antirrhinoid Rhinanthoid ? Antirrhinoid Rhinanthoid Rhinanthoid ... ? Antirrhinoid

Tetranema Veronica

? Radial

5 54

Adaxial None

4 2

5 4

? Late

Antirrhinoid Rhinanthoid

Note. References for data in this table are as follows: 1 Armstrong and Douglas 1989; 2 Awasthi et al. 1984; 3 Bentham 1846; 4 Bentham 1876; 5 Cronquist 1981; 6 Endlicher 18361840; 7 Endress 1992; 8 Endress 1999; 9 Fernandez-Alonso 1995; 10 Harms and Reiche 1895; 11 Hoggard et al. 2003; 12 Hong 1984; 13 Kampny and Canne-Hilliker 1988; 14 Kampny et al. 1993; 15 Leme 1929; 16 Payer 1857; e 17 Philbrick and Anderson 1992; 18 Pilger 1913; 19 Prakash and Singh 1997; 20 Rahn 1981; 21 Rahn 1984; 22 Rahn 1985; 23 Rahn 1996; 24 Ronse-Decraene 1992; 25 Ronse-Decraene and Smets 1994; 26 Saunders 1939; 27 Schrock and Palser 1967; 28 Singh 1979; 29 Singh and Jain 1979; 30 Takhtajan 1997; 31 Thompson 1988; 32 Walker-Larsen and Harder 2000; 33 Weber 1972; 34 Weberling and Troll 1998; 35 Wettstein 1895; 36 Wunderlin 1992; 37 Yamazaki 1957; 38 this study.

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Table 3
Summary of Floral Development in Aragoa and Some Related Genera Taxa Aragoa Digitalis Heliohebe Plantago Veronica Seal(2) Seal(2) Seal(2) Seal(2) Seal(2) Sepl(2) Sepl(2) Sepl(2) Sepl(2) Sepl(2) Sep(1) Sep(1) * * * Sequence of oral organ development St(4) Pea(1) St(2) St(4) St(2) Pea(1) Stal(2) Pea(1) Pea(1) Pea(1) Peal(2) Peal(2) Peal(2) Peal(2) Peal(2) Pead(1) Stpl(2) Pead(1) Pead(1) Pead(1) Pepl(2) * Stmad(1) * * * Ca Ca Ca Ca Ca References This study Chatin 1873; Singh 1979; Wunderlin 1992 This study Payer 1857; Prakash and Singh 1997; this study Payer 1857; Chatin 1873; Erbar 1991; Kampny et al. 1993

Note. Sequence of events is from left to right. a abaxial; al abaxial-lateral; Ca carpels; p adaxial; Pe petals; pl adaxial-lateral; Se sepals; St stamens; Stm staminode. Numbers in parentheses indicate number of whorl parts. An asterisk indicates missing organs.

1979; Wunderlin 1992; table 3), which indicates the high residual lability of this character in closely related genera. The emergence of the stamens is simultaneous in (Veronica (Plantago Aragoa)), Collinsia (Schrock and Palser 1967) and Heliohebe (g. 6C) and unidirectional (from abaxial to the adaxial side) in Angelonia (Srinivasan 1940), Antirrhinum (Endress 1999), Digitalis (Singh 1979; Wunderlin 1992), and Globularia (Chatin 1873). In the Aragoa-Angelonia clade the simultaneous emergence of the stamens appears independently at least twice and could be derived from a unidirectional pattern (table 2). In Antirrhinum, Digitalis, and the ((Chelone Collinsia) Tetranema) clade, ve stamen primordia emerge, but the adaxial one remains rudimentary (Schrock and Palser 1967; Singh 1979; Endress 1999; Walker-Larsen and Harder 2000; table 2). In (Aragoa Plantago), Angelonia, Bacopa, Globularia, Gratiola, and Hemiphragma only four stamens emerge, although in Gratiola the two abaxial are rudimentary (Chatin 1873; Wettstein 1895). This implies that the tetramerous androecial condition has evolved at least three times in the Aragoa-Angelonia clade (g. 8). In Amphianthus, Heliohebe, and Veronica there are only two adaxial-lateral stamen primordia (g. 6; table 2), a condition proposed as synapomorphic in Veroniceae (Kampny and Dengler 1997), which apparently evolved twice in the Aragoa-Angelonia clade. However, a plesiomorphic condition for tetramery cannot be ruled out because of the absence of information about androecial development in genera such as Bacopa, Gratiola, and Hemiphragma (see also Reeves and Olmstead 1998). The dimerous and/or tetramerous androecial condition has been proposed as derived from a pentamerous state in Asteridae, including Lamiales (Ronse Decraene 1992). However, in Lamiales, the tetramerous androecium is independently derived in different groups (Ronse Decraene 1992; Endress 1994, 1996; Walker-Larsen and Harder 2000), probably related to different constraints, including specic pollination syndromes, reduction in oral apex size, zygomorphy, and unilateral mechanical constraints during ontogeny (Delpino 1886; Ronse Decraene 1992; Kampny and Dengler 1997; Endress 1998).

in Digitalis and Heliohebe (Kampny et al. 1993) (gs. 3H, 6C, 7B; table 3). This character strongly suggests early zygomorphy in the corolla of Aragoa and Plantago, despite its predominantly actinomorphic symmetry during later development of the oral buds. The corollas in these genera become actinomorphic during enlargement as the tube grows uniformly and the lobes reach the same size and shape (g. 3L, 3M; g. 7F; g. 9). The actinomorphic and tetramerous corolla of Aragoa is consistent with Endresss (2003) hypothesis that merosity reduction of the perianth is related to a change from monosymmetry to polysymmetry. In Lamiales secondary actinomorphy occurs in a number of taxa (Endress 1999). The clade (((Aragoa Plantago) Veronica) Hemiphragma), in which the corolla is actinomorphic at maturity, is nested within a group of genera with strongly zygomorphic corollas. Thus, within the Aragoa-Angelonia clade, the actinomorphic corolla in mature owers could be interpreted as a derived condition. The zygomorphic corollas of some species of Plantago and Veronica could represent a symplesiomorphy. It has been found that mutation of the

Corolla
Petal primordia emerge in an abaxial-adaxial direction in all members of the clade ((Aragoa Plantago) Veronica) and

Fig. 9 Differences in symmetry of calyx and corolla during different stages of oral development in Aragoa and related genera. O organogenesis; E enlargement; A anthesis; open circle actinomorphic; gray ellipse zygomorphic.

BELLO ET AL.FLORAL MORPHOLOGY OF ARAGOA genes CYCLOIDEA and DICHOTOMA in Antirrhinum produces polysymmetric owers (Coen and Nugent 1994; Coen 1996; Luo et al. 1996, 1999), so it is possible that mutation of orthologous genes in related genera could be responsible for the secondary actinomorphy within the Aragoa-Angelonia clade. The prevalent actinomorphic tetramerous corolla in Aragoa and Plantago during late stages of development and the tetramerous androecium are apparently synapomorphies of these two genera (g. 3M, 3O; g. 4A, 4B; g. 7B7G). Although Hemiphragma possesses four stamens and an actinomorphic corolla at anthesis, its corolla is pentamerous. In Heliohebe and Veronica, the corolla is strongly zygomorphic during development, which results from the presence of three abaxial and one adaxial petal primordia and only two stamens (g. 6C, 6D). If ontogeny frequently recapitulates phylogeny, differences in timing of a character state during development, such as changes of symmetry of calyx and corolla in Antirrhinum, Aragoa, Digitalis, Plantago, and Veronica (g. 9), would indicate that zygomorphy of both whorls is the plesiomorphic condition for these genera. However, heterochronic shifts may well reect complex developmental constraints and can undermine apparently straightforward ontogenetic sequence transformations (Hufford 1996). A reduction from pentamerous to tetramerous corollas in Lamiales can occur by fusion of the adaxial-lateral petals. For example, in Digitalis (Wunderlin 1992), Orobanche, and Pedicularis (Meier-Weniger 1977), the adaxial corolla lobe is produced by two basally united primordia; the resulting corolla lobe is frequently broader than the others (Endress 1998). Assuming that a pentamerous corolla is ancestral in Lamiales (Endress 1994; Reeves and Olmstead 1998), reduction in petal number has occurred at least three times in the Aragoa-Angelonia clade: (1) in Callitriche and Hippuris, in which the corolla is totally absent (Leins and Erbar 1988); (2) in Amphianthus, with a tetramerous corolla; and (3) in ((Aragoa Plantago) Veronica), also with tetramerous corollas. During oral development of Aragoa, Plantago, and Veronica, there is no direct evidence of fusion of two primordia to form the adaxial petal, although the adaxial petal primordium is slightly broader than the others in early development of Aragoa (g. 3H), Plantago (g. 7C; see also Eichler 1878; Saunders 1939; Endress 1998), and Veronica (Hufford 1992; Kampny et al. 1993). The tetramerous corolla could be synapomorphic for the clade ((Aragoa Plantago) Veronica) (g. 8). The presence of ve petals in some species of Plantago (Rahn 1996) and Veronica (Saunders 1939;Yamazaki 1957) could be homoplastic because species-level phylogenies of these genera show that their basalmost members have tetramerous corollas (Rahn 1996; Albach and Chase 2001; Rnsted et al. 2002). Within the Aragoa-Angelonia clade, late sympetaly occurs in Antirrhinum, Aragoa, Collinsia, Globularia, Digitalis, Plantago, and Veronica (Schrock and Palser 1967; Singh 1979; Erbar 1991; this study). In these genera, the petal primordia emerge independently and become interconnected by interprimordial growth of the alternate stamens (g. 3K3M). However, there are some differences between the late sympetaly of Digitalis and Veronica; Erbar (1991) showed that in Digitalis the two upper petals arise separately and fuse later,

735

whereas in Veronica sympetaly occurs during formation of the upper and lower lips. Erbar (1991) suggested that in these variations, the original condition may become blurred by events such as dorsiventrality and abortion in the androecium. In order to analyze sympetaly in a phylogenetic context, this character needs to be carefully studied in other genera of Aragoa-Angelonia clade such as Amphianthus, Angelonia, Bacopa, Chelone, Collinsia, Gratiola, and Tetranema. Corolla aestivation has been considered an important diagnostic character in Scrophulariaceae s.l. (Pennell 1935) because in most Antirrhinoideae, the adaxial lobes are external, whereas in Rhinanthoideae, the abaxial lobes are external in bud. Armstrong and Douglas (1989) showed that this character is evident even during early ontogeny; stronger growth occurs in the adaxial lobes in Antirrhinoideae and in the abaxial lobes in Rhinanthoideae. As Aragoa exhibits rhinanthoid aestivation (g. 4A4C), it has traditionally been included in the latter subfamily (cf. Takhtajan 1997). The distribution of this character in the Aragoa-Angelonia clade shows that antirrhinoid aestivation occurs in most of the basal genera (Amphianthus, Angelonia, Antirrhinum, Bacopa, Chelone, Collinsia, Globularia, and Gratiola) (g. 8; table 2). Thus, rhinanthoid aestivation could be a synapomorphy of the clade (Digitalis (Hemiphragma (Veronica (Plantago Aragoa)))), although this character needs detailed ontogenetic study, particularly in Plantago.

Gynoecium
The general pattern of gynoecial structure and development seems to be relatively uniform in Lamiales (Kampny and Canne-Hilliker 1988; Kampny et al. 1993; Endress 1994). The unilocular gynoecium (in Gratiola, Hippuris, and some species of Plantago) and tetralocular gynoecium (present in Callitriche by formation of a false septum; see Lemee 1929; Leins and Erbar 1988) are apomorphic in the AragoaAngelonia clade, whereas the bilocular gynoecium, which occurs in Aragoa, is plesiomorphic. Seeds of Aragoa and Plantago are remarkably similar because they are attened, winged, and reticulate (M. A. Bello, personal observation; cf. also Fernandez-Alonso 1995 and Rahn 1996), thus providing a set of possible synapomorphies between the two genera. As for the ringlike nectariferous disk with glandular-ciliate border found in Heliohebe raoulii, it is remarkably similar to that found in Heliohebe hulkeana (F. Muell.) Garn.-Jones (cf. g. 1A in Garnock-Jones 1993).

Interspecic Differences
Fernandez-Alonso (1995) found that gross morphological characters of the mature owers of Aragoa are useful to propose ve species groups. The species examined here represent three of these groups: Aragoa abietina belongs to the Abietina group; Aragoa cleei and Aragoa cupressina belong to the Cuppressina group; and Aragoa cundinamarcensis belongs to the Lycopodioides group. However, we did not observe differences in early development that would suggest deep ontogenetic differentiation between these species groups. The slight differences that we have detected are evident only during late stages, when the corolla reaches its denite shape and indumentum; corollas of A. cupressina, A. cleei, and A.

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INTERNATIONAL JOURNAL OF PLANT SCIENCES Armstrong 1985). By contrast, in Digitalis, the staminode is not vascularized (Henslow 1890), which indicates that, in this case, the vasculature disappeared before the loss of the external organ. Floral vascularization of Aragoa does not show vestigial traces of lost oral parts (g. 5). It seems that the reduction of vascular traces versus external organs is distinct in different taxa (Ronse-Decraene 1992). This requires that such reductions be evaluated with care (Sattler 1973), especially in Lamiales, in which oral synorganization of sepals, petals, and stamens is expressed in diverse combinations of reduction and fusion.

abietina are subrotaceous and have a dense indumentum on the inner side of the tube, whereas in A. cundinamarcensis, it is rotaceous with a glabrous tube. This observation is consistent with Tuckers (1984, 1997) theory that oral characters that are variable at a suprageneric level often arise during early stages of development, whereas those that are variable at generic or infrageneric levels emerge during medium and late developmental stages. In other genera of Lamiales such as Agalinis Raf., the appearance of characters that are variable at a specic level also occurs during late development (Kampny and Canne-Hilliker 1987; see also Armstrong and Douglas 1989 for further examples). However, this theory requires more rigorous testing; such hierarchical levels are articial and may not be equivalent in different groups.

Acknowledgments
We thank the staff of the Micromorphology Section, Jodrell Laboratory, Royal Botanic Gardens, Kew, especially Chrissie Prychid and Peter Gasson, for their kind assistance during the development of this study. We are grateful to J. Armstrong, Larry Hufford, and an anonymous reviewer for their comments on the manuscript. We also thank the Kew Latin American Research Fellowship program for a grant awarded to Favio Gonzalez, which partly nanced this study.

Floral Vasculature
Floral vascularization provides contradictory evidence in members of the Aragoa-Angelonia clade. In some species of Veronica and Veronicastrum, the upper lip of the tetramerous corolla is irrigated by two traces, which could be taken as evidence that this lip is produced from two fused petal primordia (Saunders 1933; Yamazaki 1957; Nishino 1983;

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