American Journal of Botany 87(10): 1408–1424. 2000. THE QUESTIONABLE RELATIONSHIP OF MONTINIA (MONTINIACEAE): EVIDENCE FROM A FLORAL ONTOGENETIC AND ANATOMICAL STUDY1 LOUIS P. RONSE DECRAENE,2,4 H. PETER LINDER,3 2 AND ERIK F. SMETS2 Laboratory of Systematics, Botanical Institute, Katholieke Universiteit Leuven, Kardinaal Mercierlaan 92, B-3001 Leuven, Belgium; and 3Bolus Herbarium, University of Cape Town, 7701 Rondebosch, South Africa The systematic position of Montiniaceae remains uncertain: a relationship with Cornales has been suggested on phytochemical and embryological evidence, while molecular data point to a relationship with Solanales. We investigated the floral development and anatomy of the South African Montinia caryophyllacea to add a new set of characters for clarifying the systematic position of the family Montiniaceae. Pistillate inflorescences show a higher degree of reduction than staminate, with flowers set terminally on short lateral branches. Flowers have an irregular initiation sequence, with frequent abortions of organs. In Montinia, petals grow rapidly, and no zonal growth takes place. The gynoecium develops as a pit surrounded by a girdle. Placentation is basically parietal and becomes axillary by the postgenital fusion of placental lobes; unitegmic ovules are arranged in two parallel rows with adjacent ovules partly overlapping each other. Unisexuality is respectively attained at the stage of anther development and carpel initiation. The floral anatomy of pistillate and staminate flowers is illustrated and discussed. Observations on Montinia are compared with data of taxa from Saxifragaceae sensu stricto, Cornales, and Solanales. The absence of sympetaly in Montinia is discussed. Morphological and anatomical evidence points to a high similarity with Escalloniaceae. Although a position in the asterids is most probable, there is little support for the relationship with Solanales indicated by molecular data. Key words: sympetaly. Cornales; Escalloniaceae; floral anatomy; floral ontogeny; Montiniaceae; placentation; Saxifragaceae; Solanales; Montinia caryophyllacea Thunb. is a dioecious woody shrub endemic to southern Africa, from the Cape Peninsula north to southern Angola and east to Port Elisabeth. There are no vegetative differences between male and female plants (Engler, 1930). Male inflorescences are described as cymose with several terminal flowers, whereas pistillate flowers are described as terminal and solitary (Takhtajan, 1997). Two or three genera are currently placed in Montiniaceae: Montinia, Grevea, and Kaliphora (Dahlgren, 1980; Thorne, 1992; Skowno, 1996; Takhtajan, 1997). Kaliphora, a monotypic Madagascan genus formerly placed in Cornaceae, was included in Montiniaceae by Capuron (1969). Thorne (1992) included a fourth genus, Melanophylla, in Montiniaceae. Takhtajan (1997) removed Kaliphora as Kaliphoraceae and Melanophylla as Melanophyllaceae. Grevea (with two species) and the monotypic Montinia share a similar wood anatomy (Carlquist, 1989; Rakouth, 1989), vegetative anatomy (Ramamonjiarisoa, 1980, cited in Al-Shammary and Gornall, 1994) and pollen (Milne-Redhead, 1955; Pastre and Pons, 1973; Hideux and Ferguson, 1976). Similar pollen was also reported for Kaliphora by Hideux and Ferguson (1976). The systematic position of Montiniaceae is far from settled. De Candolle (1828) placed Montinia in the family Onagraceae (Myrtales); this was followed by Bentham and Hooker (1867). Later, Montinia was most commonly allied with members of Saxifragaceae sensu lato (subfamily Montinioideae: Engler, Manuscript received 6 October 1999; revision accepted 4 January 2000. The authors thank Anja Vandeperre for technical assistance with light microscopy preparation, photographic processing, and artwork. This research was supported by a grant from the Research Council of the KUL (OT/97/23) and a travel grant for LRDC to South Africa in February 1998 from the Fund for Scientific Research—Flanders (F.W.O.). The leading author is postdoctoral fellow of the F.W.O. 4 Author for reprint requests (e-mail: louis.ronsedecraene@bio.kuleuven. ac.be). 1 1930; Schulze-Menz, 1964), Grossulariaceae (Cronquist, 1981; Mabberley, 1987), or Escalloniaceae (Hutchinson, 1973). The family Montiniaceae was erected by Nakai in 1943 (see Appendix). Carlquist (1989) found that similarities of wood anatomy of Montiniaceae with Myrtales (viz. Thymelaeceae, although this family is associated in Malvales by most recent systems: e.g., APG, 1998) were greater than with Rosales. Dahlgren (1975) first suggested that Montiniaceae are related with Celastrales, although with uncertainty, but later shifted the family to Cornales (Dahlgren, 1980). An affinity with Cornales was maintained by Thorne (1992), while the family was placed in the closely related Hydrangeales by Takhtajan (1997). Montinia is reported to have unitegmic ovules (Schulze-Menz, 1964), but the author does not state where he obtained this information. Unitegmic, tenuinucellate ovules are characteristic of Escalloniaceae and Hydrangeaceae, which differ in this from saxifragalean or rosalean genera. Serological evidence (Grund and Jensen, 1981), storied wood, and presence of iridoid compounds (Dahlgren, Jensen, and Nielsen, 1977; Dahlgren, Rosendal-Jensen, and Nielsen, 1981) support a cornalean affinity of these families and an asterid rather than a myrtalean link (Dahlgren, 1983). Recent molecular studies (Chase et al., 1993; Morgan and Soltis, 1993; Soltis and Soltis, 1997; APG, 1998) indicate that Montiniaceae belong to the asterids. Phylogenetic hypotheses based on cpDNA break up the original Cornales of Dahlgren into several orders belonging to different supergroups. The closest relatives of Montinia, based on rbcL and 18S rDNA data, are Convolvulaceae and Solanaceae, both placed in Solanales (Euasterids I), far away from the Cornales (basal in the asterids). Interestingly, in the study of Cosner, Jansen, and Lammers (1994) and Fay et al. (1998), Montinia appears as sister to Sphenoclea (Sphenocleaceae) and Hydrolea (Hydrophyllaceae), thus forming a sister clade to the Solanales. However, the argumentation for a solanad affinity appears meager 1408 October 2000] RONSE DECRAENE ET AL.—FLORAL DEVELOPMENT AND ANATOMY OF besides the molecular data, as Morgan and Soltis (1993) enumerate more differences than similarities with members of the Solanales. We investigated the floral ontogeny of pistillate and staminate flowers of Montinia caryophyllacea to question the systematic position of the family. The absence of a corolla tube in Montinia especially demands clarification, given the systematic importance attached to it (e.g., Erbar, 1991; Erbar and Leins, 1996). The systematic importance of the floral anatomy has been repeatedly demonstrated by several authors (e.g., Eyde, 1967; Bensel and Palser, 1975a, b; Armstrong, 1986; Ronse Decraene and Smets, 1991, 1999a, b; Ronse Decraene, De Laet, and Smets, 1998). A limited morphological study of mature flowers of Montinia has been made by Skowno (1996). We studied the floral anatomy of staminate and pistillate flowers of Montinia, in addition to floral ontogeny. MATERIALS AND METHODS Flower buds of Montinia were collected from the base of Nursery Ravine, Table Mountain, Cape Town. Material was prepared with customary methods (see Ronse Decraene, De Laet, and Smets, 1998). Voucher material is kept at the botanical institutes of the Katholieke Universiteit Leuven (LV) and the University of Cape Town (BOL). Material was fixed in FAA (85 mL ethanol 70%, 10 mL acetic acid, 5 mL formaldehyde 40%). The buds were transferred to 70% ethanol and dissected under a Wild M3 dissecting microscope. The material was washed repeatedly in 70% ethanol and dehydrated by putting the buds in a 1:1 mixture ethanoldimethoxymethan (DMM or formaldehyde-dimethylacetal) for 5 min and in pure DMM for 20 min. Buds were critical-point dried using liquid CO2 in the CPD 030 (Balzers, Liechtenstein). The dried material was mounted on aluminium stubs using Leit-C (after Göcke) or double tape and coated with ;180 nm of gold (Spi-moduleTM Sputter coater of Spi Supplies, West Chester, Pennsylvania, USA) before observation with the Scaning Electron Microscope (SEM.) For light microscopy, preanthetic buds were analyzed and customary methods of preparation were used. The material was run through an alcohol as well as an alcohol-tertiary butyl alcohol series and was next embedded in paraffin. Serial sections, ;8–11 mm thick, were stained with safranin and counterstained with fast green or cotton blue using the automatic staining machine Varistain 24–3 (Shandon, Runcorn, UK). Camera-lucida drawings and photographs were made under a Leitz Dialux 20 equipped with a Wild MPS 45/51 photoautomat. RESULTS Organography—Flowers have an inferior ovary, large creamy-white petals, and small sepals. Pistillate flowers possess a stout bisected style ending in four stigmatic lobes. Antesepalous staminodes bear sterile anthers. The fruit is a loculicidal woody capsule with an axile placentation. Ovules are arranged in four rows, with adjacent ovules of two neighboring rows overlapping each other as tiles. Staminate flowers have four alternipetalous stamens surrounding a cuplike disc. Carpellodes can still be seen as shallow protuberances in the center of the flower. Organogeny—Unless otherwise stated, male and female inflorescences will be considered together. Inflorescences arise terminally. They consist of a single terminal flower, with a few associated flowers arising basipetally on lateral branches. In staminate inflorescences, a number of lateral flowers are associated with the top flower and arise basipetally and spirally in the axil of a bract (Figs. 4, 6, 26). A smaller lateral flower may be visible within a secondary bract (Fig. 4, arrow), but generally there is no further branching and no bracteoles. Low- MONTINIA 1409 er down the axis lateral branches may be formed with a similar development. The inflorescence is thus basically monotelic and paniculate (sensu Weberling, 1989). In pistillate plants there are similarly top flowers, but the inflorescence differs from the staminate inflorescence in that the top flower stands alone, except for some isolated cases of a lower flower in the axil of a bract. More generally a number of spirally inserted lateral branches are formed basipetally, each ending with another flower. Bracts subtending the lateral branches become progressively leaflike from top to bottom of the main inflorescence. On each lateral branch a variable number of smaller second-order bracts arise in a spiral sequence approaching a decussate pattern (Figs. 1, 2, 3, 9, 16). The base of the leaves and larger bracts are swollen and contain a large number of multicellular uniseriate hairs on the inside. A first pair of bracts emerges sequentially in transverse position (Fig. 2). While they curve over the growth apex of the lateral shoot, unicellular trichomes start to grow on their upper surface. Another pair of bracts arises higher up in the median region but is obliquely inserted (Figs. 1, 2, 16). A third pair may be initiated in a latero-transverse position, and finally a last one in latero-median position (Figs. 1, 2). In several cases this number was not attained (e.g., Figs. 3, 16), and in an extreme case only the first two bracts were initiated (Fig. 5). As a result it was sometimes difficult to decide at what moment initiation of the flower really starts. The orientation of the terminal flower on the lateral branches relative to the main axis is not orthogonal, but clearly diagonal (Fig. 16). The former case would occur when the last formed bracts are more or less in median position. No true bracteoles are produced. Four sepals emerge sequentially but with a variable initiation sequence. Often the lateral sepals emerge first and successively (Figs. 4, 6, 8, 9), followed by the abaxial median sepal and finally the adaxial median. Very often the initiation of the adaxial sepal is retarded, and this leads to an asymmetric shape of the receptacle and an irregular initiation of the upper floral organs (Figs. 10, 14, 20). In other cases a regular tetramerous arrangement was rapidly attained by the equal development of the floral bud. In some cases growth of the adaxial sepal precedes that of the abaxial (Fig. 5), or one of the lateral sepals is conspicuously smaller than the others (Fig. 14). In other cases the median sepals emerge before the laterals (Figs. 5, 7, 12). The sepals are inserted on the radii relative to the main axis in case there is only one bract subtending the flower, as in staminate inflorescences (Figs. 4, 6, 8). Sepals grow rapidly. The difference in size of the sepals remains visible during later stages of development, and the outer sepals are inserted lower and externally of the median sepals (Figs. 19, 20); aestivation is imbricate. At anther formation the sepals are exceeded in size by the petals and are pushed away from each other. Finally sepals remain visible as small triangular lobes arranged side by side on the periphery of the flower. During later stages of development the sepal bases become lifted by zonal growth linked with the development of epigyny. Sepal lobes are inserted on a shallow rim (Figs. 38, 40) and bear numerous stomata adaxially. The petals arise as hemispherical primordia directly on the floral meristem. No ring primordium has been detected in early stages (Figs. 7–9). The order of initiation of the petals can be variable: in some cases initiation is almost simultaneous on the four corners of a square floral primordium (Figs. 8, 9), but in others it is sequential (Figs. 10, 11, 15). Growth is generally unequal, even in those cases with a simultaneous initiation; 1410 AMERICAN JOURNAL OF BOTANY [Vol. 87 Figs. 1–9. Inflorescence, sepal, and petal initiation of Montinia caryophyllacea. 1. Early initiation of lateral shoot, abaxial view. Note the presence of three decussate pairs of bracts. 2. Other view of slightly older shoot, abaxial view. Note the displacement of the bract pairs relative to each other. The decussate pattern is disturbed above. 3. Adaxial view of young shoot. An odd primordium arises between two bracts. 4. Early sepal initiation (staminate); the lateral sepals precede a median pair. Note the presence of an odd bract with lateral primordium (arrow). 5. Other staminate bud showing the presence of two external bracts surrounding the sepals, abaxial view. 6. Partial view of staminate inflorescence developing basipetally. Note flowers subtended by a single bract. Upper flower October 2000] RONSE DECRAENE ET AL.—FLORAL DEVELOPMENT AND ANATOMY OF the petals grow as two distinct pairs, with the abaxial preceding the adaxial in size (Figs. 13, 15, 17, 19). The unequal development of the floral meristem influences the shape and initiation sequence of the petals; it appears trapezoid, not square, with one side more developed than the other (Figs. 10, 11, 14, 15). In some cases the abaxial petals attain the size of the last formed sepal (Figs. 15, 20). The petal primordia rapidly become dorsiventrally flattened and triangular in section (Figs. 14, 15, 18). Aestivation is most often imbricate ascending: the largest abaxial petal covers all others; the adaxial petal on the opposite side is completely enclosed (Figs. 19, 20, 5, 48, 49). In other cases the largest pair covers the smaller one (Fig. 53, K-N). During growth the petals remain widely separated from each other by the alternating insertion of stamen primordia. Petals rapidly outgrow the sepals, which remain attached as small lobes. The white petals are conspicuous at maturity (about five times the size of the sepals) and have one dendricularly branching midvein (Figs. 48–49, 53–54). There are usually four stamens present, but the number can fluctuate between three and five, depending on variations of the merosity of the flower or on space limitations. As soon as the petal primordia have been initiated, stamen primordia arise in the alternating spaces, almost equidistantly from the center as the petals (Figs. 9, 13, 17). Stamen primordia can arise simultaneously, but are often unequal in size, corresponding to the unequal development of the petals and the distorted floral apex. In some cases initiation appears clearly sequential with one side of the flower preceding the other (Figs. 11, 12, 15). In a few cases flower buds were trimerous with three petals and three stamens (Fig. 18). The fourth sepal was conspicuously smaller than the three others, and one of the petals appeared as two-lobed. In other cases a fourth stamen lying opposite the smallest sepal may not arise at all due to space restrictions in the flower (Fig. 14). During development of the stamens the floral apex grows progressively from a flattened surface to a shallow depression. The stamen primordia grow horizontally towards the depression and broaden towards the apex between the intervening petal lobes (Figs. 17, 20). As the petal lobes start to grow over the bud the stamen apices have converged to the center of the flower and differentiate two broad thecae. By a stronger abaxial development two unequal pollen sacs are differentiated on each theca (Figs. 24– 27). At the same time a short filament is produced. In pistillate flowers stamen development usually ends at this stage, but there is some variation in the extent of development of the staminodes, and within the same bud size differences may be conspicuous (Figs. 25, 38). In staminate flowers stamen development is rapid with the growth of two elongated thecae separated by a broad connective (Figs. 26, 27). The filament elongates shortly before anthesis. The thecal orientation is first introrse, but becomes extrorse at maturity by a stronger growth of the ventral area of the anther (Fig. 45; see also Endress and Stumpf, 1991). At maturity the connective is inconspicuous. The anther surface and upper part of the filament consist of MONTINIA 1411 papillate cells. No orbicules were found within the anther. Pollen is tricolporate with Pole/Equator index 1, size 32 mm (Fig. 46); the ektexine is reticulate with the surface covered with small spinules (not visible in Fig. 46). Gynoecium initiation starts as a shallow depression surrounded by the stamens at the time they start differentiating anther tissue. The upper portion of the cavity region develops as a girdling primordium (Fig. 21), and finally two horseshoeshaped carpel primordia are delimited (Figs. 22, 23). The carpel primordia are situated opposite two staminodes, either in latero-median or latero-transversal position. Differences between staminate and pistillate flowers start late in ontogeny. In staminate flowers carpellary primordia soon stop growing and are rapidly overtaken by the tetrasporangiate anthers (Fig. 27). As the surrounding tissue keeps growing the carpellary lobes are soon absorbed in the receptacle and appear at maturity as two (Fig. 41) or occasionally a single (Fig. 40) shallow crescent-shaped emergence that is fully embedded in disc tissue. In some cases gynoecia are completely resorbed. In pistillate flowers the style results from upward extension of the early circumferential growth at the top of the gynoecial cavity (Fig. 23). The style forms two distinct centers of growth which extend upwards between the anthers. The basal confluent zone of the style remains very short, and most extension growth occurs within the lobed region, which shows a deep median invagination (Figs. 24, 25, 29). The upper part of each carpel curves outwards and differentiates stigmatic papillae along the slit (Figs. 30, 36). The papillae grow into long stigmatic hairs, which cover the extended U-shaped stigmatic area (Figs. 37, 38). In the upper stigmatic region the two carpels appear thus as four because the styles are deeply split (Figs. 25, 38, 53, N). Two short compressed styles are thus formed with a narrow central canal (Figs. 29, 30). The styles sit on a massive disc covering the inferior ovary. The limits between style and disc are indistinct. The basal part of the style is covered with long rectangular cells, which become smaller and papillate towards the disc. Both style and disc are interspersed with anisocytic deeply sunken stomata and have a deeply striate cuticle (Figs. 38, 39). In all probability this disc functions as a nectary, although we did not test for nectar secretion. Tissue adjacent to the disc surface is darkly staining in section (Fig. 51) and supplied with numerous veins (Figs. 51, 53, 54). In staminate flowers a similar papillate epidermis occurs on the receptacle, also covering the sterile carpel remnants. However, no stomata were found on the disc. During carpel initiation the area below the insertion of organs deepens progressively and differentiates as the ovary. Placental tissue arises within the receptacle when the stylar lobes already have a considerable development (Fig. 28). At the level of fusion of the margins of neighboring carpels, tissue becomes inflated while the ovary deepens progressively. As this tissue extends in size, it becomes coalescent with similar tissue at the opposite side and eventually fuses basally. By ← at stage of stamen development. 7. Early petal initiation, abaxial view, with one sepal removed. Note that the median sepals enclose the laterals. 8. Lateral view of early petal initiation, three sepals removed. 9. Lateral view of early petal and stamen initiation, three sepals removed. Note diagonal orientation of flower relative to the bracts. Bars 5 100 mm. Figure Abbreviations: A, stamen or staminode primordium or trace; B, second order bract of lateral inflorescence; C, sepal or sepal median trace; D, dorsal carpellary trace; K, petal or petal trace; M, marginal carpellary trace; P, placenta; SE, septum; V, ventral carpellary trace. Asterisks point to the position of the main bract subtending the lateral inflorescences 1412 AMERICAN JOURNAL OF BOTANY [Vol. 87 Figs. 10–18. Petal and androecium development of Montinia caryophyllacea. 10. Apparently trimerous bud by the unequal initiation of petals, sepals, and stamens. 11. Apical view of flower with unequal initiation of four petals and two stamens. 12. Abaxial view showing irregular flower and unequal stamen initiation. 13. Flower with slightly unidirectional development of the organs. 14. Apical view of compressed bud with three stamens initiated. 15. Unequally developing flower, abaxial view. A distinction between bract, sepals, petals, and stamens is difficult to make. 16. View of whole lateral shoot at stamen development. Note the pattern of arrangement of bracts and flower orientation. 17. Detail of flower showing the insertion of stamens and petals. 18. Trimerous flower. The right petal is larger (arrow) and stands opposite one of the sepals (only slightly visible). Bars 5 100 mm. October 2000] RONSE DECRAENE ET AL.—FLORAL DEVELOPMENT AND ANATOMY OF MONTINIA 1413 Figs. 19–27. Anther and gynoecium development in Montinia caryophyllacea. 19. Lateral view of flower with petals covering floral apex. Note unequal size of petals and lower insertion of the lateral sepal bases. 20. Lateral view of terminal staminate flower; one smaller sepal is out of sight. The stamens cover the central area of the flower. 21. Early gynoecial initiation of trimerous flower, with stamens and petals removed. 22. Later stage of gynoecium development showing two prominent growth zones. 23. Older stage showing the two strongly dissected stylar appendages. 24. View of four staminodes separated by upward growth of the styles. 25. Nearly mature flower. Note that the staminodes and perianth are lifted up by peripheral zonal growth. 26. Lateral view of young staminate flower. 27. Similar partly dissected staminate flower, showing two central carpellodes. Bars 5 100 mm. 1414 AMERICAN JOURNAL OF BOTANY [Vol. 87 Figs. 28–36. Placentation and ovule development of Montinia caryophyllacea. 28. Longitudinal section of ovary showing early placenta formation. The arrow points to the fused basal area 29. Lateral view of partly dissected ovary and first traces of ovules. Note the strongly invaginated style belonging to one carpel. 30. Lateral view of dissected ovary with ovule primordia and early stigma development. 31. Detail of three developing ovules. 32. Lateral view of two rows of older ovules belonging to different placentae. Arrow points to strong invagination of the carpel 33. Lateral view of placentation at the stage fusion has occurred. Note the narrow septal attachment and rows of intercalating ovules. 34. Detail of older ovule, showing the dorsal extension of tissue. Arrow points to the position of the micropyle. 35. Detail of a micropyle. 36. Apical view of young style with two staminodes. Note the development of stigmatic papillae. Bars 5 100 mm. October 2000] RONSE DECRAENE ET AL.—FLORAL DEVELOPMENT AND ANATOMY OF MONTINIA 1415 Figs. 37–46. Late development of the flower of Montinia caryophyllacea. 37. Partly dissected mature pistillate flower. Note the extent of development of the massive placentas. 38. Mature pistillate flower, petals removed. Note the hairy bilobed stigmas, the massive disc with stomata, and the antesepalous staminodes. 39. Detail of the papillate disc with sunken stomata. 40. Mature staminate flower with single carpellode, anthers and petals removed. 41. Detail of two carpellodes of mature staminate flower. Note the papillate surface of the disc. 42. Nearly mature ovule. Note the position of the micropyle (arrow). 43. Detail of branching multicellular trichome. 44. Longisection of teratological flower. Note the presence of a gynoecium within the ovary and two normal ovules (arrows). 45. Abaxial view of mature stamen. Note the slightly extrorse position of the anthers and the lateral dehiscence. 46. Equatorial view of pollen grain situated at the stomium of the anther. Figs. 37, 38, 40, 44, bar 5 1 mm; Figs. 39, 41, 42, 45, bar 5 100 mm; Figs. 43, 46, bar 5 10 mm. 1416 AMERICAN JOURNAL OF BOTANY [Vol. 87 Figs. 47–52. Longisections and transverse sections of Flowers of Montinia caryophyllacea. 47. Longisection of preanthetic pistillate flower. Note the deep invagination within the placental tissue and the arrangement of the ovules and their vascular traces in longitudinal rows. The letters refer to the level of sections on Figs. 53. 48–49. Successive sections of staminate flowers. 48. At level of anthers. 49. At level of disc and filaments. Arrows point to carpellodes 50. Detail of anther with one of the pollen sacs. 51. Longisection of pistillate flower showing detail of petal, style, and nectariferous disc with vascular supply. 52. Longisection of row of ovules, showing the strong dorsal expansion and lack of funiculus. Figs. 47–49, bar 5 0.52 cm; Figs. 51–52, bar 5 0.18 cm; Fig. 50, bar 5 0.06 cm. later postgenital fusion of the two placental ends a massive central placenta is formed delimiting two locular chambers (Figs. 33, 37, 47, 53). A variable number of ovules arises basipetally in two rows on each placenta, even before connection of the two placentae (Figs. 29, 30) and, depending on the flower, varies from (1)-2 to 6 per row. Ovules are globular first, but rapidly develop a short curving funiculus. A single integument envelops the nucellus, leaving a narrow micropyle (Fig. 31). The ovules become elongated and curve downwards (Fig. 32), while a dorsal ridge develops beyond the insertion of the funiculus (Figs. 33, 34, 52). The funiculus is finally absorbed in the ovular tissue (Fig. 42). The micropyle is narrow and lies along smooth placental tissue (Figs. 35, 37, 44). The ovules of neighboring placentae are turned away from each other and later cover each other back to back as tiles on a roof (Figs. 32, 33, 37, 53, F). A margin of smooth placental tissue appears next to the connection of the placenta to the ovary wall. This connection (or septum) is narrow compared to the massive placenta (Figs. 33, 37). At the base of the ovary a continuous septum is found dividing the ovary in two October 2000] RONSE DECRAENE ET AL.—FLORAL DEVELOPMENT AND ANATOMY OF MONTINIA 1417 Fig. 53. Camera lucida drawings of transverse sections of pistillate flower buds of Montinia caryophyllacea; letters correspond to the levels of sectioning shown in Fig. 47. (A) Section of pedicel just below the flower. Note invaginating tissue (arrows). (B) The invaginating tissue has rejoined the peripheral tissue at two points (arrows). (C) View of one of the placental lobes. Note two bundle complexes from which a ventral converges centrally (arrow). (D) Section with two placental lobes visible. The second ventral bundle converges to the center (arrow). (E) Section at level of first ovules. The placental lobes are supplied by numerous branching traces running horizontally to the ovules. (F) Section of middle of ovary with specific arrangement of overlapping ovules. The peripheral bundles branch profusely. (G) Section above the insertion of the upper ovules with receding placental lobes. The positioning of peripheral traces becomes more clear. (H) Top of placental lobes with ventral traces still present. The dorsals are clearly visible, as well as the position of sepal medians, staminodes and petals. (I) Section at upper level of placental lobes. (J) Section at the base of the disc. Note the position of the dorsals and numerous disc traces. Marginal traces have been derived from the peripheral tissue on either side of the dorsals. (K) Section at the level of the comon stylar part. Only the marginal traces are visible. (L) Section at the level of the stylar lobes. Note the invagination of each carpel. (M) Section at the base of the stigmatic lobes. (N) Section in the middle of the stigmatic lobes. Bar 5 0.52 cm. (un)equal locules. In some instances anomalous flowers were found with an ovule developing into an entire ovary (Fig. 44). Seeds develop as flattened, curved structures, winged on the dorsal side (Fig. 42). We did not study older stages of development. Floral anatomy—Pistillate flowers—The pedicel contains a continuous vascular cylinder, which is similar in staminate flowers (Fig. 54A). Just below the base of the flower the vascular tissue breaks apart in several bundles, while a number of traces are given off tangentially and converge to the central pith (Fig. 53A). The bundles rearrange at the periphery in broad masses of vascular tissue. About 8–9 peripheral bundles were counted, but some are highly dissected; two bundles are conspicuously bigger and are formed by the convergence of tissue that has been separated to the center at lower level (Fig. 53B–C). From the latter bundles small traces split off and converge to the narrow ledge separating the two locules (Fig. 53C–D). These bundles, which we name the ventrals, converge to each other at the level of the common placental lobe. They branch profusely and eventually fuse (Fig. 53D–E). At this level the peripheral bundles have split in a higher number of 1418 AMERICAN JOURNAL OF BOTANY [Vol. 87 with sepal median traces. At a lower level, the stamen traces and sepal median traces are fused. In between one finds several smaller traces, which could be confused with deposits of tannins but are mostly the supply to the disc and the lateral supply of the sepals (Fig. 53H–I). The lateral sepal supply is either derived from the sepal medians, from the petals, or from independent bundles. The placental lobes diminish in size higher up, becoming shallow emergences before disappearing from sight (Fig. 53I–J). At that level the ventral supply stops short. Next to each of the dorsal traces two marginal traces appear between the smaller disc traces (Fig. 53J). The peripheral sepal median traces give off two lateral traces and at this level the stamen trace is well separated. At a higher level just below the style the central stylar canal is surrounded by a girdle of smaller traces together with the dorsals and accompanying marginals. Slightly higher the dorsals and marginals converge to the slit, while the disc traces spread out below the disc surface (Fig. 53J). The distance between staminode traces, sepal medians, and sepal laterals increases. At the level of separation of the style the dorsal trace ends and the marginal traces continue up the style (Fig. 53K). Slightly higher the style is split apart in two horseshoe-shaped appendages containing two marginal traces each (Fig. 53L–M). At this level stigmatic papillae are visible abaxially. Further up four stigmatic triangles can be seen opposite the petal lobes; each contains one of the marginal traces (Fig. 53N). Within the sepal ring several traces can be seen; higher up only the median remains visible before fading out (Fig. 53K–L). A single trace runs into the petal and splits into three traces and higher up into several traces (Fig. 53J–N). Fig. 54. Camera lucida drawings of transverse sections of staminate flower buds of Montinia caryophyllacea. (A) Section through pedicel with eight main interconnected bundles. (B) Section in the ovary region with differentiation of eight peripheral bundles and two dorsal bundles. Note common bundles for sepal medians and stamens.; arrow points to division of sepal median and stamen trace. (C) Section at separation of staminal traces from the common bundles with the sepal median traces. Arrow points to division of sepal median and stamen trace. The petal bundles give off sepal lateral traces. (D) Section below the level of the disc. Note the presence of numerous disc traces and the expansion of sepal lateral traces. (E) Separation of carpellodes, petals and sepals. Bar 5 0.52 cm. traces running through the ovary wall. Towards the level of ovule insertion the ventrals, or derivatives, branch profusely and provide a horizontally running supply to each ovule (Figs. 47, 52, 53E–F). In longisection, superposed tiers of vascular supply can be seen, corresponding to the supply of the ovules (Fig. 47). At the level of ovule supply the placenta is almost rectangular. The peripheral bundles remain unchanged during most of their course through the ovary wall. Above the level of insertion of the last ovule the placentae separate while the ventral supply continues upwards in each placental lobe (Fig. 53G, H). At this level the peripheral bundles rearrange in two girdles of traces that will be connected to the organs of the flower. More centrally two dorsal traces can be seen, positioned perpendicular to the ventrals; these can eventually be followed down the ovary wall (Fig. 53H). The peripheral traces consist of large petal traces and intervening stamen traces Staminate flowers—The supply to the staminate flowers is very similar to that of the pistillate, although it is more simplified. The central part of the anther is broad with the pollen chambers inserted laterally. The connective is inserted dorsally. The endothecium is single layered, and the dehiscence of the anther occurs laterally (Fig. 50). The filaments are inserted peripherally around a central disc that follows the concave shape of the flower (Figs. 48–49). Two hemispherical carpellary lobes can be seen (Figs. 49, 54E). Below the disc, no remnants of the ovary are found, except for the two dorsal traces, which continue down the whole inferior part of the flower before converging and fusing with the peripheral bundles (Fig. 54B–D). At this level several smaller traces to the disc can be seen. The median traces of the sepals get connected to the stamen traces (Fig. 54B–C), while the laterals fuse either with the sepal median traces, converge directly to the receptacle, or become connected to the petal traces. Lower in the receptacle all traces converge into a ring and form a siphonostele (Fig. 54A). Compared to pistillate flowers the vasculature of staminate flowers is less confusing, and eight peripheral bundles can be easily followed down the flower. DISCUSSION The pistillate inflorescence of Montinia presents the reduction of a cymose configuration. A description of the pistillate inflorescences as restricted to a terminal flower (e.g., Engler, 1930) is incorrect, since the inflorescence consists of a terminal branch surrounded by a number of partial inflorescences. Each lateral branch, (including the top flower on the main branch) ends with a single flower. A number of decussately inserted bracts precedes flower initiation (Figs. 1–3). These October 2000] RONSE DECRAENE ET AL.—FLORAL DEVELOPMENT AND ANATOMY OF MONTINIA 1419 bracts enclose a flower only exceptionally (Fig. 4). No true bracteoles (prophylls) are present in Montinia. It could be hypothesized that the ancestral inflorescence was a system of cymes with all flowers on the branches reduced, except for the upper one. This characteristic arrangement is partly retained in staminate inflorescences. Similar cases of sterilization, but with the eventual incorporation of the upper bracts as an epicalyx are found in Dipsacales (Hofmann and Göttmann, 1990; Roels and Smets, 1996). For example, Patrinia (Valerianaceae) and genera of the Linnaeeae (Caprifoliaceae) have flowers subtended by three pairs of decussately arranged phyllomes. Both pistillate and staminate flowers possess evidence of the other sex. However, the process of sterilization has gone further in staminate flowers: the gynoecium is limited to small carpellodes with their dorsal traces. There are no traces of placental or ovular tissue. This implies that the onset of sterilization always occurs at the same developmental level, viz. the initiation of carpel primordia. Pistillate flowers have stamens developing until the stage of anther formation prior to meiosis. The trend to dioecy also involves changes in other characters, such as inflorescence development (see above) and pollinator attraction. The trend to unisexuality implies the existence of a progressive process in this case, not a sudden mutation leading to highly different morphotypes. At a certain stage of development genes become arrested in their expression and an organ fails to attain maturity. This may indicate that unisexuality is a phylogenetically recent development in Montinia. Similarly, the acquisition of dioecy occurs at different rates in pistillate and staminate flowers of Rhus hirta (Anacardiaceae) (Gallant, Kemp, and Lacroix, 1998), although early developmental stages are similar between both sexes. Once a male or female reproductive organ has been sterilized, it either aborts completely in a next evolutionary step or it gains a new function. In the latter case sex determination occurs before the initiation of the flowers, when there are no traces of the other sex. This has been illustrated for Carica papaya (Ronse Decraene and Smets, 1999b) where the pistillode has the function of a nectary in staminate flowers. We often observed that the floral apex becomes distorted by the unequal development of the sepals. This leads to a retardation of organ initiation, unidirectional inception of primordia, and eventually to the loss of primordia. Flowers were occasionally trimerous in petals and stamens (Figs. 10, 18). This distortion happened almost exclusively on lateral flowers, suggesting that pressures of bracts are important. Alternatively, loss of organs could also be caused by insufficient nutrient allocation in lateral flowers, which arise at a later stage. Lack of space on the irregular floral apex is related with the loss of some organs. One (often broader) petal stands opposite a sepal (Fig. 18), which suggests that it has become derived by fusion. A similar variation in development has been observed in Chrysosplenium (Saxifragaceae) by Ronse Decraene et al. (1998). This case illustrates the potential for the derivation of trimerous flowers from tetramerous forms (reviewed in Ronse Decraene and Smets, 1994). Interestingly, staminate flowers of Grevea are described as trimerous in the corolla and androecium (Hutchinson, 1967). were available for comparison. Descriptions by Baillon (1884) and Hutchinson (1967) of Grevea demonstrate important additional similarities in the absence of stipules, the existence of tufts of hairs at the base of the leaves, dioecy with similar pattern of distribution of sexual organs in the unisexual flowers, and subextrorse anthers. The same distinction of cymose staminate flowers and solitary pistillate flowers is described as for Montinia. Grevea differs in the fewer (4–5) erect ovules on parietal (Baillon, 1884; Engler, 1930) or axile (Hutchinson, 1967) placentae. Kaliphora shares unisexual tetramerous flowers (not dioecious) and two recurved styles. However, contrary to Montinia staminodes are absent in pistillate flowers, and there is a solitary ovule in each locule (Hutchinson, 1967). Other differences are enumerated by Takhtajan (1997). A fourth genus, Melanophylla, placed in Montiniaceae by Thorne (1992) and sharing a similar pollen (Hideux and Ferguson, 1976), differs in its bisexual flowers, glandular trichomes, presence of bracteoles, obscure or absent nectary, and single pendulous ovule in each locule (Hutchinson, 1967; Takhtajan, 1997). Trichome anatomy of Montinia has been investigated by AlShammary and Gornall (1994). The spaghetti-like trichomatic mass consists of uniseriate eglandular hairs. Flowers do not possess any trichomes. Contrary to their report that trichomes occur on the petiole of the leaf and on the nodes only, we found trichomes on the margin and top of the leaf, and, occasionally, scattered along the main vein on the upper surface. We also found that the trichomes are occasionally bi- to multiseriate, especially at the edges of leaves (Fig. 43). Grevea has the same kind of trichomes scattered all over the leaf surface. No information is available on Kaliphora. Al-Shammary and Gornall (1994) pointed to an affinity of Vahlia with Montinia, based on similar uniseriate eglandular trichomes, the opposite, exstipulate leaves, the one-trace unilacunar nodes, a bicarpellate ovary with tenuinucellate ovules, and the presence of iridoids. Vahlia appears to be in the same group of asterid families that contains Montinia (Morgan and Soltis, 1993). However, Vahlia differs in possessing bitegmic ovules (Takhtajan, 1997). A floral ontogenetic investigation of the monospecific Vahliaceae would be worthwhile. Due to lack of data we will not include Vahlia in our discussion. Several potential relatives for Montinia have been proposed in the past (see introduction). In the following we will analyze the potential relationships of Montinia with Solanaceae, Sphenocleaceae, Saxifragaceae, Cornaceae, Hydrangeaceae, and Escalloniaceae. A comparison of characters is summarized in Table 1. The study of floral ontogeny of putatively related taxa of Montiniaceae is instructive and can throw light on the relationships of Montinia by the presence of shared synapomorphies. A comparison with the development of other tetramerous flowers can especially give evidence of a comparable initiation sequence. Floral ontogeny of taxa with inferior ovaries originally grouped in a polymorphic Saxifragaceae sensu lato has been carried out in a number of species (e.g., Ribes—Payer, 1857; Hydrangeaceae—Roels, Ronse Decraene, and Smets, 1997; Escallonia—Payer, 1857; Itea—Vandeputte, 1993; Saxifragaceae—Klopfer, 1973, Gelius, 1967; Ronse Decraene et al., 1998). The systematic relationships of Montinia—Montinia, Grevea, and Kaliphora make up the family Montiniaceae. In the introduction we already pointed to several shared characters. However, no floral materials of Grevea and Kaliphora Saxifragaceae sensu stricto—Differences in the early ontogeny of Saxifragaceae sensu stricto are obvious (e.g., Payer, 1857; Klopfer, 1973, Ronse Decraene et al., 1998), including the slow growth (even absence) of the petal primordia, sta- 1420 TABLE 1. AMERICAN JOURNAL OF BOTANY [Vol. 87 Distribution of some characters in Montiniaceae and putatively related families.a Cornaceaeb Montinia Solanaceae Escalloniaceaec Hydrangeaceae Saxifragaceae s.s Presence of tetramery Sympetaly, presence of common stamen-petal tube Haplostemony Broad intrastaminal nectary disc on top of ovary Vascular supply of nectary derived from peripheral traces Single seed per locule Occurrence of epigyny Scanty vasicentric parenchyma 1 1 2 1/2 1/2 1/2 2 2 1 2 2d 2 1 1 1 1/2 1 1/2 1 1 2 1 1 1 well developed absent absent well developed well developed absent 2 1 2 2 2 2 Perforation plates in primary xylem Vestured pits Presence of secoiridoid substances Endosperm development Endothecium c 1 1 2 1 1/2 1/2 1 2 2 2 1/2f 2 simple scalariform ? scalariform ? 1 1 1/2 1 1 (Cestrum) 2 ? 1 simple/ scalariformg 1/2 1 cellular? cellular cellular nuclear cellular (helobial) one-layered ? one(2)-layered 1-2-3 layered one-layered Unitegmic tenuinucellate ovules Axial system of ventral strands Ventral bundles 1 2 nuclear/cellular (helobial) one-to multilayered 1 1 1 2 1 2 2 1 1 1 fused in complex free, and in pairs Sepal laterals branching of midvein, and supply from petals Placentation basically 1 parietal Stylar supply marginal traces, with dorsals at the base Plants dioecious 1 Presence of prophylls 2 Petal growth Nodal vasculature rapid unilacunar Pollen nuclear number binucleate Aluminium accumu- 2 lation Presence of orbicu2 lesh ? 2 free and in 2 fused in complex free, or variously fused in complex pairs, or 2 fused fused bundles absent—branching branching of sepal branching of sepal absent—branching branching of comof petal supply, midvein, and midvein and of sepal midmissural bunsepal midvein, commissurals commissurals, vein and comdles or both or fused with missurals petal supply 1 2 1/2 1 1/2 dorsal traces dorsal traces 1/2 2 1/2 1 dorsal and ventrals, or only ventrals 2 1/2 dorsals and ventrals, or only dorsals 2 1 ventrals, occasionally with dorsals 2 1 rapid slow tri-(multi-) lacunar unilacunar rapid rapid tri-(multi-) lacunar unilacunarf binucleate 1 binucleate 2 ? 1 trinucleate 1 slow tri-(uni-, multi-) lacunar binucleate ? ? 1 1 1 1 a Data are from Armstrong (1986), Bensel and Palser (1975a, b), Carlquist (1989), Dahlgren (1990), Erbar (1997), Endress and Stumpf (1991), Eyde (1966), Huysmans, El-Ghazaly, and Smets (1998), Jansen, Smets, and Baas (1998), Klopfer (1973), Metcalfe and Chalk (1979), Murray (1945), Philipson (1967), Wilkinson (1944), Stern (1974), personal observation on Cornus sanguinea and Hydrangea quercifolia. Absence of aluminium accumulation was tested for Montinia by the first author; other information was provided by S. Jansen (K. U. Leuven, personal communication). bAccording to Takhtajan (1997), including Cornus, Afrocrania, cynoxylon, and Swida. cAccording to Takhtajan (1997), including Polyosma, Anopterus, Cuttsia, Forgesia, Quintinia, Escallonia, and Valdivia. Admittedly the family as described here, is heterogeneous. Our information is based on Polyosma and Escallonia. dThe statement that Escalloniaceae show a tendency for sympetaly (e.g., Dahlgren, 1983; Morgan and Soltis, 1993) is based on an erroneous affirmation of Krach (1977). eall Solanaceae are hypogynous, except for Nothocestrum latifolium, which is perigynous (Armstrong, 1986). fBased on Escallonia (Stern, 1974) gBased on Polyosma (Metcalfe and Chalk, 1979) hRonse Decraene (personal observation) on Hydrangea and Escallonia. October 2000] RONSE DECRAENE ET AL.—FLORAL DEVELOPMENT AND ANATOMY OF mens arising on a ring primordium, and the gynoecium, which is not initiated along a depression, but with two free primordia arising on a flattened apex. No circular gynoecial primordium occurs and the ovary never closes completely, leaving a transverse slit separating the two styles (also visible anatomically). Some anatomical similarities with Montinia exist within the core Saxifragaceae (Bensel and Palser, 1975a) in the presence of compound ventral bundles, separated from the peripheral bundles in the base of the flower, and the arrangement of peripheral bundles in the ovary wall. However, contrary to Montinia, ventrals continue into the style, the placentae often have a longitudinal division on top of the ovary, and the nectary has no vascularization. More differences are listed in Table 1. Solanales—Morgan and Soltis (1993) argued that few morphological and anatomical features are shared by Montinia and Solanales and that there are, indeed, several differences (viz. free stamens, choripetaly, inferior ovaries, lack of internal phloem and iridoid production). However, to accommodate Montinia in the Solanales, Chase et al. (1993) suggested that the sympetalous corolla of Montinia has been secondarily lost. There are several floral ontogenetic investigations on Solanaceae (reviewed by Huber, 1980). Most studies are limited to one or a few species, except for Huber (1980) who investigated the ontogeny of 20 species in the family. Solanaceae differ ontogenetically from Montiniaceae in a number of striking features: flowers are always pentamerous with sepals usually arising in a 2/5 sequence and soon becoming connected by interprimordial growth; petals arise (almost) simultaneously and lag behind in development compared to the androecium; all species of Solanaceae are characterized by zonal growth lifting the androecium and petals; the gynoecium emerges as a ring primordium on a flattened floral apex. Later development of the gynoecium is characterized by the development of two horseshoe-shaped carpel primordia separated by a median ‘‘bar’’-like septum. This is caused by an equal growth of the placental region and the ovary wall. Floral anatomy is highly variable in Solanaceae (e.g., Murray, 1945; Armstrong, 1986). However, all taxa lack a cavity in the style and there is a tendency for free ventrals to become fused. An affinity of Montinia with Sphenoclea (Sphenocleaeae) and Hydrolea (Hydrophyllaceae) has been proposed on the basis of rbcL sequences (Cosner, Jansen, and Lammers, 1994; Fay et al., 1998) and confirmed by ndhF sequences (R. G. Olmstead, University of Washington, personal communication). Sphenoclea has traditionally been related with Campanulales (e.g., Cronquist, 1981; Takhtajan, 1997) and its relationship with Solanales appears surprising. However, a cladistic study of morphological and chemical characters by Gustafsson and Bremer (1995) suggests that the Sphenocleaceae occupy a weakly supported basal position in the Campanulaceae. A floral developmental investigation by Erbar (1995) does not contradict an affinity of Sphenoclea with Campanulales, but shows that Sphenoclea is different from Montinia in a number of features, such as the initiation of an early ring primordium at the onset of petal initiation and the presence of sympetaly. However, it shares an imbricate corolla aestivation and semi-inferior ovary with Montinia; the placentation is axile (apparently axile, but basically parietal in Montinia) with narrow septal connections. Floral anatomical data would be a welcome addition for comparison. Secondary loss of sympetaly, a necessary step if Montinia is related to Solanales, is rare in the Asteridae. Apart from MONTINIA 1421 Rubiaceae where it has been reported in Mastixiodendron and some other genera (Darwin, 1977), it has been illustrated ontogenetically for Besseya (Scrophulariaceae) by Hufford (1995). In Besseya the presence of a corolla tube depends on the expression of zonal growth below the insertion of corolla lobes and filament insertion. However, contrary to Montinia, the corolla is initiated on a ‘‘corolla plateau’’ or ring primordium in Besseya and the development of a tube depends on the later growth of this plateau. Erbar (1991) and Erbar and Leins (1996) attached great importance to the early initiation of sympetalous corollas in distinguishing between ‘‘early’’ and late ‘‘sympetaly.’’ In early sympetaly ‘‘the petals arise on a ring primordium or are connected already at initiation’’ (Erbar and Leins, 1996, p. 428); in ‘‘late sympetaly’’ the petals arise separately, and become connected by interprimordial extensions (Erbar and Leins, 1996, p. 428). In other words the corolla is expressed as the independent initiation of petal primordia (late sympetaly), or the corolla develops as a ring that lacks, at least initially, independent primordia (early sympetaly). Solanales are characterized by late sympetaly; the petal primordia are small and become connected by a lateral connection of the petal bases. Interestingly, this division between early and late sympetaly corresponds roughly with the growth pattern of the corolla: in early sympetaly petals grow rapidly and soon cover the stamen primordia; in late sympetaly petals lag behind stamen development. Moreover, it appears clear that those species termed ‘‘early sympetalous’’ all have a ring primordium at the stage of petal initiation, viz. a collar delimiting the concave floral apex (cf. Roels, 1998). One should also remark that the stamen primordia arise on the inner slopes or inside the ring primordium (in the case of early sympetaly), while the petal primordia are situated peripherally or on top of it. The limits between a corolla tube proper and the stamenpetal tube common to sympetalous taxa is difficult to discern, viz., is the interprimordial region behind the stamen part of the petal tube sensu stricto or part of the basal meristem common to stamens and petals? This distinction is not made by Hufford (1995) who describes growth below the corolla lobes and stamens, as well as the confluent growth above the stamen insertion, as ‘‘zonal growth.’’ His description is essentially developmental without taking issues of homology into account. We believe that neither the approach of Leins and Erbar of a basic corolla-derived nature of the ring primordium, nor a generalization of a ‘‘corolla tube‘‘without distinguishing between its parts is wholly satisfactory. As clearly set out by Nishino (1978) and Ritterbusch (1991), the initial ring primordium ultimately gives rise to both petal primordia and stamen primordia. The corolla tube sensu stricto arises later on top of this ring by lateral extension and linking of marginal meristems of the originally free petal primordia (e.g., Nishino, 1983a, b), while the ring primordium should merely be interpreted as the early initiation of the stamen-corolla tube or even an early onset of hypanthial growth. While Erbar and Leins (1996, p. 428) explicitly state that there are two different parts, viz. a stamen-corolla tube and a corolla tube s.s. (sensu stricto), they restrict their ontogenetic observations of early and late sympetaly to the corolla tube s.s., without considering the functioning of the former, merely stating that it ‘‘results from the activity of a circular intercalary diffuse meristem under the insertion area of stamens and corolla tube s.str.’’ However, in Montinia this kind of development is absent, and there are no indications that it had a corolla tube in ancestral forms. 1422 AMERICAN JOURNAL Cornales—The broad circumscription of Cornales by R. Dahlgren (1980) and G. Dahlgren (1989) includes several families and is based mainly on embryological and phytochemical characteristics, of which the iridoids are a main feature. Besides their presence in the Cornales, iridoids occur also in taxa currently grouped as asterids (Jensen, Nielsen, and Dahlgren, 1975). In the asterids the distribution of iridoids is linked with the distribution of tenuinucellate and unitegmic ovules and coincides with that of sympetaly. However, Montinia occupies an awkward position in being strictly choripetalous as are most other Cornales. This is also a first unequivocal evidence for unitegmic ovules in Montinia. Similarly, Loasaceae have the same embryological features (Dahlgren, Rosendal-Jensen, and Nielsen, 1981), but sympetaly is present in several species. Early petal initiation is correlated with the development of circumspherical growth of a ring in Eucnide. Development of a common stamen-petal tube is linked with vertical zonal growth, but fusion of corolla lobes in suprastaminal regions is weakly expressed (Hufford, 1988). Montinia shares epigyny with various members of Cornales. The occasional occurrence of perigyny in Hydrangeaceae was interpreted as derived from ancestors with complete epigyny (Soltis, Xiang and Hufford, 1995). Data on the ontogeny of members of the Cornaceae are limited to a few species. Cornus alternifolia and two species of Cornus (C. kousa, C. sanguinea) were investigated to some extent by Payer (1857) and Roels (1998), respectively. As in Montinia, corolla aestivation is imbricate in bud; no ring primordium occurs prior to petal initiation, and petal growth is rapid. No ring primordium develops below the stamen primordia. However, the development of the ovary differs strongly from that of Montinia. All taxa of the Cornales s.s. studied show broad petal primordia on the flanks of a depression, with lateral extensions. Data from floral anatomy (Wilkinson, 1944; Eyde, 1967, 1988) indicate that Cornaceae differ from Montinia in several characteristics. Eyde (1967, 1988) attached much importance to the absence of central vascular traces to delimit Cornaceae from other genera with doubtful position. The placentation in Montinia is basically parietal, but the axial vascular system is strongly developed. Escalloniaceae and Hydrangeaceae have been connected by some authors (e.g., Escalloniales—Krach, 1977; Hydrangeales—Takhtajan, 1997). Montinia shares a scanty vasicentric axial parenchyma with some genera of Hydrangeaceae (Carlquist, 1989) and some species of Escallonia (Stern, 1974). A number of ontogenetic similarities exist between Montinia and the Hydrangeaceae (see, e.g., Klopfer, 1973; Roels, Ronse Decraene, and Smets, 1997; Roels, 1998): the formation of a central pitlike depression prior to the initiation of a gynoecial ring primordium, the rapid growth of the petals, the initiation of massive placental bands prior to ovule initiation, the short bifid style (cf. Deutzia), and identical placentation fluctuating from axile below to parietal higher up (fusion of septa in lower part). Early ontogeny of tetramerous Philadelphus resembles Montinia also in the sequential initiation of the two upper bracts or bracteoles with oblique orientation of the floral bud (Roels, Ronse Decraene, and Smets, 1997). Hydrangeaceae also possess massive (but U-shaped) placentae, a complex composed of cymes as partial inflorescences (e.g., Kirengeshoma, Philadelphus), and rapid initiation of petals. Transfer of protection of flower bud from calyx to corolla is common, contrary to the situation in Solanales. Philipson (1967) and Eyde (1966) report the existence of ‘‘axial’’ strands supplying the ovules in Escalloniaceae and OF BOTANY [Vol. 87 Corokia, respectively; these axial bundles are connected proximally with one or more peripheral bundles and are absent from Cornaceae (Wilkinson, 1944; Eyde, 1966). It is interesting that the ventral vascular system is constructed similarly in Montinia. Other similarities of Montinia with Corokia are the fact that lateral traces of the sepals are occasionally derived from the petal bundle and the existence of a short region above the ovules in which the carpel margins are not united into a septum. Flowers of Corokia and Montinia contain tannin cells, which are conspicuously absent from Cornales s.s. However, Corokia does not appear to be related with either Escalloniaceae or Montiniaceae on the basis of rbcL data (Morgan and Soltis, 1993; Cosner, Jansen, and Lammers, 1994), or morphological and chemical data (Gustafsson and Bremer, 1995). The Escalloniaceae are a poorly known, probably unnatural family containing several tribes with a high intrinsic morphological variability (e.g., Krach, 1976; Hideux and Ferguson, 1976). Molecular studies (Soltis and Soltis, 1997, Xiang and Soltis, 1996) place Escalloniaceae far apart from Hydrangeaceae and Montiniaceae. Analyses of Escalloniaceae suggest that the family is polyphyletic. Contrary to Solanaceae and Hydrangeaceae, Escalloniaceae have not been studied ontogenetically, except for the study of Payer (1857) on Escallonia floribunda. Data presented by Payer mostly accord with those of Montinia: inflorescences are cymose and terminal; there is a rapid growth of free petals with imbricate aestivation (however, descending); and gynoecium development shows numerous similarities. The gynoecium arises as a ring primordium around a central receptacular concavity; there is a similar development of continuous carpel primordia, each ending in a free dorsal carpel part. Escallonia possesses a similar strongly developed nectary with nectarostomata surrounding a stout style, and a number of ovules ranging between 2 and 12. The disc on top of the ovary is vascularized by small branches derived from carpel-wall bundles. The placentation in Escalloniaceae is intruding parietal. As in Montinia, the separation of the placental lobes is perpendicular to the carpels (Bensel and Palser, 1975b: Polyosma, Escallonia). In Montinia placentation is pseudo-axillary to nearly the top and is connected with the ovary wall by a narrow connection. The placenta itself is massive and bears two lateral rows of interconnected ovules. The placenta breaks apart in the upper half from the intrusion of a broad stigmatic canal. This breaking-up is different from Saxifragaceae (e.g., Bensel and Palser, 1975a; Ronse Decraene et al., 1998), as it does not go along the suture of the two carpels. Escalloniaceae and Montiniaceae differ also from Saxifragaceae in that no ventral carpel bundles enter the style (they do so in Saxifragoideae). Escalloniaceae mostly possess unicellular, eglandular trichomes (Escallonia, Forgesia, and Quintinia also possess glandular hairs), but shapes and surface morphology are variable. Interestingly, Grevea shares similar eglandular, uniseriate trichomes with Anopterus (tribe Anoptereae: Al-Shammary and Gornal, 1994). From their phenetic analysis Hideux and Ferguson (1976) conclude that the pollen of Montiniaceae corresponds most closely with that of Escalloniaceae. Montinia shows striking similarities with Polyosma (Escalloniaceae) at the level of floral anatomy. Polyosma differs from other Escalloniaceae in a number of anatomical characters (see Bensel and Palser, 1975b), which are also shared by Montinia: tetramerous flowers, deeply intruding parietal placentae (postgenitally fused in Montinia except for the upper part), an interconnection between peripheral bundles and cen- October 2000] RONSE DECRAENE ET AL.—FLORAL DEVELOPMENT AND ANATOMY OF tral tracheary elements in the pith, compound ventrals running well above the ovule insertion, two placentae with two rows of ovules each, petal traces providing extreme laterals of the sepals and carpel wall bundles, dorsal traces derived from the sepal-plane traces, and a canal extending through the style into the stigma. However, in Polyosma leaves are opposite and fruit development is very different. Polyosma also differs from Escalloniaceae and Montiniaceae in its seed anatomy (Krach, 1976: presence of starch in the endosperm and small undifferentiated embryo) and pollen morphology (Pastre and Pons, 1973; Hideux and Ferguson, 1976). However, similarities with Montiniaceae include a three-layered seed coat and a very small embryo. Conclusions—Neither ontogenetical nor anatomical evidence presented in this paper (see also Table 1) provides strong evidence for the phylogenetic relationships of Montinia, but the closest affinity of Montinia lies probably with some members of Escalloniaceae. There is almost no morphological support for a sister-group relationship with Solanales as suggested by molecular data. However, the aim of this study was not to compare Montinia with outgroups cladistically, as information on these potential outgroups is highly insufficient. An asterid affinity of both Montiniaceae and Escalloniaceae, as suggested by molecular data, is corroborated in this study. 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SOLTIS. 1996. rbcL sequence divergence and phylogenetic relationships of Cornaceae sensu lato. In D. E. Boufford and H. Ohba [eds.], Sino-Japanese floristic region: its diversification and characteristics. Tokyo University Press, Tokyo, Japan. APPENDIX Montinia Thunb. (1776)—Montiniaceae Fam. Montiniaceae Nakai, Chosakuronbun Mokuroku: 243. 20 July 1943. Subfam. Montinioideae (DC.) Beilschm., Flora (Bieb.) 16(2): 96, 108. 14 June 1833 (Montinieae). Tribe Montinieae DC., Prodromus 3: 35. mid March 1828.