American Journal of Botany 92(11): 1836±1852. 2005.

REPRODUCTIVE DEVELOPMENTAL COMPLEXITY IN THE AFRICAN OIL PALM (ELAEIS GUINEENSIS,)1

HEÂ LEÁ NE ADAM,2 STEFAN JOUANNIC,2 JACQUES ESCOUTE,3 YVES DUVAL,2 JEAN-LUC VERDEIL,3,4 AND JAMES W. T REGEAR2,4

2IRD/CIRAD Palm Biology Laboratory, UMR 1098, Centre IRD Montpellier, BP 64501, 911, Avenue Agropolis, 34394 Montpellier, France; and 3CIRAD-AMIS, UMR 1098, Avenue Agropolis, 34398 Montpellier Cedex 5, France

Species of the palm family (Arecaceae) are remarkably diverse in their in¯orescence and ¯oral morphologies, which make them a particularly interesting group for studies of reproductive development and its evolution. Using light and scanning electron microscopy, we describe in¯orescence and ¯ower development in the African oil palm Elaeis guineensis from the initiation of the in¯orescence meristem to ¯ower maturity. In mature palms, the in¯orescence develops over 2±3 years and is characterized by individual stages within which differentiation may be either relatively slow, as in the case of early in¯orescence meristem development, or rapid, as in the case of ¯ower organogenesis. The female in¯orescence bears ¯oral triads composed of single pistillate ¯owers ¯anked by two abortive staminate ¯owers, whereas the male in¯orescence contains single functional staminate ¯owers. This suggests a possible evolutionary movement from an ancestral hermaphrodite in¯orescence form containing fully functional ¯oral triads to the situation of temporal dioecy observed at present. Wild type ¯owers are compared to those bearing an epigenetic homeotic abnormality, known as mantled, involving an alteration of the identity of the organs in the fertile and sterile androecium.

Key words: development; ¯oral triad; ¯ower; in¯orescence; mantled; oil palm.

Despite practical constraints associated with their often A wide range of ¯oral ontogenetic studies has been reported large size, the palm family (Arecaceae, sole member of the for palms; notable examples include Ptychosperma species order ) is a group of of considerable interest in (Uhl, 1976); several genera of the tribe Phytelepheae (Palan- developmental biology studies. A key feature of palms, which dra, , , and ; Uhl and Moore, lends itself to developmental analysis, is the existence usually 1977; Uhl and Drans®eld, 1984; Barfod and Uhl, 2001); the of a single vegetative shoot apical meristem, which initiates chamaedoreoid species Hyophorbe indica (Uhl and Moore, the entire aboveground structure of the . Palms also have 1978); a number of polyandrous genera (Uhl and Moore, a number of interesting features in their reproductive devel- 1980); the calamoid Eugeissona (Uhl and Drans®eld, opment, such as variable modes of sex determination and sin- 1984); two members of the tribe Cocoseae (Beccariophoenix gle- or mixed-sex ¯ower clusters, which may de®ne speci®c madagascariensis and Polyandrococos pectinatus; Uhl, 1988); clades. Indeed, studies of in¯orescence and ¯oral ontogenesis the species Geonoma interrupta in the tribe Geonomateae are key elements to understanding evolutionary relationships (Stauffer et al., 2002); and the genus Dypsis of the tribe Are- both within the palm family and with respect to other monocot ceae (Rudall et al., 2003). The ¯owering apparatus of palm groups, as well as in angiosperms as a whole. species is a useful indicator of phylogenetic relationships and The Arecaceae were recently restructured into ®ve subfam- therefore evolutionary events. Within the family as a whole, a ilies (Drans®eld et al., 2005) as opposed to the six subfamily number of general trends in the evolution of ¯oral characters structure of the previous classi®cation (Uhl and Drans®eld, have been noted, including a progression from bisexual to uni- 1987). The new nomenclature is adopted here. Two main and sexual ¯owers and from monoecy to dioecy (Moore and Uhl, contrasting types of ¯owering behavior within a single shoot 1982). may be seen in palm species (Tomlinson, 1990). Hapaxanthy, We describe here a detailed study of reproductive devel- in general, refers to a single shoot in which there is an abrupt opment in the African oil palm (Elaeis guineensis Jacq.), a transition to the ¯owering state, with ¯owering branches ex- pleonanthic species belonging to the subfamily Arecoideae, in panding in larger numbers after vegetative extension has the tribe Cocoseae, and the subtribe Elaeidinae. The Arecoi- ceased. The ¯owering phase is thus short. Hapaxanthic shoots deae form the largest subfamily of the Arecaceae and have occur in about 5% of palms. Pleonanthy refers to a single been classi®ed into 112 genera (Drans®eld et al., 2005). Oil shoot in which ¯owering branches (in¯orescences) appear in palm is the second most important world vegetable oil crop the axils of vegetative and continue to be produced as plant after soya and is one of the most important perennial the palm continues its vegetative extension. Such shoots are crops in the tropics. Despite its economic importance, only termed pleonanthic. The ¯owering phase is extended and in- partial studies of oil palm in¯orescence and ¯ower develop- determinate. Pleonanthic shoots occur in about 95% of all ment have been reported (Beinaert, 1935; Van Heel et al., palm species. 1987). Oil palm is perennial and relatively long-lived, some- times attaining more than 100 years in age. The oil palm is a 1 Manuscript received 9 December 2004; revision accepted 21 July 2005. single-stemmed palm, which bears, like the majority of palm The authors gratefully thank Jean-Christophe Pintaud, Francis HalleÂ, and species, a single vegetative shoot apical meristem maintained three anonymous referees for helpful suggestions in the writing and improve- throughout the lifetime of the plant and localized at the center ment of this paper, FELDA Agricultural Services (Malaysia) and ASD (Costa Rica) for the generous supply of plant material, and Axel Labeyrie for pho- of the crown. Under favorable climatic conditions, this tography and logistical help. meristem is continuously active, producing a new leaf pri- 4 Authors for correspondence (e-mail: [email protected]; [email protected]) mordium approximately every 2 weeks in mature palms; at the 1836 November 2005] ADAM ET AL.ÐREPRODUCTIVE DEVELOPMENT IN ELAEIS GUINEENSIS 1837

Fig. 1. Features of reproductive development in oil palm. (A) Oil palm in¯orescence structure and axis organization. The in¯orescence unit axis (axis 1) is in the axil of a leaf. Abbreviations: F, ¯ower; fb, ¯ower bract; L, leaf; p, prophyll; pb, peduncular bract; rab, rachilla bract. (B) Chronology of the principal phases of in¯orescence and ¯ower development in oil palm. Note that this is a generalized scheme applying to in¯orescences of both sexes. (C) Diagram showing the structure of the ¯oral triad of the female in¯orescence (after Beirnaert, 1935; Van Heel et al., 1987). juvenile stage, the plastochron may be as low as 9 days (Cor- ley and Gray, 1976). The leaf takes 2±3 years to develop from initiation to the time when lea¯ets unfold in the center of the palm crown. In¯orescences are formed throughout the year in an acropetal sequence in the axils of their subtending leaves. The immature in¯orescence is protected within the crown by its subtending leaf. Moreover, as in many palms of the Co- coseae, the expanding in¯orescence axes are themselves pro- tected by a prophyll and a peduncular bract, which become thick and ligni®ed. Elaeis guineensis is monoecious, a char- acter that predominates in the Arecaceae (Drans®eld and Uhl, 1998). Oil palm produces separate male and female in¯ores- cences on the same palm in an alternating cycle of variable duration depending on genetic factors, age, and particularly environmental conditions, with the production of male in¯o- rescences generally favored by water stress (Corley, 1976a). Occasionally mixed sex in¯orescences are produced at the transition between the male and female cycles (Biradar, 1978). Mature in¯orescences are visible typically 32±36 months after germination (Corley, 1976b). As for all monocotyledoneous species, oil palm has trim- erous ¯owers. In the Arecaceae, the perianth is usually clearly distinguished into sepals and petals (Drans®eld and Uhl, 1998). In the case of oil palm however, the sepals and petals have a similar petalloid appearance and are often referred to as tepals. The reproductive organs of staminate ¯owers are composed of six stamens (two whorls of three) with connate ®laments surrounding a pistillode, whereas female ¯owers have rudimentary stamens (staminodes) and a gynoecium of three carpels. The aim of the studies described here was to provide a com- Fig. 1. Continued. plete description at the histological and cellular levels of all stages of in¯orescence development from the initiation of the primordium from the central axis through to the completion 1838 AMERICAN JOURNAL OF BOTANY [Vol. 92

Fig. 1. Continued. of ¯ower development. The study was extended to include the through a graded ethanol series (30, 70, and 100%, v/v) for 2 h and stored at ¯oral variant known as mantled, commonly observed in palms 4ЊC. produced by in vitro micropropagation and characterized by homeotic transformation of the fertile or sterile androecium Scanning electron microscopyÐFor scanning electron microscopy (SEM), (Corley et al., 1986). To our knowledge, this is the ®rst in- material was passed through three changes of absolute ethanol, critical point depth study of this type reported for palms. Mantled palms dried, mounted on stubs, and coated with platinum in a Baltec (Manchester, resemble B class mutants of model monocot and dicot species New Hampshire, USA) SCD 50 sputter coater. Samples were visualized with (CoeÈn and Meyerowitz, 1991; Zahn et al., 2005) with which a JEOL (Tokyo, Japan) 6300F scanning electron microscope at 20 kV. comparisons can be made to obtain a fuller understanding of ¯oral homeotic functions in relation to phylogeny. Histological studies and transmission light microscopyÐFor histological analysis, samples were transferred to 100% butanol for 2 days and embedded in Technovit resin (LKB Pharmacia, Uppsala, Sweden) using a previously MATERIALS AND METHODS described protocol (Buffard-Morel et al., 1992). Blocks were sectioned at 4± 5 ␮m thickness using a HISTORANGE microtome (LKB Broma, Uppsala, Plant materialÐIn¯orescence material for the characterization of stages of Sweden). Slides were double stained with periodic acid±Schiff's reagent reproductive development was collected in Coto, Costa Rica at the plantation (PAS) stain (Sigma-Aldrich, Lyon, France) to detect carbohydrate compounds of the ASD Company. All palms were 9 years old and of the tenera variety, (Buffard-Morel et al., 1992) and naphthol blue-black to detect proteins (Fisher, i.e., obtained from a cross between pisifera and dura palms (Hartley, 1988). 1968). For studies of the mantled ¯oral phenotype, normal and variant ¯owers were collected in Tun Razak, Malaysia at the plantation of the FELDA Company. RESULTS The source palms for the mantled and normal pistillate ¯owers were of iden- tical genotype and age. Palms of similar genetic origin and age were also Development and structure of the in¯orescencesÐThe in- used as the source of staminate ¯owers displaying either a mantled or a nor- ¯orescence is initiated in the axil of each leaf of the oil palm mal phenotype. The numbering attributed to in¯orescences corresponds to that from an early stage shortly after germination and continues of their axillary leaves (Fig. 1A, B). The youngest expanding leaf was num- throughout the lifetime of the plant. The development of the bered as leaf 0. The leaf produced immediately before leaf 0 is thus leaf Fϩ1 in¯orescence takes over 2 years; the organ is completely en- Ϫ and the leaf produced subsequently is leaf F 1 (this leaf is not yet expanded). closed at the base of the subtending leaf for most of this time. In this study, the in¯orescences collected individually were located between Ϫ The reproductive development of oil palm is represented sche- leaf 27 (the earliest stage at which in¯orescence isolation was possible) and matically in Fig. 1, which details the structural organization leaf ϩ18 (¯ower maturity; Fig. 1B). Earlier stages were observed without prior isolation along with the subtending partly formed leaves. Oil palms were of the in¯orescence (Fig. 1A), the key stages of in¯orescence felled in order to dissect the least accessible primordia near the shoot apical and ¯ower development (Fig. 1B), and the structure of mature meristem. In¯orescences Ϫ27 to Ϫ6 were directly ®xed. For in¯orescences ¯owers (Fig. 1C). The in¯orescence is a compound rachis Ϫ5toϩ4, the prophyll and peduncular bract were removed and the entire composed of 100±300 rachillae in the case of the male in¯o- in¯orescence ®xed (as described in next section). For in¯orescences ϩ5to rescence or approximately 150 rachillae in the case of the fe- ϩ16, rachillae were dissected from the rachis and ®xed. For in¯orescences male in¯orescence (Jacquemard, 1995). In each case, the ra- ϩ16 to ϩ18, ¯oral triads and staminate and pistillate ¯owers of normal and chis is borne by a peduncle with a typical length at maturity mantled phenotypes were dissected individually from the center of the rachilla of either 20±30 cm for the female in¯orescence or around 40 and ®xed. We used the nomenclature of Tomlinson (1990) to describe the cm for the male in¯orescence (Beirnaert, 1935). Rachillae morphological features of the oil palm material. (axis 2 in Fig. 1A) are arranged spirally around a central rachis (axis 1) emanating from the stem (axis 0). A prophyll, along Fixation of samplesÐAfter dissection, sampled material was ®xed for 1 h with the peduncular bract, tightly encloses the in¯orescence under vacuum and thereafter for8hat4ЊC in ®xation buffer (4% parafor- until approximately 6 weeks before ¯ower maturity. The pro- maldehyde, 0.1 M phosphate buffer, pH 7). Samples were then dehydrated phyll attains a ®nal length of approximately 45 cm, whereas November 2005] ADAM ET AL.ÐREPRODUCTIVE DEVELOPMENT IN ELAEIS GUINEENSIS 1839 the peduncular bract, with which it forms an essentially con- divisions will give rise to the rachilla bracts (Fig. 12). The tinuous layer of protection, is typically 2±4 cm shorter. Both basal and apical bracts are the ®rst to develop with a slight layers become ®brous as their development progresses and ®- difference in their rates of development (Fig. 13). The in¯o- nally undergo necrosis before being ruptured by the elongating rescence meristem maintains the same form and cellular or- in¯orescence. ganization as in the previous stage; however, more vascular Male and female in¯orescences differ in their rachilla mor- cells are observed in the center of the rachis and starch may phology (Figs. 2, 4), notably with respect to the numbers of be seen in these cells. During the development and growth of ¯owers that they bear. The female in¯orescence reaches a the rachis, vascularization and starch accumulation become length of 35 cm or more when fully developed (Fig. 2), and progressively more visible. The rachilla bracts of the in¯ores- each of its rachillae bears 5±30 ¯oral triads (Fig. 3). The triads cence develop in a parastichial and acropetal manner along the are arranged spirally around the rachilla axis, and each is sub- rachis axis (Figs. 14±16), the tissues at the center of lateral tended by a spiny ¯oral triad bract. The functional pistillate bracts gradually developing a vascularized structure (Fig. 14). ¯ower develops within a ¯oral triad between two nonfunc- At this stage, no differences are visible between male and fe- tional staminate ¯owers (Beirnaert, 1935). The male in¯ores- male in¯orescences (leaf stage Ϫ6). cence reaches a length similar to that of the female in¯ores- cence at maturity and is composed of rachillae, each bearing Rachilla initiation and developmentÐThe lateral rachis 400±1500 staminate ¯owers (Figs. 4 and 5). The staminate meristem (rachilla meristem) is initiated when ϳ10±15 rach- ¯owers are subtended by ¯oral bracts, which are smaller than illa bracts have been formed (Fig. 17; Ϫ6 leaf stage). Rachilla those of the female in¯orescence. meristems (Fig. 1A; axis 2) are initiated at the bases of lateral The earliest stage at which the in¯orescence bud is visible bracts by periclinal divisions in 2 or 3 cell layers underlying by light microscopy is in the axil of the fourth-youngest leaf the L1 layer (Fig. 17). At the initiation stage of the rachillae, close to the vegetative shoot apical meristem. At this stage, the cells that give rise to these structures possess a large nu- the in¯orescence consists of a group of a few cells localized cleus, as illustrated by their intense and homogeneous naphthol in the axil of the partly formed leaf (Fig. 6). These cells dis- blue-black coloration. This staining, which is speci®c to pro- play a low cytoplasm to nucleus ratio. The in¯orescence bud teins, marks the presence of histones and suggests that the is visible to the naked eye from the stage corresponding to the chromatin of these cells might be condensed (Fig. 17). When 12th-youngest leaf onwards. the rachilla primordium is formed (Fig. 18), the cells of the Figure 7 shows an in¯orescence meristem at a very young primordium have a higher nucleus to cytoplasm ratio. The nu- stage (leaf Ϫ27), corresponding to the initiation of the rachis clei are larger, with a blue, less-homogeneous coloration, sug- (Fig. 1A; axis 1). This meristem, also named the rachis mer- gesting a decondensation of the chromatin in the nucleus. The istem, is composed of a central dome, which is ¯anked by the cells underlying the L1 cells thus seem to be active, but do primordium of the future prophyll (or outer spathe). Within not divide, during the initiation stage, despite their apparent the in¯orescence dome, composed of isodiametric meriste- potential to do so (Fig. 17). The growth of the rachilla axis is matic cells, a distinct layer of cells on the surface (L1; achieved by anticlinal divisions in cells of the L2 layer (Fig. Vaughan, 1955) will give rise to the epidermal cells (Fig. 8). 19; Leaf stage Ϫ2). Rachillae develop in a basipetal manner About 10 cell layers under the L1 layer, cells with a large (Fig. 20). nucleus and small cytoplasm are apparent. Few divisions are Each rachilla will produce a series of ¯oral bracts (in the visible in this zone, suggesting that they might perform a male in¯orescence), or ¯oral triad bracts (in the female in¯o- ``stem cell'' function. A third zone of cells is visible under the rescence), each containing a ¯oral meristem in its axil. The two previously mentioned layers; in this case, the cells are rachilla meristem (axis 2) has the same basic organization as vacuolated and may later contribute to the pith of the in¯o- the rachis meristem (axis 1), namely a well-organized L1 cell rescence. At this stage, the center of the in¯orescence (rachis) layer, overlying cell layers with a high division potential. The is not vascularized. Periclinal and anticlinal divisions contrib- elongation of the rachilla is achieved by the activity of an ute respectively to the growth in depth and width of the dome. adjoining region of vacuolating cells. The rachilla meristem Figure 9 shows the in¯orescence meristem at the Ϫ26 leaf has a conical shape, the meristematic zone of the rachilla being stage, when the prophyll is more elongated along the meri- more substantial in size than that of the rachis observed pre- stem. Moreover, many more cells under the L1 layer are ac- viously, suggesting that the rachilla is at this point undergoing tively dividing at this point. At the Ϫ24 leaf stage, the prophyll an active growth phase (Fig. 21). The periphery of the rachilla entirely envelops the in¯orescence meristem, next to which the displays some periclinal divisions in cells underlying the L1 peduncular bract (or inner spathe) primordium is visible layer. These divisions will later give rise to ¯oral bracts or around the dome (Fig. 10). The prophyll and peduncular bract ¯oral triad bracts. For both sexes, bracts are more advanced elongate until they entirely envelop the in¯orescence meri- in their development at the abaxial side of the in¯orescence stem. During its subsequent development, the meristem grows axis, and the development is parastichial and acropetal (Figs. in depth along with the rachis, the number of vacuolate cells 21, 22). During the development of the rachilla, tissues in the in the center of rachis increasing. In this phase of develop- center undergo vascularization (Fig. 22; leaf stage ϩ4). At this ment, the in¯orescence meristem (Fig. 1A; axis 1) displays point, the sex of the in¯orescence can be identi®ed, the future only one functional axis, which gives rise to the prophyll and male rachilla displaying a larger number of bracts than the peduncular bract. The activity of the rachis meristem is main- future female rachilla (data not shown). tained and allows the production of rachilla bracts, which sub- tend an axillary rachilla. At the Ϫ20 leaf stage, the rachis Development and structure of the ¯oral triadÐOn the fe- meristem (axis 1) is completely enclosed by the prophyll and male in¯orescence, the rachillae bear triads of ¯owers, each peduncular bract (Fig. 11). At the ¯anks of the rachis meri- consisting of a functional pistillate ¯ower ¯anked by two non- stem, anticlinal divisions are visible under the L1 cells. These functional staminate ¯owers. The latter normally undergo ab- 1840 AMERICAN JOURNAL OF BOTANY [Vol. 92

Figs. 2±5. Macroscopic view of mature female and male in¯orescences of oil palm. 2. Female in¯orescence at anthesis after opening of the peduncular bract and prophyll. 3. Detail of female rachillae carrying pistillate ¯owers at maturity with exposed stigmas. 4. Male in¯orescence at anthesis. 5. Detail of male rachilla and ¯owers at anthesis. Abbreviations: f, ¯ower; p, prophyll; ra, rachilla; St, stigma. November 2005] ADAM ET AL.ÐREPRODUCTIVE DEVELOPMENT IN ELAEIS GUINEENSIS 1841

Figs. 6±10. Longitudinal sections (LS) of early in¯orescence development in oil palm visualized by LM. 6. In¯orescence primordium at the fourth leaf after the shoot apical meristem. 7. In¯orescence meristem at Ϫ27 leaf stage. 8. Detail of in¯orescence meristem at Ϫ27 leaf stage. 9. In¯orescence meristem at Ϫ26 leaf stage. 10. Early initiation of peduncular bract of in¯orescence meristem at Ϫ24 leaf stage. Abbreviations: pbp, peduncular bract primordium; Im, In¯orescence meristem; Ip, In¯orescence primordium; L, leaf; L1, overlying cell Layer; p, prophyll; pp, prophyll primordium; PV, provascular strands. Bar ϭ 0.1 mm. 1842 AMERICAN JOURNAL OF BOTANY [Vol. 92

Figs. 11±18. Light and SEM micrographs of rachilla development up to and including leaf stage Ϫ4. 11±16. Rachilla bract development. 17±18. Rachilla initiation and development. 11. Longitudinal section (LS) of developing in¯orescence at Ϫ20 leaf stage. 12. Detail of rachilla bract initial. 13. Upper view of in¯orescence meristem. Development of anterior and posterior rachilla bracts. 14. Longitudinal section of developing in¯orescence at Ϫ10 leaf stage. 15. Side view of developing in¯orescence at leaf stage Ϫ10. 16. LS of developing rachilla at leaf stage Ϫ10. 17±18. LS of in¯orescence to reveal successive phases of rachilla development. 17. Initiation of rachilla primordium at Ϫ6 leaf stage. Arrow points to cells at the origin of rachilla. 18. Rachilla primordium at leaf stage Ϫ4. November 2005] ADAM ET AL.ÐREPRODUCTIVE DEVELOPMENT IN ELAEIS GUINEENSIS 1843

Figs. 19±22. Rachilla and ¯oral bract development at 19. leaf stages Ϫ2, 20. 0, 21. ϩ2 and 22. ϩ4 respectively. Abbreviations: ant rab, anterior rachilla bract; Im, In¯orescence meristem; Fb, ¯oral bracts; fpb, peduncular bract; p, prophyll; post rab, posterior rachilla bract; r, rachis; rab, rachilla bract; rabi, rachilla bract initium; rabp, rachilla bract primordium; Ram, rachilla meristem; Vb, vascular bundles. Bar ϭ 0.1 mm, except for Fig 15, bar ϭ 1 mm. scission before complete maturity is reached (discussed later). staminate ¯owers elongate, until they are above the pistillate The ¯oral triad is the most common ¯owering unit in palms ¯ower (Fig. 26). (Moore and Uhl, 1982) and is designated as a cincinnus (Uhl, Floral meristems have a similar form to that of the rachis 1976). In the case of oil palm, the developmental sequence and rachilla meristems, consisting of a L1 layer on the surface, observed illustrates that the ¯oral triad is a sympodial ¯ower overlying 5 or 6 layers of active cells (Fig. 24). The two sta- cluster, i.e., involving a branching system where the main axis minate ¯owers are asynchronous in their development (Figs. is determinate and side branches are produced successively via 25, 26). axillary meristems. At leaf stage ϩ6, a new meristem arises in the axil of the Structure of normal ¯owersÐThe trimerous ¯owers of oil ¯oral triad bract initially formed by the rachilla meristem (Fig. palm contain a perianth consisting of three petals surrounded 1A, axis 2; Fig. 23). This new meristem forms an apex which, by three sepals. Since these two whorls are similar in appear- after producing bracteole 1, develops into staminate ¯ower 1 ance, they are frequently referred to as tepals (Hartley, 1988). In staminate ¯owers, the perianth surrounds an androecium, (Fig. 24). Triad development continues in the axil of bracteole usually consisting of two whorls of three stamens, themselves 1, where a new apex emerges and gives rise to bracteole 2 enclosing a rudimentary gynoecium. In the case of pistillate before developing into staminate ¯ower 2 (Fig. 25). Note that ¯owers, the perianth surrounds a sterile androecium of usually the section in Fig. 25 does not include bracteole 2. A group six rudimentary stamens (staminodes) enclosing a syncarpous of meristematic cells is present between asf1 and asf2. These gynoecium composed of three carpels. cells will subsequently give rise to bracteole 3 and the pistillate ¯ower in the same manner as observed for the staminate ¯ow- Development and structure of normal ¯owers on the fe- ers (Fig. 26). During their development, the peduncles of the male in¯orescenceÐAt leaf stage ϩ10, organ development 1844 AMERICAN JOURNAL OF BOTANY [Vol. 92

Figs. 23±31. Light and SEM micrographs of ¯oral triad development. 23. Longitudinal section (LS) showing the initiation of the triad meristem (i.e., the apex that will produce the ®rst staminate ¯ower) in the axil of ¯oral bracts. 24. Transverse section of ®rst staminate ¯ower primordium surrounded by bracteole 1. 25. LS of staminate ¯owers 1 and 2 showing sepal development. 26. LS of ¯oral triad of ¯owers at leaf stage ϩ10. 27. LS of accompanying staminate ¯ower and initiation of stamens. 28. LS of pistillate ¯ower development of sepals at leaf stage ϩ12. 29. LS showing development of staminate ¯ower 1. 30. LS of triad of ¯owers at leaf stage ϩ15. November 2005] ADAM ET AL.ÐREPRODUCTIVE DEVELOPMENT IN ELAEIS GUINEENSIS 1845 begins in the accompanying staminate ¯owers with the initi- ®eld (1987), the ovule appears to be orthotropous. Although ation and development of sepals (Fig. 25). The elongation of the ovules of many palm species contain two integuments, in these organs is achieved by anticlinal divisions at their base. our studies we observed only one. Nevertheless, because none The cells localized at the distal end of sepals are already vac- of our sections cut perfectly through the micropyle, the exis- uolate at this stage (Figs. 26, 27). After the initiation and de- tence of two integuments cannot be excluded. Within the oil velopment of the three petals, the ¯oral meristem rapidly ini- palm ovule, a nucellus surrounds a megaspore mother cell tiates stamen primordia through periclinal cell divisions in the (Fig. 37). Just before anthesis, the staminodes accumulate L2 cells at the ¯anks of the ¯oral meristem (Fig. 27). While polyphenols at their distal end, and the stigmatic region be- the organs of the staminate ¯owers are initiated, the pistillate comes ligni®ed. A canal is formed in the carpels between their ¯ower meristem does not develop any organs. However, the distal end and base by modi®cation of the middle lamella of cells of the pistillate ¯ower meristem have a high nucleus to the epidermal cells (Fig. 36). This canal may represent the cytoplasm ratio, suggesting that they remain active (Fig. 26). future path taken by the pollen tube prior to fertilization as The development of the ¯oral triad accelerates after leaf observed in Geonoma interrupta (Stauffer et al., 2002). At stage ϩ10. At leaf stage ϩ12, the sepals of pistillate ¯owers maturity (leaf stage ϩ18), the prophyll and peduncular bract are elongating by anticlinal division of the L2 cells. Floral protecting the in¯orescence rupture. The length of the mature organs undergo elongation through cell divisions at their bases female in¯orescence is about 40 cm by this time. The perianth (Fig. 28). At the same stage, the third whorl of the staminate and stigmas of the mature pistillate ¯ower are essentially ¯owers shows the ®rst signs of differentiation of the future white, becoming progressively pink or violet after pollination. stamens (Fig. 29). The development of stamens is achieved by Subsequent to fertilization, development and ripening the division of active cells in their upper regions, which will take place over a period of approximately 6 months. give rise to anthers. The accompanying staminate ¯owers are larger than the pistillate ¯ower at this point; however, during Development and structure of normal ¯owers on the male the next phase of development, the pistillate ¯ower will in- in¯orescenceÐThe ¯oral meristem of functional staminate crease in size, whereas the accompanying staminate ¯ower ¯owers is visible in the axil of the ¯oral triad bracts at leaf will stay the same (Figs. 34, 35). stages ϩ3orϩ4 (Fig. 38). As for the pistillate ¯ower, the At leaf stage ϩ15 (Figs. 30, 31), the ¯oral triads are com- meristem is elliptical and all cells display a high nucleus to posed of accompanying staminate ¯owers bearing six initiated cytoplasm ratio. Whorl initiation appears to follow the same stamens entirely enveloped by the perianth. The pistillate ¯ow- developmental pattern as that in the accompanying staminate er at this stage is composed of only the perianth. At leaf stage ¯owers. The ®rst sepal is initiated at the base of the ¯oral ϩ16, the reproductive organs of pistillate ¯owers are devel- meristem by periclinal divisions (Fig. 39) and is localized on oping, the androecium and gynoecium forming concomitantly the external side of the rachilla. The elongation of the sepals (Fig. 32). Organs are initiated by periclinal divisions in L2 is achieved by anticlinal divisions at the base of these organs. layer cells (not shown). Three free carpels are formed in the Later the petals are initiated as observed for the sepals. Peri- innermost whorl. In contrast to the perianth, the development anth organs completely envelop the meristematic zone (Fig. of ¯oral organs in the inner whorls is achieved by cell divi- 40). During the development of the staminate ¯owers, the sions in their distal regions (not shown). The development of rachilla undergoes elongation. accompanying staminate ¯owers continues with the differen- At about the leaf stage ϩ15/ϩ16, stamens are initiated on tiation of the ®lament and anther regions at leaf stage ϩ16 the ¯anks of the ¯oral meristem by periclinal cell divisions in (Fig. 33). The cells of the ®laments are seen to be vacuolate, the L2 cell layers (Fig. 40). Stamen development and elon- unlike those at the distal end of anthers, which have a high gation is relatively rapid compared with the development of nucleus to cytoplasm ratio. The meristematic zone of the sta- the perianth organs. The functional staminate ¯ower of oil minate ¯ower is completely exhausted (Fig. 33); the cells are palm carries six stamens, each consisting of a bilobed anther vacuolate and will not give rise to a gynoecium. supported by a ®lament. Each stamen is composed of four By leaf stage ϩ17, the anthers of the accompanying sta- pollen sacs grouped in pairs on each side of the ®lament (Fig. minate ¯owers are completely developed and vacuolar poly- 41). Anthers are formed from three concentric layers: an epi- phenol accumulation is seen within the cells of the petals. The derm, a transitory middle layer, and an inner zone representing anther consists of a pollen sac surrounded by the tapetum (not the future microspore mother cells (Fig. 42). Polyphenols ac- shown). The pollen sac contains a mass of dividing cells, the cumulate in cells of the conducting tissues of the stamens microsporocytes. However, the anther will not give rise to pol- (Figs. 42, 43). Figure 43 shows the formation of microspores, len because the microsporocytes will degenerate in the pollen arranged in tetrads, after the second meiotic division at the sac. Subsequently, the stalk of the staminate ¯owers will ab- ϩ18 leaf stage. Once stamen development is complete, the scise, and they will shed before anthesis. ¯oral meristem no longer contains dividing cells, and the pis- During the following stages (Fig. 36), the reproductive or- tillodes remain rudimentary. The male in¯orescence matures gans of pistillate ¯owers develop. Organs in the sterile an- in same way as the female in¯orescence. The mature sepals droecium, known as staminodes, are arrested in their devel- of the functional staminate ¯ower are scariose and yellow or opment and do not develop into stamens. The carpels formed brown, whereas the petals are colorless. Staminate ¯owers, in the innermost whorl are joined at their bases. At the base like their pistillate counterparts, produce an anise-like odor at of the carpels, a group of meristematic cells (Fig. 36) will give maturity, probably through the action of papillae (Beirnaert, rise to the ovary. Subsequently, the stigma of the carpel grows 1935; Genty et al., 1986). No septal nectaries have been ob- and the region in the center of the stigma becomes vascular- served in ¯owers of either sex. ized. Some polyphenols accumulate at the distal end in the center and in the lateral faces. The ovary is localized at the Development and structure of mantled ¯owersÐHomeotic base of the carpel. As previously indicated by Uhl and Drans- transformation of oil palm ¯owers has been observed in sev- 1846 AMERICAN JOURNAL OF BOTANY [Vol. 92

Figs. 31±37. 31. LS of pistillate ¯ower at leaf stage ϩ15. 32. LS showing development of the pistillate ¯ower at leaf stage ϩ16. 33. LS showing developing stamens in staminate ¯ower at leaf stage ϩ15. 34. Side view of ¯oral triad at leaf stage ϩ12. 35. Side view of ¯oral triad at leaf stage ϩ17. 36. LS of pistillate ¯ower at leaf stage ϩ17. 37. LS of ovary of pistillate ¯ower at ϩ18 leaf stage. Abbreviations: a, anther; asf, accompanying staminate ¯ower; B, bracteoles; B1, B2, B3, bracteole 1, 2 and 3; c, carpel; Fb, ¯oral bract, ff, female ¯ower; Fm, ¯oral meristem; m, megaspore mother cell; mi, micropyle; n, nucellus; ov, ovary; ovp, ovary primordium; pd, peduncle; pe, petal; ps, pollen sac; Ra, rachilla; rg, rudimentary gynoecium; s, sepal; sc, stylar canal; sta, staminodia; stp, stamen primordium; tg, integument; Bar ϭ 0.1 mm for LM, ϭ 1 mm for SEM. November 2005] ADAM ET AL.ÐREPRODUCTIVE DEVELOPMENT IN ELAEIS GUINEENSIS 1847

Figs. 38±43. Staminate ¯ower development. 38. Longitudinal section (LS) of staminate ¯ower meristem enclosed by ¯oral bracts. 39. LS showing elongation of ®rst sepals. 40. LS showing initiation of stamens. 41. Transverse section (TS) of elongating stamen. 42. TS showing detail of pollen sac. 43. TS of microspore after meiosis. Abbreviations: ep, epiderm; Fb, ¯oral bract; fm, ¯oral meristem; ml, mechanical layer; msp, microspore; pe, petal; ps, pollen sac; s, sepal; st, stamens; stp, stamen primordium; te, tetrad; vt, vascular tissue. Bar ϭ 0.1 mm. 1848 AMERICAN JOURNAL OF BOTANY [Vol. 92 eral cases, notably in the case of the somaclonal variant man- apical meristem. Our results show that the longest phase in oil tled induced during micropropagation in vitro (Corley et al., palm reproductive development is that involving the devel- 1986), but also in seed-derived palms (Hartley, 1988). In man- opment of the in¯orescence meristem (Fig. 1B). For about 20 tled palms, the staminodes of the pistillate ¯owers and the months, the in¯orescence meristem expands and gives rise to stamens of staminate ¯owers develop as carpel-like structures rachilla bracts, then the rachilla meristem (Fig. 1B). The sub- (Figs. 44±47). The organ of the innermost whorl does not sequent development of staminate and pistillate ¯owers takes undergo a homeotic transformation; thus the carpel is con- place more rapidly during a further 10 months. Our results served in the pistillate ¯ower, and the gynoecium remains ves- show that oil palm reproductive development consists of sev- tigial in staminate ¯owers (Fig. 47). eral growth phases, which are characterized by slow and fast In the case of the mantled pistillate ¯ower, ¯oral triad ini- growth rates. The two distinct types of growth rates are as- tiation is similar to that in normal ¯owers, and the perianth sociated with two different cell types. The ®rst is characterized organs are not visibly altered (Fig. 48). As in normal palms, by a small nucleus and a low nucleus to cytoplasm ratio. These the reproductive organs of pistillate ¯owers are initiated at cells are prevalent in the slow growth phases, when they allow about leaf stage ϩ15. No difference is visible at this stage, the conservation of the meristematic potential of the cells. The organs in the third whorl being initiated in the same way as second type of cell is characterized by a large nucleus and a for normal pistillate ¯owers (Fig. 49). However, at the next higher nucleus to cytoplasm ratio, suggesting a decondensation stage observed, organs resembling carpels (Fig. 50) are de- of the chromatin. These cells are associated with the reacti- veloping in the mantled pistillate ¯owers in place of stami- vation of meristematic potential and thus the development of nodes. Ovules do not develop in these structures, which are the in¯orescence and ¯oral organs. thus sterile. A key feature highlighted in this study is the degree of pro- In the case of the mantled staminate ¯ower, the ¯oral mer- tection afforded to the in¯orescence, which is initially pro- istem develops in a similar way to the normal ¯ower, as does tected within the crown, especially by its subtending leaf. The the perianth (Fig. 51). At this stage, it is not possible to dis- expanding in¯orescence axes are themselves protected by the tinguish the identity of the organ in the androecium (Fig. 52). prophyll and a peduncular bract, which enlarge to become During their subsequent elongation, the organs of the androe- massive and ligni®ed. Direct protection of the reproductive cium develop the central vascularization characteristic of car- organs is provided by the imbricated bracteoles, sepals, and pels (Fig. 53), as opposed to that of the peripheral vasculari- petals. The ovules are additionally protected by the ovary zation of the stamens. As for the mantled pistillate ¯ower, walls, which are ligni®ed and contain some polyphenol and ovules do not develop in the carpelloid structures of mantled idioblasts of calcium oxalate crystals in the apical parts (Uhl staminate ¯owers. In some cases, not all stamens are observed and Moore, 1973). The extreme protection afforded to the re- to be transformed into carpels. Interestingly, the homeotic productive apparatus is likely to constitute an adaptation to the transformation of stamens to carpel-like structures may be ob- long period of development of the in¯orescence. In more de- served not only in staminate ¯owers of the male in¯orescence, rived Arecoideae such as Geonoma interrupta, the perianth but also in the accompanying staminate ¯owers of ¯oral triads and the staminodial tube are considered to constitute the most on the female in¯orescence (data not shown). important structural protective devices for the gynoecium (Stauffer et al., 2002). DISCUSSION Even though palm in¯orescences are often elaborate and massive structures, relatively few terms are needed to describe Studies of the reproductive development of palm species are them, in that they are largely made up of repeating units, each often hampered by the morphological characteristics that typ- with the same construction, and with gradual but progressive ify the family, notably the large size of the adult plant and the changes along any given axis (Tomlinson and Moore, 1968; deep embedding of the in¯orescence in the palm crown during Moore and Uhl, 1982). Palm in¯orescences are panicles with its development. Consequently, relatively few developmental up to six orders of branching, each branch subtended by a studies have covered the entire sequence of events during in- bract. As with many other palm species, the oil palm in¯ores- ¯orescence and ¯ower organogenesis. We report here the ®rst cence has a reduction in the number of its branch orders com- comprehensive analysis at the cytological level of in¯ores- pared with the basic in¯orescence unit described by Tomlinson cence and ¯ower development in the oil palm. Using scanning (1990), re¯ecting morphological changes during the evolution- electron micrographs and sectioned material, we have identi- ary diversi®cation within Arecaceae. The oil palm in¯ores- ®ed a series of morphological landmarks that de®ne each stage cence may be considered to have a fundamental two-order of reproductive development. structure on the basis of its two distinct types of meristem. The ®rst order of complexity is governed by the rachis meri- In¯orescence developmentÐA summary of the time course stem, which produces the prophyll and peduncular bract and of reproductive development in oil palm, as revealed in this afterwards the rachilla bracts. The structure and form of the study, is shown in Fig. 1B. In¯orescence initiation was ob- rachis meristem is always the same, rachilla bracts being con- served in the axils of leaves only a few plastochrons after their tinuously produced. However, it is interesting to note that dur- separation from the shoot apical meristem. A small group of ing the development of this unit, the functioning of the rachis cells is visible along the base of the leaf at this stage. It is not meristem changes in terms of the organ phyllotaxies that it known if these cells will give rise directly to the rachis or if produces (opposite in the case of the prophyll and peduncular they are the precursor cells of the in¯orescence meristem. It bract to spiral in the case of the rachillae). The second order will be interesting to study the development of these cells of complexity is governed by the rachilla meristem formed in more closely in order to identify individual cell lineages. This the axil of the rachilla bract. The rachilla meristem has the may be possible with the aid of in situ hybridization in order same form and structure as the rachis meristem and has the to detect mRNA transcripts of genes which regulate the shoot same form in both male and female in¯orescences. However, November 2005] ADAM ET AL.ÐREPRODUCTIVE DEVELOPMENT IN ELAEIS GUINEENSIS 1849

Figs. 44±53. Micrographs showing whorl structure of developing and mature oil palm ¯owers (light micrographs unless otherwise stated). 44. Side view of normal pistillate ¯ower revealed by SEM. 45±53. Mantled ¯owers. 45. Side view of mantled pistillate ¯ower revealed by SEM. 46. Upper view of mantled staminate ¯ower revealed by SEM. 47. Longitudinal section (LS) of staminate mantled ¯owers with stamen and pseudocarpels. The fourth whorl is not developed. 48. LS of pistillate mantled ¯ower with sepals developed. 49. LS of pistillate mantled ¯ower at initiation of inner whorls. No difference with the normal pistillate ¯ower is visible. 50. Transverse section showing supernumerary carpels of mantled pistillate ¯ower. 51. TS of staminate mantled ¯ower with perianth organs developed. 52. LS of staminate mantled ¯ower at stage of initiation of reproductive organs. 53. Transverse view of staminate mantled ¯ower with pseudocarpels developed. Abbreviations: c, carpel; fm, ¯oral meristem; pc, pseudocarpel; pe, petal; rg, rudimentary gynoecium; s, sepals; sc, surpernumerary carpel; st, stamen; stap, staminodia primordia; stp, stamen primordium. Bar ϭ 0.1 mm for LM, ϭ 1 mm for SEM. 1850 AMERICAN JOURNAL OF BOTANY [Vol. 92 the divergent modes of functioning of the male and female Calderon-Urrea, 1993; Li et al., 1997). In staminate ¯owers of rachilla meristems is evident in the differences in the number A. of®cinalis, which is dioecious, ovule development in the and form of rachilla bracts and ¯oral meristems they produce. sterile gynoecium resembles that of the fertile gynoecium of pistillate ¯owers until degeneration starts in the cells of the Sexual specializationÐFemale in¯orescences of oil palm nucellus and integuments. Anther development is initially the display ¯oral triads with a central pistillate and two lateral same in pistillate as in staminate ¯owers, but the tapetum de- staminate ¯owers. The triad characterizes the subfamily Are- generates precociously followed by the abortion of the micro- coideae in which it represents the ultimate unit of the rachilla spore mother cells (Lazarte and Palser, 1979). In the second (Uhl and Drans®eld, 1987). Floral triads have been reported group of species that produce unisexual ¯owers, one or other within 138 of the 212 palm genera listed by Moore (1973). In of the sex organs is never initiated. For example, in staminate oil palm the ¯oral triads are borne in groups consisting of ¯owers of Cannabis sativa (Mohan Ram and Nath, 1964) and lateral staminate ¯owers and a central pistillate ¯ower with in both staminate and pistillate ¯owers of the dioecious species three associated bracteoles. Elaeis guineensis displays, like Mercurialis annua (Durand and Durand, 1991), Spinacia oler- species of the genus Ptychosperma (Uhl, 1976), the initiation acea (Sherry et al., 1993), Rumex acetosella (Ainsworth et al., of the two staminate ¯owers ®rst, followed by the pistillate 1995), and Philondendron acutatum (Boubes and BarabeÂ, ¯ower. 1996), either the gynoecium or the androecium is initiated but It has been suggested that the functional staminate ¯owers not both. Amongst those species that produce unisexual ¯ow- of the male oil palm in¯orescence are equivalent to the ®rst ers, E. guineensis falls into the ®rst group mentioned; never- monoaxial part of the female in¯orescence triad (Van Heel et theless, the primordia of reproductive organs lose their meri- al., 1987). Our observations support the conclusion that the stematic potential very early just after their initiation. As with triad is a sympodial ¯ower cluster originating from a single date palm (De Mason et al., 1982), the early development of meristem. It is interesting to note that the accompanying sta- staminate and pistillate ¯owers of oil palm is identical up to minate ¯ower is borne upon a pedicel, whereas the functional and including the stage of staminode/stamen initiation, diver- staminate ¯ower is sessile. The ¯oral triad is considered to gence being observable for the ®rst time when carpel initiation represent the ancestral con®guration of the Cocoseae (Tomlin- occurs. son, 1990). Within this tribe, a number of different modes of in¯orescence organization have been characterized. Of partic- The mantled homeotic variantÐOur results provide the ular interest is Beccariophoenix madagascariensis, which be- ®rst clear demonstration at the cytological level that the man- longs to a genus presumed to constitute the least specialized tled phenotype involves a homeotic transformation of the ¯oral group within the tribe (Uhl, 1988). In¯orescences are of mixed organs of the sterile or fertile gynoecium. Homeosis may be sex, with triads of ¯owers nearly throughout except for a few de®ned as the complete or partial replacement of one part of pairs of staminate ¯owers at the distal end. A second and more an organism with another (Lehmann and Sattler, 1992). The specialized member of the Cocoseae tribe characterized by Uhl variant phenotype mantled is regularly observed amongst clon- (1988) was Polyandrococos pectinatus. The latter species has al palms (Corley et al., 1986). In mantled oil palms, stami- a more marked spatial separation of staminate ¯owers (at the nodes and stamens of pistillate and functional staminate ¯ow- distal end of the in¯orescence) and ¯oral triads (at the basal ers develop respectively as pseudocarpels. In severe cases, the end). Polyandrococos pectinatus is protandrous, thus enabling ¯owers are sterile, although lesser-affected female ¯owers may allogamy. A similar situation is seen in coconut (Cocos nuci- be fertilized to give mantled . Although some observa- fera; Tomlinson, 1990), in which staminate and pistillate ¯ow- tions of the mantled phenotype have been reported (Corley et er function are separated both spatially and temporally within al., 1986), no studies at the histological and cellular level nec- a mixed sex in¯orescence. Oil palm might be considered to essary to obtain an in-depth understanding of this homeotic have evolved further than the aforementioned Cocoseae spe- variant have been described until now. Phenotypes resembling cies in that its functional dioecy is achieved by the separate that of mantled palms have been described in model plants production of single sex in¯orescences. Nevertheless, it is in- such as Arabidopsis thaliana or Antirrhinum majus. They are teresting to note the presence of the nonfunctional accompa- most frequently due to a disruption of the activity of B class nying staminate ¯owers on the female in¯orescence of E. gui- genes of the ABC ¯ower development model (Coen and Mey- neensis. One possible explanation of their presence is that they erowitz, 1991; Zahn et al., 2005) whereby petals are trans- represent an intermediate stage in an evolutionary progression formed into sepals and stamens are replaced by carpels. Given involving the loss of the ancestral ¯oral triads to give fully the similar appearance of sepals and petals in oil palm, it is unisexual in¯orescences. This evolutionary tendency appears dif®cult to determine whether or not the identity of whorl 2 is to occur in several palm groups (Moore and Uhl, 1982). affected by the mantled abnormality. Nevertheless, it is inter- esting to note that the expression of a key homeotic gene dis- Ontogenesis and structure of male and female reproduc- tinguishes whorls 1 and 2 in Asparagus of®cinalis (Park et al., tive organsÐIn common with about 30% of all angiosperms, 2003), a species which displays, like oil palm, morphologically oil palm produces unisexual (single sex) ¯owers rather than similar sepals and petals. Thus, although the mantled abnor- hermaphrodite ones (Richards, 1986). The ontogenesis of uni- mality does not involve any major changes to whorl 2 at the sexual ¯owers may be classi®ed into two general patterns cor- morphological level, there may perhaps be molecular alter- responding to different plant species. In the ®rst group, both ations. Ovules are not found in the developed pseudocarpels. stamens and gynoecial primordia are initiated and then arrested This suggests that the homeotic C and D functions found to at different stages of development. This mechanism is ob- specify carpel and ovule identity, respectively, in Petunia hy- served in the Leguminosae (Tucker, 1992), in Asparagus of- brida (Angenent and Colombo, 1996) might also be indepen- ®cinalis (Bracale et al., 1991), Silene sp. (Ye et al., 1991), Zea dent in oil palm. Some other naturally occurring ¯oral abnor- mays and Tripsacum sp. (De Long et al., 1993; Dellaporta and malities of a similar nature, named diwakkawakka (Hartley, November 2005] ADAM ET AL.ÐREPRODUCTIVE DEVELOPMENT IN ELAEIS GUINEENSIS 1851

1988) and coronado (unpublished data in our laboratory) have DE LONG, A., A. CALDERON-URREA, AND S. L. DELLAPORTA. 1993. Sex been observed. In diwakkawakka palms, staminate ¯owers are determination gene TASSELSEED2 of maize encodes a short chain al- normal while pistillate ¯owers have the same phenotype as cohol dehydrogenase required for stage speci®c ¯oral organ abortion. Cell 74: 757±768. those of mantled palms. This character appears to be inherited DE MASON, D. A., K. W. STOLTE, AND B. TISSERAT. 1982. Floral develop- monofactorially and to be dominant. In coronado ¯owers, both ment in Phoenix dactylifera. Canadian Journal of Botany 60: 1439± pistillate and functional staminate ¯owers are affected in the 1446. same way as for the mantled abnormality. The origin of the DRANSFIELD, J., AND N. W. UHL. 1998. Palmae. In K. Kubitzki [ed.], Families coronado phenotype, which has been observed in both E. gui- and genera of vascular plants, ¯owering plants: , vol. 4, neensis and E. oleifera, is unknown. It will clearly be of in- 306±389. Springer-Verlag, Berlin, Germany. terest to investigate the molecular causes of these three distinct DRANSFIELD, J., N. W. UHL,C.B.ASMUSSEN,W.J.BAKER,M.M.HARLEY, AND C. E. LEWIS. In press. An outline of a new phylogenetic classi®- abnormalities. cation of the palm family, Arecaceae. Kew Bulletin. TSO Publications, Studies of palm in¯orescence ontogenesis of the type de- Norwich, UK. scribed here are aimed at improving our understanding of the DURAND, B., AND R. DURAND. 1991. Sex determination and reproductive function and evolution of the ¯owering apparatus within and organ differentiation in Mercurialis. Plant Science 80: 49±65. beyond the Arecaceae. This approach should help to provide FISHER, D. B. 1968. Protein staining of ribboned epon sections for light mi- a meaningful classi®cation of the reproductive development of croscopy. Histochemie 16: 92±96. palm species as we incorporate our knowledge about the GENTY, P., A. GARZON,F.LUCCHINI, AND G. DELVARE. 1986. PolinizacioÂn entomo®la de la palma africana en America tropical. OleÂagineux 41: 99± growth, development, and functioning of in¯orescences and 112. ¯ower structures. These reference observations will be useful HARTLEY, C. W. S. 1988. The oil palm, 3rd ed. Longman Agriculture Series. in studies of the molecular determination of normal and variant Longman, London, UK. modes of ¯ower development in oil palm. They also provide JACQUEMARD, J. C. 1995. Le palmier aÁ huile. Series le technicien a useful starting point for studies on the functioning of the d'agriculture tropicale. Editions Maisonneuve & Larose, Paris, France. vegetative shoot apical meristem and the mechanisms by LAZARTE,J.E.,AND B. F. PALSER. 1979. 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