Sibbaldia: 143 The Journal of Botanic Garden Horticulture, No. 9 AMBORELLA TRICHOPODA – cultiVation of the MoSt anceStral angioSPerM in botanic gardenS Bernadette Große-Veldmann1, Nadja Korotkova2, Bernhard Reinken3, Wolfram Lobin4 & Wilhelm Barthlott5 abStract Amborella trichopoda baill., the most ancestral angiosperm, has been successfully cultivated in the botanic gardens of the university of bonn in germany (bg bonn) for more than a decade. the distribution of this plant – limited to the South Pacific island of new caledonia – and its cultivation has so far only been achieved in a few botanic gardens. this paper provides details about the culti- vation and propagation of Amborella, and information on its cultivation in botanic gardens around the world. the authors propose that the collections of this plant in botanic gardens could be used to establish ex situ conservation collections. introduction the botanic gardens of the university of bonn (bg bonn) have ten years’ experience in the cultivation of Amborella. the largest plant is 3m tall, flowers consistently and has even produced fruits. it flowered for the first time in 2003 and last flowered in october 2010. Many horticultural observations have been made during this time. the main purpose of this paper is to share our knowledge and experience with the botanic garden community in the hope that it encourages other botanic gardens to cultivate this plant. taxonoMY Amborella trichopoda baill. was described in 1869 by h.e. baillon (1869). because of its similarity to the flowers of Hedycarya, he placed it in the Monimiaceae (laurales). baillon knew only the male flowers and it was not until some 80 years later that bailey & Swamy (1948) described the female flowers. in the same year, the genus was placed into its own monogeneric family, amborellaceae, still in the laurales (Pichon, 1948). cronquist (1981) noted that it had several archaic characteristics, such as vessel-less wood, alternate leaves and essentially hypogynous flowers (with stamens, petals and 1. bernadette große-Veldmann is a student at the nees institute for biodiversity of Plants, university of bonn. address: nees-institut für biodiversität der Pflanzen, Meckenheimer allee 170, d-53115 bonn. e-mail: [email protected] 2. nadja Korotkova finished her Phd at the nees institute for biodiversity of Plants, and is now a member of the scientific staff at the botanical Museum, berlin dahlem and the institut für botanik. address: botanischer garten und botanisches Museum, Königin-luise-Str. 6–8, d-14191 berlin. 3. bernhard reinken is a horticulturist at the botanic gardens of the university of bonn and is responsible for Amborella. address: botanische gärten der universität bonn, Meckenheimer allee 171, d-53115 bonn. 4. Wolfram lobin is curator of the botanic gardens of the university of bonn. address: as above. 5. Wilhelm barthlott is director of the botanic gardens of the university of bonn and of the nees institute for biodiversity of Plants. address: as above. RB27908.indb 143 13/10/2011 08:21 144 bernadette groSSe-VeldMann ET AL. sepals attached above the ovary). takhtajan (1997) also regarded amborellaceae as being the most archaic family within the laurales and wrote, “Amborella is one of the most remarkable living fossils.” this changed following molecular studies in which Amborella was found to be sister to all other angiosperms (Qiu et al., 1999; Soltis et al., 1999). this placement as the most basal angiosperm was confirmed by subsequent molecular phylogenetic studies (borsch et al., 2003; Mathews & donoghue, 1999; Soltis et al., 2008). in 2003 the aPg classification recognised Amborella as a monotypic family, amborellaceae, within its own order, amborellales (aPgii, 2003; aPgiii, 2009). its new placement caused the species to become very well-known within the botanical establishment. the basal position of Amborella and the assumed plesiomorphic, or ancestral, state of many of its characters resulted in a further study of various aspects of its biology, such as flower morphology and anatomy (buzgo et al., 2004; endress & igersheim, 2000; Posluszny & tomlinson, 2003; thien et al., 2003; Williams, 2009; Yamada et al., 2001). Amborella was not known in cultivation before 1980 and is still rare in cultivation today. to the authors’ knowledge, a small number of botanic gardens currently grow Amborella and only three plants have flowered in cultivation (appendix 1). the culti- vation and ex situ conservation of this plant species is considered to be important at bg bonn. diStribution Amborella is endemic to new caledonia, a french dependent territory forming an archipelago in the South Pacific, off the north-east coast of australia. it is one of the smallest – although still very important – centres of biodiversity, in which very high levels of vascular plant endemism are found (Kier et al., 2009) with 76 per cent of the 3,063 native plant species being endemic (jaffré et al., 1998). furthermore, new caledonia has 108 endemic plant genera and 5 endemic plant families: amborellaceae, Paracryphiaceae, Phellinaceae, oncothecaceae and Strasburgeriaceae. Amborella only occurs on the main new caledonian island of grande terre. it grows in the understory of wet evergreen forests from about 100m to 1,000m above sea level, but most plants have been found at altitudes of between 500m and 800m. the main locations are Sarraméa (Plateau de dogy, col d’amieu), tchamba (haute tchamba) and touho. a distribution map is shown in fig. 1. new caledonia is situated between 20° and 23° latitude south and has a tropical climate which is modified by south-easterly trade winds. the temperature at about 300m ranges from 16°c to 24°c during the year. the highest precipitation occurs from january to March, whereas September, october and november are the driest months (golte, 1993). a climograph of a representative habitat of Amborella is illustrated in fig. 2. RB27908.indb 144 13/10/2011 08:21 AMBORELLA TRICHOPODA 145 fig. 1 distribution map of Amborella trichopoda. data sets taken from: herbier de nouvelle-calédonie, tropicos and Muséum nationale d’histoire naturelle, Paris. Map drawn by: laurens geffert and jens Mutke. fig. 2 climograph with precipitation (P) and temperature (t) for col d’amieu, grande terre, new caledonia. after golte (1993) (used with permission). RB27908.indb 145 13/10/2011 08:21 146 bernadette groSSe-VeldMann ET AL. Threats to amborella in the wild the already limited habitat of Amborella is threatened. among the main threats are natural hazards such as bush fires, and anthropogenic impacts such as road building, logging and mining. because of an abundance of rare metals, mining is the country’s main source of revenue (jaffré et al., 1998). furthermore, the introduction of exotic species has become a serious problem for new caledonia where about 800 neophytes currently compete with native species (gargominy et al., 1996). Conservation of amborella Ex situ conservation of this species is required. Since it has awakened great interest in the flora of new caledonia, conservation efforts on new caledonian plants are increasing (husby et al., 2010). in new caledonia, Amborella is currently propagated and culti- vated in at least one nursery as well as in the institut agronomique néo-calédonien (chad husby, pers. comm.). however, cultivation in botanic gardens is still rare. further cultivation in managed collections would act as a contribution to its conservation. deScriPtion Habit and vegetative morphology in its natural habitat the species grows up to 8m tall (jérémie, 1982), whereas at bg bonn the plant has reached a height of 3m and a width of about 2.5m (noted in 2011), making it almost certainly the largest specimen in cultivation (see fig. 3). the plant flowers consistently more or less every year; however, the actual month is not predictable and varies from one flowering event to the other. these have been noted in november 2003, august 2008, december 2009 and october 2010. Most recently the plant flowered for three months and produced fruits. Plant form: Medium-sized shrub; semi-self-supporting; maintaining its upright habit by leaning against the surrounding vegetation (thomas Speck, pers. comm.); stems divided in two branches at ground level, one 9.5cm, the other 10cm in diameter; numerous lateral scandent branches; new shoots mainly emerge at the terminal part of the branches. Leaves: evergreen; alternate; coriaceous; oblong-lanceolate; tip slightly acuminate, base obtuse; petiole up to 1cm; venation reticulate lamina 10(–16)cm × 4(–6) cm; stipules absent; leaf margin slightly incurved, undulate and serrate; leaf tip slightly recurved; marginal teeth allow the plant to hook into other branches or other plants (see figs 4 & 5). Inflorescence: cymose; up to 2.5cm long; numerous inflorescences arise from one leaf axil which gives the plant a bushy appearance; the bud is about 2mm in diameter before opening; 2 usually 5 flowers per inflorescence; inflorescences of the basal branches of the plant open first (see figs 5 & 6). Flowers: Small; dioceous; spiral; greenish to whitish; nocturnal scent absent; open at day and night. female flowers: 3–5mm in diameter; tepals whitish to slightly greenish RB27908.indb 146 13/10/2011 08:21 AMBORELLA TRICHOPODA 147 fig. 3 Amborella trichopoda specimen growing at fig. 4 Amborella trichopoda branch showing the bg bonn. Photo: Wilhelm barthlott. alternate leaves and flowering axillary inflorescence. Photo: Wilhelm barthlott. fig. 5 Amborella trichopoda stem showing the undulate leaf margin and cymose inflorescence. Photo: Wilhelm barthlott. RB27908.indb 147 13/10/2011 08:21 148 bernadette groSSe-VeldMann ET AL. fig. 6 cymose inflorescence of Amborella trichopoda with male flowers. Photo: Wilhelm barthlott. with a white margin; staminodes whitish; ovaries greenish; stigma whitish to yellow; 7–8 tepals; 4–6, mostly 5 carpels; ovary pitcher-shaped; 0–3, mostly 2 staminodes, large, with two thecae, thecae with two pollen sacs (see fig. 7). Male flowers: tepals greenish; stamens: filament greenish, anther whitish; 10–14 stamens; outer stamens larger than the inner ones; filament broad flat; anther pointed with four pollen sacs in two thecae (see fig. 8). Fruits: oval, 8–10 × 6–8mm, green when immature, ripening to red (endress & igersheim, 2000). the plant at bg bonn developed fruits for the first time after the recent flowering period in december 2010. these were green at the end of March 2011. the authors believe these to be the first fruits ever to develop in cultivation (see fig. 9). Dioecy and sex change Male flowers were observed when our plant flowered for the first time. during the subse- quent flowering events, the flowers were either functionally male or functionally female with rudimentary male organs. the functionally female flowers had up to three stami- nodes that were empty, that is, no pollen was found. our observations are confirmed by previous studies of buzgo et al. (2004), endress (2001), endress & igersheim (2000) and thien et al. (2003). RB27908.indb 148 13/10/2011 08:21 AMBORELLA TRICHOPODA 149 fig. 7 female flower of Amborella trichopoda; one staminodium can be seen. Photo: Katja rembold and eberhard fischer. fig. 8 Male flower of Amborella trichopoda. Photo: Wilhelm barthlott. fig. 9 Young fruit of Amborella trichopoda at bg bonn. Photo: annie Schulz. cultiVation the cultivation of Amborella is fairly straightforward in a cool glasshouse. temperature appears to be the most important factor for successful cultivation. high temperatures (above 25°c) should be avoided because they increase the occurrence of fungal infec- tions. changes in air humidity have a negligible effect. RB27908.indb 149 13/10/2011 08:21 150 bernadette groSSe-VeldMann ET AL. at bg bonn the plant has been grown in three environments to find out where it grows best; these are a temperate glasshouse with a minimum temperature of 13°c, a conservatory with a minimum temperature of 10°c and outside during the summer months. Amborella grows well under all of these conditions but the best results were achieved by growing it in the fern house between large tree ferns such as Cyathea sp. and Dicksonia sp. the average temperature in this glasshouse is 16–18°c. in summer the plants grow well outdoors. the city of bonn is situated in the winter hardiness zone 8 with an average annual temperature of 9.5°c (the january average is 5.9°c and the july average 17.4°c). the plants do not grow well in direct sunlight, so they are placed either between other larger plants or in a shaded place, as they are in the fern house. in winter, plants are brought inside and kept in a cool conservatory with a minimum temperature of 10°c. the compost in which the plants are grown is a peat-based mixture of 60 per cent einheitserde® ed73, 20 per cent pumice and 20 per cent lava. einheitserde® ed73 consists of 70 per cent peat and 30 per cent claydust with ph 5.8 and fertiliser with a n:P:K ratio of 14:16:18 plus a slow-release fertiliser with a n:P:K ratio of 20:10:15. fertiliser is applied every two weeks, increasing to weekly in the height of summer. We use Wuxal® super 8/8/6, consisting of 8 per cent n, 8 per cent K and 6 per cent P at a dilution rate of 0.2 per cent of product in the irrigation water. Pests and diseases the most significant problem is the oomycete Pythium splendens braun (identified by e.-c. oerke and u. Steiner, institute for Plant diseases, university of bonn, pers. comm.), which regularly infects the plants. the fungus is distributed worldwide and is also found in new caledonia (huguenin, 1966). the plant at bg bonn was infected by this fungus for the first time in 2003 during a very hot summer. the fungus thrives in warm temperatures; its optimal temperature is 38°c. in low temperatures it is not dangerous for the plant. the fungus can be controlled by the fungicide aliette® that contains 746g/kg fosetyl (as al-salt 800g/kg). Plants are spaced so that they are not touching each other and so that the fungus is not transmitted between plants. to prevent infections after pruning, the cutting areas – especially those of the larger branches – should be rubbed with charcoal powder. Propagation two propagation methods can be used for Amborella – cuttings and seeds. Propagation by seed has been successful at several botanic gardens (see appendix 1). the largest specimen at bg bonn has now developed fruits, but at the time of writing in May 2011 they have not ripened. Vegetative propagation is most frequently used at bg bonn. both soft tip and semi-ripe cuttings are possible, but it has been found that soft tip cuttings with 2–3 leaf RB27908.indb 150 13/10/2011 08:21 AMBORELLA TRICHOPODA 151 pairs are most successful. it is important that the cutting material is completely free of disease and the stems of the stock plant from which the cuttings were taken should be disinfected immediately after cutting with charcoal powder to prevent the entry of pathogens to the cuts. once ready for insertion the cuttings should be treated with a rooting hormone containing 3-indole butyric acid. after this, the cuttings can be inserted into 9cm clay pots, either singly or in groups. the number of cuttings per pot does not have a bearing on subsequent growth. the compost used for rooting cuttings is the same peat-based mix that is used for adult plants. the cuttings should be placed in a humid atmosphere and this is best achieved using a mist unit. the first roots grow in 12–14 weeks, but it takes almost one year for them to develop sufficiently to be removed from the frame and potted on into larger pots. concluSion appendix 1 shows that Amborella trichopoda is currently cultivated in at least six botanic gardens. from information obtained from botanic gardens across the world it can be seen that under the right conditions this species can be cultivated in botanic gardens; it flowers regularly and develops fruits. the authors consider this to be a good basis from which to establish ex situ conservation collections of the most ancestral angiosperm. acKnoWledgeMentS We wish to express our sincere thanks to laurens geffert and jens Mutke from the nees institute, bonn, for preparing the distribution map of Amborella, and to daud rafiqpoor for help with geographical information. We thank Winfried golte in the department of geography at the university of bonn for providing the climograph. We also thank thomas Speck from freiburg botanic garden and chad husby from Montgomery botanical center, Miami, florida for helpful information. referenceS aPg ii (2003). an update of the angiosperm Phylogeny group classification for the orders and families of flowering plants: aPg ii. Botanical Journal of the Linnean Society, 141, 399–436. aPg iii (2009). an update of the angiosperm Phylogeny group classification for the orders and families of flowering plants: aPg iii. Botanical Journal of the Linnean Society, 161(2), 105–121. baileY, i.W. & SWaMY, b.g.l. (1948). Amborella trichopoda baill., a new morphological type of vesselless dicotyledons. Journal of the Arnold Arboretum, 29, 245–254. baillon, h.e. (1869). Histoire des plantes. Vol.1. librairie de l. hachette et ciet, Paris. borSch, t., hilu, W., Quandt, d., Wilde, V., neinhuiS, c. & barthlott, W. RB27908.indb 151 13/10/2011 08:21 152 bernadette groSSe-VeldMann ET AL. (2003). non-coding plastid trnT-trnF sequences reveal a well resolved phylogeny of basal angiosperms. Journal of Evolutionary Biology, 16, 558–576. buzgo, M., SoltiS, P.S. & SoltiS, d.e. (2004). floral developmental morphology of Amborella trichopoda (amborellaceae). International Journal of Plant Sciences, 165(6), 925–947. cronQuiSt, a. (1981). An Integrated System of Classification of Flowering Plants. new York: columbia university Press. 1,262pp. endreSS, P.K. (2001). the flowers in extant basal angiosperms and inferences on ancestral flowers. International Journal of Plant Sciences, 162(5), 1,111–1,140. endreSS, P.K. & igerSheiM, a. (2000). reproductive structures of the basal angiosperm Amborella trichopoda (amborellaceae). International Journal of Plant Sciences, 161(6), 237–248. gargoMinY, o., bouchet, P., PaScal, M., jaffré, t. & tourneur, j.c. (1996). conséquences des introductions d’espèces animales et végétales sur la biodiversité en nouvelle-calédonie. Revue d’Ecologie (Terre Vie), 51, 375–402. golte, W. (1993). Araucaria: Verbreitung und Standortanspruche einer coniferengattung in vergleichender Sicht. Erdwissenschaftliche Forschung, 27. Stuttgart: franz Steiner Verlag, 167pp. huguenin, b. (1966). Micromycètes de nouvelle-calédonie. Cahiers ORSTOM, Série Biologie, 1, 61–91. huSbY, c., deterMann, r., MoYroud, r. & hall, b. (2010). les plantes de nouvelle- calédonie dans les établissements botaniques d’amérique du nord: un intérêt grandissant pour une flore extraordinairement belle et unique. Ethnopharmacologia, 46, 13–16. jaffré, t., bouchet, P. & Veillon, j.M. (1998). threatened plants of new caledonia: is the system of protected areas adequate? Biodiversity and Conservation, 7(1), 109–135. jéréMie, j. (1982). amborellaceae. in: aubréVille, a. & leroY, j.-f. (eds) Flore de la Nouvelle-Calédonie et Dépendances, 11, Muséum national d’histoire naturelle, Paris, 157–160. Kier, g., Kreft, h., lee, t.M., jetz, W., ibiSch, P.l., noWicKi, c., MutKe, j. & barthlott, W. (2009). a global assessment of endemism and species richness across island and mainland regions. Proceedings of the National Academy of Sciences of the United States of America, 106(23), 9,322–9,327. MatheWS, S. & donoghue, M.j. (1999). the root of angiosperm phylogeny inferred from duplicate phytochrome genes. Science, 286(5,441), 947–950. Pichon, P. (1948). les Monimiacées, famille hétérogène. Bulletin de la Muséum d’Histoire naturelle, 20, 383–384. PoSluSznY, u. & toMlinSon, P.b. (2003). aspects of inflorescence and floral development in the putative basal angiosperm Amborella trichopoda (amborellaceae). Canadian Journal of Botany, 81(1), 28–39. Qiu, Y.l., lee, j.h., bernaSconi-Quadroni, f., SoltiS, d.e., SoltiS, P.S., zaniS, M., ziMMer, e.a., chen, z.d., SaVolainen, V. & chaSe, M.W. (1999). the earliest angiosperms: evidence from mitochondrial, plastid and nuclear genomes. Nature, 402(6,760), 404–407. RB27908.indb 152 13/10/2011 08:21