American Journal of 88(8): 1371±1389. 2001.

HYDROPHOBIC TRICHOME LAYERS AND EPICUTICULAR WAX POWDERS IN BROMELIACEAE1

SIMON PIERCE,2,3 KATE MAXWELL,2 HOWARD GRIFFITHS,2 AND KLAUS WINTER

Smithsonian Tropical Research Institute, Apartado 2072, Balboa, Panama City, Republic of Panama

The distinctive foliar trichome of has promoted the evolution of an epiphytic habit in certain taxa by allowing the shoot to assume a signi®cant role in the uptake of water and mineral nutrients. Despite the profound ecophysiological and taxonomic importance of this epidermal structure, the functions of nonabsorbent trichomes in remaining Bromeliaceae are not fully understood. The hypothesis that light re¯ection from these trichome layers provides photoprotection was not supported by spectroradiometry and ¯uorimetry in the present study; the mean re¯ectance of visible light from trichome layers did not exceed 6.4% on the adaxial surfaces of representing a range of ecophysiological types nor was signi®cant photoprotection provided by their presence. Several reports suggesting water repellency in some terrestrial Bromeliaceae were investigated. Scanning electron microscopy (SEM) and a new techniqueЯuorographic dimensional imaging (FDI)Ðwere used to assess the interaction between aqueous droplets and the leaf surfaces of 86 species from 25 genera. In the majority of cases a dense layer of overlapping, stellate or peltate trichomes held water off the leaf epidermis proper. In the case of hydrophobic tank-forming tillandsioideae, a powdery epicuticular wax layer provided water repellency. The irregular architecture of these indumenta resulted in relatively little contact with water droplets. Most mesic terrestrial examined either possessed glabrous leaf blades or hydrophobic layers of con¯uent trichomes on the abaxial surface. Thus, the present study indicates that an important ancestral function of the foliar trichome in Bromeliaceae was water repellency. The ecophysiological consequences of hydrophobia are discussed.

Key words: Bromeliaceae; epicuticular wax; ¯uorographic dimensional imaging; SEM; trichomes; water repellency.

Bromeliaceae are ¯owering that are popular in hor- many Type 2 bromeliads are incapable of this function (Benz- ticulture and also of great ecological importance in the Neo- ing et al., 1976; LuÈttge et al., 1986). Trichome function has tropics, occupying a diverse range of habitats. One of the ®rst therefore played a pivotal role in the adaptive radiation of attempts to classify bromeliad diversity in an ecological con- Bromeliaceae via the operation of these different ecophysio- text was made by Pittendrigh (1948), who elaborated on the logical strategies. observation of Tietze (1906) that life form and the function of However, the function(s) of the trichomes of Type 1 bro- leaf hairs was re¯ected in the taxonomic relationships of gen- meliads remains enigmatic. Molecular phylogenetics indicates era. Pittendrigh's scheme was further expanded by Benzing that the genera Ayensua and are basal to the rest (2000) into the ®ve ecophysiological types summarized in Ta- of the family (Terry, Brown, and Olmstead, 1997; Horres et ble 1. al., 2000; Crayn, Winter, and Smith, unpublished data). Al- Leaf hairs or foliar trichomes (i.e., unicellular or multicel- though direct fossil evidence is negligible, mesic Type 1 Pit- lular structures arising from the epidermal tissues; Bell, 1991) cairnioideae (e.g., Ayensua, some Brocchinia, Fosterella, Pit- are almost ubiquitous in Bromeliaceae (Benzing, 1976) and cairnia) are also considered to exhibit a primitive life form are perhaps the most distinguishing vegetative feature of the (i.e., ecophysiologically they most closely resemble a hypo- family. It is well documented that the peltate trichomes be- thetical ancestor of the family). This assessment is based not longing to species with Type 3, 4, and 5 life forms support only on subfamilial characteristics such as the extensive root epiphytism by endowing the shoot with the capacity to aug- system (Tietze, 1906), but also on the presence of less ad- ment or replace the absorptive functions of roots (Schimper, vanced nonsucculent C3 physiology (see Medina, 1974) and 1888; Billings, 1904; Mez, 1904; Benzing, 1970, 1976; Benz- the simpler structure of the trichome (Benzing, 1980). Indeed, ing and Burt, 1970; Benzing et al., 1976; Nyman et al., 1987; within the Brocchinia advanced Type 4 species possess Smith, 1989; see Benzing [1980] for a detailed discussion of absorbing trichomes, while nonimpounding terrestrial species their mode of action). The trichomes of terrestrial Type 1 and possess less highly organized trichomes and are more basal within the genus (N.B. the most primitive of these, B. pris- 1 Manuscript received 27 June 2000; revision accepted 1 February 2001. matica, possesses stellate trichomes similar to those of Fos- The authors thank Harry E. Luther and Bruce K. Holst for assistance with terella species; Givnish et al., 1997). Thus, foliar trichomes of identi®cation, for generously putting the living collections and herbarium of mesic Type 1 Pitcairnioideae mediate primitive functions. the Marie Selby Botanic Gardens at our disposal, and, along with Darren M. Crayn, for providing comments on an early version of the manuscript; David Many roles other than water and nutrient absorption have H. Benzing for invaluable criticism during the review process; Jason R. Grant been ascribed to bromeliad trichomes, but these functions of- for aid with systematic issues; Jorge E. Aranda for collecting in Fortuna, ten only apply to a small number of species (such as the at- Panama; Jorge Ceballos for invaluable technical support with the scanning traction of pollinators or seed dispersers in the case of some electron microscope; and Richard Gottsberger and Aurelio Virgo for general and Billbergia species; Benzing, 2000). More gen- assistance. We gratefully acknowledge support by a grant from the Andrew eral hypotheses concerning the function of bromeliad tri- W. Mellon Foundation through the Smithsonian Tropical Research Institute. 2 University of Cambridge, Department of Sciences, Downing Street, chomes include obstruction of predators and pathogens (Benz- Cambridge, CB2 3EA, UK. ing, 2000), reduction of transpiration (Billings, 1904), and 3 Author for reprint requests. photoprotection (Benzing and Renfrow, 1971; LuÈttge et al., 1371 1372 AMERICAN JOURNAL OF BOTANY [Vol. 88

TABLE 1. Life forms or ecophysiological types of Bromeliaceae (after Benzing, 2000).

Life form Characteristics 1 Terrestrial herbs of subfamily Pitcairnioideae (and many Bromelioideae) that use roots to acquire water and nutrientsÐthe leaf hairs being nonabsorbent. 2 Terrestrial Bromelioideae with leaf bases that form a rudimentary watertight ``tank'' into which some axillary roots may grow. 3 Terrestrial or epiphytic herbs in subfamily Bromelioideae, the roots of which have reduced importance in water and nutrient acquisition with the leaf bases forming an extensive water-holding tankÐpredominantly crassulacean acid metabolism (CAM), with leaf hairs that have some capacity to take up water and nutrients.

4 Tank-forming epiphytes in subfamily Tillandsioideae and some BrocchiniaÐpredominantly C3 and with high densities of leaf hairs on the leaf bases that are highly effective at water and nutrient uptake, the roots functioning primarily as holdfasts. 5 Succulent CAM Tillandsioideae that are epiphytic or lithophytic, with leaf hairs taking up water directly over the entire leaf surface (without a tank) and possessing holdfast roots, if any.

1986). The deterrence of predators and pathogens currently has glabrous cuticle of the adaxial surface, perhaps suggesting that no experimental support. Reduction of transpiration is a xe- the trichome layer hindered absorption. Widespread occur- romorphic adaptation, and as such, it is unlikely that this rence of repellent trichome layers on the abaxial leaf blade would be an important selection pressure acting on ancestors surfaces of mesic Type 1 bromeliads would therefore suggest in mesic habitats. that hydrophobia was an important property of the foliar tri- In high densities, bromeliad trichomes produce a whitish chome in ancestral Bromeliaceae. leaf surface that re¯ects light when dry. This has been quan- Also relevant to this study are the hydrophobic waxy sur- ti®ed in Type 4 Tillandsia fasciculata (Benzing and Renfrow, faces of and Catopsis berteroniana. Tom- 1971) and semimesic Type 1 Pitcairnia integrifolia (LuÈttge et linson (1969) suggests that in the case of C. berteroniana these al., 1986) and is highly suggestive of a role in photoprotection. promote the run-off of water from the leaf blades into the tank However, in the more relevant case of Type 1 P. integrifolia, and attraction and entrapment of insect prey by these carniv- trichomes are restricted to the abaxial surface of the leaf; had orous species have also been suggested (Fish, 1976; Frank and these trichomes developed primarily to serve a photoprotective O'Meara, 1984). These species also share advanced Type 4 role, then they would be expected to occur at least in equal life forms, which usually possess hydrophilic trichomes at densities on the glabrous adaxial surface. LuÈttge et al. (1986) least lining the tank. Determinations of the occurrence of hy- note that the edges of the leaves of P. integrifolia roll inwards drophobic surfaces in Tillandsioideae and Brocchinia could to expose the trichomed abaxial surface during the dry season, shed additional light on the evolution of the Type 4 life form. perhaps to promote re¯ectance, and propose this as a form of The present study employs a novel technique, ¯uorographic regulation of light re¯ectance. However, this behavior may oc- dimensional imaging (FDI), to assess the interactions between cur simply as a consequence of drought in glabrous species aqueous droplets and the leaf blade surfaces of 86 ecologically (e.g., Pitcairnia valerii; personal observation), perhaps as a diverse bromeliad species representing 25 genera and all three response to water loss and concomitant shrinkage of water- subfamilies. Fluorographic dimensional imaging is used in storage parenchyma in the hypodermis (see Billings, 1904). conjunction with scanning electron microscopy (SEM) and More importantly, the trichomes of P. integrifolia and P. bif- spectroradiometry to reveal the mechanism by which certain rons were not found to in¯uence the heating of leaves (LuÈttge trichomes and epicuticular wax powders repel water. Fluorim- et al., 1986). Thus, a photoprotective role for trichomes re- etry is used to investigate the hypothesized role of trichomes mains without direct supporting evidence; an investigation of and wax layers in photoprotection. Nomenclature follows that photoinhibition using ¯uorimetry techniques has yet to be un- of Luther and Sieff (1998), with the exception of the recently dertaken. rejected genus Pepinia (Taylor and Robinson, 1999), which is Evidence for a further general hypothesis concerning the recognized as a subgenus of Pitcairnia (sensu Smith and role of the trichome in terrestrial Bromeliaceae is present in Downs, 1974). the literature, but has apparently been overlooked. Krauss (1948±1949) working on Ananas comosus noted that ``the tri- MATERIALS AND METHODS chomes on the lower surface of the leaf blade proper appear unwettable. Drops of water placed on this surface do not Plant material of Panamanian origin was collected from the wild, with spread, but remain unabsorbed for experimental periods of 3 voucher specimens being held at the main herbarium of the Smithsonian Trop- to 6 h.'' ical Research Institute, Panama (herbarium code SCZ) and at the University Krauss (1948±1949) also went on to observe that, whereas of Panama (PMA). Material of Trinidadian origin was obtained from the living the absorbent trichomes of Tillandsia usneoides lost their pale collections of Moorbank Botanic Gardens (Newcastle-upon-Tyne, UK). An- whitish color when wetted (Billings, 1904), those on the ab- anas comosus was grown from meristem culture, with original material pro- vided by the Centre International de Recherche en Agronomie et Development axial surface of A. comosus did not, as a consequence of air (Montpellier, France). All other material was obtained from the living collec- trapped beneath the trichomes. This implies that the trichomes tions at the Marie Selby Botanic Gardens, Sarasota, Florida, USA (accession on the abaxial surface of A. comosus repel water. Also, the numbers available on request). abaxial surfaces of Pitcairnia integrifolia and P. macrochla- Repellency was denoted by the depth of aqueous droplets on adaxial and mys leaf blades appear to be unwettable (Benzing, Seemann, abaxial leaf blade surfaces. For FDI of aqueous droplets, calibration standards and Renfrow, 1978; LuÈttge et al., 1986), and in the case of P. were prepared using glass coverslips (ϳ2 cm wide), one-half being coated integrifolia, ``water repellent.'' Indeed, Benzing (1970) dis- with a ¯at ®lm of paraplast wax (Sigma Chemical, St. Louis, Missouri, USA), covered that after 12 h of exposure the abaxial surface of P. and the other half remaining as an exposed glass surface. The thickness of macrochlamys had absorbed ϳ3.5 times less zinc65 than the these wax and glass standards was measured by micrometer, and these stan- August 2001] PIERCE ET AL.ÐWATER REPELLENT INDUMENTA OF BROMELIADS 1373 dards were lightly ®xed along one edge of a strong glass plate of ϳ40 ϫ 40 in seminatural conditions in an open-sided greenhouse at the main Smithson- cm. ian Tropical Research Institute facility in Panama. Excluding the cultivar of Leaf discs were cut from intact and surface denuded midleaf portions of Ananas comosus, these species grow in semi-exposed to exposed microhab- leaf blade (from two-thirds of the way along the blade). In many species itats and may experience several hours of direct sunlight each day (LuÈttge et denudation was achieved using sticky tape, although some species such as al., 1986; personal observations). A treatment of excessive excitation therefore Ananas comosus required careful scraping with a scalpel blade. In the case consisted of transferring plants grown in moderate sunlight (ϳ450 ␮mol pho- of apparently glabrous leaves, the procedure of denudation with sticky tape ton´mϪ2´secϪ1 at midday) to direct sunlight at midday (PPFD ഠ1700 ␮mol was conducted for the sake of consistency. Leaf discs from replicate leaves photon´mϪ2´secϪ1) for 1 h. The degree of photoinhibition was denoted by the (where possible from separate individuals) were then ®xed in rows onto the decline in the dark-adapted ratio of variable to maximum chlorophyll ¯uo- glass plate, with intact and denuded examples of both surfaces presented up- rescence (Fv/Fm) following this treatment, with intact and denuded surfaces permost. being compared.

Droplets (10-␮L each) of 0.05% (mass by volume in distilled H2O) ¯uo- For scanning electron microscopy, the majority of leaf samples were de- rescein sodium solution were quickly pipetted onto the surface of the leaf hydrated through an alcohol series, critical point dried (CPD) in CO2, and discs and calibration standards and left to stand for 40 min in a darkened then sputter-coated with gold-palladium (Hummer VI-A, Anatech, Spring®eld, room. In these darkened conditions, the leaf discs and standards were then Virginia, USA) before examination in the scanning electron microscope (Jeol illuminated with an ultraviolet (UV) transilluminator (Fotodyne, Hartland, JSM-5300LV, Jeol, Tokyo, Japan). However, samples of Catopsis were not Wisconsin, USA), and the resulting ¯uorescence from the excited ¯uoro- dehydrated in this manner, as the solvents used in CPD may destroy the chrome was photographed using a level camera mounted directly above the structure of wax surfaces (Juniper and Jeffree, 1983); samples were placed in leaf discs. Initial tests determined that the following camera settings provided the scanning electron microscope without preparation. the greatest depth of ®eld and contrast, with well-exposed ¯uorescence and a darkened background: an aperture of f/22, aperture priority (or a 9-sec ex- posure with a cable release), using ISO 100/DIN 21Њ color-reversal ®lm (Ko- RESULTS dak Elite). The depth of droplets on wax and glass standards was determined by micrometer immediately after the ¯uorograph was taken. Light re¯ectance and photoprotectionÐAn intact layer of After processing, ¯uorographs were digitally scanned (LS-2000, Nikon, dry trichomes increased the re¯ectance of visible light (400± Shinagawa-Ku, Tokyo, Japan) and the luminosity of ¯uorescein droplets was 700 nm) by an average of 6.4% on the adaxial surface of determined using Corel PHOTO-PAINT7 (Corel, Ottawa, Ontario, Canada) dactylina, although not signi®cantly on the abaxial imaging software (selecting each particular region of the image with the ``eye- surface (P Ͼ 0.05; Figs. 1±4). Re¯ectance was increased by dropper'' tool, and recording the luminosity (L) of the ``paint'' color). To 5.0 and 3.9% on adaxial and abaxial surfaces, respectively, of compensate for possible uneven lighting, eight measurements were taken from Tillandsia ¯exuosa (data not shown), 4.9 and 10.6% on adaxial each droplet, and the measurements were averaged. Luminosity and depth and abaxial surfaces of Ananas comosus (Figs. 5±8), and data from the glass and wax standards were then regressed (Excel, Microsoft, 17.8% on the abaxial surface of Pitcairnia integrifolia (but not Seattle, Washington, USA) to create a calibration equation, from which the on the glabrous adaxial surface; Figs. 9±12). Powdery epicu- depth of droplets on leaf discs was calculated using respective luminosity ticular wax increased re¯ectance of visible light by a mean of values. This technique allowed rapid, inexpensive, mass screening of samples. 6.3 and 6.6% on adaxial and abaxial surfaces, respectively, of The difference in droplet depth (⌬D) due to surface features can be summa- Catopsis micrantha (Figs. 13±16). Low densities of ®lmy tri- rized by the following equation: chomes were observed via SEM on the adaxial surface of Type 4 Werauhia sanguinolenta, but these did not alter re¯ectance ⌬Dd(b)ϭ i d(b)Ϫ e d(b) (1) (data not shown). The increased re¯ectance conferred by tri- where i ϭ droplet depth on intact surface, e ϭ droplet depth on denuded chomes or wax was not suf®cient for photoprotection, with the surface, d ϭ adaxial surface or alternatively b ϭ abaxial surface. extent of photodamage (as denoted by a percentage decline in In order to examine the effect of water surface tension on the interaction Fv/Fm) exhibited by leaves with intact surfaces equaling that between trichomes and water, the above FDI technique was also used on the of leaves denuded of trichomes or wax powders (after expo- leaves of Ananas comosus, using droplets (10-␮L each) of ¯uorescein sodium sure to an equivalent and excessive photon dose; Table 2). solution (5 mL of 0.05% ¯uorescein and 0.5 mL distilled H O); with further 2 When inundated with water, the adaxial surfaces of Aech- replicates on which 10-␮L droplets of a solution of ¯uorescein and household detergent (5 mL of 0.05% ¯uorescein and 0.5 mL neat detergent) were used. mea dactylina and Ananas comosus (Figs. 3, 7) and both sur- Re¯ectance of light by leaves was measured using an LI-1800 portable faces of Tillandsia ¯exuosa lost the re¯ectivity conferred by spectroradiometer (LI-COR, Lincoln, Nebraska, USA), via an 1800±12s ex- their trichomes. The trichomes of Pitcairnia integrifolia and ternal integrating sphere (LI-COR). Ranges of re¯ectance values were nor- those of the abaxial surface of Ananas comosus retained their re¯ectivity when treated in this manner (Figs. 8, 12). A surface malized to 100% using barium sulfate (BaSO4) as a standard; this compound has an absolute re¯ectivity of 99.3% in the wavelength range 300±800 nm ®lm of water could not be sustained on the leaves of Catopsis (Munsell Color, New Windsor, New York, USA). Measurements were taken micrantha even after several days of inundation. Indumenta of intact, water-inundated, and denuded leaf surfaces (both adaxial and ab- did not increase the re¯ectance of infrared light (800 nm) in axial). Species with water repellent trichome layers were inundated by soaking most species, except for Catopsis micrantha and Pitcairnia in water for1horuntil a surface ®lm of water could be sustained on their integrifolia. Re¯ectance of infrared wavelengths was higher removal from the water. Once again, in the case of surfaces that appeared to (40±50%) than the re¯ectance of visible light in all species have no trichomes, the denudation process was carried out with sticky tape studied. for consistency's sake. Average re¯ectance values of photosynthetically active radiation (PAR) were calculated as a mean across the wavelength range 400± 700 nm. The re¯ectance conferred by trichomes or wax powders is de®ned Leaf blade interactions with waterÐA typical ¯uorograph as the difference in mean re¯ection between intact and denuded surfaces. for a single species (Catopsis micrantha) is shown in Fig. 17. Photoinhibition of photosystem II was investigated using a PAM-2000 por- Fluorographic dimensional imaging determined that droplet table modulated ¯uorimeter (H. Walz, Effeltrich, Germany). Aechmea dacty- depth had diminished after 40 min on the intact leaf blade lina, Ananas comosus cv. Cayenne Lisse, Catopsis micrantha, Pitcairnia in- surfaces of Type 5 species when compared with surfaces de- tegrifolia, Tillandsia ¯exuosa, and Werauhia sanguinolenta were maintained nuded of trichomes (⌬D). For example, on leaf blades of Til- 1374 AMERICAN JOURNAL OF BOTANY [Vol. 88

Figs. 1±4. Aechmea dactylina leaf blade surfaces. 1±2. Scanning electron micrographs of the adaxial and abaxial surfaces, respectively. 3±4. Re¯ectance of light by the adaxial and abaxial surfaces, respectively. Re¯ectance data represent the mean Ϯ 1 SE of four replicates.

landsia nana, ⌬Dd ϭϪ732 and ⌬Db ϭϪ876 ␮m; con®rming Ϫ267 ␮m and ⌬Db ϭ 226 ␮m; Type 1 Cryptanthus whitmanii, these leaves to be highly hydrophilic. ⌬Dd ϭϪ205 ␮m and ⌬Db ϭ 407 ␮m; Type 1 Orthophytum Droplets exhibited no signi®cant difference in depth be- benzingii, ⌬Dd ϭϪ477 ␮m and ⌬Db ϭ 474 ␮m). tween intact and denuded leaf blade surfaces in most Type 4 Of the mesic Type 1 pitcairnioids, genera such as Fosterella species (P Յ 0.05; Table 3). However, there were some notable and Pitcairnia either possessed hydrophobic abaxial surfaces, exceptions; for example the hydrophobic abaxial surface of due solely to trichome cover (e.g., Pitcairnia integrifolia, ⌬Db Vriesea monstrum (⌬Dd ϭ 214 ␮m; Table 3) and both hydro- ϭ 230 ␮m), or were entirely glabrous and noninteractive (e.g., philic surfaces of Tillandsia elongata (⌬Dd ϭϪ210 ␮m and Pitcairnia patenti¯ora), with a small number possessing hy- ⌬Db ϭϪ190 ␮m). Many Type 4 taxa possessed hydrophobic drophobic adaxial surfaces (Pitcairnia arcuata, ⌬Dd ϭ 310 waxy surfaces, e.g., Catopsis micrantha (⌬Dd ϭ 800 ␮m and ␮m). The more xeromorphic pitcairnioid genera showed a ⌬Db ϭ 960 ␮m), Guzmania macropoda (⌬Db ϭ 216 ␮m), and range of trichome-mediated interactions with surface water, Werauhia capitata (⌬Db ϭ 350 ␮m). including species that possessed both hydrophilic and hydro- Trichomes, but not wax, lent subfamily Bromelioideae a phobic leaf blade surfaces (e.g., Dyckia marnier-lapostollei, range of interactions with leaf surface water. This included no ⌬Dd ϭϪ457 ␮m and ⌬Db ϭ 740 ␮m; Table 3). interaction at all (e.g., both surfaces of Type 2 Bromelia pin- Of the 16 species examined from the el®n cloud forest at guin; Table 3), hydrophilic surfaces (e.g., Type 3 Aechmea Cerro Jefe in central Panama, six possessed water-repellent dactylina, ⌬Dd ϭϪ220 ␮m and ⌬Db ϭϪ130 ␮m; Type 3 leaf surfaces (Table 3). These were either Type 1 species with A. fendleri, ⌬Dd ϭϪ130 ␮m and ⌬Db ϭϪ110 ␮m), and the repellent trichomes (Pitcairnia arcuata, Ronnbergia explo- hydrophobic abaxial surfaces of species such as Type 2 An- dens) or Type 4 species with relatively upright leaves that used anas comosus (⌬Db ϭ 160 ␮m; Fig. 8) and Type 1 Ronnbergia trichomes (Vriesea monstrum) or epicuticular wax powders explodens (⌬Db ϭ 100 ␮m). A number of bromelioid species (Catopsis micrantha, Guzmania macropoda, Werauhia capi- possessed both hydrophilic adaxial surfaces and hydrophobic tata) to shed water. A further six were Type 4 species equipped abaxial surfaces (e.g., Type 3 Aechmea nudicaulis, ⌬Dd ϭ with hypostomatous and horizontally orientated leaves. August 2001] PIERCE ET AL.ÐWATER REPELLENT INDUMENTA OF BROMELIADS 1375

Figs. 5±8. Ananas comosus leaf blade surfaces. 5±6. Scanning electron micrographs of the adaxial and abaxial surfaces, respectively. 7±8. Re¯ectance of light by the adaxial and abaxial surfaces, respectively. Re¯ectance data represent the mean Ϯ 1 SE of four replicates.

The wax powder layer of Catopsis micrantha was less pro- being highly modi®ed with an elongate wing that spirals nounced towards the tip of the leaf blade, where it still pro- around itself to form a hair-like structure (Fig. 33). moted beading up of water (Figs. 18, 19). This layer was con- Low densities of ring-peltate trichomes occurred on the hy- tinuous over the leaf blade surface (Figs. 13, 15, 20), but was drophilic surfaces of Aechmea dactylina (Figs. 1, 2, 34, 35). not present on the adaxial surface of the leaf sheath within the Individual trichomes were structurally comparable to the tri- tank of the plant. This surface is densely covered with peltate chomes comprising the continuous hydrophobic trichome lay- trichomes (Fig. 21). Powdery epicuticular wax was also pre- ers of Ananas comosus, Fosterella petiolata, Pitcairnia cor- sent on hydrophobic surfaces of Alcantarea odorata, Broc- allina, Ronnbergia explodens, and Vriesea monstrum (Figs. 6, chinia reducta, and Werauhia capitata (Figs. 22±25). 29±31, 36, 37). None of these species possessed wax powders, Surfaces that showed no trichome- or wax-mediated inter- either on the trichomes or elsewhere. action with water generally either possessed thin, ®lmy peltate On the hydrophilic adaxial surface and hydrophobic abaxial trichomes (e.g., the adaxial surface of Vriesea monstrum; Fig. surface of Cryptanthus whitmanii the trichomes appeared no 26) or lacked surface structures (e.g., the adaxial surfaces of different, although the lower densities on the adaxial surface Fosterella petiolata, Pitcairnia corallina, Pitcairnia integri- revealed the leaf epidermis proper to SEM (Figs. 38, 39). folia; Figs. 9, 27, 28). Water repellent trichomed surfaces fea- Aechmea nudicaulis also has low densities of thin, ®lmy tri- tured either high densities of large, overlapping peltate tri- chomes on the hydrophilic adaxial surface (Fig. 40) and a chomes consisting mainly of extrusive ring cells (i.e., ``ring- typical hydrophobic abaxial surface (Fig. 41). No species in peltate'' trichomes; Figs. 6, 10, 29±31) or low densities of any subfamily possessed a hydrophobic adaxial surface com- tangled stellate trichomes forming a discontinuous indumen- bined with a hydrophilic abaxial surface. Water repellent epi- tum (e.g., Pitcairnia arcuata; Fig. 32). Trichomes of Puya laxa cuticular wax powders or con¯uent layers of large ring-peltate did not signi®cantly interact with water droplets (P Յ 0.05; trichomes occurred exclusively on surfaces that possessed sto- Table 3)Ðthis species possesses two types of trichome, one mata in the species studied. 1376 AMERICAN JOURNAL OF BOTANY [Vol. 88

Figs. 9±12. Pitcairnia integrifolia leaf blade surfaces. 9±10. Scanning electron micrographs of the adaxial and abaxial surfaces, respectively. 11±12. Re- ¯ectance of light by the adaxial and abaxial surfaces, respectively. Re¯ectance data represent the mean Ϯ 1 SE of four replicates.

The addition of detergent to the ¯uorescein solution used in tance conferred was correlated with the mode of interaction FDI resulted in higher wettability of both adaxial and abaxial between surfaces and water. Hydrophobic surfaces did not lose surfaces of Ananas comosus, with aqueous droplets (10-␮L re¯ectivity when wet, whereas hydrophilic trichomes did (see volume) spreading to negligible depth (14.8 Ϯ 3.2 ␮m adax- also Billings, 1904; Krauss, 1948±1949; Benzing, Seemann, ially and 17.6 Ϯ 5.3 ␮m abaxially; Table 4) when the surface and Renfrow, 1978), and higher re¯ectivities on abaxial sur- tension of the water was reduced in this manner. faces were correlated with the presence of dense hydrophobic indumenta (e.g., Ananas comosus, Pitcairnia integrifolia). DISCUSSION Thus, the data indicate that hydrophobic and dry hydrophilic trichome layers inherently scatter light, but are unlikely to Light re¯ectance and photoprotectionÐThe data indicate have evolved primarily for the purpose of photoprotection in that trichomes and epicuticular wax powders do not have a Bromeliaceae. signi®cant photoprotective function in a range of ecophysio- The highly unusual, woolly trichomes of Puya laxa (Fig. logical types (Types 1±4). Trichomes either did not increase 33) did not interact with water droplets on the leaf surface light re¯ectance from leaf blades (e.g., Werauhia sanguinolen- (Table 3). These trichomes probably act as protection against ta) or the mean re¯ectance conferred by trichomes or wax did frost damage as exhibited by a number of Puya species grow- not exceed 6.4% on the adaxial surfaces of the species studied ing in high altitude habitats (Miller, 1994). As this example (with up to 17.8% on the abaxial surfaces). This was not suf- illustrates, distinct taxa produce trichomes that represent a ®cient to signi®cantly alter down-regulation of photosystem II more speci®c adaptation to local environmental conditions. by excess light in these species (Table 2). Indeed, trichomes Thus, dense indumenta could yet prove to furnish photopro- and epicuticular wax powders conferring re¯ectances of be- tection in the case of more extreme xerophytes (Type 5 spe- tween ϳ45 and 55% photoprotect certain desert plants (Eh- cies). A thorough investigation of the ¯uorescence character- leringer and BjoÈrkman, 1976; Robinson, Lovelock, and Os- istics of this life form was beyond the scope of the present mond, 1993). Also, the present study indicated that the re¯ec- study. August 2001] PIERCE ET AL.ÐWATER REPELLENT INDUMENTA OF BROMELIADS 1377

Figs. 13±16. Catopsis micrantha leaf blade surfaces. 13±14. Scanning electron micrographs of the adaxial and abaxial surfaces, respectively. 15±16. Re- ¯ectance of light by the adaxial and abaxial surfaces, respectively. Re¯ectance data represent the mean Ϯ 1 SE of four replicates.

TABLE 2. Decrease in Fv/Fm (the dark-adapted ratio of variable to maximum chlorophyll ¯uorescence) of six species after exposure to saturating light (PPFD ഠ1700 ␮mol´mϪ2´sϪ1) for 1 h, with the leaf blade surface either intact or denuded of surface features. Values are means Ϯ1SEof four replicates. The absence of differences in letters (a) between means of intact and denuded treatments indicates that there were no signi®cant differences at the P Յ 0.05 level as determined by Student's t test. Life forms or ecophysiological types follow Benzing (2000).

Decrease in Fv/Fm (%) Species Life form Surface Intact Denuded Aechmea dactylina 3 Adaxial 30.0 Ϯ 7.6 a 37.9 Ϯ 8.8 a Abaxial 26.9 Ϯ 5.8 a 30.1 Ϯ 5.9 a Ananas comosus 2 Adaxial 58.5 Ϯ 5.6 a 43.8 Ϯ 6.5 a Abaxial 26.3 Ϯ 4.7 a 29.2 Ϯ 3.4 a Catopsis micrantha 4 Adaxial 29.4 Ϯ 8.7 a 28.2 Ϯ 3.2 a Abaxial 22.5 Ϯ 5.0 a 29.5 Ϯ 1.7 a Pitcairnia integrifolia 1 Adaxial 52.7 Ϯ 4.9 a 48.4 Ϯ 2.7 a Abaxial 35.6 Ϯ 6.3 a 39.0 Ϯ 1.4 a Tillandsia ¯exuosa 4±5 Adaxial 11.7 Ϯ 4.1 a 21.1 Ϯ 8.7 a Abaxial 30.7 Ϯ 9.1 a 38.2 Ϯ 10.1 a Werauhia sanguinolenta 4 Adaxial 47.3 Ϯ 7.2 a 39.4 Ϯ 3.2 a Abaxial 34.1 Ϯ 1.4 a 30.9 Ϯ 2.7 a 1378 AMERICAN JOURNAL OF BOTANY [Vol. 88

Fig. 17. A typical ¯uorograph for a single species (Catopsis micrantha), used to determine the depth of aqueous droplets on leaf disc surfaces (denoting repellency) via the comparison of ¯uorescence signatures of ¯uorescein droplets against calibration droplets of known depth. In this example, epicuticular wax powder layers from the adaxial and abaxial leaf blade surfaces are either present (intact) or removed (denuded). Mean depth values presented include Ϯ 1 SE, with signi®cant differences between means (at the P Յ 0.05 level) of four replicates determined using Fisher's multiple comparison procedure.

The mechanism of water repellencyÐBrewer, Smith, and adhesion (Brewer, Smith, and Vogelmann, 1991; Watanabe and Vogelmann (1991) noted three kinds of interaction between Yamaguchi, 1993). The physics of these surface±water inter- water and the trichomes of ¯owering plants: (1) low trichome actions are outlined by Barthlott and Neinhuis (1997). This densities that do not in¯uence droplet retention or the location hydrophobic mechanism is readily demonstrable in Bromeli- of surface moisture, (2) low densities of trichomes that induce aceae. For example, a wetted pineapple leaf (Ananas comosus) surface water to aggregate into patches, and (3) high densities will lose the pale coloration of the abaxial surface only if of trichomes that lift water off the leaf surface. The leaf blade detergent is ®rst added to the water. Species with absorbent surfaces of many Type 4 bromeliads exhibit low trichome den- trichomes, on the other hand, lose this pale coloration and sities (Benzing, 1980) and did not interact detectably with sur- re¯ectivity immediately on wetting (Billings, 1904; Benzing face water in the present study (Table 3). Bromeliads that have and Renfrow, 1971; Benzing, 1980; Fig. 3). Also, droplets of low densities of attenuated stellate trichomes, such as Type 1 water will only spread on the abaxial leaf surface of pineapple Pitcairnia arcuata, appear to interact with water as described if detergent is added (demonstrated quantitatively in Table 4). by situation 2, loosely aggregating surface droplets. Consistent with the third, `lifting,' mechanism of repellency, continuous Pineapple leaves soaked overnight in a detergent solution or layers of powdery wax or ring-peltate trichomes produce an 100% acetone will regain their repellency if subsequently irregular hydrophobic surface that prevents water from coming rinsed and dried, suggesting a physical rather than chemical into contact with the epidermis proper. A summary of the prin- mechanism (personal observations). Additionally, if a pine- cipal interactions between leaf blade trichome layers and water apple leaf is partially dipped into a detergent solution rather within each ecophysiological type is presented in Table 5. than pure water, then liquid will be drawn or ``wicked'' up out In many families of ¯owering plants, water droplets bead of the solution along the trichome layer, i.e., once the surface up more readily on irregular than uniform surfaces because tension of the water is broken the leaf surface becomes strong- the droplet only contacts the tips of projections from the cu- ly hydrophilic. Thus, the physical properties of water are cen- ticle (Holloway, 1968; Juniper and Jeffree, 1983), obviating tral to the mechanism of repellency. This mechanism also August 2001] PIERCE ET AL.ÐWATER REPELLENT INDUMENTA OF BROMELIADS 1379 demonstrates, at least in part, how Type 4 species prevent wa- terrestrial lifestyle (i.e., the close proximity of vegetation and/ ter loss from the tank via capillary action. or the ground surface from which rainwater can splash up- Trichomes that characterize hydrophilic and hydrophobic wards onto the underside of the leaf) may help explain the surfaces usually share the same structure, with trichome den- evolution of hydrophobic trichome layers in Bromeliaceae. In- sity differing (e.g., the adaxial and abaxial surfaces of Cryp- deed, in the family as a whole, rosulate habits typical of genera tanthus whitmanii and hydrophilic Aechmea dactylina com- such as Fosterella and Cryptanthus tend to have hydrophobic pared with hydrophobic Ronnbergia explodens; Figs. 34±39). abaxial surfaces (Table 3). Also, terrestrial Orthophytum ben- The lower densities of peltate trichomes of Aechmea dactylina zingii has basal leaves close to the substrate that possess a and the adaxial surface of Cryptanthus whitmanii would allow repellent trichome layer on the abaxial surface, but on cauline water to come into contact with the epidermis proper, with the leaves this layer is less apparent (personal observation). interaction between the two presumably allowing water to spread and envelop trichomes. In addition, the adaxial tri- Trichome evolutionÐThe mechanism of water repellency chomes of Aechmea nudicaulis differ structurallyÐlacking the outlined above accords with the scheme of trichome structural irregular surface characteristic of the hydrophobic abaxial in- evolution detailed by Benzing (1980). In this scheme, the hy- dumentum (Figs. 40, 41). The chemical composition of hy- pothetical ancestral morphology is stellate (the simple ®la- drophilic and hydrophobic surfaces in Bromeliaceae has not mentous trichomes of some Navia species appear to be de- been investigated and the degree to which chemical vs. mor- rived; Benzing, 1980; Terry, Brown, and Olmstead, 1997). phological interactions contribute to repellency remains un- Low densities of stellate trichomes provide only discontinu- determined. Nevertheless, the physical characteristics of hy- ous, patchy repellency (e.g., extant Pitcairnia arcuata), in- drophobic trichome layers in Bromeliaceae are typical of wa- creased densities of which would maintain a greater proportion ter-repellent surfaces in other families, and the qualitative tests of the moistened leaf surface dry. Following this proposed above suggest that surface morphology is paramount to the early increase in trichome density, stellate trichomes may then operation of hydrophobia. have undergone an increase in the number of ring cells, be- coming truly peltate. This would increase the area covered by Ecophysiological consequences of a hydrophobic each trichome and thereby foster the ``lifting'' mechanism of indumentumÐIt may be signi®cant that the majority of bro- repellency (high densities of intermediate stellate/ring-peltate meliads are hypostomatous (Tomlinson, 1969; Benzing and trichomes occur in Pitcairnia corallina and P. integrifolia Burt, 1970), with stomata and hydrophobic trichome layers [Figs. 10, 31] and P. macrochlamys; Benzing, Seemann, and occurring together. Intriguingly, Barthlott and Neinhuis (1997) Renfrow, 1978). Additionally, the extrusive ring cells of such demonstrate that particulate matter will adhere more readily to peltate trichomes appear to lend the overall surface an ex- water droplets than to hydrophobic leaf surfaces, lending such tremely irregular small-scale texture (e.g., Figs. 29±31). leaves a ``self-cleaning'' capability when wetted. In concert Hydrophilic trichome layers among extant Bromelioideae with a possible function as a physical barrier to pathogens feature lower trichome densities, suggesting a decline in tri- (Benzing, 2000), this self-cleaning effect could remove path- chome density from ancestors with dense hydrophobic layers. ogens and prevent the physical blockage of stomata by partic- This perhaps re¯ects adaptive radiation into less crowded or ulates. A continuous trichome layer could also deter herbivores relatively xeric niches. Indeed, Type 1 Ronnbergia explodens from the softer underside of the leaf, although to date this has dense hydrophobic trichome layers and grows in the un- protective role is only evident in two species possessing glan- derstory of cloud forest habitats (Figs. 36, 37; Table 3). More dular trichomes (see Benzing, 2000). xeromorphic terrestrial species (CAM equipped and succulent) Benzing, Seemann, and Renfrow (1978) determined that such as Cryptanthus warasii and C. whitmanii may possess photosynthetic gas exchange was not inhibited by wetting the hydrophilic surfaces characterized by fewer trichomes (Fig. leaf blades of six species on the surfaces of which water did 38; Table 3; unpublished data), as do many Type 3 species not spread (including Pitcairnia macrochlamys). Conversely, (Aechmea dactylina, A. nudicaulis; Figs. 1, 2, 34, 40; Table the wetted trichomes of Type 5 bromeliads hold ®lms of water 3). that slow the exchange of gases between the air and the leaf Dense trichome layers in Tillandsioideae are usually hydro- (Benzing, Seemann, and Renfrow, 1978; Schmitt, Martin, and philic, unlike those of Bromelioideae and Pitcairnioideae. In- LuÈttge, 1989). Clearly, most Type 5 bromeliads must reconcile deed, Billings (1904) points out that one of the most unusual both gas exchange and water acquisition through the same features of Tillandsia usneoides is that ``unlike most similar surface, relying on temporal separation of these two processes appendages of the epidermis, the scales do not hinder the leaf by performing gas exchange when the leaf is dry. In contrast, from becoming wet.'' Dense hydrophilic trichome layers in Type 1 and Type 2 bromeliads separate the processes of gas Tillandsioideae must possess a difference that can account for exchange and water acquisition spatially between roots and their lack of water repellency. At present, differences in the leaves and tank-forming species between the leaf sheath and chemical composition of these surfaces cannot be ruled out. blade. Thus, these latter life forms do not need to compromise However, a striking structural difference between the tri- carbon gain to acquire water. In this respect, wettable tri- chomes of Tillandsioideae and those of the other subfamilies chomes on the leaf blade would not only be an unnecessary is apparent, which could also explain the different interaction investment but would be disadvantageous in mesic habitats, with water. From scanning electron micrographs published in whereas repellent trichomes would favor gas exchange, as per- other sources (Benzing, Seemann, and Renfrow, 1978; Benz- haps demonstrated by Pitcairnia macrochlamys (Benzing, See- ing, 1980; Adams and Martin, 1986), it is possible to see that mann, and Renfrow, 1978). the parts of adjacent tillandsioid trichomes that overlap one Sources of water that may moisten the underside of the leaf another are the ¯exible wings, which overlap when ¯attened may include dew and, perhaps more importantly in cloud for- (wet). Thus, when the leaf is dry and the wings are ¯exed ests, wind-borne mist. These factors in conjunction with the upwards, underlying epidermis cells are exposed (Benzing, 1380 AMERICAN JOURNAL OF BOTANY [Vol. 88 Jamaica, Argentina, ϭ ϭ L droplet of aqueous ␮ test. The photosynthetic Honduras, JA t ing UV) compared against nsi hydrophobic (wax) hydrophobic (wax) hydrophobic (wax) nsi hydrophobic (trichome) hydrophilic hydrophobic (trichome) nsi nsi nsi hydrophobic (trichome) nsi hydrophobic (trichome) nsi hydrophobic (trichome) nsi nsi hydrophilic hydrophobic (trichome) hydrophobic (trichome) nsi nsi nsi nsi hydrophobic (trichome) nsi nsi nsi hydrophobic (trichome) ϭ a 10- ct (surface structures present) owing countries: AR 74.5 b40.9 a hydrophilic 18.7 a nsi 32.2 a 31.9 a19.4 a hydrophobic56.9 (wax) a hydrophobic34.0 (wax) a nsi 90.2 a nsi 58.5 a 87.4 a 34.3 a 54.4 b 69.1 a 39.0 a 65.4 a 64.7 a 47.9 a41.0 a nsi 70.2 a hydrophobic48.7 (trichome) a 81.1 a 42.4 a 71.4 a 39.9 a 70.2 a 40.3 b 88.5 a 60.2 a 76.6 a 46.7 a 68.1 a 27.1 a 36.1 a25.8 a nsi 18.9 a hydrophobic28.4 (trichome) a 13.5 a20.9 a nsi hydrophobic (trichome) 110.2 a 109.8 a 103.6 a Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Guyana, HO m) ϭ ␮ 663.6 625.2 699.9 809.1 388.2 777.5 628.1 897.9 623.4 911.8 875.5 581.4 543.4 544.8 733.8 478.1 512.2 351.9 489.3 678.0 498.3 360.1 662.5 409.2 484.6 504.7 714.3 274.8 582.8 510.3 34.6 a58.9 a45.6 531.2 a 58.7 635.2 b 18.5 b64.3 b33.9 672.8 a26.8 708.5 a24.3 514.8 b 25.9 492.9 b 49.7 a 47.9 b 22.5 b 22.2 a 57.2 a 45.9 a 25.3 b 65.7 a14.9 b50.6 456.3 a 12.9 665.1 b 96.9 a 68.4 b 53.7 a 59.8 b 44.1 a 68.6 a 69.7 a 34.7 b 42.6 b 45.4 a 14.6 a 46.2 a 59.6 a 16.7 b 44.7 a8.6 b24.3 605.0 a 40.0 a 777.0 25.2 a17.8 b 306.9 482.8 102.2 a Depth of droplet ( Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Guatemala, GY ϭ , if not previously disclosed by Martin (²; 1994) and references ϩ 32.5 1 SE of four replicates (* denotes species of which six replicates Surface intact Surface denuded Surface type 0.05 level as determined by Student's 446.4 982.2 454.5 250.6 728.2 232.8 211.8 628.1 513.9 631.8 506.1 192.0 935.1 940.6 947.8 379.8 899.9 858.6 523.3 832.5 484.8 899.9 429.5 451.9 H 1030.3 1417.9 1171.7 1226.5 1318.9 Ϯ Յ ⌬ P Ecuador, GT ϭ abaxial abaxial abaxial abaxial abaxial abaxial abaxial abaxial abaxial adaxial abaxial abaxial 706.4 abaxialabaxial 895.9 460.0 abaxial abaxial abaxial 952.2 abaxial abaxial abaxial 983.4 abaxial 718.9 abaxial ²³³ adaxial² adaxial 284.8 ² adaxial adaxial 964.4 adaxial 476.4 ³³³ adaxial ³ adaxial adaxial ³ 512.8 adaxial adaxial ³³³ adaxial ³ adaxial ² adaxial adaxial 536.2 adaxial 258.6 Dominica, EC 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 Venezuela. Life forms or ecophysiological types follow the classi®cation of Benzing (2000). ϭ ϭ 1C 4C 1C 1C 1C 11 CAM³1C CAM³ adaxial adaxial 1C 11 Ð CAM³ adaxial adaxial 1C 1C 1C Cuba, DO ϭ Trinidad, VE ϭ Costa Rica, CU Â, Cerro Jefe, el®n ϭ Paraguay, TR ϭ . 1 CAM³ adaxial .1C sin loc sin loc. sin loc sin loc. sin loc. Peru, PG gas (880 m a.s.l.). m a.s.l.). cloud forest (1007 m a.s.l.). m a.s.l.). (450 m a.s.l.). Puenta de Coripata (1200 m a.s.l.). Consata (1200 m a.s.l.). m a.s.l.). ana. gas (800 m a.s.l.). m a.s.l.). Colombia, CR VE, La Escalera (1000 m a.s.l.). 4 C AR, BO, Dpto. La Paz, Puente Villa, BO, BR, Est. Minas Gerais, Diamanti- BO, Dpto. La Paz, Prov. Nor Yun- VE, ϭ ϭ or crassulacean acid metabolism) employed by each species was determined by  VE, Edo. Tachira (1200±1500 3  PA, Prov. Panama L.B. Panama, PE Mez VE, Kavaneyen. 4 C Brazil, CO L.B. Smith VE, , Sierra de Lema. 4 C A. Castel- (Baker) Â) Andre G.S. Varada- ϭ ϭ Mez HO, Teguicigalpa-Comayagua (1300 (Baker) Mez GY, Kaiteur Falls. 4 C Ker-Gawler* TR, Point Gourde, seasonally dry. 1 C (Beer) Baker CR, Prov. Cartago, La Suiza (1000 Rauh BO, (Brongniart) Linden & Andre (Mez) L.B. Baker VE, Hacienda Santa Elena (300 Hooker PE, (Grisebach) L.B. H. Luther BO, Dpto. La Paz, Prov. Nor Yun- Baker PG, Dpto. Paraguari, Cerro Acahay (Andre Species Origin of material Life form Carbon pathway Surface acuminata hechtioides elata ITCAIRNIOIDEAE Mexico, PA cf. cf. P Bolivia, BR cf. sp. nov. BO, Dpto. La Paz, Prov. Munecas, ϭ ϭ 3. Leaf blade surface±water interactions of species of the family Bromeliaceae, divided by subfamily. Adaxial and abaxial surfaces were either inta were used). Different letters (a±b) represent signi®cant differences between intact and denuded means at the carbon assimilation pathway (C BO ¯uorochrome after a periodstandards of of 40 measured min droplet compared depth between (¯uorochrome intact on and paraplast denuded wax surfaces. and Depth glass values surfaces). are Values derived represent from means ¯uorochrome luminosity (under excit therein, or determined via carbon isotope discrimination by Crayn, Winter, and Smith (³; unpublished data). Plant material originated from the foll ME or denuded (surface structures removed). Surface typeÐhydrophilic, hydrophobic, or not signi®cantly interactive (nsi)Ðis denoted by the depth of rajan lanos Smith L.B. Smith Smith Smith Regel ABLE UBFAMILY T S Brocchinia Brocchinia gilmartiniae Brocchinia Brocchinia reducta Deuterocohnia schreiteri Dyckia marnier-lapostollei Dyckia microcalyx Fosterella albicans Fosterella Fosterella petiolata Fosterella schidosperma Fosterella Hechtia guatemalensis Pitcairnia arcuata Pitcairnia atrorubens Pitcairnia corallina Pitcairnia echinata Pitcairnia imbricata Pitcairnia integrifolia Fosterella caulescens August 2001] PIERCE ET AL.ÐWATER REPELLENT INDUMENTA OF BROMELIADS 1381 nsi nsi nsi nsi nsi hydrophobic (trichome) nsi nsi nsi nsi nsi nsi hydrophobic (wax) hydrophobic (wax) hydrophobic (wax) hydrophobic (wax) nsi nsi hydrophobic (wax) nsi hydrophobic (wax) nsi nsi hydrophobic (wax) nsi nsi nsi nsi nsi hydrophobic (wax) nsi nsi nsi nsi 61.6 a 51.8 a 66.6 a18.6 a nsi 34.0 a nsi 39.8 a 42.0 a25.5 a nsi 34.0 a nsi 28.3 a 53.6 a40.8 a nsi 56.7 a nsi 62.7 a 23.2 b24.4 a hydrophilic 89.6 a hydrophobic38.0 (trichome) a nsi 119 a hydrophobic86.7 (trichome) a 40.3 a 36.9 a 35.2 b38.3 b hydrophilic 15.5 a hydrophilic 29.1 a nsi nsi 52.9 a 52.7 a 7.2 a 71.7 a 35.5 a 13.8 a 19.9 a 41.3 a 19.9 a 54.7 a 70.6 a 29.9 a 92.6 a 88.8 a 28.8 a 48.2 a 18.2 a 45.7 a 68.1 a 75.5 a 15.8 a 107.8 a Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ m) ␮ 591.2 791.8 865.1 711.2 758.4 446.7 747.7 607.9 465.5 498.3 684.7 915.8 753.6 521.5 991.8 618.9 521.3 553.8 547.6 546.0 618.5 316.1 638.9 673.7 569.5 690.0 503.2 822.8 743.9 353.9 814.4 702.9 1072.5 1009.3 12.7 a 55.6 a 74.5 a30.6 a44.6 441.6 a 52.2 648.0 a 33.4 a21.1 a64.1 363.3 a 38.3 510.3 b 58.0 a34.8 a22.3 495.4 a 68.1 520.0 a 12.6 a39.1 b68.4 392.7 a9.5 679.4 b70.6 484.6 a 24.3 a 714.9 27.3 a 83.1 a 8.6 a70.7 a105.9 a 232.2 82.4 347.3 a 489.3 35.8 624.6 b 4.4 b 149.2 b 32.8 b 15.8 a 19.3 a 48.2 b 47.7 a 30.2 b 8.9 a 53.9 a 7.5 b 70.9 a 44.4 a 16.7 a 29.9 a 9.4 a 11.2 b 86.6 a 119.5 a 22.7 a 10.3 a Depth of droplet ( Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Surface intact Surface denuded Surface type 444.2 497.3 476.4 522.4 647.4 926.1 854.2 958.2 422.9 342.7 662.3 597.0 618.9 848.2 671.6 495.6 794.8 658.1 555.8 454.5 640.8 427.2 547.6 824.5 831.3 686.0 737.6 890.4 604.3 240.0 1870.5 1286.7 1073.9 1971.5 abaxial abaxial abaxial abaxial abaxial abaxial abaxial abaxial abaxial abaxial abaxial abaxial adaxial abaxial adaxial abaxial adaxial abaxial adaxial abaxial abaxial 630.8 abaxial 469.9 abaxial 483.1 abaxialabaxial 844.7 892.3 abaxialabaxial 70.7 538.5 abaxial ³³³ adaxial (CAM)³ adaxial adaxial³ adaxial 295.4 ³ 363.3 ³ adaxial ³ adaxial³ adaxial 464.7 ³ adaxial adaxial 315.5 adaxial 382.7 (CAM)³ adaxial (CAM)² adaxial ²³ adaxial adaxial ³-CAM² adaxial adaxial ² adaxial 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 1C 1 CAM³ adaxial 14.7 1C 1C 1C 1C 1C 1 CAM³ adaxial 4C 4C 4C 4C 4C 4C 4C 4C 4C 4C Â, Cerro Jefe, el®n Â, Cerro Jefe, el®n Â, Cerro Jefe, el®n Â, Cerro Jefe, el®n Â, Cerro Jefe, el®n .4C  province, Mata Ahogado, sin loc Prov. Chiriqui, Fortuna, lower rimaguas (1000 m a.s.l.). cloud forest (1007 m a.s.l.). do. montane wet forest (1200 m a.s.l.). (100 m a.s.l.). City. m a.s.l.). (1000 m a.s.l.). cloud forest (1007 m a.s.l.). Cerro Iminapi, Sorata (2680 m a.s.l.). cloud forest (1007 m a.s.l.). Tipuani-Caranavi (1250 m a.s.l.). cloud forest (1007 m a.s.l.). cloud forest (1007 m a.s.l.). montane wet forest (1000 m a.s.l.). montane wet forest (1200 m a.s.l.). BR, PE, Dpto. San Martin, Tarapoto-Yu- ME, Edo. Veracruz, Playa Escondi- Sin loc. PA, Prov. Panama Sin loc. BO, Dpto. La Paz, Prov. Larecaja, PA, Cocle PA, Prov. Chiriqui, Fortuna, lower * Rauh PE, Dpto. San Martin, Tarapoto R.W. Read DO (1250 m a.s.l.). 1 C (Grisebach) (L.) Rusby (Leme) J.R. L.B. Smith L.B. Smith PA, Prov. Panama L.B. Smith EC, Prov. Manabi. 1 C Rauh PA, Prov. Panama (C. Morren) Mez & Sodiro EC, Prov. Carchi, Chical (1300 (Scheidweiler) (Ruiz & Pavon) Scheidweiler ME, Edo. Chiapas. 1 C L.B. Smith PA, Prov. Panama L.B. Smith BO, Dpto. La Paz, Prov. Larecaja, (Linden & An- L.B. Smith GT, bought in market in Guatemala Mez EC, Morona-Santiago. 1 C Standley PA, Prov. Panama (Swartz) Grisebach HO, Prov. Cortes, San Pedro, Sula musaica Species Origin of material Life form Carbon pathway Surface (Hooker) Grisebach PA, monostachia odorata ILLANDSIOIDEAE Kunth cf. T L.B. Smith Old hort. plant. 1 CAM³ adaxial 563.1 3. Continued. Â) Mez var. Grant ex Mez. var. Decaisne K. Koch Mez Mez dre ABLE UBFAMILY T Pitcairnia maidifolia Alcantarea Pitcairnia microtrinensis Pitcairnia palmoides Pitcairnia patenti¯ora Pitcairnia recurvata Pitcairnia riparia Pitcairnia rubronigri¯ora Pitcairnia undulata Pitcairnia unilateralis Pitcairnia valerii Puya ctenorhyncha Puya lanata Puya laxa S Catopsis micrantha Catopsis nitida Catopsis nutans Catopsis sessili¯ora Catopsis subulata Guzmania circinnata Guzmania coriostachya Guzmania macropoda Guzmania monostachia Guzmania musaica 1382 AMERICAN JOURNAL OF BOTANY [Vol. 88 nsi nsi nsi hydrophobic (wax) nsi nsi hydrophilic hydrophilic hydrophilic hydrophilic hydrophilic hydrophilic hydrophilic hydrophilic hydrophilic hydrophilic nsi hydrophobic (trichome) nsi hydrophobic (wax) nsi nsi nsi nsi nsi nsi nsi nsi nsi nsi hydrophilic hydrophilic hydrophilic hydrophilic hydrophilic nsi hydrophilic hydrophobic (trichome) nsi hydrophobic (trichome) nsi hydrophobic (trichome) nsi hydrophobic (trichome) 63.3 a 105.9 a 45.7 a 86.9 a 57.1 a 43.1 a 147.6 b 48.9 b 40.4 b 37.6 b 10.8 b 14.2 b 36.4 b 40.6 b 49.7 b 57.7 b 105.3 a 54.9 a 46.7 a 65.9 a 103.9 a 58.6 a 96.4 a 21.9 a 49.2 a 53.8 a 54.1 a 34.0 a 52.3 a 36.4 a 18.7 b 22.2 b 38.1 a20.9 a nsi 20.4 b hydrophobic37.9 (trichome) b 54.3 b 46.1 a 64.5 b 46.9 a 43.1 a 24.5 a 11.0 a 12.4 a 22.9 a 116.1 a Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ m) ␮ 519.0 350.4 983.5 659.1 958.5 593.8 885.4 579.4 714.8 562.0 540.1 875.0 641.0 892.5 647.2 601.3 606.7 514.3 853.1 685.3 341.8 741.8 308.4 993.8 414.5 641.4 732.1 321.3 771.0 804.3 528.3 869.5 825.3 966.0 918.8 709.3 477.3 523.5 478.1 671.4 667.7 498.8 1038.0 1028.0 44.2 a 36.1 a 72.7 a 21.0 b 92.2 a 18.8 a 5.9 a 165.5 a 22.4 a 31.3 a 79.2 a 111.3 a 0.0 a 120.9 a 0.0 a 54.0 a 44.6 a 17.1 b 46.7 a 72.0 b 87.5 a 27.9 a 51.7 a 68.9 a 52.8 a 83.9 a 45.7 a 28.6 a 59.5 a 94.9 a 56.7 a 14.5 a 24.5 a41.7 b38.7 602.3 a 45.0 587.2 a 86.4 a 61.6 a 10.5 a 77.3 b 47.7 a 11.9 b 36.5 a 13.2 b 108.2 a 16.6 b Depth of droplet ( Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ 0.0 0.0 9.4 5.7 25.1 Surface intact Surface denuded Surface type 490.2 476.4 268.7 605.6 663.3 842.8 833.5 831.3 885.5 786.8 911.8 859.0 817.5 211.1 619.5 438.3 551.8 955.2 901.8 199.2 768.3 334.1 938.3 915.5 451.6 564.5 810.0 515.5 489.4 591.0 740.5 842.2 349.0 341.2 310.5 1039.5 1020.8 1050.3 1130.3 abaxial abaxial abaxial abaxial adaxial abaxial adaxial abaxial adaxial abaxial adaxial abaxial adaxial abaxial adaxial abaxial adaxial abaxial adaxial abaxial abaxial abaxial abaxial abaxial 795.9 abaxial abaxial abaxial abaxial abaxial abaxial abaxial ³ adaxial ³ adaxial ³ adaxial 3 3 3 3 3 3 3 3 3 3 3 3 CAM adaxial 4C 4C 4C 4C 4C 3 CAM³ adaxial 617.5 2±3 CAM adaxial 4C 4C 4 CAM²5 adaxial CAM³4C adaxial 4C 31C CAM² adaxial 4±5 CAM³4±5 adaxial CAM² adaxial 32 CAM² CAM² adaxial adaxial Â, Cerro Jefe, el®n Â, Cerro Jefe, el®n Â, Cerro Jefe, el®n Â, Cerro Jefe, el®n Â, Gamboa, low- Â, Cerro Jefe, el®n Â, Gamboa, low- Â, Cerro Jefe, el®n Â, Gamboa, low- Â, Cerro Jefe, el®n Â, Cerro Jefe, el®n Â, Panama City, Â, Barro Colorado . 5 CAM² adaxial sin loc. sin loc cloud forest (1007 m a.s.l.). cloud forest (1007 m a.s.l.). cloud forest (1007 m a.s.l.). cloud forest (1007 m a.s.l.). land seasonally dry forest, cloud forest (1007 m a.s.l.). land seasonally dry forest. rimaguas (1200 m a.s.l.). seasonally dry, saxicolous on rock-faces (2700 m a.s.l). Grande (3 m a.s.l.). montane wet forest (1200 m a.s.l.). cloud forest (1007 m a.s.l.). land seasonally dry forest. cloud forest (1007 m a.s.l.). cloud forest (1007 m a.s.l.). forest (710 m a.s.l.) Cerro Ancon, lowland urban,sonally sea- dry. Island, shaded understory. saxicolous (2700 m a.s.l.). PA, Prov. Panama TR, PA, Prov. Panama ME, Edo. Chiapas (1000 m a.s.l.).PA, Prov. Panama 4PA, Prov. Panama C PA, Prov. Panama PA, Prov. Panama PA, Prov. Panama PA, Prov. Panama Agricultural clone. 2 CAM adaxial PA, Prov. Panama  Â) Â) sub- Â) J.R. Â) Andre H. Luther (Linden ex (Andre (E. Gross & (Wendland)  ex Mez TR, Textel, transitional montane (Andre L.B. Smith PA, Prov. Panama Mez PE, Dpto. Qosqo, Ollantaytambo, (L.) Grisebach PA, Prov. Bocas del Toro, Chiriqui (Grisebach) Kunth var. (Linden & An- Baker* PA, Prov. Panama Swartz* PA, Prov. Panama (Mez & Werckle (Mez) L.B. Smith PA, Prov. Panama Solander var. (Lindley) Baker BR, Andre L.B. Smith PE, Dpto. San Martin, Tarapoto-Yu- Baker* PA, Prov. Chiriqui, Fortuna, lower (Mez & Werckle (L.) Merril cv. Baker PE, Dpto. Qosqo, Ollantaytambo, Species Origin of material Life form Carbon pathway Surface ROMELIOIDEAE (Baker) L.B. Smith var. discolor B 3. Continued. Â) Mez dre stricta J.R. Grant Grant J.R. Grant Cogniaux & Marchal) J.R. Grant J.R. Grant M.A. Spencer & L.B.spiculosa Smith var. imbricata Rauh) J.R. Grant Cayenne Lisse* ex Baker* ABLE UBFAMILY T Guzmania musaica Aechmea dactylina Guzmania retusa Racinaea spiculosa Tillandsia cauligera Tillandsia elongata Tillandsia ¯exuosa Tillandsia nana Tillandisia stricta Vriesea monstrum Werauhia capitata Werauhia gladioli¯ora Werauhia hygrometrica Werauhia panamaensis Werauhia sanguinolenta Werauhia vittata S Aechmea fasciata Aechmea fendleri Aechmea magdalenae Aechmea nudicaulis Aechmea veitchii Ananas comosus Billbergia macrolepis August 2001] PIERCE ET AL.ÐWATER REPELLENT INDUMENTA OF BROMELIADS 1383 hydrophilic hydrophobic (trichome) nsi nsi hydrophilic nsi hydrophilic hydrophobic (trichome) nsi nsi hydrophilic hydrophobic (trichome) hydrophilic hydrophobic (trichome) hydrophilic nsi nsi nsi hydrophilic hydrophobic (trichome) hydrophilic hydrophilic hydrophilic hydrophilic nsi nsi nsi hydrophobic (trichome) 41.0 a 51.7 b 114.5 a 13.4 b17.8 a hydrophilic 125.4 a nsi 52.5 a nsi 22.5 a hydrophobic53.0 (trichome) a 51.2 b29.8 b hydrophilic 49.3 b hydrophilic 35.1 a 18.9 b 57.4 a 51.9 a 30.9 a 65.8 b 11.7 a 44.5 b 49.9 a 83.6 b 49.6 a 47.7 a 37.9 a 38.1 b 76.5 a 27.0 b38.5 b hydrophilic 94.2 b hydrophilic 87.8 b 10.3 b 9.1 b 48.9 a 37.1 a 36.7 a Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ m) ␮ 589.5 795.2 761.1 343.4 817.5 781.3 375.2 882.3 582.2 606.1 265.3 626.5 648.3 566.2 681.2 788.5 388.8 134.7 378.5 631.9 382.7 770.2 543.9 477.0 286.8 523.7 558.6 420.95 116.6 a 43.9 b 74.9 a28.9 a223.7 541.8 a31.6 611.4 b 358.5 43.6 a 60.2 479.2 a 62.9 a54.7 a0.0 426.9 a 58.6 535.8 a 15.5 a 33.3 b 50.3 a 47.6 a 0.0 a 37.3 b 61.2 a 17.9 b 99.4 a 41.3 a 44.9 a 36.8 a 0.0 a 97.6 b 47.7 a111.9 a0.0 286.6 a 438.9 0.0 a 39.9 a 47.5 a 76.5 a 42.9 a 51.4 a 13.6 b Depth of droplet ( Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ Ϯ 0.0 0.0 0.0 0.0 0.0 18.6 Surface intact Surface denuded Surface type 503.4 360.4 275.7 330.1 256.6 761.3 646.1 513.0 763.2 998.5 782.9 994.4 608.4 420.8 727.6 666.8 725.8 177.6 363.1 562.0 1009.3 1079.7 abaxial abaxial abaxial adaxial abaxial abaxialabaxial 578.1 786.3 abaxialabaxial 75.9 abaxialabaxial 138.9 abaxial abaxial abaxial abaxial abaxial abaxial abaxial abaxial ³ adaxial (CAM)³ adaxial 3 3 3 CAM² adaxial 1 CAM³ adaxial 1 CAM³ adaxial 47.7 3 CAM³ adaxial 2 CAM² adaxial 11C CAM³11 adaxial CAM³3 Ð3 adaxial CAM³1C CAM² adaxial adaxial adaxial 1C Â, Cerro Jefe, el®n Â, Cerro Azul, sin loc. (Distribution: VE±TR). 3 CAM² adaxial 233.3 . 3 CAM³ adaxial 169.8 Ân, near Quincemil, epi- ex hort sin loc. sin loc. Tijuca, dense forest onclay hillside, and leaf-litter substratem (30 a.s.l.). Salvacio phytic in forest. cloud forest (1007 m a.s.l.). level. partial shade (450 m a.s.l.). tropical wet forest (691 m a.s.l.). Kennedy, Praia de Maroba. (1000±1200 m a.s.l.). mantina. Martins. BR, Est. Rio de Janeiro, Barra de BR, Est. Rio de Janeiro, Rio Bonito. 3 PE, Dpto. Madre de Dios, Hacienda West Indies, BR, Est. Rio de Janeiro, near sea BR, Est. Minas Gerais, lithophyte, BR, Est. Rio de Janeiro. 3 CAM³ adaxial BR, Est. Bahia. 1 CAM³ adaxial Otto & (A. Rich- L.B. E. Gross & Weber BR, Leme & H. Leme BR, Est. Espirito Santo, Domingos L.B. Smith PA, Prov. Panama Harms EC, Prov. Napo, Tulag. 3 CAM³ adaxial 386.1 Hutchison BR, (Lemaire) Mez BR, Est. Minas Gerais, Caraca E. Pereira BR, Edo. Minas Gerais, vic. Dia- Leme BR, Est. Espirito Santo, Presidente (R. Graham) (Schott ex Beer) L.* PA, Prov. Panama Hortus ex Beer VE, bromelioides Species Origin of material Life form Carbon pathway Surface cf. 3. Continued. Dietrich R.W. Read cv. Tim Plowman Rauh ard) Mez L.B. Smith* Luther Mez Smith ABLE T Billbergia robert-readii Billbergia rosea Billbergia stenopetala Bromelia pinguin Canistrum seidelianum Cryptanthus Cryptanthus dianae Cryptanthus glaziovii Cryptanthus warasii Cryptanthus whitmanii Hohenbergia pendula¯ora Neoregelia cruenta Orthophytum benzingii Orthophytum gurkenii Orthophytum magalhaesii Quesnelia blanda Quesnelia marmorata Ronnbergia explodens 1384 AMERICAN JOURNAL OF BOTANY [Vol. 88

Figs. 18±25. Epicuticular wax powder layers of leaf blade surfaces of bromeliads. 18. Catopsis micrantha, photograph of water droplets on adaxial surface of leaf blade. 19. Catopsis micrantha, photograph of epicuticular wax powder layer on abaxial surface of leaf sheath. 20. Catopsis micrantha, scanning electron micrograph (SEM) of trichome embedded in wax layer (unprepared specimen). 21. Catopsis micrantha, SEM of trichomes on the wax-free adaxial leaf sheath surface (prepared specimen). 22. Werauhia capitata, SEM of trichome on abaxial surface. 23. Werauhia capitata, SEM of abaxial surface. 24. Alcantarea odorata, SEM of adaxial surface, 25. Brocchinia reducta, SEM of adaxial surface. August 2001] PIERCE ET AL.ÐWATER REPELLENT INDUMENTA OF BROMELIADS 1385

Figs. 26±31. Scanning electron micrographs of bromeliad leaf blade surfaces, the adaxial surfaces of which do not interact with water, the abaxial surfaces hydrophobic. 26±28. Noninteractive adaxial surfaces of Vriesea monstrum, Fosterella petiolata and Pitcairnia corallina, respectively, lacking trichomes or with ®lmy trichomes. 29±31. Hydrophobic abaxial surfaces of Vriesea monstrum, Fosterella petiolata, and Pitcairnia corallina, respectively, with well-de®ned trichomes in a con¯uent layer. 1386 AMERICAN JOURNAL OF BOTANY [Vol. 88

Figs. 32±37. Scanning electron micrographs of trichomes from bromeliad leaf blade surfaces, the indumenta of which have different interactions with water. 32. Pitcairnia arcuata, attenuated stellate trichome with radial ®laments tangled together, low densities of which form a hydrophobic indumentum. 33. Puya laxa has two types of trichome, one peltate and the other with a grossly elongate wing that spirals around itself to form a hair-like structure, the indumentum having no interaction with water. 34. Aechmea dactylina, peltate trichome in a hydrophilic indumentum. 35. Aechmea dactylina, detail of trichome shield. 36. Ronnbergia explodens, peltate trichome in a hydrophobic indumentum. 37. Ronnbergia explodens, detail of trichome shield. August 2001] PIERCE ET AL.ÐWATER REPELLENT INDUMENTA OF BROMELIADS 1387

Figs. 38±41. Scanning electron micrographs of trichomes on hydrophilic and hydrophobic surfaces of the same leaf blade. 38±39. Cryptanthus whitmanii, hydrophilic adaxial and hydrophobic abaxial surfaces, respectively. 40±41. Aechmea nudicaulis, hydrophilic adaxial and hydrophobic abaxial surfaces, respec- tively.

Seemann, and Renfrow, 1978). When the leaf is wetted, sur- face tension forces acting on the epidermis and/or the under- side of the trichome wing may permit water to spread. Thus, dense trichome layers in most Tillandsioideae have different TABLE 4. The effect of removal of water surface tension on the leaf con®gurations when wet and dry and will only form a con¯u- blade trichome-layer±water interactions of Ananas comosus. Re- ent layer after wetting. The moveable trichome wing of the pellency was denoted by the depth of a 10-␮L droplet of aqueous Type 5 life form may therefore be regarded as a device allow- ¯uorochrome after a period of 40 min. The ¯uorochrome used was ing the presence of high densities of trichomes while avoiding either ¯uorescein sodium solution (5 mL of 0.05% ¯uorescein ϩ repellency. 0.5 mL H2O) or a solution of ¯uorescein and household detergent (5 mL of 0.05% ¯uorescein ϩ 0.5 mL neat detergent). Depth values Indeed, dense layers of peltate trichomes that lack wings in are derived from ¯uorochrome luminosity (under exciting UV Tillandsioideae are hydrophobic (e.g., Vriesea monstrum; Fig. light) compared against standards of measured droplet depth (¯u- 29; Table 3). Also, the immobile trichomes of Type 3 bro- orochrome on paraplast wax and glass surfaces). Values represent meliads demonstrate that a moveable wing is not essential for means Ϯ 1 SE of six replicates. Different letters (a±c) represent absorption (Benzing, Givnish, and Bermudes, 1985). The signi®cant differences between means at the P Յ 0.05 level as determined by Tukey's multiple comparison procedure (ANOVA). moveable wing is generally associated with higher trichome densities and effective water and nutrient absorption by the Depth of aqueous droplet (␮m) leaf surface (Benzing and Burt, 1970). Leaf blade surface Fluorochrome Fluorochrome ϩ detergent Epicuticular wax powdersÐBenzing, Givnish, and Ber- Adaxial 559.3 Ϯ 81.6 b 14.8 Ϯ 3.2 a mudes (1985) suggest that Tillandsioideae and BrocchiniaÐ Abaxial 1013.1 Ϯ 41.7 c 17.6 Ϯ 5.3 a both of which include advanced Type 4 tank forms equipped 1388 AMERICAN JOURNAL OF BOTANY [Vol. 88

TABLE 5. Summary of principal leaf blade interactions with water (as determined by ¯uorographic dimensional imaging) of the different eco- physiological types of Bromeliaceae. ``Ring-peltate'' trichomes possess a shield composed mainly of ring cells, and ``wing-peltate'' trichomes possess a shield with a moveable wing. Life forms or ecophysiological types follow the classi®cation of Benzing (2000).

Life form Trichome type Trichome cover Interaction with water Example 1 stellate discontinuous hydrophobic Pitcairnia arcuata stellate/ring continuous hydrophobic Fosterella petiolata, peltate Ronnbergia explodens ring-peltate discontinuous hydrophilic Cryptanthus whitmanii 2 ring-peltate continuous hydrophobic Ananas comosus ring-peltate discontinuous hydrophilic Aechmea magdalenae 3 ring-peltate continuous hydrophobic Aechmea nudicaulis ring-peltate discontinuous hydrophilic Aechmea dactylina 4 ring-peltate continuous hydrophobic Vriesea monstrum wing-peltate continuous hydrophilic Tillandsia elongata wing-peltate discontinuous noninteractive Werauhia sanguinolenta 5 wing-peltate continuous hydrophilic Tillandsia nana with absorbent trichomesÐare derived from a common an- struct pathogens and particulates, aid in self-cleaning, and/or cestor. Indeed, in the present study only Tillandsioideae and maintain gas exchange during wet weather. Brocchinia provided examples of species in which epicuticular wax powders are produced. Waxy Catopsis species have been LITERATURE CITED shown to use wing-peltate trichomes to take up mineral ions and amino acids (Benzing et al., 1976; Benzing, 1980; Benz- ing and Pridgeon, 1983), and this probably also applies to C. ADAMS, W. W., III, AND C. E. MARTIN. 1986. Morphological changes ac- companying the transition from juvenile (atmospheric) to adult (tank) micrantha. Both leaf surfaces bear a powdery layer of epicu- forms in the Mexican epiphyte Tillandsia deppeana (Bromeliaceae). ticular wax, and this is also one of the few taxa reported to American Journal of Botany 73: 1207±1214. be amphistomatous (see Tomlinson, 1969; Figs. 13, 14). Thus, BARTHLOTT,W.,AND C. NEINHUIS. 1997. Purity of sacred lotus, or escape extensive epicuticular wax powders appear to have evolved from contamination in biological surfaces. Planta 202: 1±8. only in taxa containing Type 4 life forms, which use trichomes BELL, A. D. 1991. Plant form: an illustrated guide to ¯owering plant mor- to acquire water and minerals from tanks. phology. Oxford University Press, Oxford, UK. BENZING, D. H. 1970. Foliar permeability and the adsorption of minerals and It is likely that in many Type 4 species a combination of organic nitrogen by certain tank bromeliads. Botanical Gazette 131: 23± the horizontal orientation of the leaf and the hypostomatous 31. condition are suf®cient to keep stomata unobstructed by water; ÐÐÐ. 1976. Bromeliad trichomes: structure, function, and ecological sig- in the present study, predominantly those species that pos- ni®cance. Selbyana 1: 330±348. sessed upright leaves (e.g., Brocchinia reducta, Guzmania ma- ÐÐÐ. 1980. The biology of Bromeliads. Mad River Press, Eureka, Cali- cropoda, Werauhia capitata), and/or stomata on the adaxial fornia, USA. ÐÐÐ. 2000. Bromeliaceae: pro®le of an adaptive radiation. Cambridge surface (Catopsis micrantha) possessed hydrophobic wax University Press, Cambridge, UK. powders on the leaf blade. Possibly the upright funnelform ÐÐÐ, AND K. M. BURT. 1970. Foliar permeability among twenty species habit increases the utility of the tank as an impoundment, and of the Bromeliaceae. Bulletin of the Torrey Botanical Club 97: 269±279. tank formers face a trade-off between gas exchange and im- ÐÐÐ, T. J. GIVNISH, AND D. BERMUDES. 1985. Absorptive trichomes in poundment capacity, wax powders being a method of maxi- Brocchinia reducta (Bromeliaceae) and their evolutionary and systematic mizing both. Re¯ective epicuticular wax powders have also signi®cance. Systematic Botany 10: 81±91. ÐÐÐ, K. HENDERSON,B.KESSEL, AND J. SULAK. 1976. The absorptive been implicated in the attraction and entrapment of insects in capacities of bromeliad trichomes. American Journal of Botany 63: a small number of Type 4 bromeliadsÐCatopsis berteroniana, 1009±1014. , and B. reducta (Fish, 1976; Frank and ÐÐÐ, AND A. M. PRIDGEON. 1983. Foliar trichomes of Pleurothallidinae O'Meara, 1984; Givnish et al., 1984; Owen, Benzing, and (Orchidaceae): functional signi®cance. American Journal of Botany 70: Thomson, 1988; Owen and Thomson, 1991; Benzing, 2000). 173±180. It is possible that a slippery and re¯ective epicuticular wax ÐÐÐ, AND A. RENFROW. 1971. The signi®cance of photosynthetic ef®- ciency to habitat preference and phylogeny among tillandsioid bromeli- powder helped predispose these lineages to carnivory. ads. Botanical Gazette 132: 19±30. ÐÐÐ, J. SEEMANN, AND A. RENFROW. 1978. The foliar epidermis in Til- ConclusionsÐHydrophobic leaf surfaces of Bromeliaceae landsioidae (Bromeliaceae) and its role in habitat selection. American possess a highly irregular microrelief, thereby reducing the Journal of Botany 65: 359±365. BILLINGS, F. W. 1904. A study of Tillandsia usneoides. Botanical Gazette 38: adhesion and spread of water on the leaf blade. Hydrophobic 99±121. trichome layers occur on the abaxial leaf blade surfaces of BREWER, C. A., W. K. SMITH, AND T. C. VOGELMANN. 1991. Functional many mesic Type 1 pitcairnioids and, as these species exhibit interaction between leaf trichomes, leaf wettability and the optical prop- the putative primitive ecological condition, water repellency erties of water droplets. Plant, Cell and Environment 14: 955±962. appears to have been an important condition in early Brome- EHLERINGER, J., AND O. BJOÈ RKMAN. 1976. Leaf pubescence: effects on ab- liaceae. The trichomes of Type 4 species are specialized for sorptance and photosynthesis in a desert shrub. Science 192: 376±377. FISH, D. 1976. Structure and composition of the aquatic invertebrate com- the alternative function of water and nutrient absorption from munity inhabiting epiphytic bromeliads in south Florida and the discov- a water-®lled tank, with epicuticular wax powders employed ery of an insectivorous bromeliad. Ph.D. dissertation, University of Flor- by some species to shed water from the leaf blades. Hydro- ida, Gainesville, Florida, USA. phobic trichome layers and wax powders could potentially ob- FRANK, J. H., AND G. F. O'MEARA. 1984. The bromeliad Catopsis berter- August 2001] PIERCE ET AL.ÐWATER REPELLENT INDUMENTA OF BROMELIADS 1389

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