COMMENTARY
New evidence on the origin ofcarnivorous plantsThomas J. Givnish1
Department of Botany, University of Wisconsin-Madison, Madison, WI 53706
Carnivorous plants have fascinated scientistsand the general public since the pioneeringstudies of Charles Darwin (1). No doubtpart of their wide appeal is that carnivorousplants have turned the evolutionary tableson animals, consuming them as prey, withthe green predators often equipped with re-markable lures, traps, stomachs, and—ina few cases—extraordinary speed of move-ment. To be considered carnivorous, a plantmust be able to absorb nutrients from deadbodies adjacent to its surfaces, obtain someadvantage in growth or reproduction, andhave unequivocal adaptations for activeprey attraction, capture, and digestion (2,3). Some carnivorous species [e.g., Pinguicula(butterworts), Philcoxia] lack obvious attrac-tants; some rely on passive pitfalls [e.g.,Cephalotus (Australian pitcher plant), Sarra-cenia (American pitcher plants)] rather thanactive traps based on sticky tentacles [e.g.,Byblis, Drosera (sundews)] or snap traps[e.g., Dionaea (Venus fly-trap), Utricularia(bladderworts)]; and some lack digestiveenzymes and instead depend on commensalmicrobes or insect larvae to break downprey (e.g., Brocchinia, Darlingtonia, somespecies of Sarracenia). Based on these crite-ria, today we recognize at least 583 species of
carnivorous plants in 20 genera, 12 families,and 5 orders of flowering plants (Table 1).Based on DNA sequence phylogenies, thesespecies represent at least nine independentorigins of the carnivorous habit per se, andat least six independent origins of pitfalltraps, five of sticky traps, two of snap traps,and one of lobster-pot traps. To the extentto which molecular phylogenies have beencalibrated against the ages of fossils of otherplants, these origins of carnivory appear tohave occurred between roughly 8 and 72million years ago (Mya). In PNAS, Sadowskiet al. (4) contribute to our understandingof the origins of plant carnivory by describ-ing the first fossilized trap of a carnivorousplant, a fragment of a tentacled leaf pre-served in Baltic amber from 35 to 47Mya, and allied to modern-day Roridula ofmonogeneric Roridulaceae (Ericales) fromSouth Africa.As with most carnivorous plants, the two
living species of Roridula today grow onopen, extremely infertile, moist sites. Theoccurrence of carnivorous plants on nutrient-poor substrates has been understood sinceDarwin showed that such plants augmenttheir supply of mineral nutrients throughprey capture. The restriction of carnivorous
plants to open, infertile, moist sites, however,remained unexplained until modern cost-benefit models showed that carnivores arelikely to obtain an advantage in growth rela-tive to noncarnivores only on such sites,where nutrients and nutrients alone limit plantgrowth, and where carnivory can acceleratephotosynthesis and the conversion of photo-synthate to new leaf tissue while decreasingallocation to root tissue (2, 3, 5, 6). Wet soilsand fire can favor carnivorous plants, by mak-ing N more limiting for growth while makinglight and water less limiting (3). The wet,sandy, fireswept sites in fynbos occupied byRoridula (6) should thus favor carnivory,and indeedRoridula often grows in associationwith large numbers of carnivorous sundews.Roridula, however, is in other respects
highly unusual as a carnivorous plant. Al-though its glistening, glandular tentaclesdo trap large numbers of insects, the secre-tions are resinous rather than aqueous, andso cannot support the activities of digestiveenzymes. It does not secrete proteolytic en-zymes; several authors thus argued thatRoridula could not be carnivorous becauseit could not digest prey or absorb the min-erals released (7, 8). The resinous nature ofRoridula secretions may be an adaptation tothe summer drought in the Mediterraneanclimate it now occupies, in that they do notlose volume or stickiness during long periodsof drought; the secretions also do not dissolveduring winter rains (9). It turns out that cer-tain hemipterans (Pameridea) are capable ofnegotiating the glandular leaves of Roridulawithout becoming entangled; they eat theprey immobilized by the plant, and thenN from their excretions is absorbed byRoridula (Fig. 1). This process substantiallyaugments the N supply to the plants, withthe plants obtaining 70% or more of theirnitrogen supply in this fashion (7, 10). Themutualism appears stabilized by nonlinearinteractions: excess densities of Pamerideaturn counterproductive as the bugs switchto sap-sucking in the absence of prey, leadingto negative impacts on Roridula and, ulti-mately, on the bugs themselves (11).
Table 1. Currently recognized groups of carnivorous plants
Order Family or clade Genus/genera* No. of taxa
Poales Bromeliaceae I BrocchiniaP 2Bromeliaceae II CatopsisP 1Eriocaulaceae PaepalanthusP 1
Caryophyllales DNDD cladeDroseraceae AldrovandaS, DionaeaS, DroseraT 115Nepenthaceae NepenthesP 90Drosophyllaceae DrosophyllumT 1Dioncophyllaceae TriphyophyllumT 1
Oxalidales Cephalotaceae CephalotusP 1Ericales RS-Actinidiaceae clade
Sarraceniaceae DarlingtoniaP, HeliamphoraP, SarraceniaP 32Roridulaceae RoridulaT 2
Lamiales Byblidaceae ByblisT 6Lentibulariaceae GenliseaL, PinguiculaT, UtriculariaS 330Plantaginaceae PhilcoxiaT 1
Taxa include all members of each genus, except for the monocot genera in order Poales, where the number ofcarnivorous species within the genus is listed. Independent origins of carnivory per se are indicated by boldface entriesin the family/clade column.*Trap types indicated by superscript: L, lobster-pot trap; P, pitfall; S, snap trap; T, sticky trap.
Author contributions: T.J.G. wrote the paper.
The author declares no conflict of interest.
See companion article 10.1073/pnas.1414777111.
1Email: [email protected].
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Although the Roridula system is truly re-markable, similar kinds of complex digestivemutualisms may occur in other carnivorousplants. For example, Nepenthes bicalcarataprovides domatia for ants, despite ants beingthe most frequent prey of many Nepenthes.Givnish (5) and Hölldobler and Wilson (12)proposed that the resident ants and plantsmight have a mutualistic relationship of somekind. In fact, the resident ant Camponotusschmitzi protects N. bicalcarata from weevilsthat attack their tendrils, and in additionfacilitates the plant’s uptake of nutrients(13). The ants can swim in the pitcher fluidwithout adverse effect, retrieve large preyitems, and excrete wastes into the pitcher,accelerating nutrient uptake; ant wastes ac-count for 42–76% of total N uptake and antsprolong pitcher lifetimes (13). In other sys-tems, the prey processed by a digestive mu-tualist may not even be captured by theplant’s own traps. Nepenthes lowii attractstree shrews (Tupaia montana) to their excep-tionally large, broad traps with secretedrewards, and they defecate into the pitcherwhile marking it as their territory. Their fecesaccount for 57–100% of all leaf N (14).Nepenthes rafflesiana var. elongata, withsmaller but elongate traps, provides a roostfor a small bat and obtains nutrients fromits feces (15). Whether these systems are bestviewed as coprophagy or indirect forms ofcarnivory involving digestive mutualists thatdeliver the remains of prey is worth debating.Clearly, however, both plants benefit fromanimals whose death results in their acquisi-tion of nutrients; we might consider them
“apparent carnivorous plants,” in homage toHolt’s concept of apparent competition (16).The new fossil Roridula not only is the first
fossil trap leaf uncovered, it is one of the veryfew undoubted fossils of carnivorous plantsof any kind. Archaeamphora from Chinesesediments 112 Mya was originally describedas Sarraceniaceae, but now there is strongdoubt that it was a member of that familyor even a carnivorous plant; the unusualleaves may simply not have been traps (17).Paleoaldrovanda, putatively a member of Dro-seraceae based on a “seed,” may actually have
been a fossil insect egg (18). The remainingfossils considered legitimate remains of carniv-orous plants include one seed (now destroyed)of Byblis (Byblidaceae) from Australia (19),and palynomorphs possibly allied withNepenthaceae (20). The last two fragments,however, do not demonstrate that the plantsto which they belonged were, in fact, carniv-orous, which makes the find by Sadowski et al.(4) particularly important. The age of the am-ber Roridula, 35–47 Mya, nicely brackets thedivergence between Roridula and noncarnivo-rous Actinidiaceae roughly 39 Mya, as esti-mated from a calibrated DNA phylogeny(21). This result lends credence to the ageestimates based on molecular data, and tothe inference from phylogenetic reconstruc-tion that early Roridulaceae were carnivorous.The identity of the fossil Roridula appears tobe beyond doubt. The former occurrence ofRoridula around the Baltic—whereas its pres-ent-day distribution is restricted to the CapeFloristic Province of southwest South Africa—implies that this group was once far morewidespread. The distributions of families inthe Clethraceae-Sarraceniaceae-Roridulaceae-Actinidiaceae clade suggest that it originatedin southeastern North America or northernSouth America. In the next few years, furtherinvestigations of the Baltic amber might tell uswhat other plants grew in association withfossil Roridula, and thus the nature of thevegetation in which fossil Roridula grew.Based on cost-benefit models, the distributionof present-day Roridula, and the current dis-tributions of almost all other carnivorousplants, it seems most unlikely that fossilRoridula grew below a dense canopy of theconifer forests that produced amber!
1 Darwin C (1875) Insectivorous Plants (Appleton and Co., London).2 Givnish TJ, Burkhardt EL, Happel RE, Weintraub JW (1984)
Carnivory in the bromeliad Brocchinia reducta, with a cost/benefit
model for the general restriction of carnivorous plants to sunny,
moist, nutrient-poor habitats. Am Nat 124(4):479–497.3 Ellison M, Adamec L (2011) Ecophysiological traits of terrestrial
and aquatic carnivorous plants: Are the costs and benefits the same?
Oikos 120(11):1721–1731.4 Sadowski E-M, et al. (2014) Carnivorous leaves from Baltic amber.
Proc Natl Acad Sci USA, 10.1073/pnas.1414777111.5 Givnish TJ (1989) Ecology and evolution of carnivorous plants.
Plant–Animal Interactions, ed Abrahamson WG (McGraw-Hill, New
York), pp 243–290.6 Ellison AM, Gotelli NJ (2009) Energetics and the evolution of
carnivorous plants—Darwin’s ‘most wonderful plants in the world’
J Exp Bot 60(1):19–42.7 Ellis AG, Midgley JJ (1996) A new plant-animal mutualism involving
a plant with sticky leaves and a resident hemipteran. Oecologia
106(4):478–481.8 Juniper BE, Robias RJ, Joel DM (1989) The Carnivorous Plants
(McGraw-Hill, New York).9 Voigt D, Gorb S (2010) Desiccation resistance of adhesive
secretion in the protocarnivorous plant Roridula gorgonias as an
adaptation to periodically dry environment. Planta 232(6):
1511–1515.10 Anderson B, Midgley JJ (2003) Digestive mutualism, an alternative
pathway in plant carnivory. Oikos 102(1):221–223.
11 Anderson B, Midgley JJ (2007) Density-dependentoutcomes in a digestive mutualism between carnivorousRoridula plants and their associated hemipterans. Oecologia152(1):115–120.12 Hölldobler B, Wilson EO (1990) The Ants (Springer, Berlin).13 Bazile V, Moran JA, Le Moguédec G, Marshall DJ, Gaume L(2012) A carnivorous plant fed by its ant symbiont: A unique multi-faceted nutritional mutualism. PLoS ONE 7(5):e36179.14 Clarke CM, et al. (2009) Tree shrew lavatories: A novel nitrogensequestration strategy in a tropical pitcher plant. Biol Lett 5(5):632–635.15 Grafe TU, Schöner CR, Kerth G, Junaidi A, Schöner MG (2011) Anovel resource-service mutualism between bats and pitcher plants.Biol Lett 7(3):436–439.16 Holt RD (1977) Predation, apparent competition, and thestructure of prey communities. Theor Popul Biol 12(2):197–229.17 Brittnacher J (2013) Phylogeny and biogeography of theSarraceniaceae. Carniv Plant Newsletter 42(3):99–106.18 He�rmanová Z, Kvacek J (2010) Late CretaceousPalaeoaldrovanda, not seeds of a carnivorous plant but eggs of aninsect. J Natur Hist Mus (Prague) 179(9):105–118.19 Conran J, Christophel D (2004) A fossil Byblidaceae seed fromEocene South Australia. Int J Plant Sci 165(4):691–694.20 Kumar M (1995) Pollen tetrads from Palaeocene sediments ofMeghalaya, India: Comments on their morphology, botanical affinityand geological records. Palaeobot 43(1):68–81.21 Ellison AM, et al. (2012) Phylogeny and biogeography ofthe carnivorous plant family Sarraceniaceae. PLoS ONE 7(6):e39291.
Fig. 1. Growth form of Roridula gorgonias at Fernkloof Nature Reserve near Hermanus, showing glandular ten-tacles that immobilize insect prey. Close-up of leaves, showing a Pameridea bug (center) that eats immobilized preyand delivers nutrients to the plant via excreta.
2 of 2 | www.pnas.org/cgi/doi/10.1073/pnas.1422278112 Givnish
Carnivorous leaves from Baltic amberEva-Maria Sadowskia, Leyla J. Seyfullaha, Friederike Sadowskib, Andreas Fleischmannc, Hermann Behlingd,and Alexander R. Schmidta,1
aDepartment of Geobiology, University of Göttingen, 37077 Göttingen, Germany; bInstitute for Interdisciplinary Research on Conflict and Violence, BielefeldUniversity, 33615 Bielefeld, Germany; cBotanische Staatssammlung München, 80638 Munich, Germany; and dDepartment of Palynology and ClimateDynamics, Albrecht von Haller Institute of Plant Sciences, University of Göttingen, 37073 Göttingen, Germany
Edited by Peter R. Crane, Yale School of Forestry and Environmental Studies, New Haven, CT, and approved November 3, 2014 (received for review August1, 2014)
The fossil record of carnivorous plants is very scarce and macro-fossil evidence has been restricted to seeds of the extant aquaticgenus Aldrovanda of the Droseraceae family. No case of carnivo-rous plant traps has so far been reported from the fossil record.Here, we present two angiosperm leaves enclosed in a piece ofEocene Baltic amber that share relevant morphological featureswith extant Roridulaceae, a carnivorous plant family that is todayendemic to the Cape flora of South Africa. Modern Roridulaspecies are unique among carnivorous plants as they digest preyin a complex mutualistic association in which the prey-derivednutrient uptake depends on heteropteran insects. As in extantRoridula, the fossil leaves possess two types of plant trichomes, in-cluding unicellular hairs and five size classes of multicellular stalkedglands (or tentacles) with an apical pore. The apices of the narrowand perfectly tapered fossil leaves end in a single tentacle, as in bothmodern Roridula species. The glandular hairs of the fossils are re-stricted to the leaf margins and to the abaxial lamina, as in extantRoridula gorgonias. Our discovery supports current molecular ageestimates for Roridulaceae and suggests a wide Eocene distributionof roridulid plants.
plant carnivory | Roridulaceae | Eocene | Ericales
Plant carnivory is traditionally defined as the attraction, cap-ture, and digestion of prey by vegetative traps, with the
subsequent uptake of nutrients (1, 2). Some carnivorous plants,however, challenge the boundary of the botanical carnivory con-cept because they depend on commensal organisms for the di-gestion of their prey (2, 3). The most famous representativeof those plants is Roridula, placed in the monogeneric familyRoridulaceae that is endemic to a few localities in the south-western Cape of South Africa (4, 5).The resinous glandular leaves of both extant species, Roridula
dentata and Roridula gorgonias, capture plenty of arthropods.The sticky trapping glue of Roridula is a viscous lipophilic resincontaining triterpenoids as major component, which does notallow dissolution of digestive enzymes (6). Consequently, thesecretory glands of Roridulaceae lack enzymatic activity (7, 8).For prey-derived nutrient uptake, Roridula depends on two ob-ligately associated heteropteran Pameridea species (family Miridae,“capsid bugs”), which feed on the trapped animals (5, 9). In this“digestive mutualism” (10), the nutrient-rich fecal compounds ofthese “Roridula bugs” are incorporated by Roridula throughnanometer-sized cuticular gaps and serve for a better alimenta-tion in a nutrient-poor habitat (7, 8, 10, 11). The benefit of nu-trient uptake from captured prey is the essential criterionfor the concept of botanical carnivory (1, 2) and thus includesRoridulaceae (11, 12).Here, we report two leaf fossils from Eocene Baltic amber
possessing the relevant morphological features of an adhesiveflypaper trap plant that we assign to the Roridulaceae lineage(Figs. 1–3). Both specimens originate from the Jantarny ambermine near Kaliningrad (Russia). The amber-bearing sedimentsof this fossil site date to 35–47 million years ago (13, 14).
ResultsThe linear-lanceolate leaves are 5 and 4.5 mm long and 0.2 mmwide at the base, and they narrow gradually toward the leaf tip,which terminates in a stalked gland (tentacle; Fig. 1). The leavespossess two trichome types: tentacles and nonglandular hyalinehairs (Figs. 1 and 2). The hyaline trichomes are located on bothsides and the margins of the lamina, whereas the tentacles areexclusively found along the margins and on the abaxial sidewithout a definite arrangement (Fig. 1). The tentacles are multi-cellular, consisting of a tapering stalk and a clavate to ovoidglandular head, which shows a small pore at the center of its distalside (Fig. 1 E and F). The stalks of the glands measure between 20and 350 μm in length (Figs. 1 and 2 A and C), whereas an ex-ceptional stalk exceeds this size, reaching 1.4 mm (Fig. 3A). Aswith the stalks, the glandular heads vary in size (20–120 μm long,10–40 μm wide). Adhered organic remains as well as trichomes ofother plants attached to the glandular tentacle heads (Figs. 1A and D and 3A) indicate that they excreted a sticky secretion, asknown from adhesive traps of extant carnivorous plants. The non-glandular trichomes are hyaline, unicellular, and arcuate to straight,tapering toward an acute apex (Figs. 1C and 2E). Their lengthranges from 10 to 80 μm, and their width reaches up to 12 μm.Both leaves exhibit a well-preserved epidermis with small tetrag-
onal cells at the leaf base and elongated larger cells from the middlepart toward the leaf tip. These cells measure 3–54 × 6–18 μm.Stomata of 20–38 × 15–25 μm are present on the abaxial leaf side(Figs. 1C and 2G).Our statistical cluster analyses (SI Text) revealed that the
fossil specimens and Roridula gorgonias show the same mor-phological pattern among the tentacles. R. gorgonias exhibits five
Significance
Amber, fossil tree resin, preserves organisms in microscopicfidelity, and frequently fossils preserved in amber are other-wise absent in the entire fossil record. Plant remains, however,are rarely entrapped in amber, compared with the vast amountof insects and other animals. Our newly discovered fossils fromEocene Baltic amber are the only documented case of fossilizedcarnivorous plant traps and represent the first fossil evidenceof the carnivorous plant family Roridulaceae, which is today anarrow endemic of South Africa. Hence, our results shed lightonto the paleobiogeography of the Roridulaceae, indicating awide Eocene distribution of the roridulid ancestors and chal-lenging previous notions about a Gondwanan origin of thisplant family.
Author contributions: E.-M.S., L.J.S., H.B., and A.R.S. designed research; E.-M.S., L.J.S., F.S.,A.F., H.B., and A.R.S. performed research; E.-M.S., L.J.S., F.S., A.F., and A.R.S. analyzeddata; and E.-M.S., L.J.S., A.F., and A.R.S. wrote the paper.
The authors declare no conflict of interest.
This article is a PNAS Direct Submission.1To whom correspondence should be addressed. Email: [email protected].
This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1414777111/-/DCSupplemental.
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Fig. 1. Carnivorous leaves from Eocene Baltic amber. (A) Overview of the leaf enclosed in amber specimen GZG.BST.27310 showing the adaxial tentacle-freeside in slightly oblique view and stalked glands at the margin and on the abaxial side; arrowhead points to the exceptional long tentacle stalk with severalbranched oak trichomes attached. (B) Overview of the leaf enclosed in amber specimen GZG.BST.27311, showing abundant tentacles on the abaxial side. (C)Margin of abaxial leaf surface with tentacles of different size classes and nonglandular hyaline trichomes. (D) Leaf apex tapering into a sole tentacle. (E and F)Glandular heads with central pore (arrowheads) from both leaves. (Scale bars: A and B, 1 mm; C and D, 100 μm; E, 10 μm; F, 40 μm.)
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Fig. 2. Morphological comparison of the carnivorous leaf fossils from Baltic amber (Left) and extant Roridula species (Right). (A and B) Leaf tip ending in a soletentacle. (C and D) Stalked glands of different size classes. (E and F) Hyaline unicellular nonglandular trichomes. (G and H) Epidermal cells and stomata. (I–L) Mul-ticellular tentacles. (A, C, E, and G) GZG.BST.27310. (I and J) GZG.BST.27311. (B, D, K, and L) R. gorgonias. (F and H) R. dentata. (Scale bars: A–D, 100 μm; E–L, 50 μm.)
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size clusters of tentacle stalk lengths whereas four clusters weredetected for the amber inclusions. As an outlier, the longest ten-tacle was excluded from the cluster analyses of the fossils, but itspresence indicates that a fifth size class was present in the Eoceneleaves, as in extant Roridulaceae (Fig. 4).
DiscussionAlthough glandular secreting trichomes appear in about 30% ofvascular plants (15), the unique character combination in thefossils bears most similarities to extant representatives of theRoridulaceae. They share the long, narrow, and perfectly ta-pered leaf lamina ending in a single tentacle, the presence andmorphology of two trichome types (tentacles and nonglandularhairs), the possession of glandular hairs along the leaf marginsand on the abaxial lamina, the tentacle head with a central pore,and the size and shape of the epidermal cells and stomata. Theglabrous adaxial side of the amber inclusions and the hyalinetrichomes being located on both leaf surfaces only appear in thesepals of extant Roridula gorgonias. Besides smaller tentaclesizes, the fossils are distinguished from leaves of extant Roridulaspecies by the absence of a prominent midrib on the abaxial leafside, and are thus most similar to sepals of Roridula gorgonias.
Extant Roridula plants are very effective traps for all kinds ofarthropods due to the sticky resinous trapping glue and the hier-archical organization of the tentacles into functional units for ef-fective prey capture (16, 17). The longest tentacles make the firstcontact with the prey. Due to the high flexibility of these prom-inent tentacles, the moving prey then gets stuck to the medium-sized tentacles, which slow down the caught animal. Finally, thesmallest and stiffest tentacles immobilize the prey (16). As inmodern Roridulaceae, the leaf fossils have different size classes oftentacles that fulfill the functional roles for prey capture (entan-glement, slow-down, and immobilization) and comply with therequirements for a carnivorous nature. In addition, the pore of thetentacle heads distinguishes the fossils from any other extantcarnivorous plants with glandular adhesive traps such as sundews(Drosera) (3, 18, 19).In the fossil record, evidence of carnivorous plants is exceed-
ingly rare and macrofossils are restricted to seeds of the aquaticcarnivore Aldrovanda (Droseraceae), which are recorded sincethe Eocene (20, 21). Hence, the fossil leaves from Baltic amberare (to our knowledge) the first documented case of carnivorousplant traps being fossilized.
Fig. 3. Carnivorous leaf from Eocene Baltic amber (A and B; GZG.BST.27310) and leaves of extant Roridula gorgonias (C and D). (A) Exceptionallylong tentacle stalk (with several branched oak trichomes attached) of the fossil leaf representing the fifth size class of stalked glands. (B and C ) Overviewsshowing the tentacle-free adaxial surface and tentacles along the leaf margins. (D) Partial leaf tip showing different size classes of stalked glands. (Scalebars: A, 100 μm; B, 500 μm; C and D, 1 mm.)
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The occurrence of Eocene Roridulaceae is consistent withrecent divergence time estimates for a split of Sarraceniaceae(carnivorous American pitcher plants) from the Roridulaceae–Actinidiaceae clade about 48.6 million years ago, whereas themost recent common ancestor of Roridulaceae and Actinidia-ceae was estimated at 38.1 million years ago (22). The age ofthese Ericales lineages is further supported by Late Cretaceousfossil flowers with affinities to the Actinidiaceae and Clethraceaefamilies (23). The sediments containing the majority of Balticamber are 35–47 million years old (13, 14). Thus, the amberfossils probably represent an early representative of theRoridulaceae lineage.The geologic setting of the Baltic amber deposit and the
paleobotanical record suggest that coastal areas with carbonate-free, nutrient-poor soils and swamp depressions harbored well-structured mixed forests of angiosperm and conifer trees withintermixed open habitats growing in a subtropical to warm-temperate climate (24–27).The presence of Eocene roridulid plants in the northern
hemisphere challenges notions about the biogeographical historyof extant Roridulaceae, which were previously assumed to rep-resent “old Cape elements,” paleoendemics of Gondwanan ori-gin, dating back to up to 90 million years (28, 29). Thus, the leaffossils represent an example of pseudo-Gondwanan relicts, extinct
in Europe today and restricted to particular areas of the southerncontinents. With respect to the distinctive distribution areas of theclosely related extant families Sarraceniaceae (North and SouthAmerica) and Actinidiaceae (tropical Asia and America), the re-striction of extant Roridulaceae to small patches in the Caperegion can be regarded as relictual, probably resulting from post-Eocene extinction events.
Materials and MethodsProvenance of the Amber Piece. The leaf inclusions were discovered in anamber piece that derives from Jantarny mine near Kaliningrad (Russia).Amber in this locality is mined in the “Blue Earth” layer, which is Priabonianin age (late Eocene, 35 million years minimum age) (13, 14). The amber piecewas obtained from the collection of Christel and Hans Werner Hoffeins(Hamburg, Germany).
Microscopy and Imaging. The original 39 × 21 × 5-mm piece of amber wasground and polished manually with wet silicon carbide papers (grit from25.8- to 5-μm particle size; firm Struers). Two amber fragments measuring21 × 14 × 3 and 24 × 19 × 3 mm with one leaf inclusion each were obtainedby cutting the amber piece with a dental drill. The amber pieces are housedin the Geoscientific Collections of the Georg August University Göttingen(Göttingen, Germany) (collection numbers GZG.BST.27310 and GZG.BST.27311).Leaves of extant Roridula gorgonias and R. dentata (Roridulaceae) wereobtained from cultured specimens of Thomas Carow (Nüdlingen, Germany)and A.R.S. The leaf inclusions and the extant Roridulaceae plant materialwere examined under a Carl Zeiss Stemi 2000 dissection microscope and aCarl Zeiss AxioScope A1 compound microscope, each equipped with a Canon60D digital camera. In most instances, incident and transmitted light wereused simultaneously. The images of Figs. 1 A–D, 2, and 3 A and B are digitallystacked photomicrographic composites of up to 130 individual focal planesobtained using the software package HeliconFocus 5.0 for a better illustra-tion of the 3D structures.
Statistics. According to Voigt et al. (16), Roridula gorgonias possesses threetentacle size classes that allow a very effective capture of prey. To testwhether the tentacle size classes are present in the amber specimens, thetentacle measurements of the amber specimens, Roridula gorgonias andR. dentata were statistically evaluated, applying hierarchical cluster analyseswith the statistics package IBM SPSS 21. In total, measurement values of 103tentacles from both leaf inclusions and 103 tentacles from each extantRoridula species were used, comprising the stalk length, the stalk base andtip width, the gland length, and the gland width. The hierarchical clusteranalyses were computed using Ward’s method. The resulting clusters wereoptimized with the nonhierarchical k-means method and tested statistically,using three criteria suggested by Bacher (30), which are η-squared (η2), F-max,and the proportional reduction of error (PRE). The best cluster solution isthe one where the values of η2 and PRE do not show any considerable im-provement in the subsequent solution. Furthermore, the F value and η2 shouldbe maximal, whereas PRE is supposed to be low. The number of clusters alsoshould be selected with regard to the content and the underlying theoreticalmodel (30). Results of the statistical analyses are provided in SI Text.
ACKNOWLEDGMENTS. We thank Christel and Hans Werner Hoffeins(Hamburg) for providing the amber specimen and Thomas Carow (Nüdlingen)for providing a plant of Roridula dentata for study. We are grateful to JuliaGundlach (Bielefeld University), Dorothea Hause-Reitner, and GerhardHundertmark (University of Göttingen) for assistance, and to two anonymousreviewers for constructive suggestions.
1. Givnish TJ, Burkhardt EL, Happel RE, Weintraub JD (1984) Carnivory in the bromeliadBrocchinia reducta, with a cost/benefit model for the general restriction of carnivo-rous plants to sunny, moist, nutrient-poor habitats. Am Nat 124(4):479–497.
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Amber fossils
Size class 5 (n=5)
Size class 4 (n=9)
Size class 3 (n=3)
Size class 2 (n=18)
Size class 1 (n=61)
B
0
700
1400
2100
2800
3500 Roridula gorgonias
stal
k le
ngth
(µm
)st
alk
leng
th (µ
m)
0
300
600
900
1200
1500
Size class 5 (n=1)
Size class 4 (n=8)
Size class 3 (n=18)
Size class 2 (n=28)
Size class 1(n=43)
A
Fig. 4. Tentacle size classes of the fossil leaves and extant Roridulagorgonias based on the results of the cluster analyses and the tentaclestalk length. (A) Size classes of the fossil leaves, including the outlierwhich we interpret to represent size class 5. (B) Size classes of Roridulagorgonias. n indicates the number of tentacles per size class.
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