A novel resource-service mutualism between bats and pitcher plants

Abstract

Mutualistic relationships between vertebrates and plants apart from the pollen and seed-dispersal syndromes are rare. At first view, carnivorous pitcher plants of the genus Nepenthes seem to be highly unlikely candidates for mutualistic interactions with animals, as they form dimorphic terrestrial and aerial pitchers that trap arthropods and small vertebrates. Surprisingly, however, the aerial pitchers of Nepenthes rafflesiana variety elongata are poor insect traps, with low amounts of insect-attractive volatile compounds and low amounts of digestive fluid. Here, we show that N. rafflesiana elongata gains an estimated 33.8 per cent of the total foliar nitrogen from the faeces of Hardwicke's woolly bats (Kerivoula hardwickii hardwickii) that exclusively roost in its aerial pitchers. This is the first case in which the faeces-trapping syndrome has been documented in a pitcher plant that attracts bats and only the second case of a mutualistic association between a carnivorous plant and a mammal to date.

Full text

doi: 10.1098/rsbl.2010.1141
published online 26 January 2011Biol. Lett.
T. Ulmar Grafe, Caroline R. Schöner, Gerald Kerth, Anissa Junaidi and Michael G. Schöner
plants service mutualism between bats and pitcher−A novel resource
Supplementary data
ml
http://rsbl.royalsocietypublishing.org/content/suppl/2011/01/21/rsbl.2010.1141.DC1.ht
"Data Supplement"
References #ref-list-1
http://rsbl.royalsocietypublishing.org/content/early/2011/01/21/rsbl.2010.1141.full.html
This article cites 11 articles, 3 of which can be accessed free
P<P Published online 26 January 2011 in advance of the print journal.
Subject collections
(2389 articles)evolution  (2110 articles)ecology 
Articles on similar topics can be found in the following collections
Email alerting service hereright-hand corner of the article or click
Receive free email alerts when new articles cite this article - sign up in the box at the top
publication.
Citations to Advance online articles must include the digital object identifier (DOIs) and date of initial
online articles are citable and establish publication priority; they are indexed by PubMed from initial publication.
the paper journal (edited, typeset versions may be posted when available prior to final publication). Advance
Advance online articles have been peer reviewed and accepted for publication but have not yet appeared in
http://rsbl.royalsocietypublishing.org/subscriptions go to: Biol. Lett.To subscribe to
This journal is © 2011 The Royal Society
on January 26, 2011rsbl.royalsocietypublishing.orgDownloaded from

Evolutionary biology
A novel resource–service
mutualism between bats
and pitcher plants
T. Ulmar Grafe1,*, Caroline R. Scho
¨ner2,
Gerald Kerth3, Anissa Junaidi1
and Michael G. Scho
¨ner2
1
Department of Biology, University Brunei Darussalam, Tungku Link,
Gadong 1410, Brunei Darussalam
2
Department of Animal Ecology and Tropical Biology, University of
Wu
¨rzburg, Biozentrum, Am Hubland, 97074 Wu
¨rzburg, Germany
3
Zoological Institute and Museum, Greifswald University, Johann-
Sebastian-Bach-Strasse 11/12, 17489 Greifswald, Germany
*Author for correspondence (grafe@biozentrum.uni-wuerzburg.de).
Mutualistic relationships between vertebrates
and plants apart from the pollen and seed-
dispersal syndromes are rare. At first view,
carnivorous pitcher plants of the genus
Nepenthes seem to be highly unlikely candidates
for mutualistic interactions with animals, as
they form dimorphic terrestrial and aerial pitch-
ers that trap arthropods and small vertebrates.
Surprisingly, however, the aerial pitchers of
Nepenthes rafflesiana variety elongata are
poor insect traps, with low amounts of insect-
attractive volatile compounds and low amounts of
digestive fluid. Here, we show that N. rafflesiana
elongata gains an estimated 33.8 per cent of the
total foliar nitrogen from the faeces of
Hardwicke’s woolly bats (Kerivoula hardwickii
hardwickii) that exclusively roost in its aerial
pitchers. This is the first case in which the
faeces-trapping syndrome has been documented
in a pitcher plant that attracts bats and only the
second case of a mutualistic association between
a carnivorous plant and a mammal to date.
Keywords: Kerivoula hardwickii;Nepenthes rafflesiana
variety elongata; mutualism; nitrogen acquisition
1. INTRODUCTION
Carnivorous pitcher plants of the genus Nepenthes grow
in nutrient-poor soils and rely on trapping arthropods
to acquire sufficient nitrogen [1–3]. In many pitcher
plant species, pitcher morphology, fluid viscoelastic
properties, extent of epicuticular wax crystals and
peristome design predict, to a large extent, prey com-
position (e.g. [4,5]). In Borneo, Nepenthes rafflesiana
has five distinct growth forms, one of which is extra-
ordinary in several ways. Nepenthes rafflesiana elongata
(figure 1a) possesses aerial pitchers that, compared
with the typical variety Nepenthes rafflesiana typica,
are up to four times longer, produce fewer human-
perceptible fragrances, exhibit a unique UV light
absorption spectrum and capture insects at rates up
to seven times lower [6,7]. Surprisingly, we regularly
found woolly bats (Kerivoula hardwickii hardwickii
[8]) roosting above the digestive fluid in the aerial
pitchers of N. r. elongata, and hypothesized that pitcher
plants entice bats to roost in their pitchers in return for
nitrogen that they sequester from bat faeces or urine.
Other bat species use a variety of daytime roosts,
including furled leaves and bamboo culms [9], but
none are known to roost in pitcher plants. The aim
of this study was to quantify pitcher use by woolly
bats and to determine how much nitrogen the plant
derives from woolly bat faeces. We hypothesized that
a mutualistic relationship exists between Hardwicke’s
woolly bats and N. r. elongata.
2. MATERIAL AND METHODS
From 14 June to 30 July 2009, Hardwicke’s woolly bats were caught
by daily patrols of 423 pitchers of N. r. elongata in a peat swamp
forest in Brunei Darussalam (4835016.8000 N, 114830048.8000 E).
Using skinbond adhesive (Manfred Sauer GmbH), we fixed trans-
mitters (Holohil Systems Ltd; weight: 0.4 g) onto bats’ backs,
which fall off after 3–12 days. To localize the bats, we used a
TRX-1000S Wildlife Materials receiver (Carbondale, IL, USA).
We compared the isotopic signature of leaf blades from pitchers
used as roosts with those from control pitchers that were never
occupied, which had been monitored daily since they had opened.
Occupied (n¼38) and control pitchers (n¼17) and their associated
leaf blades were collected after three to six weeks of daily monitoring,
for analysis of total nitrogen and stable isotope ratios of
15
N/
14
N
(
d
15
N). The contribution from bats to total foliar nitrogen was
estimated using a two-member isotopic mixing model [10] using
mean
d
15
N values for leaf blades of bat-occupied aerial pitchers (n¼
38), aerial leaf blades without faecal input (n¼17) and woolly bat
faeces (n¼3, see details of the model in the electronic supplementary
material).
For the analysis of total N and
d
15
N, samples were sent to the
Cornell isotope laboratory (COIL; Ithaca, NY, USA), which used
a Thermo Delta V isotope ratio mass spectrometer (IRMS) inter-
faced to an NC2500 elemental analyser. The primary reference
scale for
d
15
N was atmospheric air. An internal standard (s.d. ¼
0.09‰ for
d
15
N) was analysed after every 10 samples to ensure accu-
racy. We measured pitcher length and fluid levels in N. r. elongata and
N. r. typica to evaluate the available roosting space. Means and s.d.
are given as descriptive statistics.
3. RESULTS
We found 14 male and 18 female Hardwicke’s woolly
bats roosting in aerial pitchers of N. r. elongata in
2008 and 2009 (figure 1b). Of these, we radio-tracked
17 males and females over an average of 6.1 +3.3 days
(range 1–12 days) and found that each tagged individ-
ual exclusively used pitchers of N. r. elongata as
daytime roosts, despite the widespread abundance of
other potential roosting sites (furled leaves, hollow
trees and other Nepenthes (N. r. typica,N. bicalcarata,
N. ampullaria)). During our 6.5 week study period in
2009, 87 (20.8%) of 418 monitored N. r. elongata
pitchers were occupied by a total of 25 bats. In total,
64 plants (out of 223) harboured at least one bat in
one of its pitchers. Thus, the incidence of plant occu-
pancy was 28.7 per cent over the total study period of
6.5 weeks, compared with a 20.8 per cent pitcher occu-
pancy rate. On average, 1.30 +0.70% of the 423
monitored pitchers was occupied on a single day. On
two separate occasions, a juvenile bat (one male, one
female) shared the same pitcher with their mother.
The elongated pitcher in N. r. elongata provided bats
ample room to roost. The distance between the lower
rim of the peristome and the pitcher fluid was
151.4 +37.8 mm (n¼58; range: 49.7–228.5 mm).
With the bats’ body length averaging 36.6 +0.8 mm
(n¼4), most N. r. elongata aerial pitchers provided
enough roosting space for two bats stacked above
Electronic supplementary material is available at http://dx.doi.org/
10.1098/rsbl.2010.1141 or via http:// rsbl.royalsocietypublishing.org.
Biol. Lett.
doi:10.1098/rsbl.2010.1141
Published online
Received 30 November 2010
Accepted 4 January 2011 This journal is q2011 The Royal Society
on January 26, 2011rsbl.royalsocietypublishing.orgDownloaded from

each other. In contrast, N. r. typica aerial pitchers,
which were not occupied by bats, provided signifi-
cantly less roosting space (t-test, t¼2912, d.f. ¼68,
p,0.001) with average distances of only 50.5 +
12.3 mm (n¼12, range: 31.6 –67.7 mm) between
peristome and fluid (figure 1c). Fluid levels were sig-
nificantly lower in N. r. elongata (25.0 +23.7 mm)
than in N. r. typica (54.0 +12.0 mm; t-test, t¼4.12,
d.f. ¼68, p,0.001).
To test the hypothesis that pitcher plants sequester
nitrogen from bat faeces or urine, we compared the
total nitrogen and
15
N/
14
N(
d
15
N) stable isotopic sig-
nature of leaf blades from pitchers used as roosts with
those of leaf blades from pitchers never occupied by
bats. We found that total foliar nitrogen was signifi-
cantly higher in leaf blades of pitchers in which
bats had been found roosting (1.48 +0.31%) than in
control leaf blades (1.31 +0.19%; Mann–Whitney
U-test, U¼208, n
1
¼38, n
2
¼17, p,0.05;
figure 2a). Moreover, foliar
d
15
N, used to infer the nitro-
gen source (i.e. its trophic level; [11]), was significantly
higher when pitchers harboured bats (1.30 +1.53‰)
than when they did not (0.51 +1.09‰; Mann–
Whitney U-test, U¼181.5, n
1
¼38, n
2
¼17, p,0.01;
figure 2b). Using a two-member mixing model [10],
we estimate that between 11 and 56 per cent (95% CI;
n¼38) or on average 33.8 per cent of foliar nitrogen in
N. r. elongata is derived from woolly bat faeces.
4. DISCUSSION
The results of the radio-telemetry and the stable iso-
tope analyses suggest a resource –service relationship
between a bat and a carnivorous pitcher plant, with
clear benefits to both mutualistic partners. The
woolly bats (K. h. hardwickii) that we observed solely
used aerial pitchers of N. r. elongata as daytime
roosts, providing the plant with nitrogen in an
nitrogen-deprived environment. This is an unusual
case of an animal –plant mutualism in which nutrients
are supplied by the animal and not vice versa [12].
Both morphological and physiological characters of
N. r. elongata aerial pitchers facilitate bat roosting. Bats
depend on shelter for survival, reproduction and pro-
tection from predators [13]. The morphology of the
N. r. elongata pitchers offers protection for K. hard-
wickii (figure 1b). Bats are hardly visible through the
pitcher wall and they are safe from rain or direct
solar radiation. Moreover, the elongated form of the
pitchers, which distinguishes them from the typical
form [1,7,6], ensures that not only single bats but
also mother–juvenile pairs comfortably fit into the
pitchers. This might be a critical feature, as mothers
and juveniles in many bat species remain associated
for several weeks to months [14]. Another conspicuous
feature of N. r. elongata aerial pitchers is that they taper
distinctly after a lignified ‘girdle’ in the lower half of the
pitcher. Woolly bats were usually wedged head first in
the pitcher with their heads below the girdle,
suggesting that this girdle in combination with the
slender pitcher form obviates the need for bats to
cling to the slippery pitcher wall or peristome. More-
over, fluid levels are much reduced in aerial pitchers
of N. r. elongata compared with its terrestrial pitchers
or those of the typical form (own observations),
suggesting that physiological modifications regulating
fluid production are in place to accommodate bats.
Our findings show that the enigma of N. r. elongata’s
lower arthropod capture rate compared with the typical
form [7] is solved when the role of bats is considered.
Attracting bats appears to be facilitated by both pitcher
morphology and physiology: narrow and cylindrical
aerial pitchers in combination with reduced liquid
levels. Bat faecal pellets can provide high rewards, with
an estimated 33.8 per cent of the total foliar nitrogen pro-
vided suggesting that trapping faeces in N. r. elongata has
been successful and might even be superior to trapping
insects in areas in which bats are abundant.
woolly bat
leaf blade
peristome
digestive
fluid level
girdle
(a)(b)(c)
digestive
fluid level
5 cm
Figure 1. Service benefit provided by N. r. elongata to K. h. hardwickii.(a) Aerial pitcher of N. rafflesiana var. elongata.(b) The
same pitcher with the front tissue removed to reveal a roosting Hardwick’s woolly bat. (c) The shorter aerial pitcher of
N. rafflesiana variety typica.
2 T.U.Grafeet al. Mutualism between bat and pitcher plant
Biol. Lett.
on January 26, 2011rsbl.royalsocietypublishing.orgDownloaded from

Recently, it has been suggested that several large
montane Nepenthes also have reduced abilities to
catch arthropod prey and obtain most of their nitrogen
critical for growth from treeshrew (Tupaia montana)
faeces [15,16]. Our study shows that the faeces-
trapping syndrome occurs not only in montane
Nepenthes but also in pitcher plants growing in lowland
forests, suggesting that low arthropod densities at
mountain sites cannot be the sole factor responsible
for the origin and maintenance of this nitrogen acqui-
sition strategy. Our study supports recent evidence
that simple modifications of pitcher morphology and
levels of digestive fluid can open up novel nitrogen
acquisition strategies [15]. However, giving up the
insect-capturing strategy completely may be risky
from an evolutionary perspective, since the probability
of attracting a bat to a particular individual aerial
pitcher was only 22.8 per cent in our study area.
Instead, it should be beneficial for pitcher plants to
pursue a dual strategy by retaining the ability to trap
insects, especially if bats are absent from some habitat
patches. Such conditional asymmetries in interspecific
interactions are common.
Our study shows that the interaction is mutualistic,
with the bat possibly more dependent on the pitcher
plant than vice versa. This mutualistic relationship
seems to be restricted to Borneo, involving one particu-
lar subspecies of K. hardwickii and one variety of
N. rafflesiana. Our study shows that woolly bats consist-
ently use pitchers as daytime roosts. However, since
woolly bats in other areas of Borneo make occasional
use of other pitcher plant species (N. bicalcarata
and N. ampullaria) that are clearly less well suited as
roosts (own observations and A.-M. Seibert 2010,
personal communication), Nepenthes –woolly bat
associations are prime candidate models to study the
evolution of mutualistic relationships with opportunities
to investigate varying degrees of exploitation.
We thank L. Othman, C. Schwitzke and M. Dahmen for
assistance in the field, and M. Struebig for introducing us
to the bats of Borneo. D. W. Davidson, D. S. Edwards,
L. Gaume, T. H. Kunz, P. M. Narins and three anonymous
referees kindly reviewed the manuscript. The University of
Brunei Darussalam (RG/1(105) & RG/1(137)) and the
German Academic Exchange Service (DAAD) funded this
project; the Forestry Department of Brunei Darussalam
granted permits to work in the field.
1 Clarke, C. 2006 Nepenthes of Borneo. Kota Kinabalu,
Malaysia: Natural History Publications (Borneo).
2 Ellison, A. M., Gotelli, N. J., Brewer, J. S., Cochran-Stafira,
D. L., Kneitel, J. M., Miller, T. E., Worley, A. C. & Zamora,
R. 2003 The evolutionary ecology of carnivorous plants.
Adv. Ecol. Res. 33,1–74.(doi:10.1016/S0065-2504(03)
33009-0)
3 Phillipps, A., Lamb, A. & Lee, C. C. 2008 Pitcher plants
of Borneo, 2nd edn. Kota Kinabalu, Malaysia: Natural
History Publications (Borneo).
4 Di Giusto, B., Grosbois, V., Fargeas, E., Marshall, D. J. &
Gaume, L. 2008 Contribution of pitcher fragrance and
fluid viscosity to high prey diversity in a Nepenthes carni-
vorous plant from Borneo. J. Biosci. 33, 121 –136.
(doi:10.1007/s12038-008-0028-5)
5 Moran, J. A., Booth, W. E. & Charles, J. K. 1999 Aspects
of pitcher morphology and spectral characteristics of six
Bornean Nepenthes pitcher plant species: implications
for prey capture. Ann. Bot. 83, 521–528. (doi:10.1006/
anbo.1999.0857)
6 Gaume, L. & Di Giusto, B. 2009 Adaptive significance
and ontogenetic variability of the waxy zone in Nepenthes
rafflesiana.Ann. Bot. 104, 1281– 1291. (doi:10.1093/
aob/mcp238)
7 Moran, J. A. 1996 Pitcher dimorphism, prey composition
and the mechanisms of prey attraction in the pitcher plant
Nepenthes rafflesiana in Borneo. J. Ecol. 84, 515 –525.
(doi:10.2307/2261474)
8 Payne, J. & Francis, C. M. 2005 A field guide to the mam-
mals of Borneo. Kota Kinabalu, Malaysia: The Sabah
Society.
9 Kunz, T. H. & Lumsden, L. F. 2003 Ecology of cavity and
foliage roosting bats. In Bat ecology (eds T. H. Kunz &
M. B. Fenton), pp. 3 –89. Chicago, IL: University of
Chicago Press.
10 Treseder, K. K., Davidson, D. W. & Ehleringer, J. R.
1995 Absorption of ant-provided carbon dioxide and
nitrogen by a tropical epiphyte. Nature 375, 137 –139.
(doi:10.1038/375137a0)
11 West, J. B., Browen, G. J., Cerling, T. E. & Ehleringer,
J. R. 2006 Stable isotopes as one of nature’s ecological
recorders. Trends Ecol. Evol. 21, 408– 414. (doi:10.
1016/j.tree.2006.04.002)
12 Solano, P. J. & Dejean, A. 2004 Ant-fed plants: com-
parison between three geophytic myrmecophytes.
total nitrogen (%)
bat control
(a)
1.9
1.7
1.5
1.3
1.1
0.9
0.7
δ
15
N (% )
bat control faeces
(b)4
2
3
1
0
–1
–2
–3
Figure 2. Resource benefit provided by roosting K. h. hardwickii to N. r. elongata.(a) Box-plot of total foliar nitrogen of pitchers
used as roosts (n¼38) and of control pitchers (n¼17). (b) Box-plot of foliar
d
15
N of pitchers used as roosts (n¼38), control
pitchers (n¼17) and bat faeces (n¼3). The scarce stable isotope
15
N accumulates from one trophic level to the next and thus
stable isotope ratios of
15
N/
14
N(
d
15
N) can be used to indicate the nitrogen source. Faeces from insectivorous bats have a
higher
d
15
N signature than the insects they feed on or the insects that are trapped by the pitcher plant.
Mutualism between bat and pitcher plant T. U. Grafe et al. 3
Biol. Lett.
on January 26, 2011rsbl.royalsocietypublishing.orgDownloaded from

Biol. J. Linn. Soc. 83, 433– 439. (doi:10.1111/j.1095-
8312.2004.00381.x)
13 Kerth, G. 2008 Causes and consequences of
sociality in bats. BioScience 58, 737–755. (doi:10.1641/
B580810)
14 Kunz, T. H. & Hood, W. R. 2000 Parental care
and postnatal growth in the Chiroptera. In
Reproductive biology of bats (eds E. G. Crichton & P. H.
Krutzsch), pp. 415–468. San Diego, CA: Academic
Press.
15 Chin, L., Moran, J. A. & Clarke, C. 2010 Trap geometry
in three giant montane pitcher plant species from
Borneo is a function of tree shrew body size. New
Phytol. 186, 461–470. (doi:10.1111/j.1469-8137.2009.
03166.x)
16 Clarke, C., Bauer, U., Lee, C. C., Tuen, A. A., Rembold,
K. & Moran, J. A. 2009 Tree shrew lavatories: a novel
nitrogen sequestration strategy in a tropical pitcher
plant. Biol. Lett. 5, 632 – 6 35. (doi:10.1098/rsbl.2009.
0311)
4 T.U.Grafeet al. Mutualism between bat and pitcher plant
Biol. Lett.
on January 26, 2011rsbl.royalsocietypublishing.orgDownloaded from