?And then there were three?: highly efficient uptake of potassium by foliar
trichomes of epiphytic bromeliads
Uwe Winkler1,* and Gerhard Zotz1,2
1Functional Ecology Group, Universita?t Oldenburg, D-26111 Oldenburg, Germany and 2Smithsonian Tropical Research
Institute, Balboa, Panama
* For correspondence. E-mail u.winkler@uni-oldenburg.de
Received: 17 February 2010 Returned for revision: 19 April 2010 Accepted: 11 May 2010
?Background and Aims Vascular epiphytes have to acquire nutrients from atmospheric wash out, stem-flow,
canopy soils and trapped litter. Physiological studies on the adaptations to nutrient acquisition and plant utiliz-
ation of nutrients have focused on phosphorus and nitrogen; potassium, as a third highly abundant nutrient
element, has received minor attention. In the present study, potassium uptake kinetics by leaves, within-plant dis-
tribution and nutrient accumulation were analysed to gain an improved understanding of physiological adap-
tations to non-terrestrial nutrient supply of plants.
?Methods Radioactively labelled 86RbCl was used as an analogue to study uptake kinetics of potassium absorbed
from tanks of epiphytes, its plant distribution and the correlation between uptake efficiency and abundance of
trichomes, functioning as uptake organs of leaves. Potassium in leaves was additionally analysed by atomic
absorption spectroscopy to assess plant responses to potassium deficiency.
?Key Results Labelled rubidium was taken up from tanks over a wide range of concentrations, 0.01?90 mM,
which was achieved by two uptake systems. In four tank epiphytes, the high-affinity transporters had average
Km values of 41.2 mM, and the low-affinity transporters average Km values of 44.8 mM. Further analysis in
Vriesea splenriet showed that high-affinity uptake of rubidium was an ATP-dependent process, while low-affinity
uptake was mediated by a K+-channel. The kinetic properties of both types of transporters are comparable with
those of potassium transporters in roots of terrestrial plants. Specific differences in uptake velocities of epiphytes
are correlated with the abundance of trichomes on their leaf surfaces. The main sinks for potassium were fully
grown leaves. These leaves thus function as internal potassium sources, which allow growth to be maintained
during periods of low external potassium availability.
?Conclusions Vascular epiphytes possess effective mechanisms to take up potassium from both highly diluted
and highly concentrated solutions, enabling the plant to incorporate this nutrient element quickly and almost
quantitatively from tank solutions. A surplus not needed for current metabolism is stored, i.e. plants show
luxury consumption.
Key words: Epiphytic bromeliads, potassium uptake, luxury consumption, plant nutrition, Vriesea splenriet.
INTRODUCTION
Vascular plants that grow epiphytically in tree crowns cannot
directly trap terrestrial nutrient pools, but have to acquire nutri-
ents from atmospheric sources in precipitation, stem flow,
throughfall, and from canopy soils and litter trapped in
tanks. Nutrient supply from these sources is generally low
and highly intermittent (Benzing and Renfrow, 1974; Clark
et al., 1998), a situation that may be directly linked to morpho-
logical peculiarities such as the velamen radicum in orchids or
bromeliad tanks with foliar uptake by absorbing trichomes
(Benzing, 1990). Remarkably, more than a century after the
early studies on epiphyte ecophysiology by Schimper (1888),
the physiological properties of the uptake structures of
bromeliads are still not well understood. For example, it was
demonstrated only recently that bromeliad tanks possess
membrane transporters for a number of different N-sources
(Inselsbacher et al., 2007), which should enable plants to
take advantage of the diversity of nutrient sources delivered
by the metabolism of tank organisms (Benzing, 1970;
Inselsbacher et al., 2007). Membrane transporters for
N-sources (Inselsbacher et al., 2007), but also those for phos-
phate (Winkler and Zotz, 2009), are characterized by high sub-
strate affinities and uptake velocities. Although there is indirect
evidence for effective uptake of potassium as well (Richardson
et al., 2000; Lin and Yeh, 2008), the physiological and bio-
chemical properties of K+-transporters in trichomes have not
yet been analysed, although next to N, potassium is usually
the second most abundant nutrient element in plant tissues.
Unlike most other plant nutrients, K+ is not a constituent of
biomolecules and is highly mobile in plants. In the cytoplasm,
potassium acts as a counter ion to the large excess of nega-
tively charged proteins and nucleic acids. Most of the cellular
potassium is located in vacuoles, functioning as a principal
vacuolar osmoticum. The potassium concentration (?[K+])
in this cell compartment can be highly variable, reflecting
the potassium status of the plant ? in times of sufficient
supply the vacuolar pool is stocked up, while under limiting
conditions the vacuolar potassium store is depleted to sustain
metabolic functions in the cytoplasm. Such ?functional
storage? periodically occurs in perennial trees (Rosecrance
et al., 1998). In accordance with its significance for plant
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growth, uptake of potassium by roots is a well-regulated and
efficient process. Membrane transporters for potassium were
first characterized by their kinetic properties (La?uchli and
Epstein, 1970). In roots of terrestrial plants there is a biphasic
uptake system, consisting of high-affinity transporters with Km
values of 10?50 mM and low-affinity transporters with Km
values of 10?80 mM (Rodriguez-Navarro, 1999; Szczerba
et al., 2009). With respect to the mechanism of transport,
three types of membrane transporters have been identified in
terrestrial plants. Transport can be mediated by secondary
active K+ : H+ symport, K+ : H+ antiport or diffusion via
K+ channels (Lebaudy et al., 2008; Szczerba et al., 2009).
In contrast, based on numerous studies using plant roots, evi-
dence for foliar uptake of potassium is restricted to carnivorous
pitcher plants (Adlassnig et al., 2009).
Because of its experimental advantages, labelled 86Rb is often
used as an analytical analogue for measuring acquisition and
tissue-redistribution of potassium. Pioneering studies using
different tissues have demonstrated that uptake kinetics for K+
and Rb+ are comparable and that both ions compete for the
same membrane transporter (La?uchli and Epstein, 1970;
Rodriguez-Navarro, 1999). Exceptions are skeletal muscles
(D?rup and Clausen, 1994) or excised maize roots (Maas and
Leggett, 1968), where the use of 86Rb+ as a reliable tracer for pot-
assium was questioned. The properties of membrane transporters
of foliar trichomes in bromeliads are not known, and therefore
control experiments to compare uptake of potassium and 86Rb+
were included in the present experiments.
The aims here were three-fold. First, the hypothesis that
leaves of epiphytic bromeliads show highly efficient uptake
of potassium was tested with a detailed analysis of the
uptake kinetics for Rb+. Second, uptake efficiency was
studied in relation to trichome abundance of leaves of different
epiphyte species as an additional step towards a more detailed
understanding of trichome function. Third, the distribution of
potassium within plants after a nutrient pulse was determined
and the long-term effect of K+ supply on plant growth was
studied to quantify the functional consequences of changing
availability of this nutrient element for epiphytes.
MATERIAL AND METHODS
Plant material
Plant material was kindly supplied by a commercial nursery
(Corn. Bak B.V., Asseldelft, Holland: Vriesea splenriet,
Aechmea fasciata, Vriesea duvaliana), or was collected in the
tropical lowland forest in Panama (all other species).
Experiments to characterize rubidium uptake in tanks were
mainly carried out with V. splenriet plants (Bromeliaceae).
Kinetic experiments were also conducted with other tank bro-
meliads, namely A. fasciata, V. duvaliana and field-collected
Werauhia sanguinolenta. All plants listed above had 12?16
leaves, were 12?16 cm tall and had a tank volume of approx.
2 mL. Other plants, which had also been collected from
natural populations, were Guzmania monostachia, Tillandsia
flexuosa, T. fasciculata and T. subulifera. These bromeliads dif-
fered in size as indicated in the figure legends. Additional plants
of the three Tillandsia species were grown from seed in
Oldenburg, which had also been collected in the field. These
plants were ,4 cm tall. Plants were kept in the greenhouse at
25 8C, at a relative humidity of approx. 60 % and a light?dark
regime of 14/10 h, being illuminated by natural sunlight, sup-
plemented with artificial light (Metal halid lamps, 400 W,
master HPI-T plus; Philips, The Netherlands) when necessary
to achieve a photosynthetic photon flux density of at least
150?180 mmol m22 s21 at the level of the plants.
A nutrient solution containing 3.2 mmol L21 NH4NO3,
0.3 mmol L21 KH2PO4, 0.65 mmol L
21 K2HPO4, 0.35 mmol
L21 K2SO4, 0.2 mmol L
21 MgSO4 and 0.34 mmol L
21
CaCl2 was applied by spraying the plants until tanks were
completely filled. Potassium-free nutrient solution was
obtained by omitting K2SO4, while K-phosphates were
replaced by Na-phosphates. Plants were fertilized once a
week, and were otherwise irrigated daily with water.
K+ deficiency experiment. Plants, which had been fertilized before
the experiment, were grown under otherwise identical conditions
in the greenhouse in complete nutrient solution and potassium-
free nutrient solution as described above. As the water content
of the plants remained constant during the 250 d of culture with
and without potassium fertilization, relative growth rate (RGR)
(Evans, 1972) could be calculated by the equation: RGR ? (ln
f. wtfinal ? ln f. wtinitial)/(Dt), where f. wt is plant fresh weight
and Dt is the duration of the experiment in days.
Rubidium uptake
During experiments in the radionuclide laboratories of the
University of Oldenburg, plants were kept in Plexiglas boxes
under conditions comparable with those in the greenhouse.
Plant uptake was measured using aquatic solutions of 86RbCl
(Hartmann Analytic, Braunschweig, Germany) with a specific
activity of 720 MBq mg21 Rb+. Radioactivity of aliquots used
for plant incubation was measured by liquid scintillation
counting, using lume gel save cocktail (Lumac, Rodgau,
Germany) in a Wallac 1415 (Wallac, Uppsala, Sweden)
liquid scintillation counter.
86Rb+ uptake from tanks. Before starting experiments, tanks were
repeatedly thoroughly rinsed with 1 mL of incubation solution,
containing 1 mM Mes-buffer, pH 6.1, and 1 mM CaCl2.
Experiments were started by adding 1?1.5 mL of incubation sol-
ution, 5?10 mL labelled 86RbCl to obtain an initial activity of
2.5 ? 107 d.p.m. mL21, and 10?90 mM RbCl to tanks. Before
sampling, plants were weighed, and weight loss during exper-
iments was compensated for by adding water. After standardiz-
ing the fluid volume in this way, tank fluids were mixed with
Pasteur pipettes and radioactivity of 5-mL aliquots was measured
by liquid scintillation counting. For kinetic measurements,
samples were taken every 30 min up to 6 h and uptake rates of
unlabelled Rb+ were calculated from the initial linear phase of
86Rb+ depletion. Tank depletion obtained for Rb+ at an initial
concentration of 0.5 and 50 mM was compared in separate exper-
iments with the decrease of potassium in tank solutions using the
same starting concentrations and plants of similar size.
Potassium concentrations were measured by atomic absorption
spectroscopy. Comparable rates of tank-depletion of Rb+ and
K+ (Table 1) indicated that Rb+ may indeed be used as tracer
for the study of the acquisition and tissue-redistribution of pot-
assium in these epiphytic plants.
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Uptake of 86Rb+ into plant tissue. Tanks of Vriesea splenriet
were supplied with 0.5 mM RbCl and 86Rb+ to obtain an
initial activity of 2.5 ? 107 d.p.m. mL21. Radioactivity in
tissue was measured from crushed plant material (approx.
0.25 g f. wt) by liquid scintillation counting after 1 and 9 d.
After the initial Rb+ pulse, plants were watered only.
Uptake of 86Rb+ by whole plants. Small plants of 1?5 cm were
transferred to screw cap tubes and submerged in incubation
solutions containing 0.5 mM unlabelled RbCl, and 0.5?
1.0 mL mL21 labelled 86RbCl to obtain an initial activity of
2.5 ? 106 d.p.m. mL21. Depending on plant size, aliquots of
10?40 mL were sampled after 6?12 h to measure radioactivity
by liquid scintillation counting. The concentration of Rb+ used
is regarded as an average value of naturally occurring [K+]
available to bromeliads from rain water, throughfall and stem
flow waters ranging from 0.1 to 0.5 mM (Benzing, 2000;
Richardson et al., 2000).
Uptake of 86Rb+ by leaf discs. Discs of 30 mm diameter were
cut from large leaves and tightened in Plexiglas tubes sealed
with O-rings to prevent uptake by cut tissue. Leaf discs were
overlaid with 2 mL of incubation solution as described above
for uptake by whole plants. Samples of 50 mL were taken
after 6?12 h to measure radioactivity.
Inhibitor experiments. In another set of experiments, concen-
tration dependencies of inhibition of Rb+ uptake of V. splenriet
by 1 mM carbonyl cyanide m-chlorophenylhydrazone (CCCP)
and 1 mM tetraethylammonium chloride (TEA) were analysed.
CCCP is a well-known ATPase inhibitor that is used to separate
active from passive uptake (Dahlmann et al., 2004), whereas
TEA acts as inhibitor of K+-channels. Experiments were started
by filling tanks with 1 mL of incubation solutions as described
under 86Rb uptake from tanks containing additionally 1 mM
CCCP or 1 mM TEA.
All other experimental details regarding nutrient uptake,
measurement of radioactivity in plant tissue and calculation
of uptake rates were as described by Winkler and Zotz (2009).
Other measurements
Trichome numbers for whole plants and leaf discs were
counted under a microscope after tissue-staining with 1 %
methyl blue. Trichome numbers of whole plants were averaged
from counting basal, middle and distal parts of the upper and
lower side of 4?8 leaves. Leaf areas were estimated from
scanned leaf pictures by using the measuring tools in Adobe
Photoshop CS 3 extended. Potassium was determined from
oven-dried (36 h, 85 8C) and milled plant samples. Briefly,
10?20 mg was digested in screw-cap tubes in a 10 : 1 : 4
mixture of nitric, sulfuric and perchloric acid for 8 h at
80 8C (Jackson, 1958), and digests were analysed in a
Perkin-Elmer atomic absorption photometer.
Data analysis
All statistical analyses were performed with R 2.6.0
(R Development Core Team, 2007). Error terms are standard
deviations. The Michaelis constant (Km) and maximal enzyme
velocities (Vmax) were determined via non-linear regression of
the Michaelis?Menten equation for four replicate runs.
RESULTS
In an initial experiment, 1.5 mL of labelled 86Rb+ and 75 nmol
unlabelled Rb+ were filled into tanks of V. splenriet. From the
decrease in radioactivity, it was calculated that tank depletion
decreased with time according to the decreasing concentration
in the tank. About 70 nmol of Rb+ was removed from the
tanks after 28 h (Fig. 1). Even at this low initial Rb+ concen-
tration, probably favouring surface adsorption, about 79+ 8 %
(mean+ s.d., n ? 4) of unlabelled Rb+ was recovered in plant
tissue, as calculated from the radioactivity estimated in the
plant material. This recovery is regarded as being sufficient
to study uptake kinetics and mechanisms for 86Rb+ by its
tank depletion, especially if initial uptake rates are measured.
The concentration dependence of Rb+ uptake, determined
based on the decrease of radioactivity in the tanks at concen-
trations ranging from 0.01 to 90 mM RbCl, indicated a biphasic
uptake system (Fig. 2). A first transporter system was saturated
at Rb+ concentrations of approx. 0.2 mM, and a second one at
Rb+ concentrations that were about two orders of magnitude
higher. Both uptake systems also differed in maximal uptake
rates (Table 2). The high-affinity transport system was charac-
terized by a low maximal uptake velocity, whereas the low-
affinity system was able to accumulate high amounts of Rb+
very rapidly. Applying the Michaelis?Menten plot, Km
values of 41.3+ 8.7 mM and 56.5+ 13.7 mM were calculated
TABLE 1. Depletion of potassium and rubidium from tanks of
Vriesea splenriet (n ? 4)
Depletion from tanks (mmol h21 g21
d. wt mL21 tank volume)
Nutrient concentration (mM) Potassium Rubidium
0.5 0.25+0.019 0.23+0.015
50 4.03+0.44 4.25+0.32
Means were not significantly different (t-tests, t . 0.83, P . 0.05).
Time (h)
0 5 10 15 20 25
R
ub
id
iu
m
 in
 th
e 
ta
nk
  (n
mo
l 1
?5 
mL
?
1 )
0
20
40
60
80
Recovery in tissue: 79 ? 8 %
y = 2?8 (1?e?2?9 ? 104x)
r 2 = 0?99
FI G. 1. Time course of rubidium depletion from tanks of Vriesea splenriet.
Tanks were filled with a mixture of 2.5 ? 107 d.p.m. 86Rb+ and 75 nmol
unlabelled RbCl in 1.5 mL Mes-buffer, 1 mM, pH 6.1, containing 1 mM
CaCl2. Tissue recovery was calculated from the label reanalysed in plant
tissue. Data are means+ s.d.; n ? 4. The regression equation is indicated.
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for the two types of transporters, respectively. Kinetic proper-
ties obtained for Rb+ uptake in three other tank bromeliads
were comparable with those of V. splenriet (Table 2).
The physiological mechanisms behind low- and high-
affinity Rb+ uptake were studied with well-known inhibitors
in V. splenriet. Rates for Rb+ uptake in the low molecular
range of substrate decreased in the presence of the ATPase
inhibitor CCCP (Fig. 3). At concentrations above approx.
10 mM, uptake of RbCl was inhibited by the K+-channel
inhibitor TEA. Saturation of inhibition with TEA was obtained
above approx. 30 mM RbCl.
The relationship between nutrient uptake rates and trichome
density (trichomes mm22) was analysed for a range of plant
species covering different plant habits, i.e. tanks and atmos-
pherics, and different plant sizes. All juvenile plants were of
the atmospheric habit, and larger plants were either atmos-
pherics (e.g. Tillandsia subulifera, T. flexuosa), or had a tank
(e.g. V. splenriet, Werauhia sanguinolenta). Figure 4 shows
the positive, albeit non-linear, relationship between trichome
density and uptake rate for Rb+ (r2 ? 0.86).
The subsequent fate of acquired Rb+ within a plant was ana-
lysed by subjecting V. splenriet to a radioactivity pulse, fol-
lowed by supply with distilled water only (Fig. 5). Irrespective
of incubation time, most leaves and roots received only a
small portion of the incorporated Rb+. One day after labelling,
86Rb+ was only found in tissue that had direct contact to the
tank solution or was adjacent to these leaves, i.e. in several
actively growing leaves and the stem. Nine days after the
pulse, more than 70 % of the 86Rb+ was found in just two
actively growing leaves (G13 and G14 in Fig. 5), while 86Rb+
had decreased substantially in all other leaves and the stem.
Radioactive Rb+ may be used as a potassium analogue in
uptake processes but does not necessarily replace potassium
functionally. Therefore, foliar [K+] was analysed directly
(Fig. 6). Plants of V. splenriet had been cultivated in the green-
house under different K+ fertilization treatments, but other-
wise identical conditions, for 250 d. In plants receiving
fertilizer including potassium, [K+] increased only in the
younger, fully developed leaves (A8?A12) and the younger,
actively growing leaves (G13?G16). The highest [K+] was
found in the same leaves that had accumulated most of the
labelled Rb+ in the short-term experiment (G13 and G14;
Fig. 5). Long-term K starvation hardly affected [K+] in
mature leaves (A1?A7). Arguably, the observed [K+] of
4.2+ 1.2 mg K+ g21 d. wt in older adult leaves (A1?A4) of
starved and fertilized plants represents the minimum [K+]
necessary to sustain metabolic functions of this leaf type. In
younger adult and growing leaves, a culture time of 250 d
may still have been too short to reach this minimum, explain-
ing why K+ deficiency had no significant effect on RGR.
As the potassium content of the entire plant did not significantly
decrease during the 250 d without fertilization with potassium
(7.7+1.4 mg K+ at the start, and 8.1+0.7 after 250 d of fertili-
zation, and 7.9+1.1 mg K+ after 250 d of potassium starvation;
n ? 4), decreasing [K+] in the growing leaves must be related to
an increasing biomass of these leaves, and potassium must have
been transferred from the younger mature leaves to the growing
leaves to maintain growth. Notably, potassium starvation over a
period of 250 d did not reduce growth rates of Vriesea plants com-
pared with fertilized plants. Indeed, cumulative RGRs were even
slightly higher: 4.0+0.3 ? 1023 d21 versus 3.6+0.3 ? 1023
d21 (t-test, t ? ?2.3, P ? 0.04).
DISCUSSION
In many epiphytic bromeliads, roots serve only as hold-fast,
their original functions in water and nutrient uptake being
adopted by leaves with absorbing trichomes (Benzing, 2000).
Together with the findings of earlier studies on N
(Inselsbacher et al., 2007) and P uptake (Winkler and Zotz,
2009), the present results provide compelling evidence that
these trichomes and the fine roots of terrestrial plants, although
very different in morphology and anatomy, share comparable
biochemical properties and uptake mechanisms. Highly effi-
cient and biphasic uptake systems for potassium in all
studied species of tank bromeliads are characterized by high-
Lo
g 
up
ta
ke
 ra
te
 (?
mo
l h?
1  
g?
1  
d.
 w
t)
Rubidium chloride (mM)
10 30 50 70 900?5 1?00
0?001
0?01
1
0?1
10
y  = 9?77 (1?e?5?64 ? 10?3x)
r2 = 0?98
y  = 2?87 (1 ?e?1?2 ? 106x)
r 2 = 0?98
FI G. 2. Biphasic kinetics of rubidium uptake from tanks of Vriesea splenriet
in the presence of 0.01?90 mM substrate. Experimental detail as in Fig. 1
except that different concentrations of unlabelled RbCl were used as indicated
in the diagram. Uptake rates were calculated from the decrease of 86Rb+ radio-
activity during the first 1?2 h of uptake. Data are means+ s.d.; n ? 4. The
regression equations for the low and high concentration ranges are indicated.
TABLE 2. Kinetic properties of rubidium uptake in tanks of different tank bromeliads in low and high concentration ranges
,1 mM rubidium .10 mM rubidium
Species Km (mM) Vmax (nmol h
21 g21 d. wt mL21) Km (mM) Vmax (mmol h
21 g21 d. wt mL21)
Vriesea splenriet 41.3+8.7 14.9+1.3 56.5+13.7 5.1+0.9
Vriesea duvaliana 32.4+14.2 21.8+5.1 25.0+12.1 18.2+2.2
Werauhia sanguinolenta 38.3+5.2 40.2+14.9 ? ?
Aechmea fasciata 52.4+10.6 24.8+4.8 52.8+12.1 18.7+3.8
Data are means+ s.d. (n ? 4); ? , not determined.
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affinity transporters with Km values ranging from 32 to 52 mM
and by low-affinity transporters with Km values between 25
and 57 mM. These values compare favourably with those
obtained from fine roots of terrestrial plants, in which Km
values of high-affinity transporters range from 20 to 50 mM
and those of low-affinity transporters from 10 to 80 mM
(Rodriguez-Navarro, 1999; Szczerba et al., 2009). Inhibitor
studies with V. splenriet suggest that the high-affinity uptake
is ATP dependent, indicating a secondary active transporter.
The low-affinity uptake of K+ in this bromeliad was not
affected by the ATPase inhibitor, but uptake rates were
reduced by inhibitors of K+-channels. Vmax also differed in
these biphasic uptake systems: active uptake allowed only
low uptake velocities, whereas diffusion through membrane
channels was a much faster process, as long as a sufficiently
large concentration gradient existed. All these characteristics
fully agree with the uptake systems for K+ found in roots.
Rubidium chloride (mM)
In
hi
bi
tio
n 
of
 ru
bi
di
um
 u
pt
ak
e 
(%
)
0
20
40
60
80
100
Inhibition by CCCP
0 1 2 3 10 20 30 40 50
Inhibition by TEA
FI G. 3. Inhibition of rubidium uptake from tanks of Vriesea splenriet by car-
bonyl cyanide m-chlorophenylhydrazone (CCCP) and tetraethylammonium
chloride (TEA) at different Rb+ concentrations. Uptake rates are expressed
as percentage inhibition by CCCP or TEA compared with a control without
inhibitors. Tanks were supplied with 2.5 ? 107 d.p.m. 86Rb+ and 1.5 mL
Mes-buffer, 1 mM, pH 6.1, containing 1 mM CaCl2, rubidium chloride as indi-
cated and either 1 mM CCCP or 1 mM TEA. Other experimental details are as
described in Fig. 1. Data are means+ s.d.; n ? 4. The regression equations for
inhibition by TEA and by CCCP are: y?2.87 (1?e21.2?106x), r2?0.98;
y?6.07 (1?e20.13x), r2?0.97.
Tissue (Leaf and other types)
A1 A2 A3 A4 A5 A6 A7 A8 A9A1
0
A1
1
A1
2
G1
3
G1
4
G1
5
G1
6
Ste
m
Ro
ots
R
ub
id
iu
m
 in
 ti
ss
ue
 (n
mo
l)
0
20
40
60
80
100
120
140
160
After 1 d
After 9 d 
FI G. 5. Distribution of labelled rubidium in leaves, stems and roots of Vriesea
splenriet. Experimental details for the uptake from tanks as described in Fig. 1,
except that tanks were supplied with 0.5 mM unlabelled RbCl. After complete
uptake of Rb+, tanks were watered only. After 1 and 9 d, plants were harvested
and cut into small sections. Amounts of rubidium were calculated from the
label in these sections. Leaves are numbered according to their age (A1 ?
oldest, non-senescent leaf ). During ontogenetic development, the length of
successive leaves is increasing in Vriesea. Thereby, fully grown adult leaves
(A1?A12) can be distinguished from growing leaves (G13?G16) by compar-
ing the length of leaves. Data are means+ s.d.; n ? 4.
A1
 
A2 A3 A4
 A5 A6
 
A7 A8
 
A9
 
A1
0
A1
1
A1
2
G1
3 
G1
4 
G1
5 
G1
6 
Po
ta
ss
iu
m
 in
 le
av
es
 (m
g g
?
1  
d.
 w
t)
0
5
10
15
20
25
30 + Potassium
? Potassium
Succession of leaves
FI G. 6. Distribution of potassium in leaves of Vriesea splenriet after 250 d of
growth with and without potassium. Plants, which had been fertilized before
the experiment, were grown under otherwise identical conditions in the green-
house. Leaves are numbered according to their age. A1 was the oldest adult
and A12 the youngest fully developed leaf. G13?G16 are still growing
leaves. Data are means+ s.d.; n ? 4.
WrS
0 10 20 30 40 50
R
b+
 
u
pt
ak
e 
ra
te
 (n
mo
l c
m?
2  
6 h
?
1 )
0
2
4
6
8
10
12
14
16
VrL
GzS
GzL
GzM
VrT
TsM
WrL
TfL
TxS
Trichome density (mm?2)
TsS
TfS
FI G. 4. Relationship between rubidium uptake and trichome density in differ-
ent bromeliads. All plants except large plants of Guzmania monostachia and
Werauhia sanguinolenta were measured completely submerged in label sol-
ution containing 0.5 mM rubidium chloride and 1 ? 107 d.p.m. mL21 86Rb+.
Trichome densities were averaged by counting basal, middle and distal parts
of the upper and lower side of 4?8 leaves. Uptake rates for large plants
from Guzmania and Werauhia were measured in the same label solution
using discs prepared from basal, middle and distal sections of upper and
lower sides of older and younger leaves. Trichome density is given as
number per mm22. Abbreviations: VrL ? Vriesea splenriet (plants of the
same size as used in other experiments); VrT ? Vriesea splenriet, basal seg-
ments of tank leaves (plants of the same size as used in other experiments);
GzS ? Guzmania monostachia, plant size 2?4 cm; GzM ? Guzmania monosta-
chia, plant size 8?12 cm; GzL ? Guzmania monostachia, plant size 22?
25 cm; WrS ? Werauhia sanguinolenta, plant size 2?4 cm; WrL ? Werauhia
sanguinolenta, plant size 16?18 cm; TfS ? Tillandsia fasciculata, plant size
1?3 cm; TfL ? Tillandsia fasciculata, plant size 14?18 cm; TsS ? Tillandsia
subulifera, plant size 1?3 cm; TsM ? Tillandsia subulifera, plant size 8?
12 cm; TxS ? Tillandsia flexuosa, plant size 2?4 cm. The regression equation
is: y?7.45x2103 (1?e22?8?1024x), r2?0.99.
Winkler & Zotz ? Potassium uptake by bromeliad tanks Page 5 of 7
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In agreement with findings for K+, uptake of P (as phosphate)
and N (in various nitrogen compounds) in bromeliads is similarly
mediated by secondary active transporters (Inselsbacher et al.,
2007; Winkler and Zotz, 2009) and this type of transport is also
common in roots (Szczerba et al., 2009). Taken together, the
current evidence suggests that fine roots and trichomes share
transporters with comparable substrate affinities and transport
mechanisms. In situ, nutrient solutions are believed to be highly
diluted (Benzing, 1990), and nutrients can ? in general ? only
be taken up by the high-affinity transporters, while K+ channels
may allow the plants to take advantage of occasional, high exter-
nal concentrations of K+, for example when larger plant or animal
parts decompose in a tank. Unfortunately, our understanding of
the nutrient fluxes in the canopy is too rudimentary to put these
speculations to the test.
Nutrient transfer into leaf tissue not only depends on the
biochemical properties of membrane transporters, but also on
their abundance. This abundance, in turn, should correlate
with the density of trichomes, which are composed of dead
wing cells, covering the leaf surface to suck in water and nutri-
ents, and of central stalk cells. These stalk cells are alive and
their membranes constitute the supposed metabolically active
site in nutrient transfer into the plant (Benzing, 1970;
Benzing and Pridgeon, 1983). Trichome density alone explains
a surprisingly large proportion of the interspecific and intra-
specific variation in the uptake rates of K+. The non-linear
relationship between trichome number and the uptake rates
of K+ suggests that the density and/or the specific type of
active transporter in the cell membranes of the stalk cells
may vary among species and or ontogenetic stage.
Are epiphytes limited by the supply of K under natural con-
ditions? A survey of published data of the foliar [K+] in field-
collected epiphytic bromeliads revealed an average [K+] of
1.1+ 0.6 % g21 d. wt (Table 3), which is close to the
general minimum requirement for eutrophic vegetation (1 %
K; Epstein, 1972). In epiphytes K+ is recycled before leaf
abscission to a considerable degree (approx. 40 % of green
leaf concentrations), comparable with N (35 %) and P (62 %)
(Zotz, 2004). This may indicate that K+ is indeed in short
supply. In contrast, at least in the time frame of weeks or
months, increased supply of K does not stimulate growth.
This finding of the present study is consistent with the
results of a greenhouse study with the tank-bromeliad
Guzmania lingulata, in which potassium fertilization did not
lead to higher final plant dry weight either (Lin and Yeh,
2008). The most likely explanation for these surprising
results is an accumulation of K+ in the vacuoles beyond the
immediate metabolic needs for this element in protein syn-
thesis, enzyme reactions or stomatal regulations, during
times when availability of K+ is high. Because the most pro-
minent role of K+ is its osmotic function, where K+ can be
(partly) replaced by other cations such as Mg2+, Na+ or
Ca2+, internal transfer to growing organs during periods of
insufficient supply may allow these bromeliads to keep up
growth for a considerable amount of time. In turn, efficient
uptake when K+ is highly available can be considered a
type of luxury consumption (Rosecrance et al., 1998;
Gierth and Ma?ser, 2008), similar to, for example, the situation
in P, where storage forms (phytin) exist (Winkler and Zotz,
2009).
TA
B
L
E
3.
P
ub
li
sh
ed
po
ta
ss
iu
m
co
nc
en
tr
at
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ns
of
16
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n
sp
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of
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. 1
6
+
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%
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w
t,
m
ea
n
+
s.
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)
Sp
ec
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s
%
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V
ar
ia
ti
on
(%
K
)
Si
te
de
sc
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pt
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n
So
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A
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hm
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ca
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is
1.
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m
ai
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ow
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1.
57
0.
42
(s
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.,
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na
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re
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Pi
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.
(2
00
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uz
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a
1.
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ow
(1
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ac
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(s
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.,
n
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na
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et
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lia
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.(
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or
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1.
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(s
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(1
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(0
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;
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ri
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ra
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ra
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(s
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(2
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Pe
rc
en
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K
re
pr
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ts
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ea
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or
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ar
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ti
on
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gi
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s.
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fo
r
n
re
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ic
at
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w
ith
in
a
st
ud
y,
or
as
ra
ng
e
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he
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om
se
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no
t
av
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bl
e.
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There is further evidence that the laboratory results are of
ecological relevance. In seven species of tank bromeliads, col-
lected in the Sao Paulo state forest, [K+] of old, mature and
young leaves was not significantly different in all but one
species (Elias et al., 2008). The distribution of leaf-potassium
in most of these naturally grown bromeliads resembled the
within-plant distribution in starved glasshouse cultures of
V. splenriet and thereby probably reflects the low and intermit-
tent nutrient supply in situ. In contrast to actively growing
leaves, [K+] in old leaves of V. splenriet did not decrease
after potassium starvation. As these leaves showed no sign of
senescence, [K+] of approx. 4 mg K+ g21 d. wt possibly reflects
the minimum value necessary to sustain metabolic functions in
this species. In the field-grown bromeliads analysed by Elias
et al. (2008), [K+] varied from 0.7 to 3.9 g21 mg d. wt in old
leaves, possibly indicating species-specific minimal potassium
demand. Different minimal concentrations and functional
storage could explain much of the interspecific variation in
[K+] in bromeliads. These values therefore are not suitable par-
ameters to predict the potassium status of these plants. To deter-
mine a possible potassium limitation in naturally growing tank
bromeliads, comparison of potassium contents between old
and growing leaves seems to be a more appropriate method.
In conclusion, the results of the present experiments shed
new light on the physiological basis that allows tank epiphytes
to cope with the low and intermittent nutrient supply in situ.
Characterization of the uptake kinetics of the major nutrient
elements, nitrogen (Inselsbacher et al., 2007), phosphorus
(Winkler and Zotz, 2009) and potassium (present study),
clearly demonstrate that leaf trichomes are as efficient as
fine roots in their function as uptake organs. Finally, the
results should also be relevant for other plant groups that use
similar strategies for nutrient acquisition via leaf glands,
such as carnivorous plants (Ellison, 2006).
ACKNOWLEDGEMENTS
We thank the Republic of Panama for making its natural
resources available for study (SEX/AP-1-09) and Corn. Bak
B.V., Asseldelft, the Netherlands, for the supply of
V. splenriet plants. We acknowledge the skilful help and excel-
lent assistance of Peter Kanje and Brigitte Rieger in perform-
ing the experiments.
LITERATURE CITED
Adlassnig W, Steinhauser, Peroutka M, et al. 2009. Uptake of potassium,
iron and manganese by carnivorous pitcher plants. Applied Radiation
and Isotopes 67: 2117?2122.
Benzing DH. 1970. Availability of exogenously supplied nitrogen to seedlings
of Bromeliaceae. Bulletin of the Torrey Botanical Club 97: 154?159.
Benzing DH. 1990. Vascular epiphytes. General biology and related biota.
Cambridge: Cambridge University Press.
Benzing DH. 2000. Bromeliaceae. Profile of an adaptive radiation.
Cambridge: Cambridge University Press.
Benzing DH, Pridgeon AM. 1983. Foliar trichomes of Pleurothallidinae
(Orchidaceae): functional significance. American Journal of Botany 70:
173?180.
Benzing DH, Renfrow A. 1971. Significance of the patterns of CO2 exchange
to the ecology and phylogeny of the Tillandsioideae (Bromeliaceae).
Bulletin of the Torrey Botanical Club 98: 322?327.
Benzing DH, Renfrow A. 1974. The mineral nutrition of Bromeliaceae.
Botanical Gazette 135: 281?288.
Caballero-Rueda LM, Rodr??guez N, Martin C. 1997. Dina?mica de elemen-
tos en ep??fitos de un bosque altoandino de la Cordillera Oriental de
Colombia. Caldasia 19: 311?322.
Clark KL, Nadkarni NM, Gholz HL, Schaefer DS. 1998. Atmospheric
deposition and net canopy in a tropical montane forest. Journal of
Tropical Ecology 14: 27?45.
Dahlmann L, Persson J, Palmquist K, Na?sholm T. 2004. Organic and inor-
ganic nitrogen uptake in lichens. Planta 129: 459?467.
D?rup I, Clausen T. 1994. 86Rb is not a reliable tracer for potassium in skel-
etal muscle. Biochemical Journal 302: 745?751.
Elias C, Fernandes EAN, Franca EJ. 2008. Chemical changes in bromeliad
leaves at vegetative stages. Journal of Radioanalytical Nuclear Chemistry
282: 111?115.
Ellison AM. 2006. Nutrient limitation and stoichiometry of carnivorous plants.
Plant Biology 8: 740?747.
Epstein E. 1972. Mineral nutrition of plants: principles and perspectives.
New York: John Wiley and Sons.
Evans GC. 1972. The quantitative analysis of growth. Oxford: Blackwell
Scientific Publications.
Gierth M, Ma?ser P. 2008. Potassium transporters in plants?Involvement in K+
acquisition, redistribution and homeostasis. FEBS Letters 581: 2348?2356.
Husk GJ, Weishampel E, Schlesinger WH. 2004. Mineral dynamics in
Spanish moss, Tillandsia usneoides L. (Bromeliaceae), from Central
Florida, USA. Science of the Total Environment 321: 165?172.
Inselsbacher E, Cambui CA, Richter A, Stange CF, Mercier H, Wanek W.
2007. Microbial activities and foliar uptake of nitrogen in the epiphytic
bromeliad Vriesea gigantea. New Phytologist 175: 311?320.
Jackson ML. 1958. Soil chemical analysis. Englewood Cliffs, NJ: Prentice-Hall.
La?uchli A, Epstein E. 1970. Transport of potassium and rubidium in plant
roots: the significance of calcium. Plant Physiology 45: 639?641.
Lebaudy A, Very A-A, Sentenac H. 2008. K+ channel activity in plants:
genes, regulations and functions FEBS Letters 581: 2357?2366.
Lin C-Y, Yeh D-M. 2008. Potassium nutrition affects leaf growth, anatomy
and macroelements of Guzmania. Hort Science 43: 146?148.
Maas EV, Leggett JE. 1968. Uptake of 86Rb and K by excised maize roots.
Plant Physiology 43: 2054?2056.
Pagano SN, Sartori AA. 1980. Annual variation of nitrogen phosphorous and
potassium in the leaves of 2 epiphytic Bromeliaceae. Revista Brasileira de
Biolog??a 40: 25?30.
Pierce S, Winter K, Griffiths H. 2002. The role of CAM in high rainfall
cloud forests: an in situ comparison of photosynthetic pathways in
Bromeliaceae. Plant, Cell and Environment 25: 1181?1189.
R Development Core Team. 2007. R: A language and environment for stat-
istical computing. Vienna: R Foundation for Statistical Computing.
Richardson BA, Rogers C, Richardson MJ. 2000. Nutrients, diversity, and
community structure of two phytotelm systems in a lower montane
forest, Puerto Rico. Ecological Entomology 25: 348?356.
Rodriguez-Navarro A. 1999. Potassium transport in fungi and plants.
Biochimica et Biophysica Acta 1469: 1?30.
Rosecrance RC, Weinbaum SA, Brown PA. 1998. Alternate bearing affects
nitrogen, phosphorus, potassium, and starch storage pools in mature pis-
tachio trees. Annals of Botany 82: 463?470.
Schimper AFW. 1888. Die epiphytische Vegetation Amerikas. Jena: Gustav
Fischer Verlag.
Schlesinger WJ, Marks L. 1977. Mineral cycling and the niche of Spanish
moss, Tillandsia usneoides L. American Journal of Botany 64:
1254?1262.
Szczerba MW, Britto DT, Kronzucker HJ. 2009. K+ transport in plants:
physiology and molecular biology. Journal of Plant Physiology 166:
447?466.
Winkler U, Zotz G. 2009. Highly efficient uptake of phosphorus in epiphytic
bromeliads. Annals of Botany 103: 477?484.
Zotz G. 2004. The resorption of phosphorus is greater than that of nitrogen in
senescing leaves of vascular epiphytes from lowland Panama. Journal of
Tropical Ecology 20: 639?696.
Zotz G, Richter A. 2006. Changes in carbohydrate and nutrient contents
throughout a reproductive cycle indicate that phosphorus is a limiting
nutrient in the epiphytic bromeliad, Werauhia sanguinolenta. Annals of
Botany 97: 745?754.
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 at sm
ithsonia7 on June 11, 2010 
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