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Wou
ter Patberg Solute tran
sport in
Sphagnum
dom
inated
bogs T
he ecop
hysiological eff
ects of mixin
g by convective fl
ow
Wouter Patberg
Solute transport in Sphagnum dominated bogsThe ecophysiological effects of
mixing by convective flow
Solute transport in Sphagnum dominated bogsThe ecophysiological effects of
mixing by convective flow
Wouter Patberg
Colophon
Graphic design – Mirjam Patberg
Pictures – Wouter Patberg
Cover: Diepveen, Dwingelderveld
page 116: Veerles Veen, Dwingelderveld
Printing – Grafimedia, Facilitair bedrijf RuG
The research reported in this thesis was carried out at the Laboratory
of Plant Physiology, which is part of the Centre of Ecological and
Evolutionary Studies (CEES) of the University of Groningen,
P.O. Box 11103, 9700 CC Groningen, The Netherlands.
This research was financially supported by ALW grant 815-02-014.
ALW (Earth and life sciences) is part of NWO, the Netherlands
Organization for Scientific Research.
This thesis was printed with financial support from the University
of Groningen.
ISBN book: 978-90-367-5241-1
ISBN print: 978-90-367-5242-8
Solute transport in Sphagnum dominated bogsThe ecophysiological effects of mixing by convective flow
Proefschrift
ter verkrijging van het doctoraat in de
Wiskunde en Natuurwetenschappen
aan de Rijksuniversiteit Groningen
op gezag van de
Rector Magnificus, dr. E. Sterken,
in het openbaar te verdedigen op
vrijdag 16 december 2011
om 11:00 uur
door
Wouter Patberg
geboren op 12 april 1976
te Hoorn
RIJKSUNIVERSITEIT GRONINGEN
Promotores Prof. dr. J.T.M. Elzenga
Prof. dr. A.P. Grootjans
Copromotor Dr. A. J. P. Smolders
Beoordelingscommissie Prof. dr. R. van Diggelen
Prof. dr. J.G.M. Roelofs
Prof. dr. H. Joosten
Contents
Chapter 1 General introduction 7
Chapter 2 The transport of solutes by buoyancy-driven water flow 17
in a water-saturated Sphagnum layer; laboratory and field evidence
Wouter Patberg, Gert Jan Baaijens, Christian Fritz, Ab Grootjans,
Rodolpho Iturraspe, Alfons Smolders and Theo Elzenga
Chapter 3 Field characteristics of buoyancy-driven water flow 33
and its global occurrence
Wouter Patberg, Erwin Adema, Myra Boers, Gert Jan Baaijens, Christian Fritz,
Ab Grootjans, Rodolfo Iturraspe, Alfons Smolders and Theo Elzenga
Chapter 4 Physiological evidence for internal acropetal transport of nitrogen 45
in Sphagnum cuspidatum and S. fallax
Wouter Patberg, Bikila Warkineh Dullo, Alfons Smolders,
Ab Grootjans and Theo Elzenga
Chapter 5 The importance of groundwater carbon dioxide 57
in the restoration of Sphagnum bogs
Wouter Patberg, Gert Jan Baaijens, Alfons Smolders,
Ab Grootjans and Theo Elzenga
Chapter 6 Photosynthesis of three Sphagnum species after acclimatization 75
to high and low carbon dioxide availability
Wouter Patberg, Jan Erik van der Heide and Theo Elzenga
Chapter 7 Summary and synthesis 91
References 105
Samenvatting 119
Dankwoord 127
Chapter 1
General introduction
General introduction – 9
Background
Bog ecosystems
Bogs are wet, acidic, peat forming ecosystems which generally have a low cover of vascular plants
and are dominated by mosses of the genus Sphagnum (peat moss or ‘veenmos’ in Dutch; Rydin
& Jeglum, 2006). Sphagnum mosses play an important role in creating their own environment,
thereby gaining competitive advantage over other plant species (Kilham, 1982; Malmer et al., 1994;
Van Breemen, 1995).
Sphagnum mosses consist of densely clustered developing and expanding branches at the top
of the plant, the so-called capitulum (plural: capitula), and the stem with fully developed branch-
es. During growth, the stem elongates from the capitulum and the branches become distributed
along the new stem. The lower portion of Sphagnum plants gradually die and will form peat. The
capitulum is the part of the plant where the main production of biomass takes place. Also, the
highest metabolic activity and nutrient uptake in Sphagnum mosses was measured in the capitu-
lum (Aldous, 2002a; Johansson & Linder, 1980; Malmer, 1988; Malmer et al., 1994; Robroek et al.,
2009; Rydin & Jeglum, 2006). For example, the contribution of photosynthetic activity of the ca-
pitulum in dense Sphagnum hummocks and lawns has been estimated to be 98% in Sphagnum
fuscum and 60% in S. balticum (Johansson & Linder, 1980).
Looking at a cross section of a Sphagnum bog, it can be divided into two layers (Clymo, 1984;
Ingram, 1978). The upper 10 to 40 cm is called the acrotelm. This layer contains the living part of
the Sphagnum mosses and is a highly permeable layer where the groundwater table fluctuates. The
spongy acrotelm has a high hydraulic conductivity and the ability to retain water in dryer periods,
thus having a strong self-regulating effect on the depth of the water table (Ingram, 1978). The layer
below is called the catotelm, a slowly permeable, permanently water-saturated anaerobic layer
which contains most of the peat (Ingram, 1978).
Bog ecosystems are ombrotrophic, which means that they receive their nutrients solely by
atmospheric deposition and are therefore characterized by a low nutrient availability (Rydin
& Jeglum, 2006). The ability of Sphagnum mosses to deal with low nutrient availability is often
attributed to their high nutrient retention capacity. The bog surface appears as a layer of densely
packed Sphagnum capitula that efficiently intercept nutrients coming from the atmosphere (Aldous,
2002a; Woodin & Lee, 1987). Sphagnum mosses lack vascular tissue for water and nutrient uptake,
but they can take up water and nutrients over the entire surface of the plant because they lack a cu-
ticle (Brown, 1982; Brown & Bates, 1990). Sphagnum mosses are able to survive under low nitrogen
conditions due to their very efficient nitrogen utilization (Bridgham, 2002; Li & Vitt, 1997) ranging
from 50 to 90% (Aldous, 2002a; Li & Vitt, 1997). Woodin & Lee (1987) even measured a retention
of 100% of inorganic nitrogen at an unpolluted site, whereas chloride and sulphate were passing
freely through the moss mat. That the efficiency of nitrogen retention by Sphagnum results in a
competitive advantage of Sphagnum over vascular plants was shown by Aldous (2002a): vascular
plants received <1% of N recently added by wet deposition. Moreover, the ability of Sphagnum spe-
cies to grow in an environment with very low nutrient concentrations is often attributed to their
pronounced capacity to exchange hydrogen ions for mineral cations (Daniels & Eddy, 1985). By
doing so, releasing H+ ions in exchange for dissolved cations, Sphagnum species have the ability
to acidify their environment (Clymo, 1963; Clymo & Hayward, 1982). Another important feature
10 – Solute transport in Sphagnum dominated bogs
of Sphagnum to survive in nutrient poor habitats is the ability to recycle nutrients from senescent
Sphagnum tissue very efficiently (Aldous 2002a, 2002b; Malmer, 1988; Van Breemen, 1995).
In contrast to vascular plants, Sphagnum mosses lack stomata. Consequently, they cannot control
their water loss actively. Water lost by evaporation must be replaced by rain or by the water from
the peat below. Since there are no specialized cells for water transport in the stem, the upwards
water transport takes place externally by capillary movement facilitated by a network of spaces
between leaves, stems and branches (Rydin & Jeglum, 2006).
Sphagnum mosses are characterized by their high water-holding capacity (Clymo & Hayward,
1982). Much water can be retained in the hyaline cells, which are large, dead cells, which make up
about 80% of the plant’s volume. Hyaline cells can rapidly absorb water through their pores (with
have diameters from 5-20 μm), and water can be retained against suction pressures of 10-100 kPa
(Van Breemen, 1995). The hyaline cells are enclosed in a network of narrower chlorophyllose cells,
the cells that contain chlorophyll and enable the mosses to photosynthesize.
The microtopography of a Sphagnum bog is characterized by a diversity of wet depressions (pools
and hollows), relatively dry lawns and dry hummocks (Andrus et al., 1983). Each microhabitat is
occupied by a different group of Sphagnum mosses, broadly defined by its water retention capacity
(Andrus et al., 1983; Hayward & Clymo, 1982). Sphagnum species observed at increasing height
above the water table have an increasing capacity to conduct water by capillary action (Clymo &
Hayward, 1982).
Due to the wet, anoxic and acidic conditions in the catotelm, the production of Sphagnum
exceeds the decomposition of organic material, resulting in the accumulation of organic material
or peat (Clymo & Hayward, 1982; Clymo et al., 1998). Peatlands store more carbon than any other ter-
restrial ecosystem. It has been estimated that the accumulation of peat has led to a carbon pool that
is about one-third of the global soil carbon pool. This is quite remarkable since peatlands occupy
only about 3% of the world’s total land area (Rydin & Jeglum, 2006). Peatlands, including Sphagnum
bogs, function as a net sink for CO2 (Clymo et al., 1998; Gorham, 1991) and as a consequence, Sphag-
num bogs play an important role in global carbon cycling (Bridgham et al., 2001a; Clymo et al., 1998;
Gorham, 1991).
Due to the extensive exploitation for fuel, agriculture and forestry over many centuries, and
the ongoing global warming, living (peat forming) peatlands have become endangered ecosystems
throughout the world (Rochefort & Price, 2003). Even nowadays (extensive) peat extraction activities
take place for commercial use in, for example, Canada, Scandinavia, Ireland and the Baltic states
(Joosten, 2009). Due to the important role of peatlands in the global carbon cycle, and because
of their unique ecological values, conservation and restoration of these ecosystems is necessary,
preventing stored CO2 being released into the atmosphere, which will lead to accelerated global
warming.
Globally much effort is dedicated to the restoration of damaged peatlands. However, the
restoration of bog remnants in particular, has proven to be fairly complicated and not always
successful (Grootjans et al., 2006; Money & Wheeler, 1999; Money et al., 2009). For the successful
conservation and restoration of Sphagnum-dominated bogs, knowledge about environmental
constrains for Sphagnum growth is necessary.
General introduction – 11
Nutrient supply in Sphagnum bogs
For their nutrient supply, Sphagnum bogs mainly depend on wet and dry atmospheric deposition.
However, it has been shown that under non-polluted conditions the annual input of nutrients
from atmospheric deposition is often insufficient to sustain the observed primary production in
these systems (Aerts et al., 1999; Aldous, 2002a, 2002b; Bowden, 1987; Bridgham, 2002; Damman,
1978, 1986; Morris, 1991; Pakarinen, 1978; Rosswall & Granhall, 1980; Urban & Eisenreich, 1988).
Therefore, other nutrients sources must be involved. Under natural conditions Sphagnum bogs
are often nitrogen-deficient (Aerts et al., 1992; Bragazza et al., 2004; Bridgham et al., 2001b;
Gunnarsson & Rydin, 2000; Li & Vitt, 1997) and therefore the availability of nitrogen is of special
interest. However, Sphagnum growth has also been shown to be limited by phosphorus (Aerts et al.,
1992; Bridgham et al., 1996), potassium (Damman, 1978; Pakarinen, 1978) and carbon dioxide (Rice
& Giles, 1996; Smolders et al., 2001).
Sources of nitrogen include N-fixation by cyanobacteria associated with Sphagnum and other
plants (Gerdol et al. 2006), the internal reallocation of nitrogen from older senescent tissues to the
metabolically active capitula (Malmer, 1988) and the mineralization of senescent Sphagnum plants
at the border of the acrotelm and catotelm. The mineralization of N has been shown to be the most
important nitrogen source for Sphagnum (Aldous, 2002a; Bowden, 1987; Bridgham, 2002; Urban &
Eisenreich, 1988). Gerdol et al. (2006) showed that direct retention of N from precipitation is less
important than recycling of mineralized N to support Sphagnum growth (Aldous, 2002b; Bowden,
1987; Bridgham, 2002; Urban & Eisenreich, 1988). The importance of re-mineralization of nitrogen
for Sphagnum growth has been demonstrated in situ by Urban & Eisenreich (1988). They calculated
the assimilation of nitrogen by plants (primarily Sphagnum) to be 66 kg ∙ ha-1 ∙ yr-1, whereas only
14.6 kg N ∙ ha-1 ∙ yr-1 was supplied by total inputs. The remainder was supplied by mineralization of
the peat.
In Sphagnum bogs the highest mineralization rates are found in the aerobic zone at the transition
zone from acrotelm to the anaerobic catotelm (Bridgham et al., 1998; Malmer, 1993; Waddington et
al., 2001). In contrast, the highest metabolic activity and nutrient uptake in Sphagna takes place in
the upper part of the plant, the capitulum (Aldous, 2002a; Johansson & Linder, 1980; Malmer, 1988;
Malmer et al., 1994; Robroek et al., 2009; Rydin & Jeglum, 2006). The spatial separation between
the actively growing photosynthesizing capitula and the mineralization of nutrients requires an
efficient nutrient transport system. Several nutrient transport mechanisms have been described
for Sphagnum bogs. Throughout the water layer nutrients are passively distributed by diffusion.
Above the water layer solutes might be transported upwards to the capitula through the extracellu-
lar capillary spaces between pendant branches and stems (Hayward & Clymo, 1982). Rydin & Clymo
(1989) demonstrated the internal acropetal transport of carbon and phosphorus. Complementary
to the abovementioned transport mechanisms, Baaijens (1982) and Rappoldt et al. (2003) reported
on a phenomenon called buoyancy-driven water flow as a possible mechanism for the external
transport of nutrients in a water-saturated Sphagnum layer.
Buoyancy-driven water flow
Buoyancy-driven water flow is the vertical convective flow of water in a water-saturated peat moss
layer, driven by the temperature difference between day and night. Due to the temperature drop
during the night the surface of the water layer will cool down, resulting in a relative cold layer on
12 – Solute transport in Sphagnum dominated bogs
top of a warmer layer. Because of the difference in density between these two layers, the cold water
will sink and the warm water will rise. Evidence for the occurrence of buoyancy-driven water flow
in a water-saturated Sphagnum layer, based on theoretical and experimental grounds, was provided
by Rappoldt et al. (2003).
The development of buoyancy flow in a water-saturated Sphagnum layer is determined by the
Rayleigh (Ra) number of that layer. Rappoldt et al. (2003) calculated that buoyancy flow occurs if
the system’s Ra number exceeds 25. For a typical peat moss layer, a temperature difference of 10
degrees between day and night will result in a Ra number of 80 which is suitable for the quick
development of buoyancy flow (Rappoldt et al., 2003). Adema et al. (2006) provided evidence for
the occurrence of buoyancy flow in the field; based on the hydraulic conductivity (k) of the Spha-
gnum layer and a temperature difference between day and night of 8˚C, in a small Sphagnum do-
minated peat bog in the Netherlands, the calculated Ra number was sufficiently high to induce
buoyancy flow.
The convective flow of water will result in the mixing of solutes as well. However, direct
evidence for nutrient transport is lacking. It is hypothesized that nutrients originating from
decomposition in the lower acrotelm, will be transported upwards by buoyancy flow and may
become available for the growing Sphagnum capitula, thereby contributing to the nutrient supply
of the Sphagnum plants. Moreover, oxygen produced by photosynthesis in the upper Sphagnum
layer will be transported downwards resulting in increased decomposition rates. In turn, nutrients
will become available to the growing Sphagnum when transported upwards by buoyancy flow.
Consequently, buoyancy flow might be an essential mechanism in the efficient recycling of
nutrients in Sphagnum bogs.
Aim and outline of this thesis
Part I: Buoyancy-driven water flow as a transport mechanism
The main objective of the first part of this thesis is to determine the importance of buoyancy-driv-
en water flow in the nutrient distribution in Sphagnum bogs. In Chapter 2 (The transport of solutes
by buoyancy-driven water flow in a water-saturated Sphagnum layer; laboratory and field evidence),
we asked ourselves the basic question whether nutrients are indeed transported by buoyancy flow.
To answer this question a straightforward, but effective mesocosm experiment was performed
in a temperature-regulated climate chamber. Buoyancy flow was generated in a water-saturated
Sphagnum matrix and the transport of solutes by buoyancy flow was visualized by the addition
and subsequent monitoring of a coloring dye. It became evident that buoyancy flow can act as a
fast and efficient transport mechanism (Chapter 2). In accordance with Rappoldt et al. (2003), a
reversal of the nutrient gradient due to the occurrence of buoyancy flow was possible and thereby
induce a stepwise increase in the nutrient concentration near the capitula. Consequently, the im-
portance of buoyancy flow as a transport mechanism in supplying the capitula is also determined
by the ability of Sphagnum capitula to enhance the uptake and assimilation by (and thus benefit
from) this increased nutrient availability. The amount of nutrients taken up by Sphagnum depends
on the nutrient concentration and the affinity of the uptake mechanism for the substrate. In the
case of, for example, temporary high ammonium concentrations in the upper Sphagnum layer due
General introduction – 13
to buoyancy-driven water flow, Sphagnum must have a suitable uptake mechanism to benefit op-
timally from the situation. Therefore, we determined the uptake kinetics of ammonium by the
capitula of S. cuspidatum and S. fallax. The possible role of the cation-binding sites in the uptake of
nutrients is taken into consideration as well. In Chapter 2 also the findings of a field experiment,
in which the transport of labeled nitrogen (15N) in a Sphagnum layer by buoyancy-driven water flow
and the subsequent uptake by the Sphagnum capitula are reported.
Whereas the laboratory experiments (Chapter 2; Rappoldt et al., 2003) are all conducted under
controlled conditions and with homogeneous samples, in the field temporal and spatial variation
in temperature and hydraulic conductivity might occur and influence the occurrence and size of
the buoyancy cells. Therefore, in Chapter 3 (Field characteristics of buoyancy-driven water flow and
its global occurrence), a series of vertical temperature profiles were recorded in a pristine Sphagnum
bog to validate the theoretical predictions in a natural situation. Based on the measured hydrau-
lic conductivities and ambient day and night air temperatures, the Ra numbers of the Sphagnum
sites were calculated. Based on these Ra numbers, the occurrence of buoyancy flow in the field
could be predicted using the model described by Rappoldt et al. (2003). Additionally in Chapter 3,
the possible occurrence of buoyancy-driven water flow in Sphagnum bogs throughout the world
was determined. Geographical Information System software was used to analyze worldwide daily
temperature data and model the occurrence of buoyancy flow in peatlands throughout the world.
The importance of buoyancy-driven water flow in the nutrient supply of Sphagnum and nutri-
ent cycling in bogs depends on the transport rate relative to other transport mechanism. To date,
diffusion and internal transport were the known mechanisms by which nutrients were transported
throughout a water-saturated Sphagnum layer. Note that capillary transport is often mentioned as a
nutrient transport mechanism (Hayward & Clymo, 1982), but this type of transport is only possible
above the water table and therefore not taken into account here.
Diffusion and internal transport are both slow processes. For example, the diffusion coefficients
for oxygen and ammonium are respectively 1.96 ∙ 10-5 and 1.95 ∙ 10-5 cm2 ∙ s-1 at 20°C (Boudreau,
1997). Internal transport is estimated to distribute solutes throughout the plant with a half time of
about 11 days, an estimation based on the symplasmic apical transport of 14C (Rydin & Clymo, 1989).
Moreover, in a review on internal transport in non-vascular plants (Raven, 2003) it was stated that
there is no evidence for symplastic transport in Sphagna faster than can be accounted for by dif-
fusion. As a consequence, we expect buoyancy-driven water flow to play an important role in the
nutrient distribution in Sphagnum bogs. In earlier studies (Aldous, 2002b; Bridgham, 2002) the
contribution of translocation to the nitrogen supply of the capitula was shown to be significant.
Although, the ability of Sphagnum to transport nitrogen internally was widely assumed (Bonnett et
al., 2010; Bragazza et al., 2005; Gerdol et al., 2006; Limpens & Berendse, 2003; Limpens & Heijmans,
2008; Malmer, 1988), internal transport of nitrogen had never been demonstrated. The assumption
that N is transported internally is mainly based on the observations of internally transported car-
bon and phosphorus (Rydin & Clymo, 1989). This idea was supported by the often observed higher
C:N ratios in stems than in capitula (e.g. Malmer, 1988) and are taken as an indication for the inter-
nal reallocation of N from the stem to capitula. However, experimental evidence for the internal
reallocation of N was lacking. Chapter 4 (Physiological evidence for internal acropetal transport of
nitrogen in Sphagnum cuspidatum and S. fallax) deals with the contribution of internal transport
in the upward translocation of mineralized nitrogen in Sphagnum bogs. Two Sphagnum species,
14 – Solute transport in Sphagnum dominated bogs
Sphagnum cuspidatum and S. fallax, were used in experiments in which diffusion and capillary
transport were excluded and the internal transport of nitrogen was monitored. For both Sphagnum
cuspidatum and S. fallax, a slow but significant acropetal transport of nitrogen through an internal
mechanism was observed. Moreover, the rate at which nitrogen was transported internally was
estimated and its importance relative to buoyancy-driven water flow, is discussed.
Part II: The importance of carbon dioxide for the growth of Sphagnum
The second part of this thesis focuses on the importance of carbon dioxide for the growth of
Sphagnum. In contrast to vascular plants, Sphagnum mosses lack a cuticle and stomates to regu-
late photosynthesis (Rydin & Jeglum, 2006), but are surrounded by an external water film through
which gas exchange for photosynthesis is taking place. The photosynthetic rate of Sphagnum
mosses has been shown to be a compromise between external water content and the availability
of CO2 (Schipperges & Rydin, 1998; Silvola, 1990; Titus et al., 1983). At low water contents, dehydra-
tion inhibits photosynthesis whereas at very high water contents Sphagnum species may suffer
from carbon limitation due to very thick boundary layers (Jauhiainen & Silvola, 1999; Rice & Giles,
1996; Silvola, 1990; Titus et al., 1983; Williams & Flanagan, 1996). Since the diffusion of CO2 is about
104 times lower in water than in air, external water films can form large barriers for gas exchange,
reducing the supply of CO2 towards the carbon assimilating cells resulting in a reduced photosyn-
thetic rate (Bowes & Salvucci, 1989; Rice & Giles, 1996; Silvola, 1990; Williams & Flanagan, 1996).
To overcome this problem many aquatic plant species make use of carbon concentrating
mechanisms (CCM), which enhances the accumulation of carbon under water (Maberly & Madsen,
2002). The most frequently used mechanism is the use of bicarbonate (HCO3
-) as a carbon source
in photosynthesis (Prins & Elzenga, 1989). Sphagnum mosses lack such a CCM. Like most aquatic
bryophytes (Raven et al., 1985), Sphagnum mosses are known to be obligate CO2 users (Bain &
Proctor, 1980) and are therefore solely dependent on the diffusive supply of CO2 to the site of carbon
fixation. In obligate CO2 users high rates of underwater photosynthesis can only be sustained when
the leaves are exposed to high concentrations of CO2 (Jauhiainen & Silvola, 1999; Silvola, 1990).
Because CO2 is continuously produced by aerobic and anaerobic decomposition processes
dissolved CO2 concentrations are normally much higher in upper peat layers than atmospheric
ones (16 μmol ∙ L-1 vs. 100-5000 μmol ∙ L-1; Bridgham & Richardson, 1992; Lamers et al., 1999; Silvola,
1990; Yavitt et al., 1997; Glatzel et al., 2004; Smolders et al., 2001; Waddington et al., 2001). High CO2
concentrations can compensate for low diffusion rates and ensure the substrate delivery for pho-
tosynthetic carbon fixation to be sufficient (Maberly & Madsen, 2002; Silvola, 1990). The refixation
of CO2 from decomposition processes has been unambiguously demonstrated by Rydin & Clymo
(1989) and Turetsky & Wieder (1999). This so-called substrate-derived CO2 has been shown to be
an important carbon source for aquatic and emergent Sphagnum mosses (Baker & Boatman, 1990;
Paffen & Roelofs, 1991; Riis & SandJensen, 1997; Roelofs, 1983; Smolders et al., 2001; Smolders et al.,
2003). As a consequence, increased ambient atmospheric CO2 concentrations (up to twice ambient)
appeared to have limited effect on the growth of Sphagnum as outlined by Smolders et al. (2001).
Chapter 5 (The importance of groundwater carbon dioxide in the restoration of Sphagnum bogs)
focuses on the importance of CO2 for Sphagnum in a field situation. The study was performed
in the “Dwingelderveld”, a nature reserve in the Netherlands characterized by numerous small,
General introduction – 15
damaged Sphagnum bogs, distributed throughout the area. Since the start of restoration measures,
the developmental success between bogs has varied significantly; some bogs developed well,
whereas others did not. Peat extraction has removed the bulk of organic material and the highly
decomposed, humified peat which is left behind, has only limited CO2 production rates (Bridgham
& Richardson, 1992; Glatzel et al., 2004; Tomassen et al., 2004; Waddington et al., 2001). Therefore,
for the successful restoration of cut-over Sphagnum bogs an additional carbon source might be
essential for the re-establishment of Sphagnum mosses. It is hypothesized that in these hydrologi-
cally degraded bog remnants the restoration of Sphagnum growth is limited by the availability of
CO2. Bog waters analysis showed that the well-developed bogs received C-rich water from outside
the bogs. It was concluded that high CO2 availability is a pre-requisite for the successful re-esta-
blishment of Sphagnum mosses and subsequent bog development. Despite the obvious importance
of a high CO2 availability for Sphagnum, the physiological background of this apparent high CO
2
requirement of Sphagnum has never been established. Therefore, the physiological background of
carbon uptake by Sphagnum was investigated as well.
In Chapter 5 the plants used in the experiments were grown under ambient CO2 conditions.
However, For Sphagnum fuscum, a hummock forming species, acclimation to CO2 levels has
been shown (Jauhiainen & Silvola, 1999). Culturing plant under high CO2 availability resulted in
lower photosynthetic rates compared to plants that were grown under CO2-limiting conditions
(Jauhiainen & Silvola, 1999). Therefore, we expected physiological adaptations in carbon dioxide
uptake in response to the CO2 concentration during the culturing period. Therefore in Chapter
6 (Photosynthesis of three Sphagnum species after acclimatization to high and low carbon dioxide
availability) the physiological characteristics of carbon uptake by three different Sphagnum species
was investigated for plants grown for a long period at high and low CO2 availability.
Chapter 7 (Summary and synthesis) is a combined summary and synthesis of this thesis. The
main focus in this chapter lies on the ecological importance of buoyancy flow in Sphagnum-dom-
inated bogs. For different Sphagnum habitats (hollows, lawns and hummocks) the contribution of
buoyancy flow in the nutrient supply of Sphagnum will be compared to other nutrient transport
mechanisms. Finally, some findings in this thesis will be discussed in relation to the restoration
and conservation of Sphagnum bogs.
Chapter 2
The transport of solutes by buoyancy-driven water flow in a water-saturated Sphagnum layer; laboratory and field evidence
Wouter Patberg
Gert Jan Baaijens
Christian Fritz
Ab Grootjans
Rodolpho Iturraspe
Alfons Smolders
Theo Elzenga
18 – Solute transport in Sphagnum dominated bogs
Abstract
Sphagnum bogs depend for their nutrients mainly on atmospheric deposition.
Yet, the main nutrient source for Sphagnum growth has been shown to be
the mineralization of organic material. The highest mineralization rates are
found in lower peat layers at the border between acrotelm and catotelm.
The highest metabolic activity and nutrient uptake, however, takes place in
the capitula, found at the top of the living Sphagnum layer. This separation
between the actively growing capitula and the site of nutrient mineralization
requires an efficient nutrient transport system. Several nutrient transport
mechanisms in bogs have been described; diffusion, internal transport and
capillary transport. Complementary to these mechanisms, buoyancy-driven
water flow was proposed as an external nutrient transport mechanism in
a water-saturated peat layer. Buoyancy flow is the vertical movement of
water driven by the difference in density between water layers, which is the
result of nocturnal cooling. This chapter shows, by means of a mesocosm
experiment, that solutes are rapidly and in large quantities transported
by buoyancy-driven water flow in a Sphagnum matrix. As a consequence,
nutrient concentrations are stepwise increased around the capitula.
Consequently, the importance of buoyancy flow in the nutrient supply of
Sphagnum is also determined by the nutrient uptake capacity of Sphagnum.
The uptake kinetics of ammonium by Sphagnum indicates that Sphagnum is
able to benefit efficiently from a stepwise increase in ammonium availability.
Therefore, compared to diffusion and internal transport, buoyancy flow
seems to be a quantitatively important transport mechanism for nitrogen in
a water-saturated bog. Additionally, the transport of N by buoyancy-driven
water flow and subsequent uptake by the capitula was shown in a field
situation. Other solutes like carbon dioxide and oxygen will be redistributed
as well by buoyancy flow and this presumably also plays an important role
in ecosystem functioning.
The transport of solutes – 19
Introduction
One of the main characteristics of Sphagnum bogs is the accumulation of peat (Clymo et al.,
1998). Due to the wet and acidic environment in bogs the production of Sphagnum exceeds the
decomposition of organic material, resulting in the accumulation of organic material or peat
(Clymo & Hayward, 1982). Sphagnum is functioning as an ecosystem engineer by contributing to the
creation of this environment (Van Breemen, 1995). Consequently, peatlands function as a net sink
for CO2 (Clymo et al., 1998; Gorham, 1991). Sphagnum dominated bogs receive nutrients mainly by
atmospheric deposition (e.g. Van Breemen, 1995). Sphagnum mosses are able to very efficiently in-
tercept nutrients from precipitation and recycle them (Li & Vitt, 1997; Malmer, 1988; Rudolph et al.,
1993; Woodin & Lee, 1987). Consequently, they are able to dominate these low nutrient ecosystems
(Aldous, 2002a; Li & Vitt, 1997; Malmer, 1988; Van Breemen, 1995).
However, it has been shown that under non-polluted conditions the annual input of nutri-
ents by precipitation is often insufficient to sustain the observed primary production in these
systems (Aerts et al., 1999; Aldous, 2002a, b; Bowden, 1987; Bridgham, 2002; Damman, 1978, 1986;
Morris, 1991; Pakarinen, 1978; Rosswall & Granhall, 1980; Urban & Eisenreich, 1988). The release of
minerals by decomposition of organic material has been shown to be an important nutrient source
to support Sphagnum growth (Aldous, 2002a; Bowden, 1987; Bridgham, 2002; Gerdol et al., 2006;
Urban & Eisenreich, 1988).
The highest mineralization rates are found in the aerobic zone of the acrotelm at the border of
the anaerobic catotelm (Bridgham et al., 1998; Malmer, 1993; Waddington et al., 2001). In contrast,
the highest metabolic activity and nutrient uptake in Sphagna takes place in the upper part of the
plant, the capitulum (Aldous, 2002a; Johansson & Linder, 1980; Malmer, 1988; Malmer et al., 1994;
Robroek et al., 2009; Rydin & Jeglum, 2006). Together, these processes result in a gradient in nutri-
ent concentration, from low in the upper layer of a Sphagnum bog to high in the lower parts were
decomposition takes place. In addition, the oxygen produced by the photosynthesizing capitula in
the top layer and the consumption of oxygen by decomposition processes in deeper acrotelm layer
will result in decreasing oxygen levels with increasing depths (Lloyd et al., 1998; Redinger, 1934).
The spatial separation between the actively growing, photosynthesizing capitula and the
mineralization of nutrients requires an efficient nutrient transport system. Several nutrient
transport mechanisms have been described for Sphagnum bogs. Nutrients are passively distributed
throughout the water layer by diffusion. Above the water layer solutes might be transported
upwards to the capitula through the extracellular capillary spaces between pendant branches and
stems (Hayward & Clymo, 1982). Nutrients are also transported internally by Sphagnum. Rydin &
Clymo (1989) demonstrated the internal acropetal transport of carbon and phosphorus. In Chap-
ter 4 of this thesis evidence for internal transport of nitrogen is provided. Complementary to
these mechanisms, Baaijens (1982) and Rappoldt et al. (2003) reported on a phenomenon called
buoyancy-driven water flow, as a possible external nutrient transport mechanism in a water-
saturated peat moss layer.
Buoyancy flow is the vertical redistribution of water, generated by nocturnal cooling. During
the night the upper water layer cools down, leading to relatively dense cold water on top of warmer
water. If the temperature drop is sufficiently large, the cold water sinks leading to mixing of the
water column. Evidence for the occurrence of buoyancy-driven water flow in a water-saturated
20 – Solute transport in Sphagnum dominated bogs
Sphagnum layer was provided, based on theoretical and experimental grounds, by Rappoldt et
al. (2003). They showed that for a typical peat moss layer a temperature difference of about 10
degrees between day and night will result in a Rayleigh number (Ra) suitable for the development
of buoyancy flow. Adema et al. (2006) provided field evidence for buoyancy-driven water flow in
a Sphagnum dominated peat bog. Hydraulic conductivity and temperature measurements in a
pristine bog in Tierra del Fuego, Argentina, indicated that buoyancy flow events occur regularly.
Moreover, daily air temperature data collected around the world indicate that buoyancy-driven
water flow is very likely to occur on a regular basis in peat lands throughout the world (Chapter 3).
Flow of water will result in the mixing of solutes and this has been proposed to be the most im-
portant ecological consequence of buoyancy-driven water flow (Adema et al., 2006; Rappoldt et al.,
2003). However, direct evidence for nutrient transport is lacking. This study focuses on buoyancy-
driven water flow as a nutrient transport mechanism in a water-saturated Sphagnum layer. It is
hypothesized that nutrients originating from decomposition will be transported upwards and may
become available for the growing capitula of Sphagnum and thereby contributing to the nutrient
supply of the Sphagnum plants.
A mesocosm experiment was set up to trace the transport of solutes during the occurrence of
buoyancy-driven water flow in a Sphagnum matrix. The contribution of buoyancy flow in the nutri-
ent supply of Sphagnum also depends on the nutrient uptake capacity of Sphagnum. Therefore, the
uptake kinetics of ammonium by the capitula of S. cuspidatum and S. fallax were determined.
Additionally, in a pristine Argentinean bog a 15N source was placed in the deeper acrotelm and the
uptake of labelled nitrogen by the capitula was measured with and without the obstruction of con-
vective flow. The importance of buoyancy-driven water flow for several nutrients and its impor-
tance with respect to other nutrient transport mechanisms in a Sphagnum bog will be discussed.
Materials and methods
Mesocosm experiment
A container (h=11.5 cm d=20 cm) was completely filled with demineralized water and Sphagnum
magellanicum mosses to create a water-saturated Sphagnum matrix. The container was insulated
with a ten centimetre thick layer of foam to prevent radial heat loss and placed on a vibration free
foundation in a climate controlled room. “Day” temperature was set at 20°C (14 hrs) and “night”
temperature at 8°C (10 hrs). Light conditions were 50 µmol ∙ m-2 ∙ s-1, continuously (Hansatech Quan-
titherm Light meter). The occurrence of buoyancy flow in the Sphagnum matrix was monitored by
continuously measuring the vertical temperature profile using an array of five chromel-alumel
thermocouples placed in the centre of the bucket at 5, 20, 40, 70 and 110 mm depth connected to
a Graphtec GL200 midi logger (Graphtec Corp., Yokohama, Japan) on which data were logged and
stored with one minute intervals. A vertical temperature profile was also monitored continuously
in a similar container filled with wet potting soil (providing a matrix in which no convective flow
was expected to occur).
A blue dye (Coomassie Brilliant Blue G, No. B0770, Sigma Aldrich) was used to mimic the transport
of dissolved compounds by buoyancy flow. A volume of 500 mL of water was extracted from the
The transport of solutes – 21
Sphagnum matrix by a siphon followed by the resettlement of the matrix for an hour. Subsequently,
at the onset of “night” (t=0), 200 mL of Brilliant Blue solution (66 mg/L) was layered through a tube
(3*5mm*40 cm) on the bottom of the Sphagnum matrix from a 500 mL Erlenmeyer flask with a flow
rate of approximately 1.3 mL · s-1. The temperature of the Brilliant Blue solution was 4°C to assist the
positioning of the layer at the bottom of the container. The homogeneous Sphagnum matrix had
a density of approximately 4 g DW ∙ L-1, which is comparable to a natural, green and growing peat
moss layer (Clymo, 1970).
After 15 minutes (t=15) the first water samples were taken from the mesocosm at 5, 55 and
105 mm depth, using an array of three black norprene tubes (l = 400 mm, 4.8 mm outer and 1.6
mm inner diameter; Saint-Gobain Performance Plastics, Verneret, France) in combination with a
peristaltic pump (Masterflex L/S model 7519-25, Cole Parmer Instrument company). Water samples
were 1 mL each and collected in disposable polystyrene cuvettes (10 ∙ 4 ∙ 45 mm; Sarstedt, Nüm-
brecht, Germany). Water samples were taken at 15 minutes intervals during the first 90 minutes of
the experiment and at increasingly longer intervals during the remainder of the experiment. The
experiment lasted for 25 hours. Care was taken that the dead volume in the tubes was excluded
from the samples taken. After 2.5 hours additional samples (1 mL) were taken with a pipette at the
periphery of the Sphagnum matrix at 5 mm depth. Immediately after sampling the extinction of the
water samples was measured on a double-beam spectrophotometer (Uvikon 940, Kontron Instru-
ments, Germany) at 580nm with demineralized water as a reference. Potential effects of tempera-
ture on the extinction values of the water samples were determined by simultaneously sampling a
solution with a known concentration of Brilliant Blue. No temperature effects were observed.
The abovementioned experiment was repeated eight times. Data were plotted and fitted using
graphing software (Prism version 4.03, 2005; GraphPad Software, Inc., San Diego, CA, USA).
The Sphagnum magellanicum plants used in the mesocosm experiment were collected in
August 2009 in a small bog located in the “Dwingelderveld” (N52°50’, E6°26’), a nature reserve in the
north of the Netherlands. The mosses used in this experiment were solely used to create a Sphag-
num matrix with acrotelmic characteristics. To minimize the adsorption and/or uptake of Brilliant
Blue during the experiment by the Sphagnum mosses they were incubated for at least 5 days in a
Brilliant Blue solution (66 mg ∙ L-1) prior to the experiment. Before the plants were used to set up the
matrix they were rinsed twice with demineralized water. The Sphagnum matrix was set up at least
12 hours before the start of the experiment to allow equilibration, thereby avoiding possible leak-
age of Brilliant Blue from the Sphagnum plants into the matrix solution during the experiment.
Uptake kinetics
Experimental design and analysis
To determine the uptake kinetics of ammonium by the capitula of Sphagnum cuspidatum and S. fal-
lax, the upper 2 cm of the plants were incubated for two hours in solutions containing 0, 5, 10, 25,
50 and 100 µmol 15NH4Cl ∙ L-1 and 20mM MES at pH = 4. Per concentration and species, three capitula
were incubated in 85 mL solution in a square Petri-dish (120*120mm; Greiner bio-one GmbH), in
triplicate. The capitula were rinsed three times with demineralized water before and after the treat-
ment. The experiment was performed in a climate chamber (light 185 µmol ∙ m-2 ∙ s-1; 18˚C).
To distinguish between the amount of ammonium taken up internally (assimilated) and
bound to the cell wall, an additional experiment was performed. The experimental set up was
22 – Solute transport in Sphagnum dominated bogs
exactly the same as the experiment described above except, to rinse of the 15NH4
+ from the cell wall,
in this experiment the plants were rinsed for two minutes with 100 mL 1mM KCl + 0.5 mM CaCl2
solution (at 150 rpm) after incubation. Subsequently, the plants were dried for at least 48 hours
at 80°C. From each Petri-dish the capitula were pooled and grinded to a fine powder using a ball
miller (Retsch MM2, Haan, Germany). Per sample the %N and %15N were measured at the Univer-
sity of California Davis Stable Isotope Facility, Davis, California, USA, using a PDZ Europa ANCA-
GSL elemental analyzer interfaced to a PDZ Europa 20-20 isotope ratio mass spectrometer (Sercon
Ltd., Cheshire, UK). The data are expressed as the amount of 15N taken up (assimilated and bound to
the cell wall) by the plants, in µmol ∙ g DW-1 calculated by the formula (%15N * %N / 100)*10000/15,
where %N is the percentage of total N in the sample, %15N is the percentage of 15N of total N and 15
is the molecular weight of the stable N isotope.
Plant material
Sphagnum cuspidatum Ehrh. Ex Hoffm. plants were collected in a pool in a small bog in the forestry
“Gasselterveld”, the Netherlands (N52°57.420’, E6°43.857’). Sphagnum fallax (klinggr.) Klinggr. plants
were collected in a small bog in the “Dwingelderveld” (N52°49.135’, E06°29.491’). Before being used
in the experiment, the plants acclimated for two weeks in a greenhouse. In the greenhouse, natural
light was supplemented with high pressure sodium lamps to obtain a 14 hour photoperiod. During
this period the plants were kept wet with demineralized water. No nutrients were supplied to the
plants.
Field experiment Rancho Hambre, Tierra del Fuego, Argentina
In February 2009 the transport of labeled nitrogen (15N) by buoyancy-driven water flow in a field
situation was examined. The location was a pristine Sphagnum magellanicum bog complex, named
“Rancho Hambre” (54°45’S 67°49’W), located in the Argentinean province of Tierra del Fuego and
characterized by the presence of numerous water hollows of different size (Mataloni & Tell, 1996).
See Grootjans et al. (2010) for a more detailed site description.
The experiment was performed in two contrasting Sphagnum habitats; a pool dominated by the
aquatic moss S. fimbriatum, growing with their capitula at the level of the water table, and a lawn
completely dominated by S. magellanicum with the capitula growing approximately 20 cm above
the water table. See figure 2 for the experimental design. Per site, two treatments were applied and
compared to a control situation. In both treatments a 15N source was introduced by placing an agar
cube (50*50*50 mm) containing 10 mg 15NH4Cl ∙ g-1 agar at 10 cm depth below the capitula. In the
first treatment, convective transport in the Sphagnum layer was excluded by the placement of a two
centimeter thick agar disc (d=30 cm) just below the capitula. Before positioning the agar disc, the
upper two centimeter of the Sphagnum plants were removed and placed back on top of the disc. In
the second treatment convective flow was not blocked. The upper two cm of the Sphagnum mosses
were cut off and immediately placed back, to rule out any effect of cutting. Cutting of the top 2 cm
also will block possible internal transport of 15N. Control values were obtained from a third treat-
ment in which the Sphagnum plants were kept intact and no 15N source was applied. All treatments
were performed in triplicate. The distance between the experimental plots was at least one meter.
All plots were sampled twice: just before and 10 days after application of the labeled nitrogen (10
February 2009). Per sample ten capitula were collected and transported in polyethylene bags to the
The transport of solutes – 23
laboratory where they were dried for at least 24 hours at 80°C. The samples were ground to a fine
powder using a ball miller (Retsch MM2, Haan, Germany). The stable nitrogen isotope composi-
tion was measured for each sample with a Carlo Erba NA 1500 elemental analyzer (Thermo Fisher
Scientific Inc.) coupled online via a Finnigan Conflo III interface with a Thermo-Finnigan Delta-
Plus mass-spectrometer. Values of 15N are expresses as percentage of total nitrogen concentration.
Statistical differences in 15N concentrations between treatments and species were tested using
a two-way ANOVA with species and treatment as fixed factors (SPSS for Windows version 16.0.1,
2007; SPSS Inc., Chicago, IL, USA). The assumption of homogeneity of variance was not met, not
even after transformation of the data. According to Heath (1995), the analysis of variance appears
not to be greatly affected by heterogeneity in variance if sample sizes are more or less equal. There-
fore, we decided to continue our analysis using non-transformed data.
Air temperature data of the months January and February were obtained from a climate station
located adjacent to the Rancho Hambre bog complex.
Results
Mesocosm experiment
A clear difference between the vertical temperature profiles in the Sphagnum matrix and the
container with potting soil was observed (figure 1a and b). Due to absence of convective flow in the
potting soil, heat transfer takes place solely by conduction, resulting in a stratified temperature
pattern and gradually decreasing temperatures during the nocturnal cooling period. The cooling
of the Sphagnum matrix proceeds differently. Most remarkable is the small temperature increase
during the nocturnal cooling in the upper Sphagnum layer after approximately 95 minutes.
According to Rappoldt et al. (2003) and Adema et al. (2006) a telltale sign of the upwards transport
of warmer water by buoyancy flow. Moreover, in the presence of convective flow the layers will
mix and the appearance of stratification will be less, which is clearly shown by the smaller
temperature differences between the layers in the Sphagnum matrix when compared to the potting
soil temperature profile.
Buoyancy flow has also been shown to result in a more efficient nocturnal cooling than the
diurnal heating, indicated by a decreasing average temperature with increasing depth (cf. Rappoldt
et al., 2003; Adema et al., 2006). This effect was also observed in the present study (figure 1d). In
the Sphagnum matrix, the average temperature at 110 mm depth is about 0.75˚C lower than at the
surface of the matrix. In contrast, the average daily temperature in the container filled with potting
soil is slightly increasing with depth.
The addition of the Brilliant Blue solution at the beginning of the night, has led to a steep Brilliant
Blue gradient in the Sphagnum matrix, with extinction values of 0.948, 0.056 and 0.063 at 105, 55
and 5 mm depth, respectively (figure 1c). The concentration of Brilliant Blue is reaching equili-
brium at the end of the experiment (t=1500). The course to equilibrium, without the occurrence
of buoyancy flow, can be fitted by a single exponential function, as plotted in figure 1c using the
data from the first 95 minutes (before the occurrence of buoyancy flow) and the equilibrium value
at t=1500.
24 – Solute transport in Sphagnum dominated bogs
However, between 90 and 120 minutes the Brilliant Blue concentration at 105 mm depth suddenly
drops from where it slowly increases to the equilibrium after 25 hours. The sudden drop at t=120
follows the small temperature increase in the upper Sphagnum matrix indicative for the occur-
rence of buoyancy flow. Up to that moment the levels of Brilliant Blue at 5 and 55 mm depth were
at a constant low level but reach equilibrium relatively fast after that moment. Striking is the high
concentration of Brilliant Blue at the edge of the upper matrix layer following the occurrence of
buoyancy flow, reaching a concentration much higher than the fitted line based on exponential
decay. If no mass flow mixing occurs and Brilliant Blue is solely redistributed by diffusion, the
equilibrium value is the maximum concentration that will be reached in the upper Sphagnum layer
and the minimum concentration for the layer at 105 mm. However, in the Sphagnum matrix, after
the occurrence of buoyancy flow, the Brilliant Blue gradient is reversed, indicating the transport of
Brilliant Blue by mixing of the different layers.
Figure 1. Temperature profiles and Brilliant Blue course during the nocturnal period of the mesocosm experiment. (a) and (b)
show the course of the vertical temperature profile in the container with wet potting soil (control) and in the Sphagnum matrix,
respectively. The numbers in the temperature courses in (a) indicate the depths at which the temperature was measured. In (b)
the occurrence of buoyancy flow is clearly visible by the small temperature increase after approximately 95 minutes (indicated
by the vertical grey line). (c) shows the course of the Brilliant Blue solution throughout the Sphagnum matrix. The solution was
added at the bottom of the Sphagnum matrix at t=0. The amount of Brilliant Blue at depths of 5, 55 and 110 mm is shown by re-
spectively closed, grey and open circles. Additional samples taken with a pipette in the upper layer of the Sphagnum matrix are
indicated by an open square. The initial concentration of Brilliant Blue at 110 mm depth is indicated by an open diamond. The
The transport of solutes – 25
black curve indicates the mixing of the Brilliant Blue throughout the Sphagnum matrix following a single exponential decay
using the data from the first 95 minutes of the experiment (before buoyancy flow starts) and the equilibrium value reached at
t=1500 (r2=0.9970). At the moment of buoyancy flow development (vertical grey line), a sudden decrease of Brilliant Blue at 110
mm depth was observed. (d) shows the average 24-hrs temperature with depth relative to the temperature at 5 mm depth for
the Sphagnum matrix (closed circles) and the potting soils container (open circles) The points refer to the measurements in (a)
and (b). Buoyancy flow results in a more efficient nocturnal cooling relative to the diurnal heating, indicated by a decreasing
average temperature with depth. This is not the case in the container filled with potting soil.
Uptake kinetics of NH4
+
The uptake of 15N as function of the concentration 15NH4Cl, for Sphagnum cuspidatum and S. fallax,
are shown in figure 2. A distinction is made between the total amount of ammonium taken up
(the fraction bound to the cell wall and the fraction taken up internally), the fraction 15N taken up
internally only (which are the 15N concentrations after rinsing) and the fraction bound to the cell
wall only (adsorption) which is derived from the difference between the total amount and adsorp-
tion. Hyperbolic curves according to the formula A=Vmax
*[15N]/(Km
+[15N]) + c were fitted to the data.
Except for the internal assimilated 15N which shows a linear response to the 15N concentration. The
uptake kinetic parameters for total uptake and adsorption are shown in table 1.
Table 1. Kinetic uptake parameters of ammonium (±SD) for Sphagnum cuspidatum and S. fallax
S. cuspidatum S. fallax
Total uptake Adsorption Total uptake Adsorption
Vmax
(µmol 15N ∙ g DW-1 ∙ hr-1) 31.3 (±4.1) 13.7 (±1.8) 20.3 (±2.4) 8.1 (±0.6)
Km
(µmol 15N ∙ L-1) 125.6 (±28.8) 60.0 (±18.4) 102.1 (±22.8) 49.5 (±10.3)
c 0.02 (±0.3) -0.15 (±0.3) -0.1 (±0.2) -0.2 (±0.1)
Figure 2. The amount of 15N (µmol ∙ g DW-1) taken up by Sphagnum cuspidatum and S. fallax when incubated for two hours at
different concentrations of 15NH4Cl at 18˚C. A distinction is made between overall uptake of 15N (assimilated and adsorption –
solid circles) and assimilated only (open circles) and 15N bound to the cell wall (adsorption (grey circles). Shown are means and
standard deviations of three replicates. Data of overall uptake and adsorption are fitted to the hyperbolic curve A=Vmax
*[15N]/
(Km
+[15N]) + c and assimilation only data to the curve y=ax+b.
26 – Solute transport in Sphagnum dominated bogs
Figure 3. The Rancho Hambre experiment. (a) Temperature difference between day and night from January 1st to February
28 2009, measured adjacent to the Rancho Hambre bog complex. The experimental period is indicated by the grey box.
A temperature difference of 8˚C is indicated by the dotted line. During the experimental period at least four times a difference
of 8˚C between day and night was measured, which has been shown to be sufficient for the formation of buoyancy-driven
water flow in Sphagnum fimbriatum sites. (b) The relative increase in 15N concentration in capitula of Sphagnum magellanicum
(open symbols) and S. fimbriatum (closed symbols) 10 days after placing the 15N source in the acrotelm. The solid line gives the
average value of all non-buoyancy flow treatments (all treatments except the 15N – BF treatment at the S. fimbriatum site) and the
dashed lines indicate the 95% interval of these samples. (c) A cross section of the upper part of a Sphagnum bog showing the
schematic view of the experimental set-up. The experiment consisted of three treatments; control, 15N-no BF and 15N – BF. Each
treatment was performed in two different habitats; a site dominated by S. magellanicum and a site dominated by S. fimbriatum.
The capitula (asterisks) of these mosses growing respectively 10 and 0 cm above the water level as indicated by the grey area.
In the control treatment the plants were left untreated. In the other treatments a 15N source was placed in the acrotelm at 10 cm
below the capitula as indicated by the small grey boxes. In the second treatment convective flow was obstructed by placing an
agar disc below the capitula (thick black horizontal line in the 15N – no BF treatment). In the third treatment, convective flow
was not obstructed (15N – BF).
The transport of solutes – 27
15N experiment Rancho Hambre
The temperature difference between day and night adjacent to the Rancho Hambre bog complex in
February 2009 are presented in figure 3a. During the experimental period a difference between day
and night temperature of at least 8˚C occurred several times in the Rancho Hambre bog complex.
In Chapter 3 it is demonstrated that buoyancy flow develops in the Sphagnum fimbriatum sites in
the Rancho Hambre bog complex at temperature differences of ≥ 8˚C between day and night. Figure
3b shows the relative increase of 15N in the capitula of Sphagnum magellanicum and S. fimbriatum
capitula in all treatments after 10 days. Individual values per plot are shown. Main effects of spe-
cies and treatment on 15N concentration in the capitula were non-significant, (F(1,10)= 0.635, F(2,10)
= 1.246, respectively; p>0.05). Also no significant interaction effect of species * treatment was
found (F(2,10) = 0.177, p>0.05). However, in two of the non-obstructed Sphagnum fimbriatum plots
(15N - BF) a much higher increase of 15N in the capitula was observed (2.28 and 2.72%,) compared
to the average increase (0.97%) in the other treatments in which transport by buoyancy flow was
not to be expected due to either too low water tables (all S. magellanicum sites) or obstruction of
transport of 15N by convective flow (S. fimbriatum, no BF).
Discussion
The mesocosm experiment clearly demonstrates that solutes are transported by buoyancy-driven
water flow in a Sphagnum matrix. These findings indicate that buoyancy-driven water flow acts as
an external nutrient transport mechanism in water-saturated Sphagnum habitats and thereby con-
tributes to the supply of nutrients to the Sphagnum capitula in the upper bog layer and thereby to
the recycling of nutrients. To date, diffusion and internal transport were the known mechanisms
by which nutrients were transported throughout a water-saturated Sphagnum layer. Note that cap-
illary transport is often mentioned as an (important) nutrient transport mechanism (Hayward &
Clymo, 1982), but this type of transport is only possible above the water table and therefore not
taken into account here. The mesocosm experiment presented here clearly shows buoyancy flow
to distribute solutes more rapidly and in larger amounts than is possible by diffusion (figure 1)
with the reversal of the Brilliant Blue gradient indicates mixing of the layers.
It was assumed that diffusion was the only mechanism by which the Brilliant Blue was redis-
tributed throughout the Sphagnum matrix before buoyancy flow started to occur. However, based
on the concentration Brilliant Blue at t=0 at a depth of 105 mm, and the diffusion coefficient for
Brilliant Blue, the decaying values in the first 95 minutes of the experiment and the equilibrium
values at t=1500 are too high to be explained by diffusion alone. Other mechanisms play a role in
the redistribution of Brilliant Blue in this phase of the experiment. This could involve small scale
convective flow, adsorption by the Sphagnum plants and/or diffusion into the hyaline cells.
However, the importance of buoyancy flow as a transport mechanism in supplying the capitula is
co-determined by the nutrient uptake capacity of the capitula. Due to buoyancy flow, nitrogen gra-
dients can be reversed in a water-saturated Sphagnum layer and thereby induce a stepwise increase
in the nitrogen concentration near the capitula. In case of a full reversal of layers the maximum
ammonium concentration in the surroundings of the capitula reflects the ammonium concentra-
tion in the deeper acrotelm. Literature values for bog water ammonium concentration are about
28 – Solute transport in Sphagnum dominated bogs
3 µmol ∙ L-1 for a pristine bog in Ireland and 105 µmol ∙ L-1 for a Dutch bog suffering high nitrogen
loads (Lamers et al., 2000).
In the observed uptake kinetics for ammonium by Sphagnum cuspidatum and S. fallax, the Vmax
of ammonium uptake when exposed to ammonium concentrations up to 100 µmol ∙ L-1 does not
seem to be reached. In a separate experiment in which the time dependence of uptake was deter-
mined for 100 µmol NH4
+ ∙ L-1 equilibration was reached after about 24 hours for both S. cuspidatum
and S. fallax (see Chapter 7, figure 1). These uptake characteristics allow Sphagnum to make full
use of the stepwise increase in ammonium supplied by buoyancy flow, considering that in every
24-hour period only one buoyancy flow event can be expected.
Interestingly, the observed uptake kinetics of ammonium by Sphagnum cuspidatum and S. fal-
lax indicate the existence of two different processes: a high affinity adsorption and a low affinity
internal uptake mechanism. The high affinity of the adsorption of NH4
+ to the cell wall implies that
the fixation of ammonium at low concentrations is mainly realized by adsorption (table 1). With
increasing concentrations the relative importance of adsorption to total uptake decreases; the cell
wall will saturate and increased uptake will take place by intracellular uptake. These observations
support the general assumption of the cell wall functioning as a temporal extension of nutrient
availability for intracellular uptake (e.g. Buscher et al., 1990; Clymo, 1963; Hajek & Adamec, 2009;
Jauhiainen et al., 1998). Sphagnum fallax shows a higher affinity for NH4
+ uptake than S. cuspidatum,
a difference that could be reflected by the different habitat of these two species. Sphagnum species
growing higher above the water level (hummocks) have higher cation exchange capacities (CEC)
than species growing in wetter habitats like lawns and hollows (Jauhiainen et al., 1998; Hajek,
2009; Clymo, 1963).
Earlier studies report various values for NH4
+ uptake rates by Sphagnum ranging from about
20 to 130 µmol NH4
+ ∙ g DW-1 ∙ hr-1(Jauhiainen et al., 1998; Rudolph et al., 1993; Twenhoven, 1992;
Wiedermann et al., 2009). However, these studies are difficult to compare since the amount of
NH4
+ applied differed.
As in our study, Jauhiainen et al. (1998) applied 50 µM NH4
+ to the capitula of seven Sphagnum
species in order to measure NH4
+ uptake rates. For S. cuspidatum and S. fallax they found uptake
rates of 67 and 78 µmol ∙ g DW-1 ∙ hr-1, respectively, which are high compared to the values found
in this study; 8.7 and 6.3 µmol ∙ g DW-1 ∙ hr-1 for S. cuspidatum and S. fallax, respectively. Nitrogen
uptake rates are suggested to be negatively correlated with nitrogen deposition rates and internal N
concentration (Limpens & Berendse, 2003). If so, the relatively low uptake rates might be explained
by the high N deposition at the sites were the plants were collected (28 kg ∙ ha-1 ∙ yr-1; RIVM, 2009),
which was reflected by the internal nitrogen concentration of 11.1 ± 0.6 and 10.1 ± 2.0 mg ∙ g DW-1
for S. cuspidatum and S. fallax, respectively. Another explanation for these low uptake rakes might
be the difference in pH between our experimental solution (4.0) and the one used by Jauhiainen et
al. (1998): between 5.0 and 5.5. Since our data show the importance of the cation exchange capac-
ity (CEC) of the cell wall in the ammonium uptake, a low pH, correlating with a low CEC (Richter &
Dainty, 1989; Rudolph et al., 1993), might have resulted in lower uptake rates.
Internal transport is estimated to distribute solutes throughout the plant with a half time of
about 11 days, an estimation based on the symplasmic apical transport of 14C (in NaHCO3; Rydin &
Clymo, 1989). Internal, apical transport of N showed even lower rates, a half time of 17 days (see
Chapter 4). According to a review by Raven (2003) on long distance transport in non-vascular
The transport of solutes – 29
plants, there seems to be no evidence for symplasmic transport in Sphagna at speeds faster than
can be accounted for by diffusion. Thus, in comparison with diffusion and internal transport,
buoyancy flow seems to be a quantitative important nutrient transport mechanism in a water-
saturated Sphagnum bog.
Nitrogen
Since Sphagnum growth in peatlands is often limited by nitrogen availability (Aerts et al., 1992;
Bridgham et al., 1996; Gunnarsson & Rydin, 2000) the supply of nitrogen to the capitula is of spe-
cial interest. The main N source for Sphagnum has been shown to be re-mineralized N (Aerts et al.,
1999; Aldous, 2002b; Bridgham, 2002; Gerdol et al., 2006; Morris, 1991; Urban & Eisenreich, 1988).
The importance of re-mineralization of nitrogen for Sphagnum growth has been demonstrated in
situ by Urban & Eisenreich (1988). They calculated the assimilation of nitrogen by plants (primarily
Sphagnum) to be 66 kg ∙ ha-1 ∙ yr-1, whereas only 14.6 kg N ∙ ha-1 ∙ yr-1 was supplied by total inputs.
The remainder was supplied by mineralization of the peat. These findings are contradictory to the
idea that Sphagnum and vascular plants utilize spatially distinct nutrient pools, with Sphagnum
relying on N from precipitation and vascular plants on mineralization of senescing organic mat-
ter in the deeper acrotelm (Malmer et al., 1994; Pastor et al., 2002). Partly due to buoyancy flow,
Sphagnum can also rely on mineralized nitrogen as a major N source. Consequently, Sphagnum
mosses may outcompete vascular plants more easily and thereby enhance their ability to engineer
the ecosystem (Van Breemen, 1995).
Gerdol et al. (2006) stated that the cycling through mineralization of senescing tissues
by heterotrophic bacteria is essential for the supply of N to the growing tissues. An interesting
observation was the positive effect of high water tables on N cycling, a result that in the original
paper was left unexplained (Gerdol et al., 2006). Significantly lower upward translocation of N to
the growing capitula at low water tables was also observed by Aldous (2002b). A likely explanation
for this phenomenon is that high water tables allow the occurrence of buoyancy flow, increasing
recycling rates of nitrogen.
The experiment performed in the Rancho Hambre bog demonstrates the transport of N by
buoyancy-driven water flow in the Sphagnum fimbriatum sites and the subsequent uptake by the
capitula in the upper Sphagnum layer. The increase in 15N concentration in the treatment with
unobstructed convective flow indicates the transport of 15N by buoyancy flow. In two of the three
replicates this increase was observed. Possibly in one case buoyancy flow did not occur, or did not
result in the upward transport of 15N. Multiple factors determine the variety in the occurrence of
buoyancy flow. Although during the experiment the temperature difference between day and night
were sufficient to induce buoyancy flow, this is not a guarantee for buoyancy flow cells to occur.
Other factors co-determine the development of these cells. For example, a heterogeneity in the
vertical hydraulic conductivity might locally hamper the development of buoyancy flow. Addition-
ally, buoyancy flow occurs as adjacent cells with warm water going up and cold water going down
(Adema et al., 2006). Sphagnum capitula residing in a downward flow do not receive nutrients from
below despite the occurrence of buoyancy flow.
Ecological implications
CO2 gradients. Lloyd et al. (1998) showed an increase with depth of the CO
2 concentration in a
30 – Solute transport in Sphagnum dominated bogs
water-saturated Sphagnum core due to the uptake of CO2 by photosynthetic activity of the capitula in
the upper layer and the release of CO2 by decomposition of organic material in the lower acrotelm.
Since submerged Sphagnum species that inhabit peat hollows have been shown to be limited by
CO2 (Rice & Giles, 1996; Rice & Schuepp, 1995), buoyancy flow might be an important mechanism
replenishing CO2 around the capitula and enhancing photosynthesis by vertical transport of CO
2
from the deeper catotelm layer.
Oxygen gradients. The photosynthetic activity results in high oxygen levels in the top Sphagnum lay-
er and decreasing levels with depth (Adema et al., 2006; Lloyd et al., 1998). Lloyd et al. (1998) measu-
red a steep oxygen gradient in the upper four centimeters of a water-saturated Sphagnum layer,
decreasing from 300 to 0 µM. Mixing of water layers by buoyancy flow will result in a downward
transport of oxygen. Adema et al. (2006) attributed a conspicuous change in oxygen concentration
at 5 cm depth in a Sphagnum layer to the occurrence of buoyancy flow. Since the aerobic decom-
position of organic material is significantly higher than the anaerobic decomposition (Bridgham
et al., 1998; Waddington et al., 2001) the transport of oxygen to the lower parts of the acrotelm will
increase decomposition rates thereby increasing the concentrations of CO2 and nutrients like N. In
turn the nutrients will become available to the growing Sphagnum when transported upwards by
buoyancy flow. Moreover, the downward transport of O2 will increase the CO
2:O
2 ratio in the upper
Sphagnum layer and will enhance photosynthetic performance even more since photosynthesis is
inhibited by oxygen (see Chapter 6; Bowes & Salvucci, 1989; Raven, 2011; Raven et al., 2008; Skre &
Oechel, 1981).
Methane. Methane is anaerobically produced in large quantities in bogs (e.g. Gorham, 1991).
Nevertheless, emissions of methane to the atmosphere are very low (Larmola et al., 2010) due to
the oxidation of methane by methanotropic bacteria (Kip et al., 2010; Raghoebarsing et al., 2005).
The mixing of methane and photosynthetically produced oxygen by buoyancy flow might results in
lower methane emission rates, and affect global carbon cycling. In addition, the released CO2 in this
oxidation process has been shown to be a significant carbon source for Sphagnum (Raghoebarsing
et al., 2005).
Overall conclusion
Net transport by buoyancy flow occurs when a vertical gradient exists. These gradients have been
shown explicitly for CO2, CH
4 and O
2 in a water-saturated Sphagnum layer (Lloyd et al., 1998) with
large concentration differences in the upper 12 cm of the Sphagnum layer for CO2 and CH
4, and O
2
decreasing to undetectable values at 2 cm depth. According to the models described by Rappoldt
et al. (2003), the size of the buoyancy cells (dependent on the Rayleigh number of the Sphagnum
layer) can be as large as 24 cm. Based on in situ temperature measurements in a pristine Sphag-
num bog in Tierra del Fuego, Argentina, cell sizes of 3 to 26 cm were predicted. Therefore, due to
buoyancy flow these solutes can be transported and gradients can even be reversed (figure 1). The
maximal transport of nutrients per buoyancy event depends on the concentration in the lower
layer. Rappoldt et al. (2003) showed in a model that the Sphagnum matrix is well mixed after 4
consecutive buoyancy events. The mesocosm experiment shows that the Sphagnum matrix was
already mixed after one occurrence of buoyancy flow.
The transport of solutes – 31
The recycling of nutrients is of great importance in the functioning of Sphagnum bogs and has been
suggested to explain the high carbon burial rates despite the low primary productivity, also known
as the “Paradox of Peatlands” (Raghoebarsing et al., 2005). This chapter shows that buoyancy flow
might be essential for the efficient recycling of nutrient by Sphagnum at least under waterlogged
conditions.
Chapter 3
Field characteristics of buoyancy-driven water flow and its global occurrence
Wouter Patberg
Erwin Adema
Myra Boers
Gert Jan Baaijens
Christian Fritz
Ab Grootjans
Rodolfo Iturraspe
Alfons Smolders
Theo Elzenga
34 – Solute transport in Sphagnum dominated bogs
Abstract
Nutrients enter Sphagnum bogs mainly by atmospheric deposition.
However, to sustain their annual production, this must be supplemented
with nutrients released by decomposition processes in the peat layer. The
capitula are the actively growing parts of the Sphagnum plants and since
they are spatially distinct from the decomposition processes, transport of
the nutrients is essential. For Sphagnum dominated bogs, several nutrient
transport mechanisms have been described; internal transport, external
transport by diffusion, capillary action and buoyancy-driven water flow. In
Chapter 2 it was shown that nutrients are transported more rapidly and in
larger quantities by buoyancy flow than would be possible by diffusion or
internal transport. Therefore, buoyancy-driven water flow is considered to be
an important nutrient transport mechanism in water-saturated Sphagnum
layers. However, this conclusion is based on laboratory experiments,
which were conducted under controlled conditions, whereas in the field
temporal and spatial variation in temperature and hydraulic conductivity
might occur and affect the occurrence and size of the buoyancy cells. This
chapter focuses on the occurrence of buoyancy flow in the field. Vertical
temperature profiles and vertical hydraulic conductivities were measured in
a pristine Sphagnum bog to validate the theoretical predictions in a natural
situation. Subsequently, Ra numbers of the Sphagnum sites were calculated
and the possible occurrence of buoyancy flow in the field was predicted
using the model described by Rappoldt et al. (2003). The calculated Ra
numbers indicate the Sphagnum fimbriatum layer to be suitable for the
development of buoyancy flow, which is supported by the course of the
vertical temperature profiles. Moreover, the predicted starting time of
buoyancy flow development very well correlates to the observed starting
time of buoyancy flow.
Additionally, worldwide daily temperature data were analyzed to model
the possible occurrence of buoyancy flow in peatlands throughout the
world. The results from this GIS analysis indicate that many peatlands are
subjected to temperature differences between day and night of 8˚C or more,
which is sufficient for the development of buoyancy flow. In the month July
about 70% of the peatlands have at least 5 days with temperature differences
between day and night suitable for the development of a buoyancy flow
event. From this analysis it is apparent that buoyancy flow is a worldwide
occurring phenomenon in peatlands.
Field characteristics of buoyancy-driven water flow – 35
Introduction
Sphagnum bogs receive nutrients mainly by precipitation (Van Breemen, 1995). However, to support
their annual production, Sphagnum mosses are supported in their nutrient supply by nutrients
released by decomposition processes in the peat (Aldous, 2002a; Bowden, 1987; Bridgham, 2002;
Gerdol et al., 2006; Urban & Eisenreich, 1988). The capitula are actively growing parts of the Sphag-
num plants (Johansson & Linder, 1980; Malmer, 1988; Malmer et al., 1994; Robroek et al., 2009;
Rydin & Jeglum, 2006) and since they are spatially distinct from the decomposition processes,
transport of the nutrients is essential. For Sphagnum dominated bogs several nutrient transport
mechanisms have been described. Internal transport (Rydin & Clymo, 1989; Chapter 4), external
transport by diffusion, by capillary action (Hayward & Clymo, 1982) or by buoyancy-driven water
flow (Rappoldt et al., 2003; Adema et al., 2006; Chapter 2).
Buoyancy-driven water flow is the vertical convective flow of water in a water-saturated
Sphagnum layer, driven by the temperature difference between day and night (Baaijens, 1982;
Rappoldt et al., 2003) . During the night the upper water layer cools down more rapidly than
the layers below, leading to relatively dense cold water on top of warmer water. Due to density
differences between these two layers the colder and denser water will sink and the warmer water
will rise. Consequently, buoyancy flow occurs as “cells” consisting of adjacent regions with up-
ward and downward flow (Adema et al., 2006; Rappoldt et al., 2003). Evidence for the occurrence of
buoyancy-driven water flow in a water-saturated Sphagnum layer was provided on theoretical and
experimental grounds by Rappoldt et al. (2003).
Whether an inversion in the vertical temperature profile leads to instability and the genera-
tion of buoyancy flow depends on the system’s Rayleigh number, a dimensionless number that
depends on several parameters, including vertical hydraulic conductivity and the temperature dif-
ference between day and night. Rappoldt et al. (2003) showed that buoyancy flow occurs if the
system’s Rayleigh (Ra) number exceeds a value of 25. For a typical peat moss layer, a temperature
difference of 10 degrees between day and night will result in a Ra number of 80 which is suitable
for the quick development of buoyancy flow (Rappoldt et al., 2003).
Adema et al. (2006) provided evidence for the occurrence of buoyancy flow in a small Sphagnum
dominated peat bog in the Netherlands. Based on the hydraulic conductivity (k) of the Sphagnum
layer and a temperature difference between day and night of 8˚C, , the calculated Ra number was
sufficiently high to induce buoyancy flow. In Chapter 2, the convective transport of solutes by
buoyancy-driven water flow is demonstrated in both a mesocosm experiment and a field situation.
In the mesocosm experiment it was shown that buoyancy flow transports nutrients more rapidly
and in larger quantities than would be possible by diffusion (Chapter 2) or by internal transport
(Chapter 4). Therefore, buoyancy-driven water flow is considered to be an important nutrient
transport mechanism in water-saturated Sphagnum layers.
Whereas the laboratory experiments are all conducted under controlled conditions and with
homogeneous samples, in the field temporal and spatial variation in temperature and hydraulic
conductivity might occur and affect the occurrence and size of the buoyancy cells. Therefore, a
series of vertical temperature profiles were measured in a pristine Sphagnum bog to validate the
theoretical predictions in a natural situation. Based on the measured hydraulic conductivities and
ambient day and night air temperatures, the Ra numbers of the Sphagnum sites were calculated.
36 – Solute transport in Sphagnum dominated bogs
Based on these Ra numbers, the possible occurrence of buoyancy flow in the field could be predicted
using the model described by Rappoldt et al. (2003).
Additionally, Geographical Information System software was used to analyze worldwide daily
temperature data and to model the possible occurrence of buoyancy flow in peatlands throughout
the world. Our findings will be discussed with respect to the supply and cycling of nutrients in
Sphagnum dominated bogs.
Materials and methods
Rancho Hambre, Tierra del Fuego, Argentina
Field measurements were performed in February 2005 in Rancho Hambre (54°45’S 67°49’W), a
Sphagnum magellanicum bog complex in the Argentinean province of Tierra del Fuego, extensively
covered by pools of different size (Mataloni & Tell, 1996). A more detailed site description is given
in Grootjans et al. (2010).
In a Sphagnum fimbriatum pool site and a S. magellanicum lawn site, respectively three and
ten intact peat monoliths were collected in 25 cm long PVC tubes with an internal diameter of 108
mm. The tubes were sealed by two PVC caps, stored at 4°C and analyzed the next day. For different
lengths of the cores the vertical hydraulic conductivity (k) was measured (in situ) using the con-
stant head method as described by Adema et al. (2006). Different lengths of the core were obtained
by removing parts of the monolith from the bottom. This process was repeated until the hydrau-
lic conductivity became too large to measure. For each length the hydraulic conductivity was
measured in triplicate. From these measurements the hydraulic conductivity of each segment was
calculated. In each core the depth of the transition from acrotelm to catotelm was determined.
For a period of thirteen days temperature profiles were measured in a Sphagnum fimbriatum pool
using eight copper-constantan-copper thermocouples connected to a Campbell Scientific AM25T
solid state multiplexer with an internal reference RTD (Resistance Temperature Detector). The
thermocouples were placed on 1, 6, 14, 25, 39, 56, 76, and 99 mm depth. The data were collected us-
ing a Campbell Scientific CR10x multi-channel data logger which measured every second of which
the average was stored every minute.
By using the temperature and the vertical hydraulic conductivity data, Ra numbers for the
upper 12 cm of the cores were calculated for each day during this 13 day period (see Rappoldt et al.,
2003). The following formula was used
Ra = k α ΔT √(t0/D
eff).
In which Ra is the dimensionless Rayleigh number, k is the vertical hydraulic conductivity (m ∙ s-1);
α is the thermal expansion coefficient (K-1); ΔT is the difference between maximum day and mini-
mum temperature of the subsequent night (˚C); t0 is the duration of the low temperature phase in
seconds (the value for t0 was 43200s (12h; Rappoldt et al., 2003) and D
eff is the thermal diffusivity
(m2 ∙ s-1). Both the thermal expansion coefficient (α) and the thermal diffusivity (Deff
) of water are
temperature dependent. For each day the values of α and Deff
were determined using the minimum,
maximum and average temperature, resulting in three calculated Ra values per day. The time of
Field characteristics of buoyancy-driven water flow – 37
onset of buoyancy flow development was derived from the telltale deviation from the monotonic
decrease in temperature in the vertical temperature profile and was compared with the predicted
time, based on the Ra numbers calculated according to Rappoldt et al. (2003).
The global occurrence of buoyancy-driven water flow
To quantify the potential global occurrence of buoyancy flow in peat bogs, air temperature data of
peatlands around the world were analyzed by using a Geographical Information System (ArcGis
software, ESRI, Redlands, CA, USA). The following digital maps were used: a map with the distri-
bution of global peatland distribution (Yu et al., 2010). Daily day and night temperature data were
obtained from NASA (https://lpdaac.usgs.gov; dataset MOD11C1 – Version 005, format: HDF-EOS;
Year: 2009; layers used: Daytime LST & Nighttime LST). Average monthly temperature data were
obtained from the same website (dataset: MOD11C3 – Version 005, format: HDF-EOS, year: 2009,
layers used: Daytime LST). Nitrogen-deposition data were derived from the Oak Ridge National
Laboratory Distributed Active Center (ORNL DAAC) site (https://daac.ornl.gov).
For all peatlands we calculated the daily temperature difference between the maximum day
temperature and the minimum temperature of the subsequent night. The minimum temperature
difference for buoyancy flow to occur is 8˚C or more (Adema et al., 2006; Rappoldt et al., 2003).
For each month we calculated the area of peatlands were ≥2, ≥5, ≥10 or ≥20 buoyancy flow events
occurred. Only peatlands with possible Sphagnum growth were included in the analysis. We
assumed that when the average monthly daytime temperature was lower than 8oC, ice formation
would prevent the development of buoyancy flow and/or plant growth would be restricted.
Additionally, global nitrogen deposition data were combined with the distribution of buoyancy
flow events. Based on the response of Sphagnum to increased nitrogen deposition as proposed by
Lamers et al. (2000), three categories were distinguished: <12, 12-18 and > 18 kg N ∙ ha-1 ∙ yr-1 (Lamers
et al., 2000).
Results
Local characteristics of buoyancy flow, Rancho Hambre, Tierra del Fuego
The hydraulic conductivity (k) values for cores of S. magellanicum and S. fimbriatum at different
depths are shown in the figures 1a and b, respectively. The hydraulic conductivity in the upper layer
of the S. fimbriatum cores is the highest (ranging from 0.3 to 0.5 m ∙ s-1) with k values decreasing with
increasing core depth. In Sphagnum fimbriatum cores the depths at which the transition between
acrotelm and acrotelm was found varied between 10 and 25 cm. In S. magellanicum the hydraulic
conductivity is close to zero and does hardly vary with depth. In two of the S. magellanicum cores
(4 and 9) the k values are higher in the upper layer (figure 1b). For all S. magellanicum cores the
transition from acro- to catotelm was found at 25 cm depth.
Temperature measurements Rancho Hambre
The temperature measurements in a Sphagnum fimbriatum pool in the Rancho Hambre bog complex
show a daily temperature cycle for several depths (figure 2a). The difference between day and night
temperature decreases with depth and results in a reversal of the temperature gradient during the
38 – Solute transport in Sphagnum dominated bogs
night. The small temperature increases in the upper Sphagnum layer during nocturnal cooling
(arrow in figure 2c) are clear indications that buoyancy-driven water flow has occurred (figure 2a).
The small temperature increases in the upper water layer are the result of upward movement of
warmer water and the sinking of cooler water (Adema et al., 2006; Rappoldt et al., 2003; Chapter 2).
Figure 1. Vertical hydraulic conductivity (in m ∙ s-1) at different depths of S. fimbriatum (a) and S. magellanicum (b) cores.
Symbols represent the average hydraulic conductivity (± SD) and are placed in the centre of the vertical line which represents
the length and depth of the associating core segment.
Figure 2b shows the calculated daily Ra numbers, ranging from 25 to 153, for each day for S.
fimbriatum core 2. The Rayleigh numbers in figure 2b describe a state of the Sphagnum matrix
calculated for a 24 hour period. However, it is very likely that the state of the Sphagnum matrix
during a particular 24 h period is also influenced by the conditions during previous day or days.
Therefore, the average Ra number for the complete period of 13 days was calculated as well. The Ra
number for the Sphagnum matrix for the complete experimental period was calculated to be 81.
Based on the calculated Rayleigh numbers, the earliest possible times of onset of buoyancy
flow were determined according to the theoretical model described by Rappoldt et al. (2003). This
model describes the relation between the Rayleigh number and the time of onset based on a 12
day/12 night temperature regime. Day/night temperature changes are either assumed to follow a
block wave or a sine wave, resulting in slightly different times of onset between these approaches.
Field characteristics of buoyancy-driven water flow – 39
Based on the sinus wave model, the times of onset were determined for the development of
buoyancy flow in the S. fimbriatum matrix during the experimental period. These are indicated by
the vertical lines in figure 2a. In general, at all days, the calculated time of onset is preceding the
temperature peaks which are indicative for the occurrence of buoyancy flow. Figure 2c is a detailed
view of the temperature profile at day 2 showing the typical temperature increase during nocturnal
cooling due to buoyancy flow (arrow) and the calculated time of onset (vertical black line).
Figure 2. a) Vertical temperature profile in a Sphagnum fimbriatum pool in the Rancho Hambre bog complex, Tierra del Fuego,
Argentina. The measuring period lasted for 13 full days. Temperature was measured at 1 (red), 6 (light blue), 14 (orange), 25 (light
green), 39 (dark blue) , 56 (dark green), 76 (brown) and 99 mm (black) depth. The dashed vertical lines indicate the times of onset
for the development of buoyancy flow determined by the theoretical model of Rappoldt et al. (2003). B) the calculated Ra numbers
for the S. fimbriatum layer during the measuring period. For each day three Ra numbers were calculated using the thermal expan-
sion coefficient (α) and the thermal diffusivity (Deff
) according to the minimum, maximum and average temperature of that day.
C) Vertical temperature profile at day 2. The vertical line indicates the calculated time of onset which is preceding the temperature
peak, pointed out by the arrow, which is indicative for the occurrence of buoyancy flow. Colors of the lines are as in figure a.
40 – Solute transport in Sphagnum dominated bogs
The global occurrence of buoyancy flow
Figure 3 shows per month the relative amount of peatlands with a) the occurrence of ≥2 and ≥5
buoyancy flow events, b) the occurrence of < 2 events, c) without sufficient temperature data and
d) without Sphagnum growth.
In the months June to September in about 80% of the peatlands a buoyancy flow event occurs at
least twice. In 40 to 70% of the peatlands it occurs at least 5 times and in 10 to 20% of the peatlands
at least 10 times. Peatlands with occurrences of 20 times or more are scarce (in general ≤ 1%; 4% in
April), however to establish that for a particular pixel ≥20 buoyancy flow events could have taken
place, the number of days with missing data must be very low. Therefore it is possible that the
number pixels with ≥20 buoyancy flow event is actually higher.
Figure 3. The relative amount of peatlands per month where buoyancy flow events occurred <2, ≥2, ≥5 or where it was assumed
that no Sphagnum growth could occur. Dark grey indicates the percentage of pixels with peatlands for which insufficient data
were available to calculate the number of occurrences of a buoyancy flow event for that particular month.
In figure 4 the distribution of peatlands throughout the world are shown. Indicated in green are the
peatlands with the occurrence of at least 5 buoyancy flow events in July 2009, in red the peatlands
with less then 5 buoyancy flow events, in orange peatlands without sufficient data for the analysis
and in grey peatlands with an average growth temperature <8˚C. In 70% of the peatlands a tem-
perature difference of at least 8˚C occurs at least 5 times (see also figure 3). In 28% of the peatlands
insufficient temperature data are available. In only 0.5% of the peatlands buoyancy flow occurs
less then 5 times. In southern Argentina and Chile the average monthly temperature is lower than
8˚C and therefore not taken into account in the analysis.
To obtain an indication whether Sphagnum growth indeed depends on the occurrence of buoy-
ancy flow for transport and cannot sustain growth on atmospheric N-deposition, we also plot-
ted the atmospheric nitrogen deposition (in kg ∙ ha-1 ∙ yr-1) in the year 1993. Based on the growth
response of Sphagnum to increasing N input (Lamers et al., 2000) a distinction is made between
areas with annual N loads of <12, 12-18 and >18 kg ∙ ha-1 ∙ yr-1. The areas with the highest nitrogen
load are North-Western Europe, East-Asia and small parts of North and South America. Remark-
ably, most peatlands do not coincide with these high N load areas and are thus likely to depend on
buoyancy flow transported nitrogen.
Field characteristics of buoyancy-driven water flow – 41
Figure 4. The occurrence of buoyancy flow in peatlands throughout the world for the month July 2009. Peatlands with the
occurrence of at least 5 buoyancy flow events are green, peatlands with less than 5 buoyancy flow events are red, in grey
peatlands the temperature was too low to allow growth and peatlands without sufficient temperature data for the analysis are
in orange. The squares indicate an atmospheric nitrogen deposition ≥ 12 (green) or ≥ 18 kg ∙ ha-1 ∙ yr-1 (grey).
Discussion
Local characteristics of buoyancy-driven water flow
Based on the vertical hydraulic conductivity of the Sphagnum fimbriatum cores and the temperature
differences between day and night, the calculated daily Ra numbers varied from 25 to 153 and
according to the model of Rappoldt et al. (2003) the S. fimbriatum layer is thus suitable for the
development of buoyancy-driven water flow. Indeed, the typical small temperature increases
during the nocturnal cooling in figure 2 very clearly indicates the regular occurrence of buoyancy-
driven water flow in the Sphagnum fimbriatum pool in Rancho Hambre.
The calculated starting time of buoyancy flow development based on the Ra number, correlates,
with exception of days 11 and 13 both with a Ra of 25, very well with time of the small tempera-
ture increases indicative of buoyancy flow. It should be noted that the calculated Rayleigh number
describes a state of the Sphagnum matrix for a single day/night period, ignoring the temperature
history during the previous days of the site. This could have been of influence at days 11 and 13,
on which the buoyancy flow event occurs much faster than the modelled time of onset. These
two days are characterized by a relatively low day temperature. As a consequence, during the cool-
ing period at night the temperature of the surface layer drops fast below the still relatively high
temperatures of the lower layers. Possibly, this explains the discrepancy with the model, which,
based solely on the relatively small diurnal temperature difference and ignoring the previous days,
predicts a very late time of onset of the buoyancy flow event.
In agreement with the model is the observation that at day 8, which has a calculated Ra number
of 153, no buoyancy flow event is apparent. At very high Ra numbers the model of Rappoldt et al.
(2003) predicts that the size of the cells become very small and as a consequence are not detectable
42 – Solute transport in Sphagnum dominated bogs
anymore with the used sensor array.
The vertical hydraulic conductivity values in the upper 12 cm of the S. magellanicum cores were in
general (with the exception of two cores) too low to result in Ra numbers suitable for the develop-
ment of buoyancy flow (data not shown). Furthermore, in the observed S. magellanicum lawns the
water table is located about 20 cm below the top of the Sphagnum plants which form an insulating
layer, preventing the development of a cool water surface layer and instability in the water column
(Van der Molen & Wijmstra, 1994). If buoyancy flow would nevertheless occur, solutes transported
from deeper layers to the upper water layer would still have to be transported to the capitula by
capillary transport. In this case, buoyancy flow only acts as an auxiliary transport mechanism and
its relative importance in the nutrient supply to the capitula is determined by the height of the
capitula above the water level. Such a situation can be found in Sphagnum lawns and the transition
zones between pools and hummocks.
Global occurrence of buoyancy-driven water flow
The results from the GIS analysis indicates that many peatlands are subjected to temperature
differences between day and night of 8˚C or more, several days each month during the growing
season. At these sites, buoyancy flow events could occur, resulting in sufficient mixing of the water
column and efficient recycling of nutrients.
In the month July about 70% of the peatlands have at least 5 days and about 90% at least 2 days,
with temperature differences between day and night suitable for the development of a buoyancy
flow event. From this analysis it is apparent that buoyancy flow is occurring in peatlands on a
global scale.
For weather stations at latitudes above 50˚, the fraction of days with Ra numbers >100 during
the months June and July was calculated by Rappoldt et al. (2003). From this analysis they con-
cluded that buoyancy flow occurs on a large scale , but is more likely in continental areas than in
coastal areas. This distinction between coastal and continental sites was not found in the present
study. This is possibly due to the fact that in the current study a temperature difference of 8 oC
was used as the sole criterion, while Rappoldt et al. (2003) calculated the Ra, taking into account
the effect of temperature on thermal expansion coefficient and thermal diffusivity. This approach
leads to different results at very low temperatures and does not affect our conclusions, since we
have included only pixels with and average monthly temperature higher than 8oC. Furthermore
our study focused on Sphagnum dominated peatlands, while Rappoldt et al. (2003) included all
weather station data available.
The mesocosm experiment in Chapter 2 has shown buoyancy flow to be an efficient and rapid
nutrient transport mechanism when compared to diffusion and internal transport. Uptake charac-
teristics for ammonium of Sphagnum (Chapter 2) imply that Sphagnum can profit from stepwise in-
creases in nitrogen availability. Therefore, even very infrequent occurrences of buoyancy flow can
make a significant contribution to the nutrient availability in a water-saturated Sphagnum layer.
The relative importance of transport of nutrients from deeper water layers to the top layer
with the capitulum, is higher when atmospheric input is low. This is especially the case for
nitrogen availability which is often considered growth limiting in Sphagnum bogs (Aerts et al.,
1992; Bridgham et al., 1996; Gunnarsson & Rydin, 2000). Since areas with a high nitrogen depo-
sition load only slightly overlap with the area covered with peatlands (figure 4), the importance
Field characteristics of buoyancy-driven water flow – 43
of buoyancy flow in the transport and recycling of nitrogen in Sphagnum bogs, is hardly dimin-
ished by increased, anthropogenic N deposition. Therefore, we conclude that buoyancy flow is
a worldwide occurring phenomenon which significantly contributes to the nutrient supply and
nutrient recycling in Sphagnum bogs.
The importance of buoyancy flow
Buoyancy-driven water flow has been shown to act as an external nutrient transport mechanism
in water-saturated Sphagnum habitats (Chapter 2), thereby contributing to the supply of nutrients
to the Sphagnum capitula in the upper bog layer and thus to the efficient recycling of nutrients.
Moreover, in comparison with diffusion and internal transport, buoyancy flow seems to be a quan-
titatively important nutrient transport mechanism (Chapter 2 and 4). The present study shows that
buoyancy-driven water flow is a worldwide occurring phenomenon in peatlands.
A Sphagnum bog can consist of hollows, lawns and hummocks. As buoyancy flow is restricted
to the water layer in a Sphagnum bog, direct supply of nutrients from deeper layers to the capitulum
by buoyancy flow only takes place in hollows. In lawns buoyancy flow can assist capillary driven
nutrient transport and in hummocks buoyancy flow probably is relatively unimportant or does
not occur. As the initial successional stage of a Sphagnum bog is the colonization of aquatic Sphag-
num species of water bodies followed by the invasion of hummock forming species, buoyancy flow
seems to be particularly important in the early stages of bog development. Efficient nutrient use
might be of great importance in creating dominance over vascular plants by keeping the nutrient
concentrations low in the lower acrotelm and thereby facilitating conditions beneficial to Sphag-
num growth.
With the regular occurrence of buoyancy flow, the nitrogen concentration in the acrotelmic
water will be determined by the decomposition rate in catotelm, the depth of the buoyancy flow
cells, the depletion in the top water layer by uptake and assimilation and dilution by rain water.
One can easily imagine how such a complicated process, that prevents the build up of high nutri-
ent levels in the deeper layers and thereby the establishment of vascular plants, can be disrupted
by high nitrogen loads.
Chapter 4
Physiological evidence for internal acropetal transport of nitrogen in Sphagnum cuspidatum and S. fallax
Wouter Patberg
Bikila Warkineh Dullo
Alfons Smolders
Ab Grootjans
Theo Elzenga
46 – Solute transport in Sphagnum dominated bogs
Abstract
Several mechanisms for acropetal or upward transport of nutrients have
been described for Sphagnum layers; diffusion, buoyancy-driven water
flow, capillary transport and internal transport. This chapter focuses on the
contribution of internal transport in the translocation of nitrogen from the
catotelm, where nutrients are released by decomposition of organic mate-
rial, to the acrotelm where growth of the Sphagnum plants takes place. The
internal transport of nitrogen has often been assumed to be present, but has
never been explicitly demonstrated before.
The ability of Sphagnum to transport nitrogen internally was investigated
by monitoring the transport of labeled nitrogen in two Sphagnum species,
S. cuspidatum and S. fallax, in experiments in which diffusion and external
transport were prevented. For both species a slow, but significant acropetal
transport of nitrogen through an internal mechanism was observed. Within
the time frame of the experiments, the internal transport of nitrogen was
only observed when nitrogen was supplied as NH4
+ and not as NO3
-. The
speed at which the internal transport took place was estimated at 5 mm ∙
day-1 and the equilibration between donor and acceptor parts of the plants
was characterized by half time value of 17 days. Relative to other transport
mechanisms, like buoyancy flow and capillary transport, internal transport
represents only a minor fraction of the upward transport of externally sup-
plied nitrogen to the capitula. The main importance of internal transport
is therefore considered to be the reallocation of internally catabolized
nitrogen compounds.
Physiological evidence for internal acropetal transport – 47
Introduction
Sphagnum mosses are the most dominant plant species in bogs. For their nutrient supply they de-
pend mainly on atmospheric deposition. Bogs are, therefore ombrotrophic ecosystems and under
natural conditions they are usually nitrogen-deficient (Bragazza et al., 2004; Bridgham et al., 2001b;
Gunnarsson & Rydin, 2000; Li & Vitt, 1997). Sphagnum mosses are able to survive due to their very
efficient nitrogen utilization (Bridgham, 2002; Li & Vitt, 1997). Sphagnum utilizes nitrogen from
atmospheric deposition with an efficiency that ranges from 50 to 90% (Aldous, 2002a; Li & Vitt,
1997). Woodin & Lee (1987) even measured a retention of 100% of inorganic nitrogen at an unpol-
luted site, whereas chloride and sulphate were passing freely through the moss mat. The efficiency
of nitrogen retention by Sphagnum clearly results in a competitive advantage of Sphagnum over
vascular plants. Aldous (2002a), for instance, showed that vascular plants could take up less than
1% of N recently added by wet deposition.
Nonetheless, under non-polluted conditions, the annual input of nitrogen by atmospheric
deposition is not sufficient to support the observed primary production in Sphagnum dominated
ecosystems (Aerts et al., 1999; Aldous, 2002a, b; Bowden, 1987; Bridgham, 2002; Damman, 1978,
1986; Morris, 1991; Pakarinen, 1978; Rosswall & Granhall, 1980; Urban & Eisenreich, 1988). Mass
balance calculations have shown that direct retention of N from precipitation is less important
than recycling of mineralized N to support Sphagnum growth (Aldous, 2002b; Bowden, 1987;
Bridgham, 2002; Urban & Eisenreich, 1988).
The highest mineralization rates have been found in the aerobic zone of the acrotelm at the border
of the anaerobic catotelm (Bridgham et al., 1998; Malmer, 1993; Waddington et al., 2001). On the
other hand, the highest metabolic activity and nutrient uptake in Sphagnum mosses was measured
in the upper part of the plant, the capitulum (Aldous, 2002a; Johansson & Linder, 1980; Malmer,
1988; Malmer et al., 1994; Robroek et al., 2009; Rydin & Jeglum, 2006). Since the capitula are spa-
tially separated from of the layer where mineralization occurs, transport of nitrogen to the capitula
is necessary for the required recycling of nitrogen in a Sphagnum bog.
Several nutrient transport mechanisms have been described for Sphagnum bogs; diffusion,
buoyancy-driven water flow (Adema et al., 2006; Rappoldt et al., 2003; Chapter 3), capillary transport
(Clymo & Hayward, 1982) and internal transport (Rydin & Clymo, 1989). Our study focuses on the
contribution of internal transport in the upward translocation of nitrogen in a Sphagnum bog.
Sphagnum plants are lacking both roots and a vascular transport system. Rydin & Clymo (1989),
however, demonstrated the internal acropetal transport of carbon and phosphorus in Sphagnum
fallax. They showed the presence of numerous plasmodesmata linking stem cells which create a
possible symplastic (cytoplasm to cytoplasm flow, contrasting apoplastic transport which indi-
cates the flow of solutes through the cell walls) transport pathway. Furthermore, Ligrone & Duckett
(1998b) found cytological evidence for nutrient translocation in Sphagnum. In a light- and electron
microscope study they revealed that the cells in the central region of Sphagnum stems have a
highly specialized cytoplasmic organization which has only been described for assumed solute-
conducting cells in mosses (Ligrone & Duckett, 1994). The presence of these cells, referred to as
“conducting parenchyma cells”, strongly suggests a cellular specialization in symplastic transport
(Ligrone & Duckett, 1998a). Internal transport of nitrogen has often been assumed (Aldous, 2002b;
Bonnett et al., 2010; Bragazza et al., 2005; Gerdol et al., 2006; Limpens & Berendse, 2003; Limpens
48 – Solute transport in Sphagnum dominated bogs
& Heijmans, 2008; Malmer, 1988) but, to our knowledge, has never been demonstrated before.
Here, the ability of Sphagnum to transport nitrogen internally is investigated. Sphagnum
cuspidatum and S. fallax are used in experiments in which diffusion and capillary transport are
excluded and the internal transport of labeled nitrogen is monitored. An attempt is made to eluci-
date the mechanism of transport, apo- or symplastic. By killing a small part of the stem cells sym-
plastic transport is excluded while the transport of solutes through the apoplast is still possible.
The contribution of internal transport in the nutrient supply to Sphagnum and to the nutrient
cycling in a Sphagnum bog with respect to other nutrient transport mechanisms in a Sphagnum bog
will be discussed.
Materials and methods
Plant material
Sphagnum cuspidatum Ehrh. Ex Hoffm. plants were collected in a pool at the edge of a small bog
in the nature reserve “Dwingelderveld” (N52°49.777’, E6°25.994’) in the north of the Netherlands.
Sphagnum fallax (klinggr.) Klinggr. plants were collected in a small bog adjacent to the
“Dwingelderveld” (N52°49.135’, E06°29.491’). The mean annual nitrogen deposition in the north of
the Netherlands is 28 kg ∙ ha-1 ∙ yr-1 (RIVM, 2009).
The plants were collected and stored overnight (dark, 4°C) in plastic containers. Before being
used in the experiment the plants were cut to a length of 10 cm and rinsed three times with demin-
eralized water. Only visually non-damaged and fully green plants were used in the experiments.
Experimental design
To measure the ability of Sphagnum to transport nitrogen internally, three experiments were per-
formed in which the transport of solutes by diffusion and capillary transport was excluded. The ex-
perimental set up for these experiments consisted of two compartments divided by a barrier created
by placing two square Petri-dishes (120*120mm; Greiner bio-one GmbH) against each other. Both
compartments were filled with 85 mL of diluted artificial rainwater (100 times) containing 20 mM
MES buffer (pH = 4.0). To one of the two compartments, the donor compartment, labeled nitrogen 25
µmol ∙ L-1 15NH4Cl (98 atom %; Sigma Aldrich Inc., product number 299251) or 10 µmol ∙ L-1 K15NO
3 (98
atom %; Sigma Aldrich Inc., 335134) was added. The other compartment was called the acceptor dish.
Both solutions were not in contact with each other. Three plants with a length of 10 cm were placed
over the rim dividing the two compartments. The 10 cm long plants had green stems and were there-
fore considered to be alive and healthy. Two Sphagnum species were used in this experiment, Sphag-
num cuspidatum and S. fallax. The compartments were covered with a lid to reduce evapotranspira-
tion. All experiments took place in a climate controlled room at 18±1°C and a 16L:8D photoperiod.
Experiment 1: acropetal transport of nitrogen
In the first experiment the internal transport of ammonium and nitrate from the stem to the capitula
(acropetal transport) was measured. The upper 2 cm of the plants (including the capitulum) were
placed in the acceptor dish. The 8 cm long stem was placed in the donor dish. Plants were harvested
after 1, 2, 4 and 8 days.
Physiological evidence for internal acropetal transport – 49
Experiment 2: Basipetal transport of nitrogen
The potential transport of nitrogen from the capitula to the stem (basipetal transport) was
determined by placing the upper 2 cm of the Sphagnum plants in the donor dish and the stem in
the acceptor dish. Nitrogen was applied as ammonium (15NH4Cl) or nitrate (K15NO
3). Plants were
harvested at day eight.
Experiment 3: Apoplastic transport of nitrogen
To distinguish between apoplastic and symplastic transport of ammonium the plants were again
placed with the stem in the donor compartment, but a small segment of the stem was killed by
steam using a modification of the method described by Rydin & Clymo (1989). Two centimeter
below the capitula a stem section of ca. 1 cm was treated with steam for 60 seconds. The adjoining
sections of the stem were protected from the steam by temporarily covering them with cork. Vital-
ity of the steam-treated and untreated stem sections was determined by FDA staining (Elzenga et
al., 1991; Heslop-Harrison & Heslop-Harrison, 1970). Vital cells were distinguished from non-vital
cells based on their fluorescein green appearances using a Zeiss fluorescence microscope. During
the experiment, the steamed part of the stem became white whereas the untreated parts remained
green. Plants were harvested at day four.
In all three experiments leakage of 15N from the donor to the acceptor compartment by capil-
lary flow along the stem was excluded by covering the stem at the rim dividing the two compart-
ments with white Vaseline (Lamers & Indemans, ‘s Hertogenbosch). The efficiency of this waxy
barrier was tested visually by using the dye Brilliant Blue (Coomassie Brilliant Blue G, No. B0770,
Sigma Aldrich) as a marker. In the acceptor compartment no Brilliant Blue was observed when wax
was used.
A second test for leakage and contamination of the acceptor dishes with 15N, three capitula
(so called “contamination control capitula”) were placed in the acceptor compartment during the
experiment and were harvested together with the other capitula and stems. These capitula indicate
a possible change in 15N concentration in the capitula during the experiment when not exposed to a
source of labeled nitrogen. All experiments were performed in triplicate resulting in 3*3 plants per
sampling day.
Analyses
Upon harvesting the Sphagnum plants were separated into the upper first centimeter (the capitu-
lum) and the undermost seven centimeters of the stem. A 2 cm segment bridging the rim was not
included in the analysis to allow the grinding of the tissue without interference of possible wax
remains. After harvesting the plants were three times thoroughly rinsed with demineralized water
and dried for at least 48 hours at 80°C. Subsequently, the plants were grinded individually to a fine
powder using a ball miller (Retsch MM2, Haan, Germany). The contamination capitula were pooled
to reduce the number of samples. For each sample the %N and %15N (stable nitrogen isotope com-
position) was measured with a Carlo Erba NA 1500 elemental analyzer (Thermo Fisher Scientific
Inc.) coupled online via a Finnigan Conflo III interface with a Thermo-Finnigan DeltaPlus mass-
spectrometer. Additionally, the natural occurring 15N concentration in the capitula and stems of
the Sphagnum plants was determined (day=0 samples).
The data are expressed as the amount of 15N assimilated in the plants (in µmol ∙ g DW-1 and was
50 – Solute transport in Sphagnum dominated bogs
calculated by the formula (%15N * %N / 100)*10000/15, where %N is the percentage of total N in
the sample, %15N is the percentage of 15N of total N and 15 is the molecular weight of the stable N
isotope.
Statistical analyses
In the acropetal experiment the 15N concentrations of the capitula and “contamination control
capitula” in the acceptor dishes at day 1, 2, 4 and 8 were compared to the values in the capitula
and stems at day zero by using a one way ANOVA, with day as independent variable. A Dunnett’s
post-hoc test was performed in case of differences within factors. In both the “basipetal” and
the “apoplastic” experiment, a t-test was used to determine differences in 15N concentrations in
capitula or stems between day 0 and the end of the incubation period.
In general for each factor we had three replicates and three plants per replicate.
Prior to analysis, all data were transformed (1/x) when necessary to meet the assumption of
homogeneous variance. All statistical analysis were performed by using SPSS for Windows (version
16.0.1, 2007; SPSS Inc., Chicago, IL, USA).
Hyperbolic curves were fitted to the ammonium uptake data using graphing software (Prism
version 4.03, 2005; GraphPad Software, Inc., San Diego, CA, USA).
Results
For both Sphagnum cuspidatum and S. fallax a slow but significant acropetal transport of ammoni-
um through an internal mechanism was observed (figure 1). A one way ANOVA showed a significant
difference in 15N concentration between days in the capitula of both S. cuspidatum (F(4,40)=12.389,
p<.01) and S. fallax (F(4,39)=8.522). In the capitula of S. cuspidatum a significant increase of 15N was
observed after four (p=0.12) and eight days (p<.01); 4.8 ± 0.4 and 7.1 ± 0.9 µmol ∙ g DW-1, respectively,
compared to 3.7 ± 0.3 µmol 15N ∙ g DW-1 at the beginning of the experiment (day=0). For S. fallax a
significant increase of 15N in the capitula was observed at day eight (p=0.001); 4.0 ± 0.3 µmol ∙ g
DW-1 compared to 2.8 ± 0.2 at day=0. The concentration of 15N in the contamination control capitula
remained very low throughout the experiment (average value; p>0.05), from which we concluded
that no contamination occurred and that the concentration of 15N in the Sphagnum mosses was
not affected by the experimental procedure itself. Most obvious is the rapid uptake of 15NH4
+ by the
stems of both Sphagnum cuspidatum and S. fallax for which within one day respectively 64 and 84%
of the maximum concentration is reached (figure 1).
For nitrate the uptake by the stems of S. cuspidatum shows a similar pattern as ammonium;
74% is taken up within the first day (figure 1). On the other hand, hardly any NO3
- was taken up by
the stems of Sphagnum fallax.
A one way ANOVA showed no significant difference in 15N concentration between days in
the capitula of S. cuspidatum (F(4,40)=1.940, p>.05) and S. fallax (F(4,40)=1.225, p>.05). Also no
significant difference between 15N concentration in the contamination control capitula between
days was observed from which we assumed that no contamination occurred.
Physiological evidence for internal acropetal transport – 51
Figure 1. Mean 15N concentration (± SE) in the stems (grey circles), capitula (black circles) and contamination capitula (open
circles; ± SD) in Sphagnum cuspidatum (a,c) and Sphagnum fallax (b,d) measured after 0, 1, 2, 4 and 8 days of incubation of the
stems in 15NH4Cl (a and b) or K15NO
3 (c and d). Because of large differences in 15N concentration between the capitula and stems
these values are plotted distinctively on respectively the left and the right y-axes. Notice that the concentration of NO3
- in the
donor compartment was lower than ammonium (10 vs. 25 µmol ∙ L-1). Significant increase of 15N in the capitula compared to
the concentration at the start of the experiment (day = 0) are indicated by an asterisk (p<0.05). When no error bar is visible the
standard error is lower than the size of the circles. For clarity the untransformed data are shown.
In the experiment designed to determine possible basipetal transport of nitrogen, the 15NO3
- and 15NH
4+ was readily taken up from the experimental solution by the capitula in both Sphagnum
cuspidatum and S. fallax within eight days (figure 2). However, no significant increase of 15N in the
stems was observed after 8 days in S. cuspidatum with either ammonium or nitrate (t(16)=-2.048
and t(16)=0.163, resp., p>0.05) and also S. fallax showed no significant increase with NH4
+ and NO3
-
(t(16)= -1.523 and t(16)=0.584, resp., p>0.05; figure 2).
When a 1 cm segment of the stem was killed by steam no significant differences in 15N con-
centration between the capitula at day 0 and day 4 were observed for both Sphagnum cuspidatum
(t(15)= -0.76, p>0.05) and S. fallax (t(15)= -0.600, p>0.05; figure 3) and the acropetal transport as
demonstrated in figure 1 is inhibited.
52 – Solute transport in Sphagnum dominated bogs
Figure 2. The average concentration of 15N (± SE) per gram dry weight in the stems (black circles), capitula (white circles) and
contamination capitula (± SD; grey circles) in Sphagnum cuspidatum and Sphagnum fallax measured after the incubation of the
capitula in 15NH4Cl or K15NO
3 for 0 and 8 days. No significant differences in 15N concentration between days in the stems of both
S. cuspidatum and S. fallax were observed. When no error bar is visible the standard error is lower than the size of the circles.
For clarity the untransformed data are shown.
Discussion
Internal transport of nitrogen in Sphagnum
For both Sphagnum cuspidatum and S. fallax a slow but significant acropetal transport of nitrogen
through an internal mechanism was observed (figure 1). In this chapter we describe that live stems
of S. cuspidatum and S. fallax do take up both, NH4
+ and NO3
- , although with different efficiencies,
and that nitrogen supplied as NH4
+ is subsequently transported acropetally to the capitulum. The
difference between NH4
+ and NO3
- uptake efficiency will also have been determined by the concen-
trations in which both N species were applied, 25 and 10 µmol ∙ L-1 for NH4
+ and NO3
-, respectively.
Yet, these values were chosen since they represent natural bog water concentrations under natural
conditions.
After uptake, both NO3
- and NH4
+ are assimilated into the amino acid glutamine (Gln) and
subsequently converted into other amino acids (Kahl et al., 1997; Rudolph et al., 1993). Therefore, it
is assumed that transport of N takes place in the form of amino acids.
When a segment of the stem is killed by steam, Sphagnum plants no longer show a significant
Physiological evidence for internal acropetal transport – 53
Figure 3. The average concentration (± SE) of 15N in the stems (black circles), capitula (open circles) and contamination capitula
(± SD; grey circles) in Sphagnum cuspidatum and S. fallax measured after the incubation of the stems in 15NH4Cl for 0 and 4 days.
A one centimeter segment of the stem was killed by steam. No significant differences in 15N concentration between days in the
capitula of both S. cuspidatum and S. fallax were observed. When no error bar is visible the standard error is lower than the size
of the circles. For clarity the untransformed data are shown.
transport of 15N to the capitula within four days. Killing a segment of the stem by steam, blocks
symplastic transport of compounds, but should still allow apoplastic transport (Rydin & Clymo,
1989). Therefore, these results are indicative for the symplastic nature of acropetal transport of
nitrogen, and are in full agreement with the findings of Ligrone and Duckett (1998b) and Rydin
& Clymo (1989) who demonstrated cellular specializations of Sphagnum for symplastic transport.
Nevertheless, the duration of four days might be too short to totally exclude a small contribution by
apoplastic transport. An alternative hypothesis is the diffusive transport of NH4
+ and NO3
- through
the cell wall and the hyaline cells present in the epidermis of the stem of Sphagnum. However, dif-
fusive extracellular transport would result in similar transport rates in both acropetal and basipetal
direction. In contrast, 15N is not being transported into the stem due to basipetal transport of 15N in
experiment 2. In our experiment we only observed net transport of nitrogen from stem to capitula.
A possible explanation for this unidirect transport might be the higher sink strength for N of the
capitulum, relative to the stem, which can be expected because growth is taking place exclusively
in the capitulum.
Since mineralized nitrogen is an important nitrogen source (Aerts et al., 1999; Aldous, 2002b;
Bridgham, 2002; Gerdol et al., 2006; Morris, 1991; Urban & Eisenreich, 1988), that becomes available
in the deeper layers, distinct from the growing capitulum in the top layer, the supply of nitrogen
depends on upward-directed transport mechanisms (Aldous, 2002b; Bridgham, 2002). Uptake by
the stem and subsequent internal transport to the growing parts is thus a possible pathway for
mineralized nitrogen.
Internal, stem-mediated transport can be involve in re-allocation of nitrogen from older,
senescing tissue, a nutrient retention mechanism common in vascular plants (Aerts, 1990, 1995;
Chapin, 1980; Vitousek, 1982) and in transport of nitrogen taken up from the external medium
by the stems. This distinction must be taken into consideration when reviewing the importance
of different processes in the transport of externally mineralized nutrients. Gerdol et al. (2006)
stated that the mineralization of N is more important than the enzymatic reallocation of N.
54 – Solute transport in Sphagnum dominated bogs
Our experimental procedure only involves the translocation of the externally mineralized nitro-
gen. Higher C:N ratios in stems than in capitula are often observed in Sphagnum (e.g. Malmer, 1988)
and are taken as an indication for the internal reallocation of N from the stem to capitula. This
assumption, in combination with the internal transport of C and P (Rydin & Clymo, 1989), were
reasons for the general acceptance of internal transport in Sphagnum (Bonnett et al., 2010; Bragazza
et al., 2005; Gerdol et al., 2006; Limpens & Berendse, 2003; Limpens & Heijmans, 2008; Malmer,
1988). However, experimental evidence for the internal reallocation of N was lacking. Our findings
provide physiological evidence for internal transport of N taken up from the external medium.
There is, however, no reason to assume that the same mechanism does not transport N internally
released from senescing tissue.
NH4
+ and NO3
-
Within the time frame of the experiments the internal transport of nitrogen was only observed
when nitrogen was supplied as NH4
+ and not as NO3
-. According to a review by Rudolph et al. (1993)
this difference might have a metabolic origin. After uptake NO3
- is reduced to NH4
+, which takes
place in the chloroplast. The subsequent assimilation into amino acids also takes place in the
chloroplast. On the other hand, the assimilation of NH4
+ can take place in the cytosol (Rudolph
et al., 1993). Taking symplastic transport into account (see below), the N present in the cytosol is
available for transport whereas the amino acids in the chloroplast are more or less ‘fixed’.
The uptake of NO3
- by the stems differed between the two species. The stems of S. fallax hardly
took up NO3
- within the eight days of the experiment. Wiedermann et al. (2009) showed NO3
- to
be taken up in small amounts by capitula of Sphagnum balticum and S. fuscum from a solution
containing in total four N forms (NH4
+, alanine and glutamine and NO3
-). According to Woodin &
Lee (1987), most absorption of nitrate takes place in the capitulum and decreases down the stem.
With the stems hardly taking up NO3
- the transport to the capitula can not be expected.
The rate of internal transport
In earlier studies (Aldous, 2002b; Bridgham, 2002) the contribution of translocation to the nitrogen
supply of the capitula was shown to be significant. However, these studies did not distinguish be-
tween different types of transport. Based on the uptake and internal transport of ammonium data by
S. cuspidatum (figure 1) the rate by which nitrogen is transported internally can be estimated. Since
already after one day the maximum concentration of 15N in the stem is nearly reached, we can as-
sume that the concentration in the stem is almost constant for the duration of the experiment. Since
there is a lag period of almost 4 days before labeled N appears in the capitulum and we know the
length of the stem segment between the donor compartment and the receiving capitulum (which is
2 cm), we can calculate the speed of the acropetal, symplastic transport process: 5 mm/d. From the
increase in the capitulum in the four days following the lag, 2.3 µmol ∙ g DW-1 or 12% of the total N
taken up by the stems, we can calculate a half time value of equilibration between donor and recep-
tor sections of the plant: 17 days. Notice that this calculation is based on the assumption that the
amount of 15N in the capitula is a function of the amount of 15N taken up by the stems in the preced-
ing four days. It should be noted that the delay and apparent transport speed taken the assimilation
of NH4
+ into amino acids is also included. This calculated half time value for N is higher than the
estimated half time value of 11 days for the internal transport of C and P (Rydin & Clymo, 1989).
Physiological evidence for internal acropetal transport – 55
Since the internal transport of nitrogen is a mechanism for efficient nitrogen use, the transport
rate is expected to be reduced under high N content in the capitula (Bragazza et al., 2004). The
plants used in these experiments were collected from an area with a high load of atmospheric
nitrogen deposition. It has been demonstrated that Sphagnum mosses subjected to a high N sup-
ply accumulate elevated amounts of N (Nordin & Gunnarsson, 2000; Van der Heijden et al., 2000;
Limpens & Berendse, 2003; Limpens et al. 2011). Moreover, The capitula of the Sphagnum cuspida-
tum plants used in our experiments also showed a relatively high N content: 15.3 ± 2.2 mg ∙ g-1. If
indeed internal transport rates are negatively affected by nitrogen supply, our estimates are likely
to be an underestimation for Sphagnum residing in non-polluted areas and internal transport
might be more important.
The importance of internal transport
The uptake of ammonium by the living stems and the subsequent transport of N to the capitula
shows that internal transport very likely functions as a mechanism in supplying the capitula with
N. Moreover, it implies that Sphagnum mosses are able to compete with microbes and vascular
plant roots for available soil nitrogen (see Bridgham, 2002) and thereby might function as a N
retention step contributing to the efficient use of N by Sphagnum in ombrotrophic bogs.
However, the importance of internal transport in the nutrient supply of Sphagnum and nutrient
cycling in bogs depends on the transport rate relative to other transport mechanism. In a review of
the internal transport in non-vascular plants (Raven, 2003) it is claimed that there is no evidence for
symplastic transport in Sphagna faster than can be accounted for by diffusion. Next, the rate at which
nitrogen is transported internally is very slow compared to the transport rates by buoyancy-driven
water flow (Rappoldt et al., 2003). Chapter 2 shows buoyancy flow to be a fast and effective nutrient
transport mechanism in bog water. The uptake kinetics of Sphagnum cuspidatum and S. fallax show
the ability of Sphagnum to take up large amounts of ammonium relatively fast, indicating buoyancy
flow to be a very effective nutrient transport mechanism in supplying the capitula with nitrogen.
Therefore, with the regular occurrence of buoyancy flow, the supply of nitrogen by internal
transport might be insignificant, compared to the supply of nitrogen by buoyancy flow. Indeed,
buoyancy flow is restricted to water-saturated Sphagnum habitats and, because of its dependence
on varying physical parameters like the difference in temperature between day and night, an
irregularly occurring phenomenon. The importance of internal transport might therefore reside in
its continuous character, continuously supplying Sphagnum with N which contrasts with the pulse
wise supply of N by buoyancy flow (and of course precipitation). For Sphagnum species that form
hummocks that extend above the water surface and do not benefit from buoyancy flow, internal
transport is, next to capillary transport, a possible acropetal pathway for nutrients. Clymo (1973)
estimated the average velocity by capillary flow to be 0.4 mm ∙ min-1. However, this rate, and the
concomitant nutrient transport, is dependent of several factors, like evaporation, plant density
and pore water nutrient concentrations (Clymo & Hayward, 1982). Moreover, during extracellular
transport nutrients may be lost to microorganism or vascular plant roots. Compared to the external
transport mechanisms (buoyancy flow and capillary transport) internal transport very likely rep-
resents a minor contribution in the upward transport of externally supplied N to the capitula. The
main importance of internal transport is therefore considered to be the reallocation of internally
catabolized nitrogen compounds.
Chapter 5
The importance of groundwater carbon dioxide in the restoration of Sphagnum bogs
Wouter Patberg
Gert Jan Baaijens
Alfons Smolders
Ab Grootjans
Theo Elzenga
58 – Solute transport in Sphagnum dominated bogs
Abstract
Essential for successful bog restoration is the re-establishment of Sphagnum
mosses. High CO2 availability has been shown to be of great importance for
the growth of Sphagnum mosses. In well-developed Sphagnum bogs large
amounts of CO2 are produced by decomposition processes in the peat layer.
In cut-over Sphagnum bogs this carbon source is often absent or strongly re-
duced. Therefore, for the successful restoration of cut-over Sphagnum bogs
an alternative, additional carbon source might be essential for the re-estab-
lishment of Sphagnum mosses. This chapter focuses on the role of CO2 in the
development of Sphagnum bogs in a field situation.
Study area is one of the largest wet heathland reserves in Western Europe
and is characterized by many small damaged Sphagnum bogs. Rewetting
measures resulted in large developmental differences between bogs; some
bogs developed markedly well, whereas others did not. Of ten small bogs the
developmental success was quantified using aerial photographs and surface
water and groundwater samples were collected. In addition, the physiologi-
cal characteristics of carbon dioxide uptake of two Sphagnum species were
determined.
Water chemistry analysis revealed that the total inorganic carbon concen-
tration (TIC) in the nearby groundwater of the well-developed bogs, is sig-
nificantly higher than that of poorly developed bogs. The CO2 availability in
the surface water of the investigated bogs was positively correlated to the in-
organic carbon in the groundwater. It is concluded that the well-developing
bogs are fed by a carbon-rich groundwater inflow from outside the bog.
The CO2 uptake kinetics of both Sphagnum species are characterized by a
high compensation point and a low affinity, both indicating an adaptation
to a high CO2 availability.
The present findings indicate that high carbon dioxide availability is a pre-
requisite for the successful re-establishment of Sphagnum mosses in peat
bog restoration projects and that carbon-rich groundwater can apparently
substitute for the decomposing peat layer as a source of CO2. Therefore, the
availability of CO2 should be included in bog restoration feasibility studies.
The importance of groundwater – 59
Introduction
Due to the extensive exploitation for fuel, agriculture and forestry over many centuries, living (peat
forming) mires have become endangered in most of North-Western Europe (Rochefort & Price,
2003). Even nowadays (extensive) peat extraction activities take place for commercial use in, for
example, Canada, Scandinavia, Ireland and the Baltic states (Joosten, 2009). Due to the important
role of peatlands in the global carbon cycle, and because of their unique ecological values, globally
much effort is dedicated to the restoration of damaged mires. However, the restoration of large bog
remnants in particular, has proven to be fairly complicated and not always successful (Grootjans et
al., 2006; Money & Wheeler, 1999; Money et al., 2009).
Essential for successful bog restoration is the re-establishment of Sphagnum mosses followed
by the re-development of a functional acrotelm, leading to a self-sustaining system (Money &
Wheeler, 1999; Money et al., 2009; Smolders et al., 2003). Since wet conditions are essential for
Sphagnum growth (Robroek et al., 2009), the creation of suitable wet conditions is a prerequisite in
restoring peatlands (e.g. Money et al. 2009). Often rewetting is realized by inundating large areas to
ensure wet conditions throughout the year (Money & Wheeler, 1999; Smolders et al., 2003). The wa-
ter layer can be colonized by aquatic Sphagnum species, especially Sphagnum cuspidatum, to form
dense mats on which peat forming species like S. magellanicum and S. papillosum might establish
(Money & Wheeler, 1999; Wheeler & Shaw, 1995). However, this method often results in large water
bodies in which Sphagnum growth is severely hampered. The lack of success in re-colonization of
aquatic Sphagnum species in rewetted bog remnants has been ascribed to the limited availability
of light and/or CO2 (Money & Wheeler, 1999; Smolders et al., 2001; Smolders et al., 2003; Wheeler &
Shaw, 1995).
Like most aquatic bryophytes (Raven et al., 1985) Sphagnum mosses are known to be obligate
CO2 users (Bain & Proctor, 1980) and are solely dependent on the diffusive supply of CO
2 to the site
of carbon fixation (Rubisco - ribulose-1,5-bisphosphate carboxylase-oxygenase). In very wet condi-
tions the Sphagnum mosses are surrounded by a thick water layer which lowers CO2 conductivity
resulting in a reduced photosynthetic rate (Silvola, 1990; Williams & Flanagan, 1996). Consequent-
ly, high rates of underwater photosynthesis can only be sustained when the leaves are exposed to
high levels of CO2 (Jauhiainen & Silvola, 1999; Paffen & Roelofs, 1991; Silvola, 1990; Smolders et al.,
2003).
In well developed bogs CO2 is produced in large quantities by decomposition processes in the
peat layer (Bridgham & Richardson, 1992; Glatzel et al., 2004; Smolders et al., 2001; Waddington
et al., 2001). Carbon dioxide concentrations in the pore water can reach up to several millimo-
lar (Smolders et al., 2001; Smolders et al., 2003), compensating low diffusion rates and ensuring
sufficient substrate delivery for photosynthetic carbon fixation (Maberly & Madsen, 2002; Silvola,
1990). This so-called substrate-derived CO2 has been shown to be an important carbon source for
aquatic and emergent Sphagnum mosses (Baker & Boatman, 1990; Paffen & Roelofs, 1991; Riis &
SandJensen, 1997; Roelofs, 1983; Smolders et al., 2001; Smolders et al., 2003). Under more reductive
conditions the methane production in the catotelm may become higher than the production of
CO2. This methane can be oxidized by methanotrophic bacteria to CO
2, which then can be used as a
carbon source by Sphagnum mosses (Kip et al., 2010).
Cut-over peat bogs often lack this source of additional inorganic carbon. This type of damage
60 – Solute transport in Sphagnum dominated bogs
can often be found in North-Western Europe (Joosten, 2009). Peat extraction has removed the bulk
of organic material. The highly decomposed, humified peat which is left behind, has only limited
CO2 (and methane) production rates (Bridgham & Richardson, 1992; Glatzel et al., 2004; Tomassen
et al., 2004; Waddington et al., 2001). Therefore, for the successful restoration of cut-over Sphag-
num bogs an additional carbon source might be essential for the re-establishment of Sphagnum
mosses.
This study focuses on the role of CO2 in the development of Sphagnum bogs in a field situation.
The study area is one of the largest wet heathland reserves in Western Europe and is characterized
by numerous small Sphagnum bogs. They have been damaged by drainage and small scale peat ex-
cavations in the past. From 1988 onwards, rewetting measures have been carried out, but the devel-
opmental success has varied significantly between bogs; some bogs developed well, whereas oth-
ers did not. It is hypothesized that in these hydrologically degraded bog remnants the restoration
of Sphagnum growth is limited by the availability of CO2. We expect that the well-developing bogs
are being fed by lateral, carbon-rich, groundwater inflow. Groundwater with high inorganic carbon
concentrations entering a bog, will release high amounts of CO2 when it comes into contact with
the more acidic water around the Sphagnum mass. The higher availability of CO2 will stimulate the
growth of aquatic Sphagnum mosses and subsequent bog development. We also determined the
CO2 requirement for two Sphagnum species (S. cuspidatum and S. fallax), which are abundant in
well-developing bogs.
Materials & Methods
Study area
Study area is the “Dwingelderveld”, one of Europe’s largest wet heathland areas (about 3500 hectares),
situated in the north of the Netherlands (52°49’6.71”N 6°27’28.48”E). The landscape consists of pine
forest, wet and dry heathland and many small peat bogs scattered throughout the area (figure 1).
The presence of boulder clay underneath the reserve is responsible for the generally wet character
of the area. Wind erosion resulted in differences of up to 5 meters in height of the Pleistocene
sand cover. During the second half of the last century most of the wet heathland and bogs became
desiccated by drainage activities both in the reserve (for the benefit of pine plantations) and also in
the surrounding brook valleys (for the benefit of agriculture). Many small bogs were also subjected
to small scale peat cutting by farmers. These activities have ended around 1950.
In 1988 large scale rewetting measures, i.e. closing of drainage ditches and cutting of trees, were
initiated with variable results in developmental success of the different bogs. Some bogs developed
well and are characterized by a luxurious growth of Sphagnum spp., whereas others did not and
mainly consist of open water with marginal occurrence of Sphagnum mosses (Grootjans et al.,
2003).
An interesting observation by Grootjans et al. (2003) was that bogs with abundant Sphagnum
growth were located in old erosion gullies, while bogs without Sphagnum-dominated succession
were found outside or at the edge of these gullies. In these erosion gullies impermeable podsolic
layers stretch out beyond the border of the bogs itself. The groundwater levels in the sandy hills are
generally higher than in the gullies. Since the vertical conductance of these podsolic layers is very
The importance of groundwater – 61
low, a horizontal sub-surface flow of groundwater towards the bogs is facilitated, prolonging the
residence time of water in the soil and allowing the groundwater to become possibly enriched with
inorganic carbon (Grootjans et al., 2003; figure 2).
Figure 1. Aerial photograph of the research area in the “Dwingelderveld”. All bogs in this part of the nature reserve are outlined
by a black line. The bogs used in this field study are indicated by a number corresponding to the numbers used in table 1. The
numbers of the poorly developed bogs are underlined. The locations of the sampling sites are indicates by black circles. The
white spot in the Diepveen (6) is open water as proven by field observations.
Classifying peat bogs
In 10 bogs 20 sampling sites were selected (figure 1; table 1). Aerial photographs of 1982 and 2006
were used to determine the developmental success of each of the sampling sites. The increase in
surface area covered by Sphagnum mosses was used as a measure for the success of bog develop-
ment. With the use of image analysis software (ImageJ, version 1.41o, National Institute of Health,
USA) the area of open water per sampling site in both 1982 and 2006 was calculated based on grey
scale differences between vegetation and open water. The developmental success was determined
by calculating the relative decrease of open water surface between 1982 and 2006. The following
formula was used: (%OW1982
- %OW2006
) / %OW1982
, where %OW is the surface percentage of the
bog occupied by open water in 1982 or 2006. In this aerial photograph analysis the vegetation was
assumed to be bog vegetation dominated by Sphagnum mosses. This was validated by field observa-
tions. In some cases, Cootjes Veen, Groote Veen East and Veerles Veen, the aerial photograph analy-
sis had to be adjusted. Here the vegetation was dominated by vascular plants (e.g. Molinia caerulea)
instead of Sphagnum mosses. Consequently these bogs were classified as poorly developed bogs.
62 – Solute transport in Sphagnum dominated bogs
Table 1. Names, coordinates, the percentage open water in 1982 and 2006, the decrease of open water and the developmental
success of the investigated bogs in the “Dwingelderveld”. The developmental success of the bogs was determined by calculating
the relative decrease of open water surface in 2006 compared to the surface open water in 1982. A “+” indicates a well developed
bog and a “-“ indicates a poorly developed bog. For three bogs the developmental success was not determined by aerial photo-
graphs as indicated by an “*”. See the materials and methods section for a more detailed explanation. The numbers in the first
column correspond with the numbers in figure 1
# Name Coordinates
Surface
open water %
Decrease
open water
(%)
Develop mental
success
1982 2006
1 Barkmans Veen N52°49.423’ E6°26.283’ 29 6 78 +
2 Groote Veen N52°49.178’ E6°25.991’ 51 0 100 +
3 Reigersplas N52°50.019’ E6°26.946’ 62 0 100 +
4 Adderveen N52°49.885’ E6°27.058’ 74 43 42 -
5 Cootjes Veen N52°49.000’ E6°26.244’ 0 0 * -
6 Diepveen N52°49.140’ E6°26.404’ 52 56 -8 -
2 Groote Veen East N52°49.171’ E6°26.243’ 71 0 * -
7 Kliploo N52°50.082’ E6°26.380’ 100 98 2 -
8 Schurenberg N52°49.551’ E6°25.951’ 100 83 17 -
9 Veerles Veen N52°49.003’ E6°25.992’ 61 0 * -
10 Zandveen N52°49.694’ E6°26.471’ 85 69 19 -
Water sampling and analysis
Groundwater and surface water samples were taken at the sampling sites in February and April
2007, August and October 2008 and September 2009. Groundwater samples were collected using
piezometers (Ø32 mm PVC tubes with nylon filters) and a peristaltic pump. The piezometers
were placed just outside the bogs on the slopes of the gully and always with the filters above the
impervious layer. The exact position of the piezometers was chosen such that they intercepted the
groundwater flowing into the bogs, based on the results of a previous hydrological study of the area
(Verschoor et al., 2003). One day before sampling the water present in the piezometers was discard-
ed to allow refilling with fresh groundwater. Surface water samples were taken in the bog close to
the piezometers. 30 mL airtight bottles were filled by gently submersing them in the surface water.
Water samples were transported in a cool box to the laboratory where pH and TIC measurements
were performed immediately. The remaining samples were stored frozen until further analyses.
Precipitation data of the “Dwingelderveld” were obtained from the Royal Netherlands Meteorologi-
cal Institute (www.knmi.nl, station number 327 “Dwingeloo”).
The concentration of Total Inorganic Carbon (TIC) in the water samples was determined by
measuring the CO2, released after acidifying the samples to a pH <3, using an Infra-Red Gas Analyzer
(IRGA, ABB Advance Optima). The pH of the water samples was determined using a combined pH
electrode with an Ag⁄AgCl internal reference (Cole Parmer Instrument Company, Illinois, USA) and
a PHM 64 pH meter (Radiometer, Copenhagen). The concentrations of CO2 and bicarbonate in the
water samples were calculated based on the pH and the TIC concentration (Prins & Elzenga, 1989).
The importance of groundwater – 63
Concentrations of nitrate (NO3), ammonium (NH
4) and chloride (Cl) were measured colourimetri-
cally according to Geurts et al. (2008) and potassium (K) by flame photometry by using an Auto
Analyzer 3 system (Bran+Luebbe, Norderstedt, Germany). Aluminium (Al), calcium (Ca), iron (Fe),
magnesium (Mg), manganese (Mn), sodium (Na), total phosphorus (P), sulphur (S), silicon (Si) and
zinc (Zn) were measured using an ICP Spectrometer (IRIS Intrepid II, Thermo Electron Corporation,
Franklin, MA).
Figure 2. A cross section of a part of the study area in the “Dwingelderveld”. The small bogs are situated in the gullies which
are surrounded by the higher Pleistocene sand cover. A layer of boulder clay is present underneath the whole area, resulting
in wet conditions throughout the area. The bogs are characterized by the presence of podsolic layers, responsible for the wet
conditions and bog development. Note the lack of peat development in the bog in the forefront which lies outside the gully.
Groundwater samples were taken adjacent to the border of the bogs, as indicated by the piezometer.
CO2 uptake characteristics
Carbon dioxide uptake characteristics of Sphagnum cuspidatum Ehrh. Ex Hoffm. and S. fallax
(klinggr.) Klinggr., two pioneer Sphagnum species important in the initial stage of bog formation
(Money, 1995; Smolders et al., 2003), were determined by measuring the photosynthetic activity
(A) at different CO2 concentrations at saturating light conditions (1500 µmol ∙ m-2 ∙ s-1; Hansatech
Quantitherm Light meter). Plants were collected in April 2008 in two small Sphagnum bogs in the
Dwingelderveld area.
A capitulum was placed in a closed thermostatic cuvette containing 1 mL of measuring
solution (see below) which was stirred continuously. The photosynthetic evolution of oxygen was
measured by a Clark electrode located at the bottom of the cuvette in combination with a millivolt
recorder. The temperature of the measuring solution was 21°C.
Any inorganic carbon present in the hyaline cells or adhering water was removed by illuminat-
ing the capitula with 1000 µmol ∙ m-2 ∙ s-1 for at least 60 minutes while keeping them in a CO2 free
medium. When the capitula showed a steady, low rate of oxygen uptake it was assumed that no sig-
nificant CO2 stores were left in the hyaline cells. The solution was then removed from the cuvette
by using a syringe and replaced by a solution with the desired CO2 concentration. Different CO
2
concentrations ranging from 0 to 800 µmol ∙ L-1 in 10 times diluted artificial rainwater (Smolders et
al., 2001) containing 20 mM MES pH = 5.5 were created of which 1 mL was used in the cuvette and
64 – Solute transport in Sphagnum dominated bogs
1 mL was used to exactly determine the Total Inorganic Carbon concentration, by using an infrared
gas analyzer (CO2 analyzer model no. S151, QUBIT Systems Inc., Kingston, ON, Canada). The pH of
the measuring solution was determined using a pH microelectrode (type MI-406, Microelectrodes
Inc., Bedford, NH, USA) in combination with an Ag⁄AgCl micro-reference electrode (type MI-401,
Microelectrodes Inc., Bedford, NH, USA) and a millivolt meter. The concentrations of CO2 and bi-
carbonate in the water samples were calculated based on the pH and the TIC concentration (Prins
& Elzenga, 1989).
Per measurement one capitulum was used and each capitulum was used for a maximum of
three measurements. Large branches were trimmed to fit in the cuvette. After usage capitula were
frozen at -80°C, ground to a powder and the chlorophyll concentration was determined according
to Lichtenthaler (1987).
Statistical analysis
Data were tested for normality using a Kolmogorov-Smirnov test and equality of variance using
Levene’s test. The assumption of homogeneity of variance was not always met, not even after
transformation of the data. According to Heath (1995), the analysis of variance appears not to be
greatly affected by heterogeneity in variance if sample sizes are more or less equal. Therefore, we
decided to continue our analysis using non-transformed data. Differences in groundwater TIC
concentration and surface water CO2 concentration between well and poorly developed bogs, were
tested using a mixed model, with bog development (well and poor) as fixed factor and sampling
date as repeated measure. Differences between sampling dates were determined by using Bonfer-
roni’s post-hoc test. The Pearson correlation coefficient was determined for the relation between
groundwater TIC and surface water CO2 concentrations (both log transformed). The mixed model
and the Pearson correlation test were performed using SPSS for Windows (version 16.0.1, 2007;
SPSS Inc., Chicago, IL, USA).
Additionally, the data were analyzed by applying principal component analyses (PCA) by using
Aabel (version 3.0.3; Gigawiz Ltd. Co., Tulsa, OK, USA). All data were normalized. A hyperbolic
curve was fitted to the CO2 uptake data using graphing software (Prism version 4.03, 2005; Graph-
Pad Software, Inc., San Diego, CA, USA).
Results
Classification of peat bogs
Based on the aerial photographs and field observations the developmental success of the selected
bogs was classified into two categories; well developed bogs, with an open water decrease of at
least 78% and poorly developed bogs with a decrease of open water less than 20% (table 1). The
“Adderveen”, however, is an exception with an open water decrease of 42%. For most sites the field
observations confirmed the classification based on the aerial photograph analysis: well developed
bogs showed luxurious and dominant growth of Sphagnum spp. without or with little open water,
whereas in poorly developed bogs marginal Sphagnum spp. growth was accompanied by the pres-
ence of vascular plants and open water. Open water surface decreased in all sites between 1982 and
2006, except for Diepveen, where the open water surface slightly increased with 8%.
The importance of groundwater – 65
Table 2. Water analysis of the groundwater and surface water of well (+) and poorly (-) developed bogs; average values for all
sampling dates are given with standard deviation (SD). Ion concentrations are given in µmol ∙ L-1
Groundwater Surface water
+ - + -
mean SD mean SD mean SD mean SD
TIC 4983.7 802.4 2765.6 1426.6 1230.3 736.8 768.1 638.4
HCO3
- 152.7 102.9 64.2 72.0 15.2 17.9 25.0 50.9
CO2
4831.0 756.2 2701.4 1389.7 1215.1 730.3 743.1 625.5
pH 4.8 0.3 4.5 0.5 4.3 0.3 4.7 0.5
NH4
94.6 54.7 84.4 78.5 76.5 70.3 71.6 80.0
NO3
7.4 14.6 8.3 12.5 8.3 13.9 9.4 14.8
K 14.1 10.6 25.5 24.2 26.4 19.2 34.2 27.7
P 2.6 4.1 1.9 1.9 4.7 7.8 3.9 7.6
Al 42.2 31.1 36.4 33.0 10.1 7.5 7.5 9.3
Ca 61.9 59.6 51.9 32.1 30.5 34.5 34.2 32.0
Cl 246.4 58.1 341.2 181.7 256.3 80.3 289.5 152.3
Fe 61.1 24.0 24.5 18.2 42.4 107.2 12.1 13.9
Mg 67.0 21.4 42.4 26.7 26.3 14.0 30.3 13.0
Mn 0.5 0.2 1.0 1.2 0.6 0.3 1.2 1.5
Na 231.4 38.2 284.8 130.2 192.3 48.0 227.0 94.6
S 39.2 18.1 53.5 55.9 22.0 15.8 27.9 21.8
Si 263.2 105.8 121.0 83.9 35.8 22.1 19.3 21.9
Zn 8.7 13.7 5.4 5.3 0.9 0.9 0.8 1.0
Water chemistry
Bog water chemistry data are shown in table 2. The difference in groundwater composition between
the well developed and poorly developed bogs is illustrated by a principal component analysis (figure
3). The groundwater samples from well and poorly developed bogs appear as clusters in the PCA
diagram along the first principal component axis that is dominated by high TIC, iron (Fe) and silicium
(Si), indicating that good Sphagnum development is associated with groundwater rich in inorganic
carbon. The groundwater is relatively acid and CO2 is the main inorganic carbon species (table 2).
The chemical composition of groundwater and surface water is clearly different, with concentrations
of most multivalent minerals and inorganic carbon being higher in the groundwater (table 2).
Groundwater TIC concentrations are shown per location in figure 4. The average ground-
water TIC concentration near the well developed bogs was 4983 ± 802 µmol ∙ L-1 and ranged from
2620 to 6215 µmol ∙ L-1. The poorly developed bogs, with an average TIC concentration of 2765 ±
1426 µmol ∙ L-1, showed a much wider range in TIC concentration both between measurements at
66 – Solute transport in Sphagnum dominated bogs
individual locations and between locations. A significant main effect of category on groundwater
TIC concentrations was found (F(1,12) = 20.246, p=0.001). In the well developed bogs the average
CO2 concentration in the surface water was 1215 ± 730 µmol ∙ L-1 and for the poorly developed bogs
743 ± 625 µmol ∙ L-1 (table 2). For surface water CO2 concentration also a significant main effect of
category was found, F(1,13) = 6.063, p=0.029.
Figure 3. Principal Component Analysis (PCA) biplot of all groundwater samples and selected environmental variables. Each
symbol represents a sampling location at one of the sampling dates. Black circles are the well developed bogs and the open
circles represent the poorly developed bogs. The first axis explains 27% of the variation and second principal component
accounts for 20 % of the variation.
Figure 4. Box plot showing the total inorganic carbon (TIC) concentration in the groundwater in µmol ∙ L-1 per location of
both well (gray boxes) and poorly (white boxes) developed bogs. Box plots are composed of minimum, maximum, 25%, 75%
quartiles and the median. Where used, north, south, east and west, indicate sampling sites at one bog.
The importance of groundwater – 67
Figure 5. Groundwater total inorganic carbon concentrations (TIC, solid lines) and surface water CO2 concentrations (dashed
lines) for the well developed bogs (filled symbols) and the poorly developed bogs (open symbols) shown per sampling date.
Symbols represent mean values in µmol ∙ L-1. Bars represent standard deviations and are one sided for readability of the graph.
Different letters mean significant differences between sampling dates, tested for groundwater TIC and surface water CO2
concentrations separately.
Figure 5 shows the changes over time in the total inorganic carbon and CO2 concentration of both
groundwater and surface water of well and poorly developed bogs. A significant main effect of
date on both groundwater TIC and surface water CO2 concentrations was found (F(4,48) = 5.734 and
F(4,52) = 6.126, respectively, p<0.01); the groundwater TIC concentration was significantly higher
on August 15 2008 than in February 8 2007 (p<0.05). For the surface water CO2 concentrations
significant higher values were found in April and September (figure 5). However, no significant
interaction effect of category * date was found for both groundwater TIC (F(4,48) = 0.566, p>0.05)
and surface water CO2 concentrations (F(4,52) = 1.317, p>0.05). In other words, groundwater TIC
and surface water CO2 concentrations in well and poorly developed bogs, respectively, were not
affected differently by date.
The logarithm of the groundwater TIC concentration was positively and significantly correlated
to the logarithm of the CO2 concentrations in the surface water. However, this correlation was the
strongest in the well developed bogs; Pearson’s r=0.521 (p<0.01) compared to a value of r= 0.286
(p<0.05) in the poorly developed bogs (figure 6).
68 – Solute transport in Sphagnum dominated bogs
Figure 6. The relation between the total inorganic carbon (TIC) concentration in the groundwater and the CO2 concentration in
the surface water for well (filled circles) and poorly (open circles) developed bogs. Water samples collected in April and August
are in black, others in gray. Lines are regression lines for well (solid line, r2=0.272) and poorly (dashed line, r2=0.082) developed
bogs. For clarity of presentation, the non-transformed data are shown.
Figure 7. Dose-response curves of photosynthetic activity as a function of the carbon dioxide concentration for Sphagnum
cuspidatum (filled symbols; r2=0.935) and S. fallax (open symbols; r2= 0.974).
CO2 uptake characteristics
The photosynthetic rate of Sphagnum cuspidatum and S. fallax as a function of CO2 concentration
are shown in figure 7. A hyperbolic curve according to the formula A=Vmax
*[CO2]/(K
m+[CO
2]) + c was
fitted to the data. From the curve the compensation point (Γ) for CO2 was calculated (table 3).
Table 3. Carbon dioxide uptake characteristic (±SD) for Sphagnum cuspidatum and S. fallax. See text for explanation of the parameters
S. cuspidatum S. fallax
Vmax
(nmol O2 ∙ s-1 ∙ mg chl-1) 19.8 ± 1.5 33.3 ± 2.6
Km
(µmol CO2 ∙ L-1) 133.2 ± 31.1 231.4 ± 52.5
Γ (µmol CO2 ∙ L-1) 10.2 7.2
c -1.4 ± 0.4 -1.1 ± 0.6
The importance of groundwater – 69
Discussion
Restoration success
The situation in the “Dwingelderveld” is characterized by a number of small Sphagnum bogs that
were subjected to identical restoration measures, but differing in groundwater influence and
developmental success. This offered us the opportunity to study the importance of CO2 in the
development of Sphagnum bogs in a field situation.
The aerial photograph analysis revealed distinct differences in restoration success. The results
were based on a quantitative approach; the occurrence of a high cover of Sphagnum species in
general. The success of the restoration was also evaluated by Everts et al. (2002) using total species
composition. Their results were in agreement with the results from our approach using aerial
photographs.
Evidence for influence of local groundwater flows
The PCA analysis using all main groundwater chemical data showed that well developed and poorly
developed bogs separate well, and that total inorganic carbon (TIC) differences are largely respon-
sible for that; the groundwater nearby the well developed bogs contains a significantly higher TIC
concentration than groundwater nearby the poorly developed bogs (figure 4).
We also found that CO2 concentrations in the surface water of the well developed bogs were
significantly higher than in the poorly developed bogs. Moreover, the CO2 availability in the surface
water of the investigated bogs is positively correlated to the inorganic carbon concentration in the
groundwater (figure 6). Carbon dioxide is very likely released from the carbon-rich groundwater
upon entering the acidic bog environment resulting in an increased CO2 availability stimulating
Sphagnum growth. Higher groundwater levels in the surrounding sandy areas will result in a flow
of the local groundwater towards the bogs. The concave shaped impermeable layer, essential for
bog formation, will result in the uni-directional flow of the local groundwater towards the bogs.
An alternative hypothesis is that the high inorganic carbon concentrations found in the
groundwater nearby the well developed bogs has its origin in the decomposition of organic material
in the bog. However, the chemical signature of the groundwater indicates an origin from outside
the bog. This was particularly clear in the silicium values, which are generally much higher in
water that has been in contact with mineral sediments for a longer period (Engelen & Jones, 1986).
Moreover, the groundwater near the Diepveen and the Zandveen which are poorly developed bogs,
contained high TIC concentrations despite the absence of accumulated organic material (figure 4).
Additionally, the placement of the piezometers on the concave shaped impermeable layers ensures
the sampling of inflowing water. Therefore, the results presented are in agreement with the hy-
pothesis that the well-developing bogs are fed by a lateral, carbon-rich, groundwater inflow.
To investigate differences between well and poorly developed bogs, a principal component
analysis was performed based on the chemical composition of the groundwater. Since the deve-
loping Sphagnum capitula are in close contact with surface water and only indirectly influenced by
groundwater this seems counter-intuitive. However, surface water composition is more influenced
by short term changes compared to the more stable composition of groundwater. Next, fluxes of
CO2 are more important than the resulting concentrations. The growth of Sphagnum mosses resul-
ting from the release of carbon rich groundwater will generate some positive feedbacks which will
70 – Solute transport in Sphagnum dominated bogs
result in an enhancement of the submerged Sphagnum growth (figure 8). The resulting accumula-
tion of organic matter will enhance the internal generation of CO2 from decomposition processes.
Furthermore, it will result in a decrease of the water depth which will increase light availability.
Figure 8. Schematic view of the positive feedbacks concerning CO2 availability in a Sphagnum dominated bog resulting from
the input of carbon rich groundwater. In a well developed bog the decomposition of accumulated organic matter results in a
high availability of CO2 stimulating Sphagnum growth, which in turn increases the accumulation of organic matter. During the
initial stages of bog development organic matter is absent and the inflow of carbon rich groundwater can substitute for the or-
ganic matter as a source of CO2, stimulating Sphagnum growth and thereby inducing the internal positive feedback mechanism
concerning CO2 availability. Additionally, the accumulation of organic matter decreases water depths, increasing light and CO
2
availability stimulating Sphagnum growth as well.
Seasonality of groundwater input
Some seasonality in groundwater input appears to be visible. In April and August we observed
increased CO2 concentrations in the well developed bogs compared to the poorly developed bogs
(figure 5). In autumn and winter we did not observe differences in CO2 in the surface water of the
bogs.
In spring and (early) summer lowered surface water levels (due to evaporation) will result in
an increase of the hydrological gradient resulting in an increased inflow of groundwater into the
bogs. In autumn and winter, this gradient will be decreased by higher surface water levels due to
an increase in precipitation. Additionally, strong rainfalls will lead to an increased run off towards
the bogs of very superficial groundwater which is relatively poor in CO2. Interestingly, this period
from April to August represent the growing season of the Sphagnum mosses (Clymo, 1970). High
CO2 consumption in the well developed and growing Sphagnum bogs, could even have caused a
lower than expected CO2 concentrations difference between well and poorly developed bogs.
Why do some Sphagnum species require high CO2 concentrations?
Smolders et al. (2003) showed very poor growth of five Sphagnum species under inundated
conditions on strongly humified, ‘black’ peat. Carbon dioxide concentrations in the water layer
remained very low in this situation (<20 µmol ∙ L-1). The same species, however, developed very
The importance of groundwater – 71
well on weakly humified, ‘white’ peat. In short, low carbon availability in combination with low
diffusion rates of CO2 in water severely reduces CO
2 availability and limitation of Sphagnum growth
is very likely to occur.
The requirement for high CO2 availability by Sphagnum can partly be explained by the
mechanisms of CO2 uptake that are determined by the physiological characteristics of both
Sphagnum cuspidatum and S. fallax (table 3; figure 7). Both Sphagnum species are characterized by
high CO2 compensation values, the CO
2 concentration at which CO
2 fixation by photosynthesis bal-
ances CO2 loss by respiration. Air-equilibrated water contains a CO
2 concentration of 10 - 20 µmol
∙ L-1 between 25 and 10°C. The high compensation values of S. cuspidatum and S. fallax imply that
under air-saturated conditions no, or extremely limited, net carbon accumulation can occur. In
the acidic bog environment, where no reservoir of bicarbonate is present to replace the CO2 that
is taken up, this is especially relevant and Sphagnum growth will not occur when CO2 is provided
exclusively through equilibration with air. The high Km
values of 231.4 and 133.2 µM CO2 for S. cuspi-
datum and S. fallax, respectively, further indicate that even when CO2 is present at a concentration
that is higher than air-saturated, carbon utilization is not optimal. In the investigated Sphagnum
species CO2 concentrations up to 400 - 500 µM are still not saturating (figure 7). Like most aquatic
plants lacking a carbon concentrating mechanism the kinetic properties of CO2 uptake of the in-
vestigated Sphagnum species indicate an adaptation to a high CO2 availability (Raven et al., 1998).
Under natural conditions the stagnant bog water will result in thick boundary layers and long
diffusion path lengths when compared to the well stirred conditions during the measurements,
lowering the apparent affinity for CO2 concentration even more. Additionally, photosynthetically
produced oxygen will accumulate in the boundary layer and because of the competition between
CO2 and O
2 at the site of Rubisco, photosynthetic rate is negatively affected by this increase in O
2
(Bowes & Salvucci, 1989). The importance of diffusion rates on CO2 availability for Sphagnum has
been shown by Baker and Boatman (1985). They showed the ability of Sphagnum cuspidatum to form
smaller and thinner leafs under low CO2 availability. This will reduce the boundary layer resistance
and facilitate CO2 uptake. Paffen & Roelofs (1991) concluded that a dissolved CO
2 concentration of
at least 750 µmol ∙ L-1 is necessary for the optimal growth of Sphagnum cuspidatum and the subse-
quent formation of floating vegetation. This was shown by the remarkable low average CO2 concen-
trations in the surface water of the poorly developing sites Schurenberg North, Cootjes Veen South
and Groote Veen East; 233, 287 and 122 µmol ∙ L-1, respectively. At those sites Sphagnum growth was
severely hampered.
For the successful re-establishment of aquatic Sphagnum species an additional carbon source
seems to be crucial. This field study shows that the inflow of carbon-rich groundwater can substi-
tute for the peat layer as a source of CO2 during the initial stages of bog development.
Other explanations for restricted Sphagnum growth in peat ponds
Of course, many factors affect the re-establishment of Sphagnum and subsequent bog develop-
ment (Money et al., 2009). In the majority of the studied bogs, Sphagnum growth appears to be
limited by the availability of CO2. However, there are several interesting exceptions. This is illus-
trated by the high CO2 availability in some of the poorly developed bogs (figure 4). Nutrient and
light limitation might be responsible for failing Sphagnum re-colonization (e.g. Money & Wheeler,
1999; Smolders et al., 2003). A shortage in nutrients would probably prevent an increase in produc-
72 – Solute transport in Sphagnum dominated bogs
tion at enhanced CO2 concentrations in natural ecosystems (Kramer, 1981). The Diepveen and the
Zandveen are two bogs lacking the re-colonization of Sphagnum under conditions of apparently
sufficient inorganic carbon (figure 4). However, nutrient limitation seems not to be responsible
for this since no clear difference in groundwater or surface water chemistry compared to the other
bogs was found (data not shown). Smolders et al. (2003) concluded that the availability of both
light and CO2 have to be sufficient to enable submerged Sphagnum to reach high photosynthetic
and growth rates. These factors might indeed affect the Sphagnum development in both Diepveen
and Zandveen South in which the water depth is generally several meters in the centre of the bog
and on average >50 cm at the edges, very probably hampering Sphagnum growth due to the reduced
light availability. Additionally, physical constraints like wind and wave action possibly severely
hamper Sphagnum growth in open water bodies (Money et al., 2009). The lack of re-colonization of
Sphagnum mosses and hampered growth of already established Sphagnum mosses has often been
ascribed to high levels of atmospheric nitrogen deposition (Lamers et al., 2000; Money & Wheeler,
1999; Twenhoven, 1992). The ammonium availability in the surface water at all sampling sites (ta-
ble 2) reflects the high nitrogen loads as present in the north of the Netherlands, on average 28 kg ∙
ha-1 ∙ yr-1 (Limpens et al., 2003; RIVM, 2009). However, since no significant differences in nitrogen
availability are found between well and poorly developed bogs (data not shown) the current ni-
trogen deposition is not the determining factor for bog developmental success. Moreover, Tomas-
sen (2004), suggested that bog vitality is much less affected by high nitrogen deposition if other
environmental factors, such as water table and the availability of other nutrients (such as CO2), are
optimal.
Thresholds in the restoration of bogs
The present study demonstrates that Sphagnum bogs in the “Dwingelderveld” are part of the
total landscape hydrology instead of being hydrologically distinct entities. This might be the
case for many damaged Sphagnum bogs and it implies a landscape approach for successful bog
restoration. The current findings clearly show that high CO2 availability is a pre-requisite for the
successful re-establishment of Sphagnum mosses and subsequent bog development. Therefore,
CO2 availability should be included in bog restoration feasibility studies.
Chapter 6
Photosynthesis of three Sphagnum species after acclimatization to high and low carbon dioxide availability
Wouter Patberg
Jan Erik van der Heide
Theo Elzenga
76 – Solute transport in Sphagnum dominated bogs
Abstract
A high CO2 availability stimulates the growth of both aquatic and emergent
Sphagnum species. As shown in the previous chapter, the physiological
parameters of CO2 uptake by Sphagnum also show an adaptation to a high
CO2 availability; a high CO
2 compensation point and a low affinity for CO
2.
However, from literature it is known that Sphagnum is able to acclimatize to
different CO2 levels. For example, culturing plants under high CO
2 availabil-
ity, results in lower photosynthetic rates compared to plants that are grown
under CO2-limiting conditions. In this chapter, the physiology of carbon
uptake by Sphagnum (substrate specificity, affinity and plasticity of carbon
assimilation) was determined for three Sphagnum species grown for long
periods at high and low CO2 availability.
The CO2 compensation point and the K
m values of the high and low CO
2
grown S. cuspidatum plants indicate that primary production is limited un-
der air-equilibrated conditions. Remarkably, in S. cuspidatum the low CO2
treated plants were capable of higher photosynthetic rates compared to the
high CO2 treated plants at similar, high CO
2 concentrations. This difference
was not found for S. fallax and S. magellanicum. Possibly, this reflects the
difference in habitat: S. cuspidatum is a submerged aquatic species, while S.
fallax and S. magellanicum are both emergent species.
Considering the high CO2 compensation point, the low affinity for CO
2, the
absence of a carbon concentrating mechanism and the limited morphologi-
cal and physiological plasticity of the plants when exposed to a low external
CO2 concentration, primary production by Sphagnum is expected to be ex-
tremely low when solely supplied with atmospheric CO2. This agrees with
our findings in Chapter 5 that when an organic layer is lacking, i.e. during
the initial stages of bog development, an alternative external CO2 source
seems to be essential for the successful (re-)establishment of Sphagnum.
Photosynthesis of three Sphagnum species – 77
Introduction
Bogs ecosystems are very wet and acidic and are often dominated by mosses of the genus Sphagnum
(Clymo & Hayward, 1982). The decomposition of organic material in bogs is slower than the photo-
synthetic fixation of CO2, resulting in the accumulation of peat (Clymo et al., 1998). Since Sphagnum
bogs function as carbon sink they play an important role in global carbon cycling (Bridgham et al.,
2001a; Gorham, 1991).
In contrast to vascular plants Sphagnum mosses lack a cuticle and stomates to regulate
photosynthesis (Proctor, 2008). Sphagnum mosses are surrounded by an external water film
through which gas exchange for photosynthesis is taking place. Since the diffusion of CO2 is about
104 times lower in water than in air, the diffusional barrier formed by the external water films
reduces the supply of CO2 to the carbon assimilating cells and, consequently, the photosynthetic
rate (Bowes & Salvucci, 1989; Rice & Giles, 1996; Silvola, 1990; Williams & Flanagan, 1996).
The photosynthetic rate of Sphagnum mosses has been shown to be a compromise between
external water content and the availability of CO2 (Schipperges & Rydin, 1998; Silvola, 1990; Titus
et al., 1983). At low water contents, dehydration inhibits photosynthesis whereas at very high
water contents Sphagnum species may suffer from carbon limitation due to very thick boundary
layers (Jauhiainen & Silvola, 1999; Rice & Giles, 1996; Silvola, 1990; Titus et al., 1983; Williams
& Flanagan, 1996). For Sphagna, a morphological difference between aquatic and non-aquatic
species was demonstrated by Rice and Schuepp (1995); aquatic Sphagnum species have, compared
to non-aquatic taxa, longer and thinner leaves and consequently a thinner boundary layer.
The growth of submersed aquatic macrophytes is often limited by CO2 (Raven et al., 1985; Rice
& Schuepp, 1995). To overcome the diffusion barrier many aquatic plant species make use of a
carbon concentrating mechanism (CCM), which enhances the accumulation of carbon (Maberly &
Madsen, 2002). The mechanism most often found is the utilization of bicarbonate (HCO3
-) as a car-
bon source in photosynthesis (Prins & Elzenga, 1989). Most aquatic bryophytes, however, lack such
a CCM and are known to be pure CO2 users (Bain & Proctor, 1980; Raven et al., 1998; Raven et al.,
1985). By performing pH drift experiments, Bain and Proctor (1980) demonstrated that Sphagnum
cuspidatum is a pure CO2 user and exclusively depends on diffusion of CO
2 to the site of carbon
fixation. Due to the diffusional barrier presented by the water layer surrounding the plants high
rates of photosynthesis can only be sustained when the leaves are exposed to high levels of CO2
(Jauhiainen & Silvola, 1999; Raven et al., 1985; Silvola, 1990).
A high CO2 availability has been shown to stimulate the growth of both aquatic and emergent
Sphagnum species (Baker & Boatman, 1990; Jauhiainen & Silvola, 1999; Paffen & Roelofs, 1991; Riis
& SandJensen, 1997; Roelofs, 1983; Smolders et al., 2001; Smolders et al., 2003). Chapter 5 describes
a field study, which demonstrates the importance of a high CO2 availability for the successful re-
establishment of Sphagnum and subsequent bog development. Despite the obvious importance
of a high CO2 availability for Sphagnum, the physiological background of this apparent high CO
2
requirement of Sphagnum has never been established.
The physiological characteristics of aquatic plants lacking a CCM indicate an adaptation to
high CO2 availability: a high CO
2 compensation point concentration and a low affinity for CO
2.
With these characteristics it is likely that Sphagnum will be limited by the diffusion of CO2 under
air equilibrated conditions (Raven et al., 1985). Chapter 5 shows the need for high CO2 availability
78 – Solute transport in Sphagnum dominated bogs
based on the physiological background of carbon uptake by Sphagnum cuspidatum and S. recurvum.
However, the Sphagnum plants used in that experiment were grown under ambient CO2 conditions.
For Sphagnum fuscum, a hummock forming species, acclimation to high CO2 levels has been shown.
Culturing plants under high CO2 availability results in low photosynthetic rates compared to plants
that are grown under CO2-limiting conditions (Jauhiainen & Silvola, 1999). In the present study,
the physiological background of carbon uptake by Sphagnum (substrate specificity, affinity and
plasticity of carbon assimilation) was determined for plants grown for a long period at high and
low CO2 availability. Three Sphagnum species (Sphagnum cuspidatum, S. fallax and S. magellanicum)
were grown for four months under high or low CO2 availability. The Sphagnum species used in this
study occupy different ecological niches. Sphagnum fallax and S. magellanicum both are emergent
and grow above the water surface, while S. cuspidatum is growing completely submerged. From an
evolutionary perspective, emergent species might be adapted to water holding capacity and less to
low CO2 levels (Rice & Schuepp, 1995).
The high and low CO2 S. cuspidatum plants were used to measure the photosynthetic response
at different CO2 concentrations. At CO
2 concentrations close to the saturation level, low CO
2
grown Sphagnum cuspidatum plants exhibited a higher photosynthetic rates compared to the
high CO2 grown plants. At this CO
2 level, the photosynthetic rate of S. fallax and S. magellanicum
was measured as well. However, differences in photosynthetic rate between treatments were not
observed. In addition, supplemental to the pH drift experiments performed on S. cuspidatum by
Bain and Proctor (1980) similar pH drift experiments were carried out to test carbon utilization by
S. fallax and S. magellanicum; both species were shown to be pure CO2 users as well.
Materials and methods
pH drift experiment
For the pH drift experiment Sphagnum fallax (klinggr.) Klinggr. and S. magellanicum Brid. were collected
in a small bog in the “Dwingelderveld”, a nature reserve in the north of the Netherlands (N52°49.178’,
E6°25.991’). The upper two cm of ten plants were incubated in a closed 250 mL Erlenmeyer flask com-
pletely filled with ten times diluted artificial rainwater (Smolders et al., 2001) supplemented with
1 mM NaHCO3. The flasks were kept at 20˚C by placing them in a water bath. The solution was con-
tinuously and slowly stirred. The flasks were illuminated by a halogen lamp (FL 103, Walz, Effeltrich
Germany) with a light intensity of approximately 350 µmol · m-2 · s-1. The pH of the solution was
measured continuously for at least 6 hours using a combined pH electrode with an Ag⁄AgCl internal
reference electrode (Cole Parmer Instrument Company, Illinois, USA) in combination with a home
made amplifier (input impedance 1011 Ohm) and a Campbell Scientific CR10X data logger. The experi-
ment was terminated when no further pH increase could be observed. At the end of the experiment,
three 1 mL samples were taken from each Erlenmeyer flask for total inorganic carbon (TIC) measure-
ments. TIC concentration was measured using an infrared gas analyzer (CO2 analyzer model no. S151,
Qubit Systems Inc., Kingston, ON, Canada). The ratio between HCO3
- and CO2 in the TIC samples was
calculated based on the pH (Prins & Elzenga, 1989). For each species the pH drift experiment was per-
formed in triplicate. Carbon uptake, CO2 compensation point and half-saturation constant (K
m) for
CO2 uptake by S. fallax and S. magellanicum were calculated according to Maberly and Spence (1983).
Photosynthesis of three Sphagnum species – 79
CO2 uptake experiment
Sphagnum cores were collected in April 2010 from two small bogs in the “Dwingelderveld”. Three
species of Sphagnum were collected, Sphagnum cuspidatum Ehrh. Ex Hoffm., S. fallax (klinggr.)
Klinggr. and S. magellanicum Brid.. Per species four homogeneous 10 cm thick sections of 18 cm
by 25 cm were cut out of the Sphagnum carpet and gently placed in a glass container (18*25 cm,
height 10 cm) and transported to a greenhouse. During transport the water table in the containers
was kept at approximately 2 cm below the capitula. In the greenhouse the cores were cut to a depth
of 5 cm and placed in plastic nets which, in turn, were mounted in the same glass container. The
sides of the containers were covered with black plastic sheets to keep out the light. The containers
were filled with artificial rainwater (according to Smolders et al. (2001) except for NH4NO
3 of which
100 µmol ∙ L-1 was used). The S. cuspidatum cores were placed in the containers with the capitula at
the water level, S. fallax with their capitula ~1 cm and S. magellanicum ~3 cm above the water table.
Growth was compensated for by lowering the nets with the Sphagnum cores in order to keep the
water level equal with respect to the top of the capitula. During the experiment the containers were
continuously fed with artificial rainwater (~20 mL ∙ hr-1) from 40 liter containers by using black
norprene tubes (l = 400 mm, 4.8 mm outer and 1.6 mm inner diameter; Saint-Gobain Performance
Plastics, Verneret, France) in combination with a peristaltic pump (Masterflex L/S model 7519-25,
Cole Parmer Instrument company). The solution left the container through an overflow located 3
cm below the rim. For each species, two containers were fed by artificial rainwater bubbled with
carbon dioxide (Carbon Dioxide 4.0, Linde AG, Munich, Germany) and two containers with air-
equilibrated artificial rainwater, resulting in a ‘high’ and ‘low’ CO2 treatment, respectively. Vascular
plants were removed from the Sphagnum cores on a regular basis. In the greenhouse, natural light
was supplemented with high pressure sodium lamps to induce a 14 hour photoperiod. The plants
acclimated for two weeks before the treatments started. After four months the photosynthetic rate
of the Sphagnum species at different CO2 concentrations was measured (see below).
During the culture period the CO2 concentration and pH of the pore water in the containers
were measured every two months. Water samples were collected using 10 cm long Teflon Rhizons
(Eijkelkamp, Agrisearch, Giesbeek, the Netherlands) which were placed diagonally in the middle of
the Sphagnum cores. The total inorganic carbon (TIC) concentrations were measured using an Infra
Red Gas Analyzer (IRGA; Li-7000 CO2 / H
2O analyzer, Li-Cor, Inc., USA); The pH was measured using a
Metrohm 780 pH meter (Metrohm, Herisau, Switzerland) together with a combined Metrohm glass
electrode (Metrohm 6.0258.010). The CO2 concentration in the water samples was subsequently
calculated based on the pH and the TIC concentration (Prins & Elzenga, 1989). During the growth
period the pigment content of the mosses was determined three times with regular intervals. Per
container five capitula were randomly collected, pooled and pigments were determined according
to Lichtenthaler (1987).
CO2 uptake characteristics were determined by measuring photosynthetic activity (A) at differ-
ent CO2 concentrations. Photosynthetic activity was determined by the photosynthetic evolution
of oxygen by a capitulum placed in a closed thermostatic cuvette containing 1 mL of measuring
buffer (see below) at 18 ± 0.2˚C and saturating light conditions (1500 µmol ∙ m-2 ∙ s-1; Hansatech
Quantitherm Light meter). The solution in the cuvette was stirred continuously. Per measurement
one capitulum was used and each capitulum was used for only one measurements. Large branch-
es were trimmed to fit in the cuvette. Oxygen was measured by a Clark electrode located at the
80 – Solute transport in Sphagnum dominated bogs
bottom of the cuvette in combination with a millivolt recorder (Kipp and Zonen BD40; Delft, the
Netherlands) connected to a Graphtec GL200 midi logger (Graphtec Corp., Yokohama, Japan) on
which data were logged every second. The measuring buffer consisted of 10 times diluted artificial
rainwater (Smolders et al., 2001) and 20 mM MES set at pH = 4.0 using NaOH. During the initial
phase of the experiment, the oxygen concentration in the measuring buffer was in equilibrium
with air (21%) which seemed to reduce photosynthetic rate considerably. Therefore, the oxygen
concentration was reduced by flushing the measuring buffer with N2 which resulted in an average
(±SD) oxygen concentration in the buffer of 9% (±3).
Different CO2 concentrations were obtained by diluting 20 to 600 µL of 0.1 M NaHCO
3 solution
with 35 mL of buffer of which 1.5 mL was injected (Hamilton 2.5 mL syringe) into the cuvette whereof
0.5 mL was instantly retaken for the immediate determination of the CO2 concentration. Total inor-
ganic carbon (TIC) concentrations were measured using an Infra Red Gas Analyzer (IRGA; Li-7000
Co2/H2O analyzer, Li-Cor, Inc., USA). After usage the fresh weight of the capitula was determined
and the plant material was subsequently frozen at -80°C for until determination of the chloro-
phyll concentration according to Lichtenthaler (1987). Photosynthetic activity is expressed as the
amount of oxygen produced (nmol O2 ∙ s-1) on a fresh weight (g FW) and chlorophyll (mg) basis.
Prior to each measurement residual inorganic carbon from the culture medium, present in the
hyaline cells or adhering water was reduced as much as possible by illuminating the capitula with
1000 µmol ∙ m-2 ∙ s-1 for at least 60 minutes while keeping them in a CO2 free medium.
Statistical analysis
Hyperbolic curves were fitted to the carbon dioxide uptake data, using graphing software (Prism
version 4.03, 2005; GraphPad Software, Inc., San Diego, CA, USA).
Per species t-tests were performed to indicate differences between high and low CO2 avail-
ability on photosynthetic performance by using SPSS for Windows (version 16.0.1, 2007; SPSS Inc.,
Chicago, IL, USA).
Results
pH Drift experiment
If only free CO2 is taken up, the final pH value of a pH drift experiment will be between 8 and 10,
whereas in case of HCO3
- uptake the pH can reach a value between 11 and 12 (Bain & Proctor, 1980;
Prins & Elzenga, 1989). So, a distinction between a CO2 and a HCO
3- user can be made, based on this
difference in maximum pH.
In the pH drift experiment maximum pH values of 8.1 ± 0.03 and 8.4 ± 0.18 were measured for S.
fallax and S. magellanicum, respectively, indicating that these Sphagnum species are not able to utilize
HCO3
- as a carbon source, and are therefore defined as strictly CO2 users. Figure 1 shows the photo-
synthetic activity (A) (expressed as relative carbon uptake compared to the maximum carbon up-
take at the beginning of the experiment) of Sphagnum fallax and S. magellanicum as a function of CO2
availability during the pH drift experiment. Hyperbolic curves according to the formula A=Vmax
*[CO2]/
(Km
+[CO2]) + c were fitted to the data (see figure 1). From the curves the CO
2 compensation point con-
centrations (Γ), the affinity for CO2 (K
m(CO2)) and photorespiration (c) were determined, see table 1.
Photosynthesis of three Sphagnum species – 81
Figure 1. Photosynthetic activity as a function of CO2 concentration of Sphagnum fallax (a) and S. magellanicum (b) during
the pH drift experiment. Photosynthetic activity is expressed as the CO2 uptake relative to the maximum CO
2 uptake at the
beginning of the pH drift experiment (in %).
Table 1. The affinity for CO2 (K
m), CO
2 compensation point (Γ) and photorespiration (c) (±SD)for Sphagnum fallax and S. magel-
lanicum determined from the relation between CO2 availability and photosynthetic activity as shown in figure 1
S. fallax S. magellanicum
Km
(µmol CO2 ∙ L-1) 14.4 ± 5.0 24.5 ± 2.9
Γ (µmol CO2 ∙ L-1) 7.2 ± 2.1 8.5 ± 0.9
c 60% ± 8.3 47% ± 3.9
CO2 uptake experiment
CO2 concentration, pH and chlorophyll content during growth phase
Per treatment there were two containers per species. However, per species and treatment plants
from only one container were used in the photosynthesis measurements. For these containers the
average, minimum and maximum CO2 concentrations and average pH values measured in the pore
water during the growth phase of the experiment are shown in table 2.
82 – Solute transport in Sphagnum dominated bogs
Table 2. Average, minimal and maximum carbon dioxide concentrations (n=3; in µmol ∙ L-1) and average pH (n between
brackets) in the pore water of the containers during the growth phase of the experiment. + = high and - = low CO2 treatment
[CO2] (µmol ∙ L-1)
pHmean min. max.
S. cuspidatum+ 8539 4930 13243 3.7 (3)
- 337 227 507 4.6 (2)
S. fallax+ 7829 4033 14126 4.2 (3)
- 184 104 276 4.4 (2)
S. magellanicum+ 5392 4562 6599 4.3 (2)
- 183 85 248 4.3 (2)
The pigment content of the three Sphagnum species after growth for four months at the two
different CO2 treatments are given in figure 2. Per species and treatment the pigment concentration
of five pooled capitula was determined. For all three species the plants grown at the low CO2 regime
contained higher pigment concentrations (chl a, b and carotene) compared to plants grown at high
CO2. This higher pigment content in the low CO
2 treated plants might be indicative of an increased
investment in the photosynthetic apparatus under more severe CO2 limitation.
Figure 2. Pigment content of Sphagnum cuspidatum, S. fallax and S. magellanicum grown at high and low CO2 availability for a
period of four months. The white bars represent chlorophyll a, light grey bars chlorophyll b and the dark grey bars carotene,
all in mg ∙ g FW-1. Per species and pigment type, the left bar represents the high (+) and the right bar the low (-) carbon dioxide
treatment. Each bar represents one sample of five pooled capitula.
Photosynthetic response to CO2
The photosynthetic rate of Sphagnum cuspidatum grown at high and low CO2 availability as a
function of CO2 concentration is shown in figure 3. Hyperbolic curves according to the formula
A=Vmax
*[CO2]/(K
m+[CO
2]) + c were fitted to the data. From the curves the CO
2 compensation point
concentrations (Γ) were calculated. The maximum photosynthetic rate (Vmax
), the affinity for CO2
(Km
) and CO2 compensation point are presented in Table 3. Based on visual inspection and to reduce
the degrees of freedom in the fitting procedure, one Km
value was fitted for both treatments.
Photosynthesis of three Sphagnum species – 83
Table 3. CO2 uptake characteristics (±SE) for Sphagnum cuspidatum grown for three months at high carbon dioxide availability
(+) and at low carbon dioxide availability (-). See text for explanation of the parameters
+ -
Vmax
(nmol O2 ∙ s-1 ∙ FW-1) 2.71 ± 0.29 2.77 ± 0.27
Km
(µmol CO2 ∙ L-1) 11.60 ± 3.52 11.60 ± 3.52
Γ (µmol CO2 ∙ L-1) 15.26 11.35
c -1.54 ± 0.19 -1.37 ± 0.16
For pure CO2 users the CO
2 compensation point is the lower limit for net C fixation. The CO
2
compensation point concentrations for the high and low CO2 grown S. cuspidatum plants are 15.26
and 11.35 µmol CO2 ∙ L-1, respectively (table 3). Considering that air-equilibrated water contains a CO
2
concentration of 10 - 20 µmol ∙ L-1 between 25 and 10°C, the CO2 compensation point found for S.
cuspidatum (both treatments) indicates that primary production is limited by CO2 diffusion under
air-equilibrated conditions. Under air-equilibrated conditions no net growth can be expected,
since the minimal carbon gain during the light period is expected to be more than offset by the
carbon loss due to respiration during the dark period.
The Km
value for S. cuspidatum grown at high and low CO2 availability is 11.60 µmol ∙ L-1 (table
3), indicative for the fixation of CO2 by Rubisco (lacking a CCM). Consequently, Rubisco is far from
being saturated in air-equilibrated conditions and high CO2 concentrations are needed for optimal
photosynthesis.
More remarkably is the difference in photosynthetic rate between high and low grown S.
cuspidatum at higher CO2 concentrations (figure 3 and 4). Based on fresh weight (figure 4a) the
photosynthetic rate of S. cuspidatum is on average significantly higher in the low CO2 treatment
(1.24 ± 0.23 nmol O2 ∙ s-1 ∙ g FW-1), than in the high CO
2 treatment (0.75 ± 0,18; t(6)=-3.411, p<0.05).
The exact CO2 concentrations at which photosynthetic rate were measured are mentioned in the
accompanying text.
The average photosynthetic rates of S. fallax in the high and low CO2 treatments were 1.47 ± 0.53
and 3.08 ± 1.45, respectively. For S. magellanicum the average photosynthetic rate was 1.66 ± 1.04 for
the high and 2.33 ± 1.29 for the low CO2 treatment. For both S. fallax and S. magellanicum the average
rates of photosynthesis were highest in the low CO2 treatment but no significant differences be-
tween treatments were observed; t(6)=-2.343, p=0.058 and t(9)=-0.934, p=0.375, respectively.
84 – Solute transport in Sphagnum dominated bogs
Figure 3. Net photosynthetic rate of Sphagnum cuspidatum grown at high and low CO2 availability, measured at different CO
2
concentrations at 18±0.2˚C. Photosynthetic rate is expressed as the amount of oxygen released based on fresh weight. The data
are fitted to hyperbolic curves of the form A=Vmax
* [CO2] / (K
m + [CO
2]) + c, where V
max is the maximum photosynthetic rate, K
m
the CO2 concentration at which half of the maximum rate of photosynthesis is reached and c is a constant.
Figure 4. Box plots showing the photosynthetic rate (in nmol O2 ∙ s-1 ∙ g FW-1) of S. cuspidatum, S. fallax and S. magellanicum
based on fresh weight (a) and based on chlorophyll content (b). Plants were grown for four months at a high (+) or low (-) carbon
dioxide availability. The average (± SD) carbon dioxide concentrations at which photosynthetic rates were measured were 177 ±
9 µmol ∙ L-1 (+) and 150 ± 10 (-) for S. cuspidatum; 151 ± 4 (+) and 153 ± 9 (-) for S. fallax and 158 ± 6 (+) and 159 ± 11 (-) for S. magellani-
cum. The box plots are composed of minimum, maximum, 25%, 75% quartiles and the median. Significant differences between
treatments are indicated by an asterisk (p<0.05). The amount of used capitula is noted between brackets in figure a.
Photosynthesis of three Sphagnum species – 85
Based on chlorophyll content (figure 4b) the photosynthetic rates also show a significant difference
between treatments for S. cuspidatum, t(6)=-2.858, p<0.05) with the low CO2 grown plants giving a
higher rate (26.57 ± 4.16 nmol O2 ∙ s-1 ∙ mg Chl-1) than the high CO
2 plants (16.77 ± 5.46). For S. fal-
lax and S. magellanicum the differences in photosynthetic rate between treatments differed not
significantly, t(6)=-0.516, p=0.624 and t(9)=1.467, p=0.177, respectively. For S. fallax the photosyn-
thetic rates based on chlorophyll content were on average lower in the high CO2 treatment than the
average rate measured in the low CO2 treated plants; 56.55 ± 22.37 and 66.54 ± 33.25, respectively.
On the other hand, the average photosynthetic rates of S. magellanicum based on chlorophyll con-
tent were highest in the high CO2 treatment, 92.98 ± 55.19 compared to 49.51 ± 43.30 in the low CO
2
treatment.
Discussion
Kinetic properties of CO2 uptake by Sphagnum
Like most aquatic bryophytes, Sphagnum has been shown to be a pure CO2 user. For S. cuspidatum
this was shown by Bain and Proctor (1980) and the results presented here show it for S. fallax
and S. magellanicum. CO2 uptake by S. cuspidatum, as shown here, is characterized by a high CO
2
compensation point and a Km
value consistent with Rubisco being the primary CO2 fixing enzyme
and with the absence of a carbon concentrating mechanism (table 1 and 2) (Raven et al., 1985). For
submerged aquatic macrophytes a low affinity for CO2 is common and K
m values are usually in the
range of 30 – 70 µM CO2 (Bowes & Salvucci, 1989; Raven et al., 1985). The K
m values for S. cuspidatum,
S. fallax and S. magellanicum found in this study, are lower (11.6, 14.4 and 24.5 µmol ∙ L-1, resp.) and
very likely a consequence of the low oxygen levels used and the vigorous stirring during the photo-
synthesis measurements. Due to the competition of CO2 and O
2 at the site of Rubisco, K
m values and
CO2 compensation points are higher with increasing O
2 concentrations. Therefore, the presented
Km
values are very likely lower compared to Km
values at ambient oxygen levels. This is supported
by the higher calculated Km
values (133.2 and 231.4 µmol ∙ L-1 for S. cuspidatum and S. fallax, resp.)
obtained from the CO2 response curve in Chapter 5, which were determined at ambient O
2 levels.
The difference in Km
values with differing O2 concentration stresses the influence of O
2 availability
on CO2 uptake. This is especially of interest when regarding the high O
2 concentrations in the up-
per Sphagnum layer due to photosynthetic activity (Lloyd et al., 1998). In the stagnant bog water,
the boundary layers will be substantially thicker, compared to the well-stirred conditions in the
presented experiments, reducing the affinity for CO2 even more. The K
m values determined from
the pH drift experiments are likely to be an underestimation, compared to the values in the natural
situation, due to the reservoir of HCO3
-, present at high pH, that can replenish the CO2 that is taken
up and which is not present in the acidic bog situation.
Despite the variability in Km
and compensation point due to O2 availability (and other
environmental conditions as light and temperature (e.g. Maberly & Spence, 1983)), these kinetic
properties are suggestive of optimization for operation at high CO2 concentrations at the active site
of Rubisco, a condition that is not being met under air-equilibrated conditions. The CO2 acquisition
characteristics are typical for an aquatic plant lacking a CCM (Raven et al., 1985).
86 – Solute transport in Sphagnum dominated bogs
Plasticity of CO2 assimilation characteristics
Main focus of this chapter, however, is the difference in CO2 uptake characteristics between
Sphagnum plants subjected to different CO2 concentrations. In principle, for a submerged plant that
can only utilize CO2 there are three possible strategies to increase carbon fixation rates at external
concentrations of CO2 that are slightly higher than the compensation concentration: 1. Increase
in the affinity of the primary CO2 fixing enzyme (lower K
m(CO2)); 2. Increase in the photosynthetic
capacity (higher Vmax
) and 3. Decrease in dark respiration (less negative c). In figure 5 the effects of
these three strategies on net photosynthesis at low external CO2 concentration are illustrated.
Figure 5. Visualization of the photosynthetic response as a function of CO2 concentration for the three possible strategies that
can be applied by a submerged Sphagnum plant, in order to increase carbon fixation rates at low external CO2 concentrations.
The strategies are: (i) increasing the affinity of the primary CO2 fixing enzyme (lower K
m); (ii) increasing the photosynthetic
capacity (higher Vmax
) and (iii) decreasing dark respiration (less photorespiration). The strategies curves are plotted relative to
a control curve, using arbitrary units on both axes. At air-equilibrated conditions, water contains a CO2 concentration of 10 - 20
µmol ∙ L-1 between 25 and 10 °C which is indicated by the grey box.
Between treatments, no (significant) difference in CO2 affinity (K
m) and compensation point (Γ) by
S. cuspidatum were observed. Increased affinity and lowering the CO2 compensation point are strat-
egies shown by micro-organisms and submersed angiosperms to gain more carbon in response to
a decreased DIC availability (Bowes, 1996). Despite the fact that Sphagnum lacks a biophysical and
biochemical mechanism to increase the concentration of C at the site of fixation by Rubisco, S.
cuspidatum shows some adaptation to CO2 availability. However, our results indicate that S. cuspida-
tum does increase its photosynthetic capacity under CO2-limiting conditions, possibly to increase
carbon fixation at low external CO2 availability. This is shown by the difference in V
max between S.
cuspidatum grown in the high and the low CO2 treatments; the low CO
2 treated plants were capable
of higher photosynthetic rates compared to the high CO2 plants at similar CO
2 concentrations. This
might be caused by an increase in the chlorophyll concentration under CO2 limiting conditions
Photosynthesis of three Sphagnum species – 87
compared to high CO2 availability. This was shown for all three Sphagnum species (figure 2). The
higher concentrations of chlorophyll probably allows the mosses to gain more CO2 (Rice, 1995) and
thus lead to an increase in photosynthetic performance. For S. magellanicum this inverse correlation
between chlorophyll content and CO2 availability was already shown by Smolders et al. (2001).
Jauhiainen and Silvola (1999) showed a reduced photosynthetic efficiency for Sphagnum fuscum
when grown under high CO2 availability, compared to plants grown under low CO
2 conditions. Our
findings are in line with the general pattern to down-regulate photosynthetic performance at high
carbon availability (e.g. Maberly & Madsen, 2002). On the other hand, increased dry weight by the
production of non-structural carbohydrates under high CO2 levels (Van Der Heijden et al., 2000)
might cause the lower photosynthetic rate based on weight (figure 4). However, these increases are
small on a dry mass base and therefore not taken into account in this study.
Possible morphological adaptation to increased CO2 uptake
Neither S. fallax, nor S. magellanicum were able to enhance photosynthetic performance in comparison
with the high CO2 grown plants, despite CO
2 limitation (indicated by an increased chlorophyll con-
tent in the low CO2 treatment). An explanation could be found in the morphological differences be-
tween the Sphagnum species. A mechanism to increase photosynthetic performance under low CO2
conditions is to enhance the supply of CO2 by reducing the boundary layer thickness. Aquatic plant
species commonly change morphology in order to reduce boundary layer resistance under CO2-lim-
ited conditions (Bowes & Salvucci, 1989; Rice & Schuepp, 1995). Sphagnum cuspidatum was shown to
be able to form thinner leaves when grown submerged at low CO2 availability (Baker & Boatman, 1985;
Rydin & McDonald, 1985). In the present study, leaf morphology was not determined. However, since
S. cuspidatum was grown submerged, a morphological adaptation of S. cuspidatum to low CO2 levels
very likely contributes to the significant higher photosynthetic rate of plants grown under low CO2
levels compared to the high CO2 plants. When growing emergent, like S. fallax and S. magellanicum in
this experiment, the formation of thinner leaves is not to be expected, since this will not results in a
reduced boundary layer thickness when growing under low CO2 availability. Moreover, for emergent
growing Sphagnum plants, plant morphology must compromise the possible conflicting require-
ments of water holding capacity (and conduction) and free gas exchange for photosynthesis (Proctor,
2008). Since the formation of thinner leaves will result in a decreased water holding capacity, this
morphological adaptation is not to be expected in S. fallax and S. magellanicum.
However, when based on chlorophyll content, the photosynthetic rate of S. fallax and
S. magellanicum grown at low CO 2 availability were on average lower than the rate of the high
CO2 plants, which is opposite to the differences between treatments when based on fresh weight
(figure 3), indicative for an increased photosynthetic performance at low CO2 availability probably
due to the increased investment in the photosynthetic apparatus.
There is a substantial difference between species, and to a lesser extent between treatments, in
the variation of the photosynthetic rate (figure 4); S. fallax and S. magellanicum show a variation of
at least 300% whereas S. cuspidatum exhibits a more narrow range. Sphagnum fallax and S. magel-
lanicum grown under low CO2 levels show a greater variation in photosynthetic rate than the plants
grown under high CO2 (figure 4). Due to their emergent growth S. fallax and S. magellanicum were
not directly in contact with the CO2 concentrations in the water, possibly leading to varying CO
2
availability throughout the containers and consequently less uniform adaptations by the plants.
88 – Solute transport in Sphagnum dominated bogs
Ecological consequences of low affinity CO2 uptake
Because of the competition between CO2 and O
2 at the site of Rubisco, photosynthetic rate, and
consequently Km
and the compensation point, is affected by the ratio between CO2 and O
2 (Bowes
& Salvucci, 1989; Maberly & Spence, 1983). For Sphagnum this was shown by Skre & Oechel (1981)
and here it became evident during the set up of this experiment; in an air-equilibrated measuring
buffer containing 21% O2 the photosynthetic rate of Sphagnum cuspidatum was severely inhibited at
a CO2 concentration of 20 µmol ∙ L-1. Reduced O
2 levels resulted in an increased photosynthetic per-
formance. Due to the production of O2 and the uptake of CO
2 as a result of photosynthetic activity in
combination with the low gas diffusion rates in water, the CO2/O
2 ratios are very likely to decrease
rapidly having negative consequences for photosynthetic performance. Methanotrophic bacteria,
globally occurring in symbiosis with Sphagnum mosses (Kip et al., 2010; Larmola et al., 2010), have
been shown to be of great importance by providing Sphagnum with substantial amounts of CO2 by
oxidizing CH4 using oxygen derived from photosynthesis (Raghoebarsing et al., 2005). We believe
that the consumption of oxygen by these methanotrophs, which reduces the O2 concentration in
the close vicinity of the photosynthesizing cells, is at least as important for the successful growth
of aquatic Sphagnum species.
Considering the low affinity for CO2, the absence of a carbon concentrating mechanism and the
limited morphological and physiological reactions of the plants to low external CO2 concentration,
primary production by Sphagnum is expected to be extremely low when solely supplied with
atmospheric CO2. Growth of Sphagnum therefore requires an additional CO
2 source. Due to aerobic
decomposition processes in the peat layer, additional CO2 will be produced in Sphagnum bogs
(Bridgham & Richardson, 1992; Glatzel et al., 2004; Lamers et al., 1999; Smolders et al., 2001; Wad-
dington et al., 2001) and as a consequence pore water CO2 concentrations can reach up to several
millimoles (Smolders et al., 2001; Chapter 5), thereby ameliorating the effects of low diffusion rates
(Maberly & Madsen, 2002; Silvola, 1990). This agrees with our findings in Chapter 5 that when an
organic layer is lacking, i.e. during the initial stages of bog development, an alternative external
CO2 source seems to be essential for the successful (re-)establishment of Sphagnum.
Chapter 7
Summary and synthesis
Summary and synthesis – 93
The importance of buoyancy-driven water flow in Sphagnum dominated bogs
The requirement of nutrient transport
The motivation for this thesis is based on the findings by Baaijens (1982) and Rappoldt et al. (2003).
They report on a phenomenon called buoyancy-driven water flow, which is the occurrence of
convective flow in water-saturated Sphagnum layers when the temperature difference between day
and night is sufficiently large. During the night, the surface of the peat moss layer cools and results
in a relatively denser and colder water layer on top of warm water. When the density difference
become large enough the cold water in the top layer sinks and warm water rises. It was hypothesized
that in this flow of water, solutes will be transported as well. Therefore, buoyancy-driven water flow
was proposed as a newly discovered mechanism for the translocation of nutrients in a Sphagnum
dominated peat bog.
In bogs, the mineralization of organic matter has been shown to be the most important
nutrient source for Sphagnum (Aerts et al., 1999; Aldous, 2002a, b; Bowden, 1987; Bridgham, 2002;
Damman, 1978, 1986; Morris, 1991; Pakarinen, 1978; Rosswall & Granhall, 1980; Urban & Eisenreich,
1988). In contrast, the highest metabolic activity and nutrient uptake takes place in the capitula
(Aldous, 2002a; Johansson & Linder, 1980; Malmer, 1988; Malmer et al., 1994; Robroek et al., 2009;
Rydin & Jeglum, 2006). The spatial distinction between mineralization and capitula requires an
efficient nutrient transport system.
Diffusion, internal transport and capillary transport were the known nutrient transport
mechanisms in Sphagnum bogs. Complementary to these mechanisms, buoyancy-driven water
flow was hypothesized to be a possible external nutrient transport mechanism, redistributing
nutrients from lower Sphagnum layers to the capitula, and vice versa. Evidence for the development
of buoyancy-driven water flow in a water-saturated Sphagnum layer was provided, based on
theoretical and experimental grounds, by Rappoldt et al. (2003). However, direct evidence for the
transport of solutes was lacking and the importance of buoyancy flow in nutrient transport in
Sphagnum bogs remained unclear. This thesis provides direct evidence for the transport of solutes
by buoyancy flow. Moreover, it is demonstrated that buoyancy flow transports nutrients in such
quantities that it, relative to other transport mechanisms, plays an important role in the redistribu-
tion of nutrients in a water-saturated Sphagnum layer.
The transport of solutes by buoyancy flow
The mesocosm experiment in Chapter 2 unequivocally demonstrates the transport of solutes by
buoyancy-driven water flow in a water-saturated Sphagnum matrix. Moreover, the experiment
shows that due to buoyancy flow a reversal of the gradient can take place in a relatively short period
of time. The findings of this mesocosm experiment indicate that buoyancy-driven water flow acts
as an efficient external nutrient transport mechanism in water-saturated Sphagnum habitats and
thereby can contribute to the supply of nutrients to the Sphagnum capitula in the upper bog layer
and the recycling of nutrients.
The uptake capacity of ammonium by the capitula
In a water-saturated Sphagnum layer, a stepwise increase of solutes near the capitula can be
94 – Solute transport in Sphagnum dominated bogs
induced due to the reversal of the gradient by buoyancy flow (Chapter 2, figure 1). The importance
of buoyancy flow as a nutrient transport mechanism in supplying the capitula is also determined
by its ability to absorb and take up pulses of high nutrient concentrations.
Sphagnum has been shown to be opportunistic in its N uptake (Twenhoven, 1992; Woodin et
al., 1985). The strong cation exchange capacity of the cell wall of Sphagnum is often regarded as
an efficient mechanism to retain cations when supplied by rain water (Bates, 1992; Buscher et al.,
1990). Ammonium, the dominant form of nitrogen in bog water, has been shown to be retained
very efficiently by Sphagnum (Li & Vitt, 1997; Wiedermann et al., 2009; Williams et al., 1999;
Jauhiainen et al., 1998; Twenhoven, 1992). Additionally, the very short lag phase of the substrate
inducible enzyme nitrate reductase enables Sphagnum assimilating even short pulses of nitrogen
(Woodin et al., 1985). Moreover, Sphagnum is very well able to deal with pulse-wise supply of N by
the accumulation of a surplus of N in N-rich amino acids like arginine and asparagine (Baxter et al.,
1992; Karsisto, 1996; Limpens & Berendse, 2003; Nordin & Gunnarsson, 2000).
The observed uptake kinetics for ammonium by Sphagnum cuspidatum and S. fallax (Chapter
2) very well fit the opportunistic nitrogen uptake characteristics. Ammonium uptake is not satu-
rated by concentrations up to 100 µmol ∙ L-1 (Chapter 2, figure 2). The time over which the uptake
rates can be maintained also determines the ability of Sphagnum to benefit from the high nutrient
availability caused by buoyancy flow. In a separate experiment the time dependence of uptake was
determined for NH4
+ (figure 1). It was shown that, when exposed to NH4
+, for 190 hours, the capitula
of S. cuspidatum and S. fallax take up 17 and 13% of the final value within one hour and 76 and
64% within 24 hours, respectively. Together, the uptake characteristics enable Sphagnum to benefit
from a stepwise increase in ammonium (or cation) availability which is the case after precipitation
and buoyancy flow events.
Figure 1. The increase of 15N in capitula of S. cuspidatum (open symbols) and S. fallax (filled symbols; in µmol ∙ g DW-1) over
time when incubated in 100 µmol 15NH4Cl ∙ L-1. Each symbol represents the average of three capitula, which were incubated
in a square Petri-dish filled with 50 mL of experimental solution (see materials and methods section in Chapter 4) containing
the labeled nitrogen, 20 mM MES (pH=4.0) and 100 times diluted artificial rainwater. Error bars represent standard deviations.
The experiment took place in a climate controlled room at 18±1 °C and a 16L:8D photoperiod and a light intensity of 185 µmol
∙ m-2 ∙ s-1).
Summary and synthesis – 95
In Chapter 2 the role of the cation exchange sites in ammonium uptake was demonstrated as
well. The adsorption of ammonium by the cell wall is most important at lower concentrations.
With increasing concentrations the relative importance of adsorption to total uptake decreases;
the cell wall will saturate and increased uptake will take place by intracellular uptake. Compared
to the cation exchange, the active intracellular uptake of ammonium is a slow process. These
observations support the general assumption of the cell wall functioning as a temporal extension
of nutrient availability for intracellular uptake (Buscher et al., 1990; Clymo, 1963; Hajek & Adamec,
2009; Jauhiainen et al., 1998).
The internal transport of nitrogen
The relative importance of buoyancy flow should be weighed against the contribution of the other
transport mechanisms occurring in a water-saturated Sphagnum layer: diffusion and internal
transport. The mesocosm experiment demonstrated that the transport of solutes by buoyancy flow
can be much faster than is possible by diffusion alone. Moreover, buoyancy flow can transport
more solutes upwards than is possible by diffusion. In Chapter 4 the internal transport rate of
nitrogen in Sphagnum was determined. The experiments were performed using two Sphagnum
species, S. cuspidatum and S. fallax, occupying respectively hollows and pools, both wet habitats
where buoyancy flow is likely to occur.
Until this present study it was generally assumed that nitrogen was translocated by an
internal transport mechanism, but direct evidence for such a mechanism was lacking. In Chapter
4 physiological evidence for the internal acropetal transport of nitrogen in Sphagnum is provided.
The findings in Chapter 4 are indicative for symplastic transport of nitrogen, which is in line with
the findings of Ligrone and Duckett (1998b) and Rydin & Clymo (1989) who demonstrated cellular
specializations in Sphagnum for symplasmic transport. No basipetal transport of nitrogen was
observed. Therefore, it seems to be a mechanism that supplies the capitulum with nitrogen and
thereby contributes to the efficient use of N.
The amount of N transported internally to the capitula is low compared to the amounts
potentially transported upwards by buoyancy flow and, subsequently, taken up by the capitula.
The uptake kinetics of the capitula show a much faster uptake rate for ammonium than can be
supplied by internal transport (Chapter 2, figure 2). When exposed to 25 µmol 15NH4
+ ∙ L-1 the uptake
by the capitula of S. cuspidatum is 5.6 ± 2.1 µmol ∙ g DW-1 ∙ hr-1, whereas the transport of this amount
by internal transport takes at least four days. Therefore, with the regular occurrence of buoyancy
flow, the supply of nitrogen by internal transport will be insignificant, compared to the supply of
nitrogen by buoyancy flow. Thus, in comparison with diffusion and internal transport, buoyancy
flow seems to be a quantitatively important nutrient transport mechanism in a water-saturated
Sphagnum habitats.
Buoyancy flow is restricted to water-saturated Sphagnum habitats and, because of its
dependence on varying physical parameters (i.e. the difference in temperature between day
and night) an irregularly occurring phenomenon. The importance of internal transport might
therefore reside in its continuous character, supplying the capitulum slowly, but steadily, with
N and contrasting with the pulsed supply of N by buoyancy flow and atmospheric deposition.
Moreover, during extracellular transport nutrients may be lost to microorganism or vascular plant
roots. For Sphagnum species that form hummocks that extend above the water surface and do not
96 – Solute transport in Sphagnum dominated bogs
benefit from buoyancy flow, internal transport is, next to capillary transport, a possible acropetal
pathway for nutrients. Clymo (1973) estimated the average velocity by capillary flow to be
0.4 mm ∙ min-1. However, this rate, and the concomitant nutrient transport, is dependent on several
external factors, like evaporation, plant density and pore water nutrient concentrations (Clymo &
Hayward, 1982).
Based on the uptake by the stems and internal transport to the capitula of ammonium by S.
cuspidatum, the rate by which nitrogen is transported internally was estimated at 5 mm ∙ day-1,
which is in accordance with a half time value of equilibration between the stem and capitula of 17
days. Compared to external transport mechanism (buoyancy flow and capillary transport), internal
transport is very likely of limited importance for the upward transport of externally supplied N to
the capitula. The main function of internal transport is therefore assumed to be the reallocation of
internally broken down N.
Since the internal transport of nitrogen is a mechanism for efficient nitrogen use, the trans-
port rate is expected to be reduced under high N availability (Bragazza et al., 2004). The transport
rate of solutes by buoyancy-driven water flow is independent from the internal concentration,
thus also supplying the capitula with nitrogen (and other nutrients) when there is a low demand.
The assimilation of N in amino acids by Sphagnum enables the Sphagnum plants to take up the N
and store it for later use. With the regular occurrence of buoyancy flow, and thereby the supply of N
to the capitula, the sink strength for N of the capitula will be reduced and consequently the rate of
internal transport as well. Thus, the relatively low contribution of internal transport in the trans-
port of N in a water-saturated Sphagnum layer will very likely be reduced even more in the presence
of buoyancy flow.
Ecological importance of buoyancy flow
Chapter 3 clearly shows the regular occurrence of buoyancy-driven water flow in a Sphagnum pool
in a field situation. Moreover, the theoretical models of Rappoldt et al. (2003) on the develop-
ment of buoyancy flow can be applied to the field situation. Based on the Ra numbers, calculated
from the vertical hydraulic conductivity of the Sphagnum cores and the difference in temperature
between day and night, the occurrence and starting times of buoyancy flow development was very
well predictable. The results from the GIS study indicate that many peatlands throughout the world
are subjected, several days each month during the growth season, to temperature difference be-
tween day and night, which are suitable for the development of buoyancy flow.
Sphagnum bogs can consist of a patchwork of hollows, lawns and hummocks. As buoyancy
flow is restricted to the water layer in a Sphagnum bog, direct supply of nutrients from deeper layers
to the capitulum by buoyancy flow only takes place in hollows. The transport of nitrogen by buoy-
ancy-driven water flow and the subsequent uptake by the capitula in the upper Sphagnum layer in
a field situation was demonstrated by the field experiment performed in the Rancho Hambre bog
complex, Argentina (Chapter 2; figure 2). The increase in 15N concentration in the S. fimbriatum
capitula in the treatment with unobstructed convective flow, indicates the upward transport of 15N by buoyancy flow. As expected, in the S. magellanicum sites, however, no increase in 15N in the
capitula was observed.
Moreover, in the observed S. magellanicum lawns (Chapter 3) the water table is located about 20
cm below the top of the Sphagnum plants which form an insulating layer (Van der Molen & Wijm-
Summary and synthesis – 97
stra, 1994), preventing the development of a cool water surface layer and instability in the water col-
umn. If buoyancy flow would nevertheless occur, solutes transported from deeper layers to the upper
water layer would still have to be transported to the capitula by capillary transport (or by diffusion in
case of, for example, CO2). In this case buoyancy flow only acts as an auxiliary transport mechanism
and its relative importance in the nutrient supply to the capitula is determined by the height of the
capitula above the water level. Such a situation could be found in Sphagnum lawns and the transition
zones between pools and hummocks. As the initial successional stage of a Sphagnum bog is the colo-
nization of aquatic Sphagnum species of water bodies followed by the invasion of hummock forming
species, buoyancy flow seems to be particularly important in the early stages of bog development.
An overview of the (expected) relative importance of buoyancy flow, diffusion, internal
transport and capillary transport in three different Sphagnum habitats (hollow, lawn and hummock)
are presented in figure 2.
Figure 2. A schematic cross section of three different Sphagnum habitats (hollows, lawns and hummocks) indicating the
relative importance (in %) of buoyancy flow (light grey), diffusion (dotted grey), internal transport (dark grey) and capillary
transport (medium grey) in the redistribution and cycling of nutrients. The three habitats differ in the height of the capitula
(indicated by the asterisks) relative to the water table level (indicated by the thick dashed horizontal lines). In hollows, buoy-
ancy flow is the major mechanism by which nutrients are transported. Since the capitula are at the water level in hollows,
buoyancy flow can directly contribute to the supply of nutrients to the capitula. Diffusion and internal transport take place as
well in hollows but has been shown to be less rapid and effective than buoyancy flow. Even though buoyancy flow might occur
irregularly, the opportunistic nutrient uptake capacity of Sphagnum will result in a significant contribution of buoyancy flow
even at a few occurrences of buoyancy flow. Since buoyancy flow is restricted to the water layer its contribution will be less in
lawns and very likely totally absent in hummocks. In contrast, capillary transport only takes place above the water table and
therefore only contributes to nutrient transport in lawns and hummocks. Since the plant density is higher in hummocks than
in lawns, capillary transport will be faster and consequently more important in hummocks than in lawn. Moreover, in lawns
buoyancy flow might occur, reducing the relative contribution of capillary transport in lawns compared to hummocks. With
increasing heights of the capitula above the water table the relative importance of diffusion will be less and completely reduced
to zero in hummocks. Internal transport has been shown to be a slow mechanism. As a consequence, internal transport very
likely plays a minor role in the distribution of nutrients in especially hollows and hummocks because of the presence of the
faster mechanisms buoyancy flow and capillary transport, respectively. In lawns the relative importance of internal transport
is expected to be greatest since buoyancy flow is less effective and capillary transport will be less fast due to the low plant
density in these habitats.
98 – Solute transport in Sphagnum dominated bogs
Nitrogen
The main N source for Sphagnum has been shown to be re-mineralized N (Aerts et al., 1999; Aldous,
2002b; Bridgham, 2002; Gerdol et al., 2006; Morris, 1991; Urban & Eisenreich, 1988). The importance
of re-mineralization of nitrogen for Sphagnum growth has been demonstrated in situ by Urban &
Eisenreich (1988). They calculated the assimilation of nitrogen by plants (primarily Sphagnum) to
be 66 kg ∙ ha-1 ∙ yr-1, whereas only 14.6 kg N ∙ ha-1 ∙ yr-1 was supplied by total inputs. The remainder
was supplied by mineralization of the peat.
Maybe one would expect that vascular plants may be better competitors for this source of
nitrogen by scavenging the peat for mineralized N with their fine roots (Backeus, 1990; Jackson
et al., 1990) and as a consequence vascular plants will outcompete Sphagnum mosses. This is,
however, not the case. Instead, Sphagnum is capable of very efficient (re-)use of mineralized
nitrogen, creating a low nutrient environment which consequently contributes to their domi-
nance over vascular plants.
As already mentioned by Gerdol et al. (2006), these findings are contradictory to the general idea
that Sphagnum and vascular plants utilize spatially distinct nutrient pools, with Sphagnum relying
on N from precipitation and vascular plants on mineralization of senescing organic matter in the
deeper acrotelm (Malmer et al., 1994; Pastor et al., 2002).
The importance of nutrient transport for efficient nutrient recycling is generally accepted.
For example, Aldous (2002b) mentions the translocation of nitrogen as a key process in bog nu-
trient cycling. Blodeau et al. (2006) states that Sphagnum mosses are the dominant species in
northern peatlands, in part because they have the capability to conserve nitrogen by transferring
it from lower, inactive parts of their stem to apices where new biomass is formed (Aldous, 2002a,
b; Malmer, 1988). However, the contribution of the different nutrient transport mechanisms in
nutrient recycling was never determined. In this thesis we demonstrate that buoyancy-driven
water flow is an important mechanism contributing to the recycling of mineralized nutrients in
Sphagnum bogs. Consequently, Sphagnum mosses may outcompete vascular plants more easily
and thereby enhance their ability to engineer the ecosystem (Van Breemen, 1995).
Carbon dioxide
In waterlogged Sphagnum habitats, the stagnant bog water will result in thick boundary layers and
long diffusion path, lengths which reduces the supply of CO2 to the carbon assimilating cells and,
consequently, the photosynthetic rate (Bowes & Salvucci, 1989; Rice & Giles, 1996; Silvola, 1990;
Williams & Flanagan, 1996). Consequently, high rates of underwater photosynthesis can only be
sustained when the leaves are exposed to high levels of CO2
(Jauhiainen & Silvola, 1999; Paffen &
Roelofs, 1991; Silvola, 1990; Smolders et al., 2003). Submerged Sphagnum species that inhabit peat
hollows have been shown to be limited by CO2 (Chapter 5; Rice & Giles, 1996; Rice & Schuepp, 1995).
On the other hand, high carbon dioxide concentrations haven been shown to stimulate Sphagnum
growth (e.g. Smolders et al., 2001; Chapter 5). However, doubling atmospheric CO2 concentrations
appear to have rather limited effects on the growth of Sphagnum (Heijmans et al., 2001; Hoosbeek
et al., 2001; Jauhiainen et al., 1994; Toet et al., 2006).
The high CO2 requirements for Sphagnum is determined by physiological characteristics of the
CO2 uptake mechanism of Sphagnum, as shown in Chapters 5 and 6. Sphagnum mosses have been
shown to be pure CO2 users (Bain & Proctor, 1980; Chapter 6) and therefore exclusively depend on
Summary and synthesis – 99
diffusion of CO2 to the site of carbon fixation. The kinetic properties of CO
2 uptake indicate an ad-
aptation to a high CO2 availability. The investigated Sphagnum species are characterized by a high
CO2 compensation value, the CO
2 concentration at which CO
2 fixation by photosynthesis balances
CO2 loss by respiration. Air-equilibrated water contains a CO
2 concentration of 10 - 20 µmol ∙ L-1
between 25 and 10°C. The high compensation value of Sphagnum implies that under air-saturated
conditions no, or extremely limited, net carbon accumulation can occur. In the acidic bog envi-
ronment, where no reservoir of bicarbonate is present to replace the CO2 that is taken up, this is
especially relevant and Sphagnum growth will not occur when CO2 is provided exclusively through
equilibration with air. Furthermore, The high Km
values of 231.4 and 133.2 µM CO2 for S. cuspidatum
and S. fallax, respectively, further indicate that even when CO2 is present at a concentration that is
higher than air-saturated, carbon utilization is not optimal.
Remarkably, Sphagnum cuspidatum was shown to be able to form thinner leaves when grown
submerged at low CO2 availability (Baker & Boatman, 1985; Rydin & McDonald, 1985), which
will reduce the boundary layer resistance and facilitate CO2 uptake. Moreover, in Chapter 6 it is
demonstrated that S. cuspidatum is able to increase its photosynthetic capacity under CO2-lim-
iting conditions, possibly to increase carbon fixation at low external CO2 availability. Very likely
due to an increased Rubisco concentration under CO2 limiting conditions compared to high CO
2
availability.
Buoyancy flow will result in net transport when a vertical gradient exists, as for example is the
case for nutrients. Uptake and assimilation of CO2 will result in depletion in the zone of the capitula
where most photosynthetic activity takes place. In contrast, in the lower acrotelm CO2 is released
as a consequence of the decomposition of organic material. An increasing CO2 concentration with
depth has been shown in a water-saturated Sphagnum layer (Lloyd et al., 1998). Therefore, buoyancy
flow is very likely an important mechanism also in replenishing CO2 in the upper Sphagnum layer
and enhancing photosynthesis.
Oxygen
Photosynthetic activity in the top layer of the Sphagnum matrix will also result in the production of
oxygen. Due to the thick boundary layers and the low diffusion rates of oxygen in water the oxygen
will accumulate in the upper Sphagnum layer during the day, resulting in a decreasing gradient with
depth (Adema et al., 2006; Lloyd et al., 1998). Lloyd et al. (1998) measured a steep oxygen gradient
in the upper four centimeters of a water-saturated Sphagnum layer, decreasing from 300 to 0 µM.
The mixing of the water layers by buoyancy flow will result in a net downward transport of oxygen.
Adema et al. (2006) attributed a conspicuous change in oxygen concentration at 5 cm depth in a
Sphagnum layer to the occurrence of buoyancy flow.
Because of the competition between CO2 and O
2 at the site of Rubisco, photosynthetic rate is
negatively affected by the accumulation of O2 in the surroundings of the capitula (Chapter 6; Bowes
& Salvucci, 1989; Raven, 2011; Raven et al., 2008; Skre & Oechel, 1981). Together with the upwards
transport of carbon dioxide by buoyancy flow, the CO2:O
2 ratio in the upper Sphagnum layer will
increase and thereby enabling the mosses to photosynthesize. However, during the day the CO2:O
2
ratio decreases again due to photosynthetic activity. Therefore, photosynthetic rates of aquatic
Sphagnum species might be highest in the beginning of the day.
The downward transport of oxygen very likely will increase decomposition rates since
100 – Solute transport in Sphagnum dominated bogs
the aerobic decomposition of organic material is significantly higher than the anaerobic
decomposition (Bridgham et al., 1998; Waddington et al., 2001). Consequently, this might result
in a positive feedback mechanism in which the concentrations of CO2 and nutrients like N and
P will be increased by the downward supply of oxygen. These nutrients will become available to
the growing Sphagnum when transported upwards by buoyancy flow which in turn will stimulate
photosynthesis. Consequently, under N-limiting conditions, the transport of oxygen by buoyancy
flow might even determine primary production.
Moreover, the negative effect of oxygen on photosynthesis might give rise to another impor-
tant feature of the symbiosis between Sphagnum and methanotrophic bacteria. The consumption
of oxygen by the methanotrophs, which reduces the O2 concentration in the close vicinity of the
photosynthesizing cells and thereby enhance photosynthesis. This might at least be as important
for the successful growth of aquatic Sphagnum species as the concomitant CO2 release.
Methane
Inseparable from the availability and redistribution of O2 and CO
2 in Sphagnum bogs is the presence
of methane. Under more reductive conditions the methane production in the catotelm may become
higher than the production of CO2. This methane can be oxidized by methanotrophic bacteria to
CO2, which then can be used as a carbon source by Sphagnum mosses (Kip et al., 2010; Raghoebars-
ing et al., 2005). Methane is anaerobically produced in large quantities in bogs (e.g. Gorham, 1991).
Nevertheless, emissions of methane to the atmosphere are relatively low (Larmola et al., 2010) due
to the activity of methanotropic bacteria. The mixing of methane and photosynthetically produced
oxygen by buoyancy flow might have an important role in 1) the CO2 supply to the capitula and 2)
the low methane emission rates, thereby playing a significant role in the global carbon cycling.
Other determinants for the importance of buoyancy flow
Basically, the nutrient concentration in acrotelmic water will be determined by the decomposition
rate in the catotelm and the depletion in the top water layer by uptake and assimilation and
dilution by rain water. As discussed above, buoyancy flow has its effect on the distribution of
multiple nutrients which mutually interact. Not mentioned are phosphorus and potassium, which
have been shown to limit Sphagnum growth (Aerts et al., 1992; Bridgham et al., 1996; Hoosbeek et
al., 2002) and the organic peat layer being an important source for these nutrients (Bates, 1992;
Damman, 1978, 1986).
The effect of buoyancy flow on the nutrient concentration and distribution will be determined
by two other factors. First, the depth of the buoyancy cells: the deeper the cells the more mixing
takes place which very likely results in a higher nutrient availability. Second, the frequency of
buoyancy flow events. Because of its dependence on the difference in temperature between day
and night, buoyancy flow is an irregularly occurring mechanism. The more buoyancy flow events
occur the more the water layer will be mixed, which results in higher decomposition rates, the
amount of transported nutrients and photosynthetic activity (see above).
At low frequencies, gradients are allowed to be build up and relatively large amounts of solutes
might be transported at once. Consequently, the transport and nutrient availability might not be in
synchrony with the requirements of Sphagnum for primary production. For nitrogen this might not
pose a problem, since the pulse-wise availability of N is buffered by the opportunistic uptake and
Summary and synthesis – 101
assimilation characteristics of Sphagnum (Chapter 2). In contrast, photosynthesis will very likely
benefit more from a frequent occurrence of buoyancy flow. Photosynthetic activity will result in
lowered CO2 and increased O
2 levels in the surroundings of the capitula, which in turn will inhibit
photosynthesis. Since this inhibition can occur within the period of one day, overall photosyn-
thetic rates will be enhanced optimally with a diurnal occurrence of buoyancy flow.
The relative importance of transport of nutrients from deeper water layers to the top layer with
the capitula, is higher when atmospheric input is low. This is especially relevant for nitrogen since
atmospheric N deposition has increased significantly during recent decades (Vitousek, 1982). An
increased N deposition results in higher ammonium concentrations in the bog water (Lamers et al.,
2000). Under such conditions the decreasing nitrogen gradient with depth will be reduced or even
be completely diminished, thereby reducing the net upward transport of nitrogen by buoyancy
flow to zero. Under high N availability, Sphagnum growth has been shown to shift from a nitrogen
to a phosphorus limitation (Aerts et al., 1992). Consequently, the importance of buoyancy-driven
water flow as a nutrient transport mechanism will also shift from nitrogen to phosphorus. More-
over, the availability of CO2 might become important under high N loads as well. However, since ar-
eas with a high nitrogen deposition load only slightly overlap with the area covered with peatlands
(Chapter 3, figure 4), the importance of buoyancy flow in the transport and recycling of nitrogen in
Sphagnum bogs, is hardly diminished by increased, anthropogenic N deposition.
Restoration and conservation of Sphagnum bogs
Throughout the world, Sphagnum bogs have become greatly endangered and consequently much
effort is dedicated to the restoration of damaged bogs and the conservation of bog remnants.
Therefore, studying the functioning of Sphagnum bogs is inseparable to the restoration and
conservation of these ecosystems. Chapter 5 focuses on the restoration of Sphagnum bogs. As
mentioned in the previous paragraph, several studies have reported on the stimulation of Sphag-
num growth by high CO2 concentrations (Baker & Boatman, 1990; Jauhiainen & Silvola, 1999; Paffen
& Roelofs, 1991; Riis & SandJensen, 1997; Roelofs, 1983; Smolders et al., 2001; Smolders et al., 2003).
Chapter 5 and 6 show the physiological background of the high CO2 needs of Sphagnum. In addition
to these studies, Chapter 5 reports on a field study in which the significance of CO2 availability for
the successful re-establishment of Sphagnum and subsequent bog development is demonstrated.
Study area was the “Dwingelderveld”, a nature reserve in the north of the Netherlands
characterized by several small damaged Sphagnum bogs throughout the area. After rewetting
measures were taken, the developmental success between these bogs varied significantly; some
bogs developed well, whereas others did not. It was shown that the poorly developed bogs were
limited in CO2, whereas the successful re-establishment of Sphagnum in the well developed bogs
was correlated with high CO2 availability.
Groundwater essential for bog development; a paradox
The findings in Chapter 5 clearly show that high CO2 availability is a pre-requisite for the successful
re-establishment of Sphagnum mosses and subsequent bog development. In well developed
bogs CO2 is sufficiently available due to the decomposition of organic material in the peat layer
102 – Solute transport in Sphagnum dominated bogs
(Bridgham & Richardson, 1992; Glatzel et al., 2004; Smolders et al., 2001; Waddington et al., 2001).
However, damaged bogs subjected to peat cuttings or drainage often lack such a carbon source
and an additional CO2 source is needed for Sphagnum growth. Therefore, one of the conclusions of
Chapter 5 is that CO2 availability should be included in bog restoration feasibility studies.
Remarkably, water chemistry analysis revealed that the well developed bogs in the “Dwingelderveld”
(Chapter 5) received carbon rich groundwater from outside the bogs, increasing CO2 concentrations
in the bog stimulating Sphagnum growth and thereby bog development. Interestingly, this
dependence of Sphagnum growth on groundwater input hides a paradox; bogs exists due to being
isolated from the groundwater. In such an ombrotrophic, low nutrient environment Sphagnum
mosses have a competitive advantage over vascular plants. Moreover, groundwater is often
characterized by a high pH and a high Ca2+ concentration and has been shown to be toxic for most
Sphagnum species (Skene, 1915; Clymo & Hayward, 1982).
The need for carbon under high N loads
A large number of studies have focussed on the negative effects of an increased atmospheric
nitrogen deposition on the growth of Sphagnum and the functioning of Sphagnum bogs (Limpens
et al., 2011). The lack of re-colonization of Sphagnum mosses and hampered growth of already
established Sphagnum mosses has often been ascribed to high levels of atmospheric nitrogen
deposition (Lamers et al., 2000; Li & Vitt, 1994; Money & Wheeler, 1999; Twenhoven, 1992). Chapter
5 demonstrates that the successful restoration of Sphagnum bogs is possible under high nitrogen
loads. It is hypothesized that CO2 can compensate the negative effects of a high nitrogen deposition
on an ecological as a physiological level.
Sphagnum mosses lack a regulatory mechanism for nitrogen uptake. As a consequence,
internal nitrogen concentration increases with increasing nitrogen deposition rates (Bragazza et
al., 2005; Lamers et al., 2000; Limpens et al., 2011). However, under high levels of N deposition
levels, Sphagnum is not able to filter out all the N from precipitation and nitrogen leaches to the
rhizosphere were it becomes available for vascular plants, thereby making the bog vulnerable
to invasions by competitive vascular plants that require a high N supply. The reduced growth of
Sphagnum is often attributed to the shading by vascular plants (Berendse et al., 2001; Heijmans et
al., 2001; Lamers et al., 2000; Limpens et al., 2011).
After uptake nitrate is reduced to NH4
+ prior to assimilation, while ammonium is directly
assimilated into glutamine (Rudolph et al., 1993). Subsequently, glutamine is converted into other
amino acids (Kahl et al., 1997; Rudolph et al., 1993). With increasing N deposition, plants are no
longer N-limited, but will still take up N (Lamers et al., 2000). Continued assimilation of N leads to
accumulation of free amino acids (Baxter et al., 1992; Karsisto, 1996; Nordin & Gunnarsson, 2000).
Under these conditions a decrease in Sphagnum growth was observed (Baxter et al., 1992; Nordin
& Gunnarsson, 2000), possibly the result of the accumulation of amino acids requiring carbon
and energy (Baxter et al., 1992; Nordin & Gunnarsson, 2000). Hence, under high N availability
an additional need of carbon is very likely. Paffen & Roelofs (1991) showed that high ammonium
concentrations in the water layer had no major effects on the growth of submerged growing S.
cuspidatum when simultaneously a high concentration of 1000 µmol CO2 ∙ L-1 was applied.
High CO2 levels might enable Sphagnum to increase their competitive strength over vascular
plants by enabling continued growth and N assimilation, keeping the nitrogen concentrations low
Summary and synthesis – 103
in the pore water and thereby gain competitive strength over vascular plants.
The importance of carbon under high N availability is supported by the following findings.
In Sphagnum, under normal conditions, NH4
+ is stored in amino acids having relatively high C:N
ratios like glutamine (5:2) (Kahl et al., 1997; Nordin & Gunnarsson, 2000; Rudolph et al., 1993).
However, under increased nitrogen loads, N is accumulated in amino acids with lower C:N ratios,
mostly arginine (Nordin & Gunnarsson, 2000), which has a C:N ratio of 3:2, the lowest all amino
acids. This shift in amino acid accumulation suggests an economical use of carbon under high N
loads. Thus, the high carbon dioxide needs of Sphagnum are very likely enhanced under high N
availability.
Moreover, the interaction between CO2 availability and high levels of atmospheric N deposi-
tion might have an effect on nutrient uptake as well. The accumulation of N in N-rich rich amino
acids has been shown to be at the expense of C rich amino acid like phenylalanine (Smolders et al.,
2001), which is a precursor of cell wall compounds like polymeric uronic acids and phenolic com-
pounds. A study performed by Richter and Dainty (1989) on the cell wall ion exchange capacity of
Sphagnum russowii suggests that polymeric uronic acids account for over half the cation exchange
capacity (CEC) and phenolic compounds for about 25%. With its high CEC, the cell wall also plays
an important role in nutrient uptake (see Chapter 2). As mentioned above, high N loads reduces the
amounts of amino acid important in cell wall assimilation and composition. This might result in
changes in CEC and consequently in nutrient uptake processes. Additionally, high concentrations
of NH4
+ in bog water (as a consequence of high N loads) directly affect cation composition at the
cell wall and possibly thereby the nutrient balance in Sphagnum.
Concluding remarks
Sphagnum mosses are known for their ability to engineer their environment (Van Breemen, 1995).
One of these engineering abilities is to maintain low nutrient concentrations in the pore water,
preventing increased vascular plant cover and keeping the competitive advantage. Depending
on Sphagnum habitat, different transport mechanism play a role in this efficient scavenging for
nutrients. In this thesis we demonstrated that buoyancy-driven water flow plays an important role
in the distribution of nutrients in Sphagnum bogs. Buoyancy flow might also have an important
influence on primary production and decomposition rates by preventing the building up of
gradients that have a negative feedback on these processes (N, CO2 and O
2). In this context, the
disruption of the nutrient balance in these ecosystems by high N atmospheric deposition loads are
easily understood.
Buoyancy flow, a worldwide occurring, but poorly studied phenomenon in Sphagnum bogs
will have to be taken account when we want to understand bog functioning.
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Solute transport in Sphagnum dominated bogs – 117
Samenvatting
Samenvatting – 121
De aanleiding van dit proefschrift was een fenomeen dat door Baaijens (1982) en Rappoldt et
al. (2003) is beschreven. Dit fenomeen, ‘buoyancy-driven water flow’ genaamd, is een convectie-
stroming in de bovenste laag van een met water verzadigde veenmoslaag, die optreedt als gevolg
van het temperatuurverschil tussen dag en nacht. ’s Nachts koelt het oppervlak van de veenmos-
laag af, wat resulteert in een relatief koude, zware laag bovenop warmer, lichter water. Als het
dichtheidsverschil tussen deze twee lagen groot genoeg is dan zinkt de bovenste, zwaardere laag
en komt de warme, ondergelegen laag naar boven.
Hoogvenen zijn voor de aanvoer van nutriënten voornamelijk aangewezen op atmosferische
depositie. Het blijkt echter dat onder niet vervuilde omstandigheden de jaarlijkse aanvoer van
nutriënten middels depositie niet voldoende is voor de waargenomen primaire productie in deze
systemen. Er moeten derhalve andere nutriëntenbronnen bij betrokken zijn. Mineralisatie van or-
ganisch materiaal is de belangrijkste bron van nutriënten voor veenmossen in deze ecosystemen.
De grootste metabolische activiteit en de grootste opname van nutriënten vindt echter plaats in
het capitulum, het bovenste deel van een individuele veenmosplant. De ruimtelijke scheiding tus-
sen mineralisatie en de capitula vereist een efficiënt systeem voor het transport van nutriënten.
Diffusie, intern transport en capillair transport waren de bekende wegen waarlangs nutriën-
ten worden getransporteerd in een hoogveen. Aanvullend op deze mechanismen werd buoyancy
flow beschreven als mogelijk mechanisme waarlangs nutriënten van lagere veenlagen naar boven
en vice versa, kunnen worden getransporteerd. Bewijs voor het optreden van buoyancy flow werd
geleverd door Rappoldt et al. (2003) op theoretische en experimentele gronden. Echter, direct
bewijs voor het transport van opgeloste stoffen ontbrak en het belang van buoyancy flow in het
transport van nutriënten in een hoogveen bleef onduidelijk. Dit proefschrift levert direct bewijs
voor het transport van opgeloste stoffen middels buoyancy flow. Bovendien laat dit proefschrift
zien dat buoyancy flow nutriënten transporteert in zulke hoeveelheden dat het, ten opzichte van
andere transportmechanismen, een belangrijke rol speelt in de nutriëntenvoorziening van een
veenmoslaag wanneer deze met water is verzadigd.
Transport van nutriënten middels buoyancy flow
Het mesocosm experiment in hoofdstuk 2 laat zien dat opgeloste stoffen snel én in grote
hoeveelheden getransporteerd kunnen worden door buoyancy flow in een Sphagnum-matrix.
Hieruit kan worden geconcludeerd dat buoyancy flow een efficiënt extern transportmechanisme is
in een waterverzadigde Sphagnum-laag en kan bijdragen aan de voorziening van nutriënten in de
bovenste laag van een hoogveen en het hergebruik van nutriënten.
De opnamecapaciteit van ammonium door veenmossen
Daarnaast laat het mesocosm experiment zien dat als gevolg van buoyancy flow er een sterke
plotselinge, of stapsgewijze, toename van nutriënten in de bovenste Sphagnum-laag kan op-
treden. Door buoyancy flow getransporteerde nutrienten zijn alleen van ecologisch belang als
Sphagnum planten het vermogen hebben om die in korte tijd toegenomen hoeveelheid nutriënten
daad werkelijk te benutten. In hoofdstuk 2 is de opnamekinetiek van Sphagnum voor ammonium
bepaald. De resultaten laten zien dat Sphagnum heel goed kan profiteren van de stapsgewijze be-
schikbaarheid van ammonium. In hoofdstuk 2 is ook de rol van de celwand als ionenwisselaar
in de opname van nutriënten gedemonstreerd. Adsorptie van ammonium aan de celwand vindt
122 – Solute transport in Sphagnum dominated bogs
vooral plaats bij lagere concentraties. Met toenemende concentraties neemt het belang van
adsorptie in verhouding tot de totale opname af; de celwand raakt dan verzadigd en een toename
in de opname zal plaatsvinden via interne opname. Vergeleken met adsorptie aan de celwand is de
actieve interne opname een langzaam proces. Deze waarnemingen steunen het algemene idee dat
de celwand functioneert als een tijdelijke bufferopslag voor nutriënten, voordat ze intern worden
opgenomen.
Het interne transport van stikstof
Het relatieve belang van buoyancy flow moet afgewogen worden tegen de bijdrage van de andere
transportmechanismen, diffusie en intern transport, die op kunnen treden in een waterver-
zadigde veenmoslaag. Uit het mesocosm experiment in hoofdstuk 2 blijkt dat buoyancy flow een
sneller transportproces is dan diffusie. Daarnaast is gebleken dat er meer voedingsstoffen middels
buoyancy flow getransporteerd kunnen worden dan via diffusie. In hoofdstuk 4 is de interne
transportsnelheid van stikstof in Sphagnum bepaald. Tot deze studie werd algemeen aangenomen
dat stikstof intern door Sphagnum wordt getransporteerd, maar direct bewijs daarvoor ontbrak.
Hoofdstuk 4 levert fysiologisch bewijs voor het opwaartse transport van stikstof in Sphagnum.
Echter, ten opzichte van de hoeveelheid stikstof die mogelijk door buoyancy flow wordt getrans-
porteerd (en vervolgens wordt opgenomen), is de hoeveelheid die intern naar de capitula wordt
getransporteerd klein). Onder omstandigheden waarin buoyancy flow regelmatig optreedt, is de
bijdrage van intern transport aan de voorziening van stikstof bijna verwaarloosbaar klein. Dus
in vergelijking met diffusie en intern transport is buoyancy flow kwantitatief het belangrijkste
mechanisme voor het transport van nutriënten.
Het ecologische belang van buoyancy flow
Hoofdstuk 3 laat zien dat buoyancy flow onder veldomstandigheden ook voorkomt. Middels een
GIS-studie is gebleken dat in vele hoogvenen over de gehele wereld, tijdens het groeiseizoen,
meerdere dagen per maand, temperatuurverschillen tussen dag en nacht optreden die geschikt
zijn voor de ontwikkeling van buoyancy flow.
De belangrijkste bron voor stikstof voor Sphagnum is de mineralisatie in de diepere lagen in
een hoogveen. Het is gebleken dat Sphagnum deze stikstof efficiënt hergebruikt. Het transport
van stikstof naar de capitula is hierbij van groot belang. In hoofdstuk 2 wordt het transport van
stikstof middels buoyancy flow en de daaropvolgende opname door de capitula aangetoond in een
veldsituatie.
Ook voor de beschikbaarheid van koolstofdioxide in een Sphagnum-veen kan buoyancy
flow van groot belang zijn. In waterverzadigde Sphagnum-habitats kan de stilstaande waterlaag
resulteren in dikke grenslagen en grote diffusieweerstanden. Hierdoor wordt de toevoer van CO2
naar de planten vertraagd, resulterend in lagere fotosynthesesnelheden en vervolgens in een ver-
minderde groei. Hoge fotosynthetische activiteit onder water kan alleen worden bereikt onder
hoge CO2- beschikbaarheid. Het is gebleken dat ondergedoken Sphagnum-soorten in hun groei
worden gelimiteerd door de CO2-beschikbaarheid. De behoefte aan hoge CO
2 beschikbaarheid
wordt bepaald door fysiologische karakteristieken van Sphagnum (Hoofdstuk 5 en 6). Veenmos-
sen kunnen alleen CO2 gebruiken als koolstofbron en zijn volledig afhankelijk van diffusie voor
hun koolstofvoorziening. De opnamekinetiek voor CO2 laat een aanpassing zien aan een hoge
Samenvatting – 123
CO2 beschikbaarheid (Hoofdstuk 5 en 6). In hoofdstuk 6 wordt bovendien gedemonstreerd dat
Sphagnum cuspidatum in staat is de fotosynthetische activiteit te verhogen onder CO2 limiterende
omstandigheden, waarschijnlijk door de koolstoffixatiecapaciteit te verhogen.
Opname en assimilatie van CO2 in de laag waarin de capitula zich bevinden resulteert in
verlaging van de CO2 concentratie. In de diepere lagen daarentegen, komt veel CO
2 vrij bij de afbraak
van organisch materiaal. Door buoyancy flow wordt CO2 in de bovenste lagen aangevuld, wat tot
een hogere fotosyntheseactiviteit kan leiden. Ook voor het transport van zuurstof kan buoyancy
flow van groot belang zijn. Bij fotosynthese wordt zuurstof geproduceerd. Aangezien fotosynthese
voornamelijk plaatsvindt in de capitula in de bovenste Sphagnum laag, zal zuurstof zich daar opho-
pen. Vanwege de competitie tussen CO2 en O
2 voor binding aan RuBisCo, zal de fotosyntese worden
geremd. Door het optreden van buoyancy flow zal de zuurstofconcentratie in de bovenste laag afne-
men. In combinatie met het opwaartse transport van CO2 zal de fotosynthese gestimuleerd worden.
Bovendien zal waarschijnlijk de afbraak van organisch materiaal in diepere lagen gestimuleerd
worden door het neerwaartse transport van zuurstof. Dit kan resulteren in een positieve terugkop-
peling waarbij door het neerwaartse transport van zuurstof de concentraties CO2, N en P onderin
toenemen, die vervolgens opwaarts worden getransporteerd middels buoyancy flow en beschikbaar
komen voor de veenmossen, leidend tot hogere fotosynthese, meer zuurstof, etc. Daarnaast zal
door de beschikbaarheid van zuurstof in diepere lagen de methaanemissie reduceren als gevolg
van oxidatie tot CO2. Op deze manier draagt buoyancy flow bij aan de Global Carbon Cycle.
Factoren die de effecten van buoyancy flow bepalen
In principe wordt de concentratie van nutriënten in het veenwater bepaald door afbraaksnelheid
van organisch materiaal en de benutting ervan in de bovenste laag door veenmossen en de verdun-
ning door regenwater. Zoals hierboven genoemd heeft buoyancy flow invloed op de verdeling van
alle opgeloste stoffen die onderling op elkaar inwerken.
De invloed van buoyancy flow op de nutriëntenconcentratie en -verdeling wordt mede bepaald
door twee andere factoren. Ten eerste de diepte van de buoyancy cells: hoe dieper de buoyancy cells des
te meer menging zal plaatsvinden, wat waarschijnlijk resulteert in een hogere beschikbaarheid van
nutriënten. Ten tweede is de frequentie van optreden van belang. Hoe vaker buoyancy flow optreedt,
des te meer zal de waterlaag gemengd zijn, resulterend in hogere decompositiesnelheden, hogere
hoeveelheden nutriënten die getransporteerd worden en hogere fotosynthetische activiteit.
Het relatieve belang van het transport van nutriënten uit diepere lagen naar de bovenste laag
waar de capitula zich bevinden, is groter naarmate de inbreng vanuit de atmosfeer laag is. Dit
geldt met name voor stikstof, waarvan de afgelopen decennia de atmosferische depositie sterk is
toegenomen. Een hoge stikstofdepositie resulteert in hoge ammoniumconcentraties in het veen-
water en vlakt de gradiënt in de stikstofconcentratie met de diepte af. Zonder concentratiegradiënt
is het netto transport van stikstof middels buoyancy flow nul. Echter, aangezien de gebieden met
een hoge stikstofdepositie slechts voor een klein deel overlappen met de gebieden waar buoyancy
flow op kan treden (Hoofdstuk 3, figuur 4), wordt het belang van buoyancy flow voor het transport
van nutriënten nauwelijks beïnvloedt door een toename in stikstofdepositie.
Herstel en behoud van hoogvenen
Over de hele wereld worden hoogvenen sterk bedreigd. Als gevolg daarvan wordt er veel moeite
124 – Solute transport in Sphagnum dominated bogs
gedaan om deze ecosystemen te restaureren en overblijfselen ervan te behouden. Hoofdstuk
5 handelt over het herstel van hoogvenen. Hierin wordt een veldstudie beschreven waarin het
belang van de beschikbaarheid van CO2 voor de groei van veenmossen en de daaropvolgende
ontwikkeling van hoogveen wordt gedemonstreerd. Studiegebied was het Dwingelderveld, een
groot natuur gebied in het noorden van Nederland. Dit gebied wordt gekenmerkt door meerdere
kleine hoogveentjes die, wat ontwikkeling betreft sterk van elkaar verschillen; sommigen zijn goed
ontwikkeld, andere niet. Het bleek dat de slecht ontwikkelde veentjes gelimiteerd werden door CO2.
Dit in tegenstelling tot de goed ontwikkelde veentjes, waarin de succesvolle groei van veenmossen
was gecorreleerd aan een hoge CO2 beschikbaarheid. De bevindingen in laten daarom zien dat een
hoge CO2 beschikbaarheid een vereiste is voor een succesvolle vestiging van veenmossen en het
daaropvolgende herstel van een hoogveen. In goed ontwikkelde, onaangetaste hoogvenen is altijd
voldoende CO2 aanwezig door de afbraak van organisch materiaal in een dik veenpakket. In aan getaste
hoogvenen daarentegen, ontbreekt deze organische laag regelmatig als gevolg van vervening en/
of drainage. Een aanvullende CO2-bron is dan nodig voor de succesvolle groei van Sphagnum. Eén
van de conclusies van hoofdstuk 5 is, dat de CO2-beschikbaarheid meegenomen moet worden in
studies naar de haalbaarheid van herstel van hoogvenen.
Opmerkelijk was dat de goed ontwikkelde veentjes in het Dwingelderveld koolstofrijk
grondwater ontvingen van buiten de veentjes, waardoor de CO2 concentratie in de veentjes stijgt en
de groei van de veenmossen en daarmee de ontwikkeling van het hoogveen, wordt gestimuleerd.
In de afhankelijkheid van de groei van de veenmossen van grondwater schuilt een interessante
paradox: veenmossen hebben een competitief voordeel ten opzichte van hogere planten in een
ombrotroof, nutriëntenarm milieu. Grondwater wordt vaak gekarakteriseerd door een hoge pH,
hoge calciumconcentraties en hoge nutriëntenconcentratie. Deze omstandigheden zijn zeer
ongunstig voor veenmossen waardoor ze doorgaans hun bestaansrecht ontlenen aan het feit dat
ze afgesloten zijn van grondwaterinvloeden.
Dankwoord
Dankwoord – 129
De afgelopen vijf jaar heb ik met veel plezier aan dit proefschrift gewerkt. Ik heb mogen werken
aan zeer interessante en intrigerende planten en ecosystemen. De combinatie van veldwerk en
laboratoriumexperimenten en het combineren van de disciplines ecologie en fysiologie maak-
ten het een afwisselend en fascinerend onderzoek. Ik heb ontzettend veel geleerd en ik heb hele
mooie en bijzondere plekken van de wereld gezien. Uiteindelijk heeft het allemaal geleid tot dit
proefschrift. Zeker niet in de laatste plaats hebben de mensen met wie ik de afgelopen jaren heb
samengewerkt daaraan bijgedragen. Sterker nog, zonder al die mensen was dit proefschrift nooit
tot stand gekomen. Iedereen ontzettend bedankt! Een aantal wil ik in het bijzonder noemen.
Allereerst mijn promotoren Theo Elzenga en Ab Grootjans. Jullie hebben mij de mogelijkheid
geboden om promotieonderzoek te gaan doen. Dank jullie wel voor het vertrouwen dat jullie in
mij gesteld hebben. Theo, dank je voor je begeleiding de afgelopen jaren. Op wetenschappelijk
gebied heb ik veel van je geleerd. Ik heb ontzettend veel prijs gesteld op de vrijheid die je mij hebt
gegeven waardoor ik mijn eigen ideeën kon ontwikkelen en uitvoeren. Bij naderende onzekerheid
wist je me altijd wel weer te overtuigen, op te peppen en te motiveren om vervolgens weer met
frisse moed aan de slag te gaan en het overzicht te bewaren. Maar bovenal wil ik je bedanken voor
de prettige samenwerking in de afgelopen jaren. Bovendien ben je de eerste die ik ken die “Fillmore
East - June 1971” van The Mothers ook een geweldige plaat vindt. Ab, ik heb grote bewondering voor
jouw positieve en kritische houding ten opzichte van de wetenschap. Onze samenwerking heeft
mijn kennis vergroot en mijn blik op de wetenschap verruimd. Dank daarvoor.
Ook mijn co-promotor Fons Smolders heeft een belangrijke bijdrage geleverd aan dit proef-
schrift. Ondanks je rol van begeleider op afstand was je altijd zeer betrokken en bereid om mee te
denken over alle facetten van het onderzoek. Ook heb je me altijd voorzien van nuttig commentaar
op mijn manuscripten.
Gert Jan Baaijens, brein achter het fenomeen buoyancy flow, bedankt voor je enthousiasme,
inspirerende ideeën, mooie verhalen en gastvrijheid.
Christian Fritz, mede dankzij jou heb ik een onvergetelijke tijd gehad in Ushuaia en omstreken.
Jacob Hogendorf, ik dank je voor je bewaarzucht en je goede zorgen voor mijn mossen. Ze
staan aan de basis van al mijn experimenten!
Robert, met de afronding van dit proefschrift komt er ook een einde aan het koffiedrinken op
de dinsdagochtend. Ondanks dat deze traditie nog niet zo lang bestaat, ga ik het zeker missen.
De afgelopen jaren heb ik mij op het laboratorium van Plantenfysiologie bijzonder thuis gevoeld.
Zonder iedereen bij naam te noemen wil ik alle labgenoten daarvoor bedanken. Een aantal in het
bijzonder. Marten, bedankt voor al je hulp en expertise tijdens het ontwikkelen en uitvoeren van
mijn experimenten, de oneliners, de overheerlijke koffie, de FC Groningen-kranten en je luis-
terend oor, maar bovenal voor je goeie gezelschap. Jan Henk, ook bij jou heb ik me altijd in goed
gezelschap bevonden. Jouw kritische kijk op de wetenschap (en de verrichtingen van de FC) en je
enthousiasme voor de biologie heb ik altijd zeer gewaardeerd en hebben ongetwijfeld hun weer-
slag gevonden in dit proefschrift.
Ook wil ik mijn mede-aio’s Fatma en Ika hier speciaal noemen. Het was mij een waar genoegen
om samen met jullie in hetzelfde schuitje te zitten. Fatma, ik heb grote bewondering voor je gedre-
venheid en vastberadenheid. En dat in combinatie met een immer goed humeur! Ika, jij was mijn
klankbord, steun en toeverlaat op het lab. En daarbuiten, gezellig biertjes drinken en concerten
130 – Solute transport in Sphagnum dominated bogs
bezoeken met Germaine en Chris. Ik ga er van uit dat we elkaar in de toekomst nog regelmatig gaan
zien!
Tijdens mijn onderzoek heb ik het voorrecht gehad om een aantal studenten te mogen begeleiden
tijdens hun bachelor of masteronderzoek. Arne, Arrie, Bikila, Charlotte, Jan Erik en Myra; jullie
hebben fantastisch werk geleverd en dat heeft dan ook geleid tot een grote bijdrage aan dit proef-
schrift. Ik ben jullie daar zeer dankbaar voor. Daarnaast hoop ik dat ik mijn enthousiasme voor het
onderzoek een beetje aan jullie heb kunnen overdragen. Wat ik wel weet, is dat ik veel plezier heb
gehad aan de samenwerking met jullie.
Steven, jouw rol in dit proefschrift is groter dan je denkt. Ik vind het geweldig dat je straks als
paranimf aan mijn zijde staat.
Mijn zus, Mirjam, bedankt voor je hulp bij de layout van dit proefschrift. Zonder jou was het
me niet gelukt!
Mijn ouders, Harry en Janny, jullie hebben altijd met mij meegeleefd en onvoorwaardelijk
gesteund op alle fronten en daar ben ik jullie ontzettend dankbaar voor.
Lieve Silke en lieve Veerle, geweldig dat jullie er zijn! Jullie hebben mij de afgelopen jaren met
beide benen op de grond gehouden; het schrijven van een proefschrift is slechts bijzaak.
Lieve Germaine, de afgelopen jaren hebben voor een groot deel in het teken gestaan van dit
proefschrift. Nu is het af. Tijd voor iets nieuws! Dank je voor je nimmer aflatende vertrouwen en je
liefdevolle steun.
Wouter
Wou
ter Patberg Solute tran
sport in
Sphagnum
dom
inated
bogs T
he ecop
hysiological eff
ects of mixin
g by convective fl
ow
Wouter Patberg
Solute transport in Sphagnum dominated bogsThe ecophysiological effects of
mixing by convective flow