26 Feather mosses, nitrogen fixation and the boreal biome

> T. H. DeLuca School of the Environment and Natural Resources, Bangor University, Bangor, UK t.h.deluca@bangor.ac.uk

Introduction

Nitrogen (N) is the primary limiting nutrient for both productivity and decomposition in boreal forests (Tamm, 1991) yet we lack a clear understanding of the influence of N inputs from nitrogen fixation on forest ecosystem dynamics, productivity and carbon (C) cycling. Although our atmosphere is 78% N2 gas, none of that N is available directly to land plants or animals; it must first be converted to ammoniacal (NH3) N by nitrogen-fixing microbes, often acting in symbiosis with plant species, and then further transformed to useful amino acids in plants or bacteria. As there is a general lack of woody or herbaceous N-fixing plant-microbe associations in upland boreal forests, the primary source of useful N supply in these ecosystems has until recently remained a mystery.

There is currently a great deal of interest in boreal forest ecosystems and the vital role they play in global N and C balance. The boreal ecosystem is the second largest biome in the world and one of the largest terrestrial C sinks on Earth, storing approximately 30% of total terrestrial carbon stocks. Feather mosses are an ever-present component of the boreal forest floor (see Figs 1, 2), accounting for as much as 95% of ground cover, and a major contributor to the annual productivity of the boreal biome. While the ecology and ecosystems of these bryophytes within the boreal forest have been widely described by numerous researchers, the contribution of bryophytes and, in particular, feather mosses, to global ecosystem models often goes unobserved.

The feather moss ‘bottom layer’ is a vital organ of the boreal forest, functioning to filter water, insulate against temperature change, regulate nutrient uptake and minimise nutrient losses. It provides the breathing, protective skin of the forest floor (Lindo and Gonzeles, 2010). Potentially

formed from both feather mosses and sphagnum mosses, this dense insulting layer controls both heat flux from incoming radiation and losses from the largely mineral boreal soils (Beringer et al., 2001), thereby directly controlling soil temperatures and indirectly regulating surface processes such as carbon cycling.

Over the past 10 years, we have investigated the notion that feather mosses also serve as hosts for N-fixing cyanobacteria, aiming to expand our knowledge of their beneficial properties. Our results show that, for the biota

Feather mosses, nitrogen fixation and the boreal biome

Figure 1. A late succession Scots pine, Norway spruce forest in northern Sweden. Feather mosses make up the light green carpet on the forest floor.

Figure 2. A close up view of Pleurozium schreberi, pic: © Uwe Drehwald

27 Feather mosses, nitrogen fixation

and the boreal biome

of the relatively pristine (low N deposition) environments of northern Scandinavia, feather mosses are a dominant source of N (DeLuca et al., 2002b; Zackrisson et al., 2004; DeLuca et al., 2007; Zackrisson et al., 2009), thus providing a first step towards a holistic understanding of the role which natural N fixation plays in these high latitude boreal forests. This research has also provided an insight into the means by which N subsequently accumulates in such pristine forest ecosystems and the following paragraphs describe some of the history of our studies and raise some future questions that have yet to be addressed.

Feather moss carpets

Upland moss communities of the boreal forest contain hundreds of species of bryophytes but are dominated by two pleurocarpous species (in which the female archegonium reproductive structures are on a short side branch rather than on the main stalk): Pleurozium schreberi and Hylocomium splendens, and one acrocarpous species (in which the archegonia are at the top of the stem), Polytrichum commune. The photosynthetic capacity of these feather mosses is reflected in an ecosystem productivity that rivals, and at times (during early to mid-succession) can surpass, that of the overstorey (Bond-Lamberty et al., 2007). Hylocomium and Pleurozium often account for 30-95% of the average cover of the boreal forest floor and yield a net primary productivity (NPP) of 200 to >400 kg ha-1 yr-1, comprising a total biomass of between 0.1–2.0 Mg C ha-1

as live moss and 2.0–4.0 Mg C ha-1 as dead moss (Vogel and Gower, 1998; Turetsky, 2003). Their photosynthetic contribution to C fixation capacity has been found to be of particular importance when the overstorey is under stress, as, for example, in the large-scale bark beetle outbreak that recently raged across the Canadian boreal forest, where bryophyte productivity remained relatively consistent with the opening of the forest canopy in the absence of fire (Olsson and Staff, 1995). With the dip in photosynthetic C fixation by trees and the net C emissions associated with dead tree decay, the carbon assimilation associated with the moss bottom layer becomes much more important for ecosystem maintenance. The carbon fixed by the mosses is slowly incorporated into the fibric materials that comprise the dense humus and debris mat often associated with boreal Spodosol soils (Beringer et al., 2001). Although feather mosses have long been known to intercept and retain nutrients from forest throughfall, there has previously been little understanding of the role they play

in creating bioavailable N in the boreal forest. As stated previously, the conspicuous lack of any significant quantity of herbaceous or woody N-fixing plant-microbe associations across much of the boreal forest had long puzzled forest ecologists, who therefore questioned the source of available N supply in fire-maintained or harvested boreal forest ecosystems (Tamm, 1991).

Repeated efforts to measure N fixation associated with feather mosses have failed because most researchers would take their samples from high latitude ecosystems in late June to mid-July, a time of uniquely low N fixation activity in forest feather moss ecosystems (DeLuca et al., 2002b; Zackrisson et al., 2004; Zackrisson et al., 2009). Also, the spatial variation of N fixation in feather mosses obligates an intensive sampling strategy, which was not the norm in surveys of bryophyte N fixation. Our work has therefore opened up the possibility that feather mosses are actually responsible for a great deal of the N accumulation resident in boreal forests and has underscored the importance of feather mosses in the total N economy of the boreal biome (DeLuca et al., 2002b; Zackrisson et al., 2004).

We have found cyanobacteria (see Fig. 3) residing in the leaf incurve of some of the pleurocarpous moss species, leaving them both difficult to observe and difficult to culture. Nevertheless, we have commonly retrieved three genera of cyanobacteria (Nostoc, Stigonema and Calothryx) associated with Pleurozium schreberi alone (Gentili et al., 2005). These cyanobacteria demonstrate several unique properties, including having biphasic temperature optima for productivity at 13oC and 22oC, which makes them ideally suited to N fixation in boreal ecosystems. The feather moss carpets are thus a uniquely convenient system for studying N fixation, as either individual shoots or whole moss carpets can be placed in a vessel for manipulation or measurement of fixation activity and this has allowed us to conduct intensive monitoring.

Distribution of N fixation in Pleurozium schreberi in Fennoscandia

Over the past 10 years, we have established sampling plots for Pleurozium schreberi and Hylocomium splendans at about 50 sites across northern Sweden, Finland and Norway (Fig.4). The majority of samples have been collected from natural forest preserves to minimise site disturbance and all sites occur within the mid- to northern boreal zone of Fennoscandia (59o–69oN; 17o–19oE). When samples from

28 Feather mosses, nitrogen fixation and the boreal biome

Figure 4. A stylized view of forest succession and N cycling in Scots pine forests of northern Sweden.

Figure 3. The cyanobacteria, Stigonema sp., taken from the leaf incurve of Pleurozium schreberi.

these plots were assessed for N fixation activity using the acetylene reduction technique, there was a clear relationship between the latitude and longitude of the sample collected and N fixation rate, wherein fixation rates were greater in the north, away from the Botinian Bay section of the Baltic Sea, likely as a result of the high rates of N deposition along this more populated coast. In comparison, the boreal forests of northern Sweden exhibit exceptionally low rates of N deposition, both compared to southern Sweden and to other more industrial regions of the world (Gundale et al., 2011). Indeed, nitrogen deposition in Sweden ranges from about 20 kg N ha-1 yr-1 in southern Sweden to less than 1.0 kg ha-1 yr-1 in Northern Sweden where most of our studies have been performed. Nitrogen deposition can interfere greatly with N fixation in cyanobacteria and this may partially account for the geographic variation in our results.

Successional influence on N-fixation rates

Fire is the primary form of chronic disturbance in the boreal forest ecosystems of northern Europe and is a potent driver of shifts in nutrient cycling and plant community composition. The recovery of organic N resources lost during severe wildfire is dependent upon biological N fixation (Smithwick et al., 2005) and, in the boreal forest, this is primarily accomplished by the feather moss-cyanobacterial associations (Fig. 5). We have found that feather mosses recover to pre-fire densities ca 30- 60 years after fire and

Figure 5. Nitrogenase activity in Pleurozium schreberi and Hylocomium splendans along a latitudinal gradient (59o N to 69o N) in Sweden (from Zackrisson et al., 2009).

?

Boreal Forest Secondary Succession and N Availablity

N 0?

2

Early

Mid

Late

Time since �re

Throughfall N

Moss N �xation Re

lativ

e ra

te

Re

lativ

e so

il co

ncen

trat

ion

NO 3 NH 4

R-NH 2

Fire

Rain / throughfallNH , NO

NO 4 3+ -

NH , NO

4 3+ -

Nitrogen �xation

Moss

NH , R-NH

4 2+ -

Shrubs N 0

CH

2

4?N

R-NH 2

2

!

Site and latitude (name and degrees north)Ty 5

9Br 5

9Bj 59

So 59Fi 59

Va 60In 60

Gr 60

Ab 60Ed 60

Lo 60Ho 61

Son 61Vat 61

Ta 62Lon 63

Ro 64Ku 64

Vi 64Mo 64

Ki 64Vi 65

De 66Ja 65

Ru 65Gu 65

Tj 65Ku 65

Nu 65Su 66

Ai1 66Ai2 66

Ai3 66Aiv3 66

Aiv1 67

Aiv2 67Ad 67

Ru 67Na 67

An 69

Acet

ylen

e re

duct

ion

(μm

ol m

-2 d

-1)

0

100

200

300

400

500

PleuroziumHylocomium

!Site and latitude (name and degrees north)Ty 5

9Br 5

9Bj 59

So 59Fi 59

Va 60In 60

Gr 60

Ab 60Ed 60

Lo 60Ho 61

Son 61Vat 61

Ta 62Lon 63

Ro 64Ku 64

Vi 64Mo 64

Ki 64Vi 65

De 66Ja 65

Ru 65Gu 65

Tj 65Ku 65

Nu 65Su 66

Ai1 66Ai2 66

Ai3 66Aiv3 66

Aiv1 67

Aiv2 67Ad 67

Ru 67Na 67

An 69

Acet

ylen

e re

duct

ion

(μm

ol m

-2 d

-1)

0

100

200

300

400

500

PleuroziumHylocomium

!

Acet

ylen

e re

duct

ion

(µm

ol m

-2-1

)d

Site and latitude (name and degrees north)

Pleurozium

Hylocomium

400

300

200

100

0

100

0

120

80

60

40

20

140

29 Feather mosses, nitrogen fixation

and the boreal biome

that N fixation rates increase linearly with time during the recovery period (DeLuca et al., 2002a; Zackrisson et al. 2004). The rates of N fixation measured in fire-damaged locations were scaled up to ecosystem level by factoring in the percentage ground cover of P. schreberi (Zackrisson et al., 2004) and using an experimental ratio of 3 moles of acetylene reduced to correspond to 1 mole of N2 fixed in the field (15N calibrated value, DeLuca et al., 2002b). Applying these assumptions, we estimated N-fixation rates to be less than 0.5 kg ha-1 yr-1 in early succession sites (aged 25–80 years since fire), between 0.4 and 1.6 kg ha-1 yr-1 on mid-succession sites (100 – 200 years since fire) and between 1.0 and 3.0 kg ha-1 yr-1 in late-succession ecosystems (>200 years since fire).

Early succession forests exhibit relatively high levels of N availability as a product of recent fire; this N being ultimately recycled through the canopy and resulting in higher rates of N deposition via throughfall (DeLuca et al., 2008). In this throughfall, water percolates through the canopy, picks up nutrients and then delivers them to the moss bottom layer. If the N content of the throughfall is high (as in early succession), N fixation in feather mosses is reduced (DeLuca et al., 2008).

Nitrogen deposition

The influence of N deposition on N fixation rates has been studied at several locales in northern Sweden. A fertilisation study at a late successional forest site (Ruttjeheden, 355 years since fire) where N fixation rates had been observed to be relatively high (Zackrisson et al., 2004) demonstrated a very clear elimination of N fixation with N applications as low as 4.25 kg N ha-1 yr-1. In a long-term N enrichment study near Vindlen, we observed the influence of 5 years application of 0, 3, 6, 12 or 50 kg N ha-1 yr-1 as ammonium nitrate, NH4NO3 (Gundale et al. 2011). Isotopic analyses associated with this study suggested that feather mosses absorbed much of the N supplied at low applications rates, with accumulation of fertiliser N in the dwarf shrub layer occurring only at the highest levels of application. These studies support the notion that N deposition directly down-regulates the N fixation associated with feather mosses (Huttunen et al., 1981; Zackrisson et al., 2004; Gundale et al., 2011).

Figure 6. Nitrogenase activity in feather mosses and throughfall N deposition (as observed in resin lysimeters) averaged across three natural fertility gradients in Northern Sweden (DeLuca et al., unpublished data).

Numerous natural fertility gradients exist in northern Sweden where groundwater recharge zones deliver nutrients and moisture to the soil surface. This results in the creation of a completely unique forest composition where groundwater discharge zones are dominated by extremely large spruce and birch trees, with an understory dominated by tall herbaceous species, in contrast to the modest sized pines and dwarf shrubs of normal upland forests. Nitrogen fixation rates in the tall herbaceous ecosystems within discharge zones are greatly reduced in line with the far greater levels of N in the throughfall (Fig. 6). In contrast, N fixation rates were maximised in lower fertility dwarf shrub stands and at intermediate levels in the low fertility lichen heath stands.

Heath Ericoid Herb

Acet

ylen

e re

duct

ion

(µm

ol m

-2 y

r-1)

0

100

200

300

400

Heath Ericoid Herb

Inor

gani

c N

dep

ositi

on (k

g N

ha

1 yr

)

0

2

4

6

8

10-

1-

30 Feather mosses, nitrogen fixation and the boreal biome

Summary and Ongoing StudiesOngoing and proposed studies are designed to test the fate of the N fixed in feather moss carpets and to assess the role of organic N uptake on the overall N balance of feather moss ecosystems. More detailed investigations are being undertaken on the interaction of a range of external N sources on N fixation rates, including studies on the influence of roadside pollution on feather moss productivity and ecology. Mosses near busy highways have been observed to have few cyanobacterial associates and suppressed nitrogenase activity, whereas mosses located 100 m or more from the highway express normal rates of N fixation. These studies suggest that feather moss N fixation could serve as a sensitive indicator of N pollution as the nitrous oxides emitted from vehicles apparently inhibit both the presence and activity of cyanobacteria.

In summary, boreal feather mosses clearly play an important role in the ecological function of boreal forests. They function to filter forest nutrient throughfall, to insulate the largely mineral-based soils, and to perform the historically important process of adding plant-available N to the ecosystem. The N provided by feather mosses remains crucial to the function of pristine boreal forest ecosystems and, given their broad ecological amplitude, Pleurozium schreberi and Hylocomium splendens, along with their cyanobacterial associates, may be the most broadly distributed N-fixing association on Earth.

References

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Bond-Lamberty, B., Peckham, S. D., Ahl, D. E. and Gower, S. T. (2007). Fire as the dominant driver of central Canadian boreal forest carbon balance. Nature 450, 89-92.

DeLuca, T. H., Nilsson, M.-C. and Zackrisson, O. (2002a). Nitrogen mineralization and phenol accumulation along a fire chronosequence in northern Sweden. Oecologia 133, 206-214.

DeLuca, T. H., Zackrisson, O., Gentili, F., Sellstedt, A. and Nilsson, M.-C. (2007). Ecosystem controls on nitrogen fixation in boreal feather moss communities. Oecologia 152, 121-130.

DeLuca, T. H., Zackrisson, O., Nilsson, M.-C. and Gundale, M. J. (2008). Ecosystem feedbacks and nitrogen fixation in boreal forests. Science 320, 1181.

DeLuca, T. H., Zackrisson, O., Nilsson, M.-C. and Sellstedt, A. (2002b). Quantifying nitrogen-fixation in feather moss carpets of boreal forests. Nature 419, 917-920.

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Gundale, M. J., DeLuca, T. H. and Nordin, A. (2011). Bryophytes attenuate anthropogenic nitrogen inputs in boreal forests. Global Change Biology 17, 2743–2753.

Huttunen, S., Karhu, M. and Kallio, S. (1981). The effect of air pollution on transplanted mosses. Silva Fennica 15, 495-504.

Lindo, Z. and Gonzeles, A. (2010). The bryosphere: an integral and influential component of the Earth’s biosphere. Ecosystems 13, 612-627.

Olsson, B. A. and Staff, H. (1995). Influence of harvesting intensity of logging residues on ground vegetation in coniferous forests. Journal of Applied Ecology 32, 640-652.

Smithwick, E. A., Turner, H. M., Mack, M. C. and Chapin, F. S. (2005). Post fire soil N cycling in northern conifer forests affected by severe, stand replacing wildfires. Ecosystems 8, 163-181.

Tamm, C. O. (1991). Nitrogen in terrestrial ecosystems. Springer, Berlin.

Turetsky, M. R. (2003). The role of bryophytes in carbon and nitrogen cycling. The Bryologist 106, 395-409.

Vogel, J. C. and Gower, S. T. (1998). Carbon and nitrogen dynamcis of boreal jack pine stands with and without a green alder understory. Ecosystems 1, 386-400.

Zackrisson, O., DeLuca, T. H., Gentili, F., Sellstedt, A. and Jäderlund, A. (2009). Nitrogen fixation in mixed Hylocomium splendens moss communities Oecologia 160, 309-319.

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31 Feather mosses, nitrogen fixation

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