Long-term effects of nitrogen deposition on epiphytic lichens
Marie Rönnqvist
Student
Degree Thesis in Ecology 30 ECTS Master’s Level
Report passed: 07-06-2013 Supervisor: Kristin Palmqvist
Long-term effects of nitrogen deposition on epiphytic lichens
Marie RönnqvistAbstract
The main aim of this study was to analyse the long-term effects on epiphytic lichen dry mass development, diversity and community structure after exposure to a simulated nitrogen deposition gradient. A whole tree experiment was set up in a boreal forest in the County of Västerbotten in Sweden, whereby 15 trees were artificially irrigated and nitrogen fertilized during seven consecutive years (2006-2012). The treatments were equal to an additional deposition of 0.6, 6, 12.5, 25 and 50 kg N ha-1 yr-1. Branches from the trees were collected in October 2012 and the lichen material was harvested and further analysed during the spring 2013. The results from this study showed that the lichens were directly affected by the long- term increased nitrogen deposition. Generally, lichen dry mass and species richness declined at high nitrogen loads and the initial positive effects of low nitrogen loads reported in a preceding study had thus depressed with time. The results from this study also showed that the composition of the lichen community changed benefitting nitrogen-tolerant species, probably not because of competition but more likely because of nitrogen-sensitive species disappearance. This study strongly indicates that it takes more than a few years to detect changes in lichen communities exposed to enhanced nitrogen loads. In addition low concentrations seem to have a cumulative impact. Consequently, this study stresses the importance of also considering the cumulative effect of low nitrogen loads when determining critical values. The critical load for lichen communities in boreal forests might be below 6 kg N ha-1 yr-1.
Keywords: simulated nitrogen deposition, boreal forest, epiphytic lichens, cumulative loads
Table of Contents
1 Introduction
... 11.1 Aim
... 11.2 Background
... 21.2.1 Nitrogen ... 2
1.2.2 Lichens ... 2
1.2.3 Nitrogen deposition effects on lichens ... 3
2 Materials and Methods
... 42.1 Field site and irrigation-fertilization
... 42.2 Lichen material
... 42.3 Weight and surface area measurements
... 52.4 Image analysis of lichen abundance
... 52.5 Statistical analyses
... 63 Results
... 73.1 Lichen dry mass
... 73.2 Species richness
... 93.3 Specific thallus mass
... 93.4 Relationships between lichen abundance and dry mass
... 114 Discussion
... 144.1 Long-term effects of nitrogen fertilization
... 144.2 Conversion of abundance to dry mass
... 164.3 Conclusions
... 165 Acknowledgments
... 176 References
... 181
1 Introduction
Over the last 150 years there has been a significant alteration of the nitrogen (N) cycle at global, regional and local scales (Galloway et al. 2004). Anthropogenic activities have increased reactive nitrogen (Nr) heavily; the average N deposition rates exceed 10 kg N ha-1 yr-1 in large parts of the world, which is more than an order of magnitude higher compared to natural input of Nr (Galloway et al. 2008). It is also predicted that the N deposition may double and some regions might even receive as much as 50 kg N ha-1 yr-1 (Galloway et al.
2004). N deposition has been a key driver of local plant extinctions along with land use and precipitation changes (McLean et al. 2011). Evidently, increased inputs of Nr have accelerated the losses of biological diversity and are considered to be one of the major threats to biodiversity (Vitousek et al. 1997, Sala et al. 2000). Nitrogen-limited biomes such as temperate and boreal forests are considered to be particularly sensitive to N deposition (Sala et al. 2000, Bobbink et al. 2010). Moreover, in temperate forests lichens are known to be among the most N-sensitive organisms and the critical load for lichen community effects has in many regions long been exceeded (Bobbink 2010, Geiser et al. 2010). The critical load concept has been brought up to establish deposition levels that can be tolerated by ecosystems without any harmful effects on sensitive elements of the environment (Nilsson and Grennfelt 1988).
Lichens play a significant role in ecosystems; they provide hiding places for invertebrates, birds uses them as nesting material, they represent a significant winter fodder for reindeers, and lichens are also biological weathering agents in the development of soils (Petterson et al.
1995, Gauslaa et al. 2008, Seaward 2008). It has been difficult to separate the direct effects of N on lichens from other factors (N deposition is often correlated with other pollutants);
and studies examining long-term effects of N deposition on lichens are rare even if recent studies suggest that cumulative loads of N might be an important factor (Van Dobben et al.
2001, Johansson et al. 2011. De Schrijver et al. 2011). However, in 2012 Johansson et al.
presented results from a long-term study of an increased N deposition to naturally established lichens in a boreal forest with low background N. They found that four consecutive years of a simulated N deposition gradient in 2006-2009 caused an alteration of the epiphytic lichen community and that N is a significant pollutant for many lichen species.
Moreover, they found that responses to an increased N load differ between lichen species and that the responses changed over time. The simulated N deposition continued for three more additional years after 2009 and was subsequently halted in October 2012.
1.1 Aim
This study is an extension of the results presented in Johansson et al. (2012) after the first four years of simulated N deposition, now presenting data from all the seven years. The results from the N fertilization experiment have so far been based on abundance data estimated using non-destructive image analysis to enable monitoring of the same twigs every year. However, lichen material collection became possible when the N fertilization experiment was halted in October 2012. My first aim was to analyse the long-term effects on epiphytic lichen dry mass, diversity and community structure after exposure to the simulated N deposition gradient for seven years. My second aim was to analyse how the morphology of the epiphytic lichen species respond when exposed to a long-term increased N load. Finally, my third aim was to investigate the possibility to convert the earlier abundance data collected by Johansson et al. (2012) to dry mass data in order to estimate how the dry masses of the different lichen species have developed over time.
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1.2 Background
1.2.1 Nitrogen
Nitrogen is a component of many biologically important molecules such as DNA, proteins and enzymes; evidently it is essential for the survival of all living things. Most of the nitrogen on the planet is however unavailable to most organisms since its most stable form is nitrogen gas (N2) which accounts for approximately 78% of the atmosphere (Emsley 2011). N2 needs to be converted into other forms to become accessible, the term reactive nitrogen (Nr) includes all biologically active, photochemically reactive and radiatively active N compounds in the biosphere and atmosphere (Galloway et al. 2008). Nr that is not the form of organic compounds (e.g. proteins, urea) is either inorganic reduced forms (e.g. NH3, NH4) or inorganic oxidized forms (e.g. NOX, NO3, N2O, NO3) (Socolow 1999).
Nr creation occurs in several ecosystems through biological nitrogen fixation (BNF), lightning, and thermal fixation. The BNF process implicates that organisms, such as cyanobacteria, can fix N2 into organic forms; thermal fixation happens when the air is heated to extreme temperatures (Follett and Hatfield 2001). However, anthropogenic activities have dramatically increased Nr input by food and energy production since the middle of the 19th century (Galloway et al. 2008). Nonreactive N2 is converted to reactive NH3 to sustain food production (fertilization of soils); also, crops (e.g. legumes, rice in association with nitrogen fixing organisms) that promote conversion of N2 to organic N through BNF are very common (Galloway et al. 2003). Energy production, essentially combustion of fossil fuels, converts both fossil N and atmospheric N2 (thermal fixation) to reactive NOX (Galloway et al. 2003).
Nr can be emitted to the atmosphere as NOX, NH3 or organic N, which later carries by winds, precipitates and forms N depositions (Neff et al. 2002).
As previously mentioned the N deposition has increased and further increases are predicted, some regions might even receive as much as 50 kg N ha-1 yr-1 (Galloway et al. 2004). In Sweden the total N deposition is approximately 1 kg N ha-1 yr-1 in the northern parts and up to 25 times higher in the southwest parts (Pihl-Karsson et al. 2003). The productivity of many ecosystems is limited by N supply, especially in temperate and boreal regions (Vitousek et al.
2002). Therefore, the alteration of N deposition (addition of a limiting nutrient) can change plant growth, which species are dominant and also decrease the ecosystems biodiversity at the landscape level and reduce the species richness within communities (Vitousek et al.
1997).
1.2.2 Lichens
Lichens are symbiotic organisms (herein denoted as species) composed of one fungal partner and one or more photosynthetic partners; the fungal partner is referred to as the mycobiont and the photosynthetic partner (green algae and/or cyanobacteria) is referred to as the photobiont (Nash III 2008 a). Fungi depend on an external supply of fixed carbon and lichenization is a common nutritional strategy, particularly among the ascomycetes (~20% of all known fungal species are lichenized), hence the photobiont provides fixed carbon (carbohydrates) originating from photosynthesis (Honegger 1998). The photobiont partner receives in return water and nutrients from the mycobiont, which also positions the photobiont cells at optimal places within the lichen thallus to facilitate gas exchange and illumination (Honegger 1991). Lichen photobionts can be found free-living, very abundant even, whilst some are rarely found outside lichen thalli (Honegger 1998). It is generally considered that lichens are mutualistic symbioses, however the relationship might be more complex, for instance if the mycobiont obtain most of the benefits and the photobiont grows more slowly than when free-living the symbioses is rather parasitic (Ahmadjan 1993, Nash III 2008 a). Even so, the lichen symbiosis is very successful occupying almost all terrestrial habitats and surviving extreme conditions such as cold, heat and drought; the number of species might be as high as 17 000 (Hale 1974).
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Most lichens are terrestrial and colonize many different substrates, they occur frequently as epiliths growing on the surface of rocks and they also occur commonly as epiphytes on living trees and shrubs (Smith 1982, Nash III 2008 a). Based on their morphology lichens are segregated into the major groups: crustose, foliose and fruticose. Crustose lichens (e.g.
Mycoblastus sanguinarius) are tightly attached to the substrate and are practically
inseparable from it; foliose lichens (e.g. Platismatia glauca) are leaf like and adhere partially to the substrate; frutiose lichens (e.g. Usnea dasypoga) are bushy or threadlike and attached to the substrate by a holdfast or are pendulous (McCune and Geiser 2009). Lichen growth may primarily be limited by water availability, whilst wet and metabolically active the growth is rather limited by light because of the photosynthesis dependence on light (Palmqvist et al.
2008). However, lichens may also be limited by N with increasing photobiont content increasing photosynthesis (Palmqvist et al. 2002) and growth (Palmqvist & Dahlman 2006).
Growth of lichens can respond to seasonal changes in light and climate, although area and weight do not always increase in parallel causing changes in specific thallus mass (STM) (Larsson et al. 2012). The dry mass gain of lichens responds mainly to its photosynthetic activity; photosynthesis is also important for the lichen area gain however, the area expansion also depends on water availability (Gauslaa 2009). Generally, accumulation of lichen dry mass is a slow process (McCune 1993).
In addition to carbon lichens also require the same nutrients as plants (nitrogen, phosphorous, calcium, magnesium etc.), though they have no roots and therefore the nutrients are generally acquired passively through wet or dry deposition (Nash III 2008 b).
Cyanobacterial photobionts can fix N2 thereby providing both reduced carbon and nitrogen to the mycobiont (Rai 2002). Lichens have developed efficient mechanisms for taking up water and nutrients from atmospheric deposition, this and the fact that most species live for a very long time, may explain why lichens have been recognised as being very sensitive to air pollution (Hawksworth 1971, Nash III b 2008). Most important are the toxic pollutants SO2 and NO2, increased concentrations of these strongly negatively affect species diversity (Van Dobben et al. 2001).
1.2.3 Nitrogen deposition effects on lichens
Different lichen species respond differently to an enhanced nitrogen deposition and more N- tolerant lichen species (nitrophytic) seems to replace the more N-sensitive ones (acidophytic) (Geiser et al. 2010, Johansson et al. 2011, Evju and Bruteig 2013). The difference in response between lichen species might depend on their ability to (1) coordinate growth between the photobiont and the mycobiont (Gaio-Olivera et al. 2005, Palmqvist and Dahlman 2006, Johansson et al. 2012), (2) cope with concomitant increases in bark pH (Van Dobben and Ter Braak 1998), (3) resist parasitic attacks (Strengbom et al. 2002, Johansson et al. 2012).
Lichens might also lack mechanisms for avoiding excessive N uptake leading to a build-up of N within the thallus and consequently to toxicity (Dahlman et al. 2003, Gaio-Oliveira et al.
2005). Increased nitrogen deposition can also stimulate the growth of free living algae on the thallus surface and lead to shading of and a reduction of photobiont photosynthesis (Scott 1960). Changes in species interaction strength is generally considered to be one of the major reasons explaining changes in plant community structure due to nutrient addition, however less is known about competitive exclusion in lichen communities (Levine et al. 1998, Strengbom 2002). Differences in lichens N optima and responses to N availability together with raised nutrient levels can affect lichen growth rates and competition between lichen species, which can lead to alterations in lichen community structure (Palmqvist et al. 2002, Welch et al. 2006).
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2 Materials and Methods
2.1 Field site and irrigation-fertilization
During the years 2004 and 2005 an experiment was set up in an old growth forest stand at Kulbäcksliden (64°11’N, 19°35’E), a part of Vindelns Experimental Forests managed by The Unit for Field-based Forest research (Swedish University of Agricultural Sciences). The experimental set up, in detail described in Johansson et al. (2010), consists of 16 old-growth spruce (Picea abies) trees with a wooden tower built around them (Figure 1). The towers were connected to an automated irrigation-fertilization system and the trees have been artificially irrigated and nitrogen fertilized from 2006 to 2012. One of the trees did not receive any irrigation or fertilization, thus constituted a dry control (DC). The other 15 trees were divided into three groups based on lichen abundance (high, medium or low), and five N- concentrations were assigned randomly to the five trees in each group. A total of eight branches per tree were marked and photo documented annually at the end of each treatment period (Johansson et al. 2012).
The fertilization consisted of nitrogen in the form of NH4NO3 at five concentrations: 0.04, 0.41, 0.81, 1.63 and 3.2 mM. The lowest concentration approximately equals the concentration of background deposition (Forsum et al. 2006) in the area to avoid dilution of N for irrigated control (IC) trees. The treatments were equivalent to a deposition of 0.6, 6, 12.5, 25 and 50 kg N ha-1 yr-1, denoted as IC, 6N, 12.5N, 25N and 50N adding 1800 liters of the respective fertilizer concentration to each tree each year, distributed over the 16 m2ground area of each tower thus corresponding to 112 mm extra “precipitation”.
Background precipitation in the area equals ca 500 mm per year (SMHI 2013). The treatment period lasted for four months each year from 2006 to 2012 and coincided with the active growth period of the forest vegetation, approximately early June to late September.
(a) (b)
Figure 1. Location of the old growth experimental forest stand at Kulbäcksliden (64°11’N, 19°35’E) in the County of Västerbotten in Sweden (a). Photo (by Kristin Palmqvist) of the experimental set up showing four of the 16 treatment trees with wooden towers (b).
2.2 Lichen material
Branches from the 16 trees included in the former study described in section 2.1 were collected in mid-October 2012. Also, branches from two trees randomly assigned to constitute additional dry controls (DC) were collected at the same occasion. Hence, 18 trees in three groups with six treatments (DC, IC, 6N, 12.5N, 25N and 50N) were included in this study creating a block design experiment with three trees acting as replicate units. To cover within-tree variation, four branches per tree were collected. These were selected to be as
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similar to the previously marked branches as possible and collected at two heights (2 and 4 meters from the ground); two branches per height. The branches (72 in total) were then stored in individual plastic bags in a freezer (-20°C) until thawed at room temperature four months later. Each collected branch was cut into a 40 cm long and 10 cm wide section and the middle of the section was marked with a plastic stip. The branches were sprayed with water for full expansion of the lichen thalli (they swell when hydrated) and then photographed from above (one photo per 20 cm twig length) with a scale grid as background (Figure 2). The photographs were taken in the laboratory with a Canon Power Shot Pro1 digital camera while the branches were illuminated with two fluorescent lamps to minimize differences in colour and appearance due to light quality. Further on, the lichen material on each section was harvested and pooled per lichen taxa. The branches harboured a total of 11 lichen taxa. Six of these were identified to the species level: Alectoria sarmentosa (Ach.) Ach., Mycoblastus sanguinarius (L.) Norman, Parmelia sulcata Taylor, Platismatia glauca (L.) W.L.Culb. & C.F.Culb., Tuckermannopsis chlorophylla (Willd.) Hale and Vulpicida pinastri (Scop.) J.-E.Mattsson & M.J.Lai. The rest of the specimens were identified to the genus level: Bryoria Brodo & D.Hawksw., Hypogymnia (Nyl.) Nyl., Parmeliopsis Nyl. and Usnea Dill. ex Adans, except for Crustose lichens (only identified to group level). Species richness was recorded as the number of species observed per branch.
Figure 2. Example of photos of a branch with a plastic strip marking the middle of the branch (one photo per 20 cm twig length) and a scale grid as background. The branch is from one of the IC trees harvested in 2012.
2.3 Weight and surface area measurements
After harvest the lichens were dried at room temperature (19.5-21.5°C; 26-37% RH) for approximately 15 hours and then weighted to the nearest 0.001 gram, recording their dry mass. However, three of the lichen taxa (M. sanguinarius, Parmeliopsis spp. and Crustose lichens) were not weighed because it was too difficult to separate them from the bark. For Hypogymnia spp., P. sulcata, P. glauca and T. chlorophylla one lichen tallus per species was first scanned to obtain their wet area (A) and then the individual dry masses (DM) were recorded after the drying. The scanned images were edited for noise reduction in Adobe Photoshop CS4 Extended ver. 11.0 (Adobe Systems Incorporated, USA) and the surface area was measured in ImageJ ver. 1.44 (National Institutes of Health, USA), using the scale grid in each image to enable scale calibration. Based on the surface area and individual dry mass measurements, the Specific Thallus Mass (STM) was computed according to Equation 1.
STM = DM/A (1)
STM was not determined for the frutiose lichens (A. sarmentosa, Bryoria spp. and Usnea spp.) because the method for obtaining the surface area was not applicable.
2.4 Image analysis of lichen abundance
Photos of the branches described in section 2.2 were analysed using ArcGIS 10.1 ESRI (Environmental Systems Resource Institute, USA). The scale in the photos was set using the scale grid background and a 0.5 × 0.5 cm2 grid was superimposed on top of each photo. Each lichen thallus larger than approximately 2 mm under each grid interception point was recorded and manually identified. The total area analysed per each branch was equal to the
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beforehand trimmed section of 10 × 40 cm2. When recording the abundance Bryoria spp, Hypogymnia spp and Usnea spp were only identified to the genus level. The abundance data (cm2 coverage m-1) was computed as the number of hits per branch intercept multiplied by 0.25 cm2 per hit and 200 intercepts per meter. The abundance analysis was made for two branches per tree from the DC, IC and 50N treatment trees and not for the whole material due to time constraints.
2.5 Statistical analyses
As previously indicated four branches per tree was collected to cover within-tree variation, though data for all samples of each species from each tree were pooled.
Differences in dry mass between treatments and species were compared by using a two-way ANOVA, the treatment units (tree) was added as an error term. When comparing differences in dry mass between treatments for each species separately, one-way ANOVA was used.
Distributions of the measured dry masses were not normal, hence logarithmic transformation was performed. The distribution of the data for species richness was fairly normal therefore differences between treatments were compared using a one-way ANOVA. Differences in Specific Thallus Mass (STM) between treatments and species were analysed using a two-way ANOVA. When the effect of treatment on STM was analysed separately for each species a one-way ANOVA was used. The STM data was normally distributed.
To be able to convert estimated lichen abundance obtained from the image analysis to lichen dry mass the relationship between dry mass and lichen abundance was examined. The lichen dry mass data from the branches used in the image analysis were fitted to the lichen abundance from the same analysis by a linear regression. However, the regression plot indicated a non-linear relationship and therefore, the dry weight was square root transformed and the linear relationship became even stronger (presented in the results). The best model was identified using the Akaike Information Criterion (AIC). To determine whether the inclusion of species and treatment gave a better fit between dry mass and abundance a two-way ANOVA was used. Finally, according to Equation 2 the estimated lichen abundance from year 2005-2009 and 2012 was converted into lichen dry mass.
y=(0.011x+0.16)2 (2)
When comparing the actual dry mass data with the converted dry mass data a one-way ANOVA was used.
All statistical analyses were performed in R version 2.15.1 (R Core Team 2012) and the significant level used was α=0.05.
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3 Results
3.1 Lichen dry mass
The total lichen dry mass varied from 1.53±0.12 to 8.48±0.65 g m-1 twig length. The dry mass on the 50N treatment trees was significantly lower compared to both the dry control (DC) and the irrigated control (IC) (one-way ANOVA, F5,12=1.68, P=0.05).
Figure 3. Total lichen dry mass ([g m-1]) for each treatment (DC, IC, 6N, 12.5N, 25N and 50N), bars denote ± 1 SE.
Data for lichen dry mass sampled from the same tree were pooled. See section 2.1 for description of the treatments and their abbreviations. The separate dry masses for the species are presented in Figure 4.
For A. sarmentosa, Bryoria spp., Hypogymnia spp., P. sulcata, P. glauca, T. chlorophylla, Usnea spp. and V. pinastri the dry mass varied between 0.03-4.14, 0.02-1.32, 0.05-1.32, 0.01-0.28, 1.07-5.36, 0-0.10, 0-0.19 and 0-0.03 g m-1 twig length, respectively (Figure 4). The lichen dry masses varied significantly between the species (two-way ANOVA, F6,72=35.6, P=2e-16) also with a significant interaction between treatment and species (two-way ANOVA, F30,72=2.6, P=0.0006). Regardless of treatment A. sarmentosa, Bryoria spp., Hypogymnia spp. or P. glauca accounted for most of the lichen biomass (92-97%). These four species were therefore further analysed (Figure 5).
The dry mass of A. sarmentosa on the 25N and 50N trees was significantly lower than both the DC and the IC treatments (one-way ANOVA, F5,12=14.1, P=0.0001). The dry mass of Bryoria spp. on the 50N treatment trees was significantly lower than both the DC and the IC (one-way ANOVA, F5,12=3.6, P=0.003). The dry mass of Hypogymnia spp. on the 50N treatment trees was as well significantly lower than both the DC and the IC (one-way ANOVA, F5,12=12.81, P=0.0002), while the dry mass of P. glauca was similar across all the treatments (one-way ANOVA, F5,12=0.2, P=1.0).
DC IC 6 12.5 25 50
[g m-1] 02468
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Figure 4. Dry mass ([g m-1]) of each species for each treatment (DC, IC, 6N, 12.5N, 25N and 50N). Data for lichen dry mass of each species of each tree were pooled. Bars denote ± 1 SE for the dry masses of all species for each treatment. See section 2.1 for description of the treatments and their abbreviations. The dry masses for A.
sarmentosa, Bryoria spp., Hypogymnia spr. and P. glauca are in detail presented in figure 5.
Figure 5. Dry mass ([g m-1]) of A. sarmentosa, Bryoria spp., Hypogymnia spr. and P. glauca for each treatment (DC, IC, 6N, 12.5N, 25N and 50N), bars denote ± 1 SE.Data for lichen biomass of each species of each tree were pooled. See section 2.1 for description of the treatments and their abbreviations.
DC IC 6 12.5 25 50
Vul Usn Tuc Pla Par Hyp Bry Ale
[g m-1] 02468
Ale Bry Hyp Pla
[g m-1]
0 1 2 3 4 5 6
DC IC 6 12.5 25 50
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3.2 Species richness
The mean species richness (number of species observed per branch) for the DC, IC, 6N, 12.5N, 25N and 50N treatments were 7.33, 7.40, 7.33, 6.75 and 5.25, respectively (Figure 6).
The variance in species richness on the 50N trees was significantly lower than on the DC trees (one-way ANOVA, F5,12=5.5, P=0.002). However, the variance in species richness on the 25N and 50N trees was significantly lower than the IC (one-way ANOVA, F5,12=5.5, P=0.04;
F5,12=5.5, P=0.001).
Figure 6. Species richness for each treatment (DC, IC, 6N, 12.5N, 25N and 50N). Median, quartiles (Q1 and Q2) and influential points for each treatment (kg N ha-1 yr-1).
3.3 Specific thallus mass
The Specific Thallus Mass (STM) (i. e. dry mass per wet thallus surface area) varied from 2.45 to 13.25 mg cm-2 (Figure 7). The variation in STM for T. chlorophylla was significantly lower than for the other species’ STM (two-way ANOVA, F8,225=10.6; P=1.1e-08).
The STM for Hypogymnia spp. varied from 2.45 to 12.77 mg cm-2 and was significantly higher on the 50N treatment trees compared to the DC trees (one-way ANOVA, F5,63=0.9, P=0.04). For P. glauca the STM varied from 3.51 to 13.23 mg cm-2 and was significantly higher on the 25N and 50N treatment trees compared to both the DC and the IC trees (one- way ANOVA, F5,66=2.5, P=0.04). The STM for P. sulcata varied from 3.91 to 12.93 mg cm-2 and was significantly higher on the 50N treatment trees compared to the IC trees (one-way ANOVA, F5,42=1.2, P=0.04). For T. chlorophylla the STM varied from 3.27 to 9.88 mg cm-2 and was significantly higher on the 6N and 12.5N treatments trees compared to the DC trees (one-way ANOVA, F4,40=2.9, P=0.03) and, the STM was significantly higher than the IC on the 6N treatment trees (one-way ANOVA, F4,40=2.9, P=0.02).
In order to visualize how the treatments affected the lichens’ visual appearance, photos of Hypogymnia spp., P. glauca and A. sarmentosa (the latter not included in the STM analysis) are presented in figure 8-10.
Species richness
DC IC 6 12.5 25 50
5678
10
Figure 7. Mean dry mass per thallus area (i.e . specific thallus mass, STM ([mg cm-2]) for Hypogymnia spp., P.
sulcata, P. glauca and T. chlorophylla for each treatment (DC, IC, 6,N 12.5N,25N and 50N). Bars denote ± 1 SE.
Figure 8. Hypogymnia spp. sampled from trees with DC, IC and 50N treatment (left to right).
Figure 9. P. glauca sampled from trees with DC, IC and 50N treatment (left to right).
Figure 10. A. sarmentosa sampled from trees with DC, IC and 50N treatment (left to right).
345678910
[mg cm-2]
DC IC 6 12.5 25 50
Hypogymnia spp.
345678910
DC IC 6 12.5 25 50
P. glauca
345678910
DC IC 6 12.5 25 50
P. sulcata
345678910
DC IC 6 12.5 25 50
T. chlorophylla
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3.4 Relationships between lichen abundance and dry mass
As already mentioned, the three most frequent taxa on the branches were A. sarmentosa, Hypogymnia spp. and P. glauca (Figure 4). On the DC branches they accounted in average for 39%, 18% and 40% of the lichen abundance, respectively. On the IC branches they accounted in average for 30%, 15% and 45%, and, on the 50N treatment branches they accounted in average for 9%, 5% and 81%. The other taxa (Bryoria spp., P. sulcata, T.
chlorophylla, Usnea spp. and V. pinastri) in total accounted for less than 2%, 3% and 9% in average on the DC, IC and 50N treatment branches, respectively.
As previously indicated in section 2.5 lichen dry mass was strongly correlated to lichen abundance (R2=0.8, P<2.2e-16) and the linear relationship was best described with the dry mass data square root transformed (R2=0.9, P<2.2e-16) (Figure 11).
(a) (b)
Figure 11. Lichen dry mass ([g m-1]) as a function of lichen abundance ([cm2 m-1]) (P<2.2e-16, R²=0.82 (y=1.028x.0.15), AIC=314), dotted line indicating a non-linear relationship (a). Square root transformed dry mass ([g0.5 m-0.5]) as a function of lichen abundance ([cm 2 m-1]) (P<2.2e-16, R²=0.90 (y=0.011x+0.160), AIC=-30) (b).
The species with the utmost dry masses and abundances are: P. glauca (diamonds), A. sarmentosa (circles), Hypogymnia spp. (crosses) and Bryoria spp. (triangles).
Despite the variation in STM, neither species nor treatment had a significant effect on the dry mass to abundance relationship (P>0.05). The lichen abundance data obtained from the image analyses could therefore be converted into lichen dry mass using Equation 2 regardless of species and treatment (Figure 12). When comparing the actual lichen dry mass data with the converted lichen dry mass data, there was no significant difference between these two approaches (P>0.05).
0 50 100 150 200 250 300
02468101214
[cm2 m-1]
[g m-1]
0 50 100 150 200 250 300
0123
[cm2 m-1]
[g0.5 m-0.5)]
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Figure 12. Actual lichen dry mass ([g m-1]) from the branches included in the image analyses and the lichen abundance from the same image analyse converted to lichen dry mass ([g m-1]). The image analyse included DC, IC and 50N treatment trees. Bars denote ± 1 SE. See section 2.5 for description of the relationship between lichen abundance and dry mass.
The model describing the relationship of lichen abundance and dry mass (Figure 11, Equation 2) was also applied to lichen abundance data from the previously made image analyses by Johansson et al. (2012). This can be justified because the data was collected (Figure 13) and the image analyses were performed with nearly the same method (see section 2.4) in this and the former study; the branches come from the same trees and the same species were recorded. The results presented in Johansson et al. (2012) were based on image analyses of eight branches per treatment tree annually documented as described in section 2.1.
However, although originating from the same trees, the converted dry masses from year 2005 to 2009 were generally lower than the here obtained dry masses from 2012 (Figure 4 and 14).
(a) (b)
Figure 13. Photographs of branches from the same tree (IC treatment) and height (2m), used in image analyses described in section 2.4. Photo (a) was taken by Otilia Johansson at the site in Kulbäcksliden in 2007 and photo (b) was taken in the laboratory by Marie Rönnqvist in 2012, see section 2.2 for description of the photo method.
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Figure 14. Lichen dry mass ([g m-1]) converted from lichen abundance from image analyses. The photos included in these analyses were taken year 2005-2009 on branches from the same trees included in this study. Bars denote
± 1 SE for the dry masses of all species for each treatment (IC, 6N, 12.5N, 25N and 50N) and year (2005-2009. See section 2.5 for description of the relationship between lichen abundance and dry mass.
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4 Discussion
4.1 Long-term effects of nitrogen fertilization
My results show that seven years of nitrogen (N) fertilization has caused changes in dry mass for the epiphytic lichen species (Figure 3), however the effects differ between them (Figure 4 and 5). In agreement with other studies, I found the dry mass of P. glauca to be similar between all treatments, supporting that P. glauca is the most N-tolerant species of the monitored lichen community (McCune and Geiser 2009, Johansson et al. 2012). Palmqvist and Dahlman (2006) recorded an increase in dry mass for P. glauca during high-intensity N- fertilization and suggested a well integrated symbiosis regardless of environmental disturbance. However, I could not detect such an increase in dry mass for P. glauca. Neither did I find any significant increases in dry mass for A. sarmentosa, Bryoria spp. and Hypogymnia spp., which is in contrast to the findings made by Johansson et al. (2012); they found that these three species increased in abundance during low N treatments. The results of Palmqvist and Dahlman (2006) and Johansson et al. (2012) are based on rather short- term studies (one growing season and four years, respectively). Hence, the results from this 7 year study indicate that initial positive or neutral effects of enhanced N loads can have negative long term effects. I also found a significant decrease in dry mass at the 50 kg N ha-1 y-1 load for both Bryoria spp. and Hypogymnia spp. further supporting the hypothesis of cumulative effects of N additions (De Schrijver et al. 2011). At the 25 and 50 kg N ha-1 y-1 loads the dry mass of A. sarmentosa significantly decreased; this is consistent with findings by several authors (McCune and Geiser 2009, Geiser et al. 2010, Johansson et al. 2012).
Considering A. sarmentosa being such an N-sensitive species, I hypothesise the main reason for its dry mass decline is the N-fertilization per se however as discussed above, effects of cumulative loads probably also play a vital role. The mechanism behind the N induced decline in A. sarmentosa remains quite unknown; Johansson et al. (2012) suggests that reduced stability of the lichen thalli or an increased vulnerability to diseases are the two most realistic explanations.
I found a significant decline in species richness for the two highest N loads (25 and 50 kg N ha-1 y-1) and no increase at lower loads. And even though not significant, the data for species richness also indicates a decline at 12.5 kg N ha-1 y-1. In contrast, Johansson et al. (2012) found a significant increase in abundance under the IC, 6N and 12.5N treatments. This supports the hypothesis that it takes longer time than a few years to detect negative changes of species richness in lichen communities exposed to an enhanced N deposition. In addition, low concentrations seem to have an impact because initial positive effects during the lower treatments were depressed with time (Johansson et al. 2012). Therefore, I suggest that even 6 kg N ha-1 yr-1 might be above the critical load of N deposition for epiphytic lichen communities in boreal forests, and thereby support previously determined critical loads (Bobbink et al. 2010). Large parts of Sweden receives an N deposition lower than 6 kg N ha-1 yr-1 (SMHI 2011). However, when bearing in mind that cumulative N addition might have negative consequences the risk of epiphytic lichen species disappearing in the boreal forests of Sweden still seems high. In southern parts of Sweden where the N deposition exceeds 6 kg N ha-1 yr-1 and old growth forests are rare A. sarmentosa is considered to be near threatened (NT) by The Red List of Swedish Species (Gärdefors 2010).
Hypogymnia spp. is just as P. glauca considered to be an N-tolerant species; Dahlman et al.
(2003) found that both P. glauca and Hypogymnia physodes survived intensive N fertilization for 16 years when phosphorus (P) and other nutrients were added. N enrichment can promote a shift from N-limited to P-limited growth in epiphytic lichens and the balance between N and P supply is important for the symbiotic interaction between the mycobiont and photobiont (Hogan et al. 2010, Johansson et al. 2011). Too low levels of P together with a cumulative load of N might therefore be the reason why, in this study, neither of these two N- tolerant species increased in dry mass, nor that the dry mass of Hypogymnia spp. was significantly and negatively affected by the 50N treatment. Speculatively, negative effects for
15
P. glauca may then occur as well after several years (10-20) of N fertilization due to P limitations.
My results of total lichen dry mass did only show a significant negative effect for the 50N treatment. However, the data indicates negative effects of the 12.5N and 25N treatments (Figure 3). An unaffected total lichen biomass does not necessarily mean that the composition of the lichen community is unchanged. Figure 4 strongly indicates that the lichen community is affected by the increased N deposition; N-tolerant species seems to replace the N-sensitive ones as previously documented by several authors (Geiser et al. 2010, Johansson et al. 2011, Evju and Bruteig 2013). Moreover, I found a significant interaction between treatment and species suggesting the lichen species have different N optima (Palmqvist et al. 2002, Johansson et al. 2012); thus indicating that the replacement of N- sensitive species is not competition induced. The proportion of P. glauca increased in the lichen community at the 25N and 50N treatments (Figure 4), this could be because of an increased competition as suggested in Welch et al. (2006). However, I found a significant decrease in dry mass for A. sarmentosa, Bryoria spp. and Hypogymnia spp. at the higher N treatments therefore I suggest, that the proportional increase in P. glauca is more likely due to N-sensitive species disappearing than due to competition between the lichen species (Dahlman et al. 2003, Gaio-Oliveira et al. 2005). I found no significant treatment effects on lichen dry mass for P. sulcata, T. chlorophylla, V. Pinastri and Usnea spp. Not necessarily because a lack of responses to the fertilization, but more likely due to the small number of occurrences (low statistical power to detect treatment effects). Accordingly, my results show that species richness distinctly declines at the two highest N loads, strongly indicating a loss of N-sensitive species supported by several other authors and in line with earlier findings (Vitousek et al 1997, Sala et al. 2000, Van Dobben et al. 2001, Rajaniemi 2002, Johansson et al. 2012, Evju and Bruteig 2013).
Only by looking at the lichen species harvested in this study one can see that seven years of an increased N load has changed the lichen species morphology (Figures 8-10). The question is how and why. The STM for P. glauca on the lower treatment trees are in line with findings made by Gauslaa and Coxson (2011); they found an STM of 7.21 (±0.15) for P. glauca and could not find a significant difference in STM between open and closed canopy sites. Hence, it is most likely because of the N fertilization that the STM for P. glauca for the 25N and 50N treatments are higher than the STM found by Gauslaa and Coxson (2011). In general the specific thallus mass (STM) increased for the higher N treatments for Hypogymnia spp., P.
glauca, P. sulcata and T. chlorophylla. (Figure 7). Dry mass gains can result in an increase in STM (see Equation 1). However, as I found a decrease in dry mass (Figure 3-5) an increase in dry mass cannot explain the STM increase. A decline in surface area in relation to dry mass is more likely the reason for the increased STM for the lichen species in this study. Results published by Dahlman et al. (2003) imply a significant shift in thallus N investments from mycobiont to photobiont tissue; this probably leads to an increase in the photobiont and a decrease of the mycobiont partner. Evidently, in this study the photobiont increases with higher N loads because the lichens turn greener with higher treatment load (Figure 8-10), hence an increased chlorophyll a concentration and probably also an increased photosynthetic capacity (Dahlman et al. 2003). The mycobiont needs hydration to form turgor pressure in order to contribute to the area growth (Money 2008), and if the photosynthesis proceed for a longer time than the mycobiont stays hydrated (i.e. when it has sufficient turgor pressure) the STM increases (Larsson et al. 2012). In addition, high water content in the thalli (owing to the artificiall irrigation further described in section 2.1) can prevent transfer of carbon from the photobiont to the mycobiont and therefore inhibit the mycobiont and indirectly hinder area growth (Johansson et al. 2011). Gauslaa et al. (2009) found an area-expanding effect of moisture in lichens; however in this study the IC trees did not increase in area (decrease in STM) compared to the lichens on the DC trees. This is also probably because of the prevented carbon transfer hypothesised by Johansson et al. (2011). It would be interesting to investigate how the STM for frutiose lichens respond to the N
16
fertilization; however it would require a different approach to be able to determine their surface area.
4.2 Conversion of abundance to dry mass
Attempting to convert earlier abundance data published by Johansson et al. (2012) I examined the relationship between lichen dry mass and abundance for the lichens collected in October 2012. Surprisingly, neither species nor treatment seemed to affect the relationship suggesting only a minor change in morphology. However, the variations in STM negate this and it should also be noted that the abundance analysis was not performed for the whole material as further described in section 2.4. By extending the relationship to include abundance analysis from all branches from all the treatment trees, a different result might be obtained. Using the relationship between lichen dry mass and the in this study obtained lichen abundance material, the actual and the converted lichen dry masses from the branches collected in 2012 were similar and I therefore consider the conversion approach to be adequate (Figure 12).
The converted lichen dry masses from year 2005 to 2009 were clearly lower than the actual dry masses from the branches collected in 2012 (Figure 12 and 14). This is likely due to a bias as it cannot be ruled out that branches with more lichens where unwittingly chosen when we collected the material from the treatment trees. The lichen dry mass data from 2012 can therefore not be combined with the converted data from 2005-2009. The converted data however is very interesting because it provides the opportunity to follow how the dry masses for the lichen species have developed over time instead of how the abundances have changed as previously published in Johansson et al. (2012). Photographs of the branches included in the study of Johansson et al. (2012) were taken directly after the simulated N deposition was subsequently halted in October 2012. Therefore, an image analysis of those and thereon a conversion to dry mass data would provide the whole long-term outcome of the seven years of N fertilization at the site.
4.3 Conclusions
I have in this study found direct long-term effects of an increased N deposition. The epiphytic lichens in the boreal forest at the study site were affected in several ways. Firstly, the lichen dry mass and species richness declined at high N loads, and initial positive effects of low N loads (Johansson et al. 2012) were thus depressed with time. This strongly indicates that cumulative loads of N are important and need to be considered when determining critical load values. The critical load for lichen communities in boreal forests might be below 6 kg N ha-1 yr-1 and if the prediction of enhanced N depositions becomes reality, many lichen species and communities on the planet are at a great risk of being severely damaged or even extinct.
Secondly, N-limited lichen species might become P-limited (Johansson et al. 2011) due to N enrichment and therefore decline in dry mass with time. Thirdly, the composition of the lichen community changed benefitting N-tolerant species, most likely because N-sensitive species disappeared. Fourthly, the foliose lichen species morphology responded to the N fertilization by an increased STM, probably due to a decrease in surface area. However, the mechanism behind the responses is not fully investigated in this study and remains unclear.
Lastly, the effects of long-term atmospheric N deposition on epiphytic lichen communities needs further investigation in order to fully understand how the different lichen species and the lichen community respond to an enhanced and cumulative N input.
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5 Acknowledgments
First of all I would like to thank my supervisor Kristin (Kikki) Palmqvist for being such a great support throughout this whole project and for always finding time to answer my questions and listen to my concerns. I would also like thank Johan Olofsson for introducing me to this project and for climbing up those high towers when the rest of us safely stayed on the ground. I also want to express my gratitude to Ann Sehlstedt for helping me with the practical work and giving smart solutions. Thereto I want to thank Per-Anders Esseen and Lars Ericson for their guidance and help with identification of some tricky lichens. I would also like to thank Otilia Johansson for contributing data and valuable information. Finally, I want to thank Mattias Lindh for all the encouragement and for challenging me to think outside the box.
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6 References
Ahmadjian, Vernon. 1993. The lichen symbiosis. Canada: John Wiley & Sons, Inc.
Bobbink, R., Hicks, K., Galloway, J., Spranger, T., Alkemade, R., Ashmore, M., Bustamante, M., Cinderby, S., Davidson, E., Dentener, F., Emmett, B., Erisman, J-W., Fenn, M., Gilliam, F., Nordin, A., Pardo, L. and De Vries, W. 2010. Global assessment of nitrogen deposition effects on terrestrial plant diversity: a synthesis. Ecological Applications 20(1):30-59.
Dahlman, Lena, Persson, Görgen, Näsholm, Torgny and Palmqvist, Kristin. 2003. Carbon and nitrogen distribution in the green algal lichens Hypogymnia physodes and Platismatia glauca in relation to nutrient supply. Planta 217:41-48.
De Schrijver, An, De Frenne, Pieter, Ampoorter, Evy, Van Nevel, Lotte, Demey, Andreas, Wuyts, Karen and Verheyen, Kris. 2011. Cumulative nitrogen input drives species loss in terrestrial ecosystems. Global Ecology and Biogeography 20:803-816.
Emsley, John. 2011. Nature's Building Blocks: An A-Z Guide to the Elements. New York:
Oxford University Press.
Evju, Marianne and Bruteig, Inga E. 2013. Lichen community change over a 15-year time period: effects of climate and pollution. The Lichenologist 45(1):35-50.
Follet, R. F. and Hatfield, J. L. 2001. Nitrogen in the Environment: Sources, Problems, and Management. The Scientific World 1(S2):920-926.
Gaio-Oliveira, Gisela, Dahlman, Lena, Palmqvist, Kristin, Martins-Loução, Maria. A and Máguas, Christina. 2005. Nitrogen uptake in relation to excess supply and its effects on the lichens Evernia prunastri (L.) Ach and Xanthoria parietina (L.) Th. Fr. Planta 220:794-803.
Galloway, James N, Aber, John D., Willem Erisman, Jan, Seitzinger, Sybil P., Howarth, Robert W., Cowling, Ellis B. and Cosby, B Jack. 2003. The Nitrogen Cascade. BioScience 53(4):341-356.
Galloway, J. N., Dentener, F. J., Capone, D.G., Boyer, E. W., Howarth, R. W., Seirzinger, S.P., Asner, G. P., Cleveland, C.C., Green, P. A., Holland, E. A., Karl, D. M., Michaels, A. F., Porter, J. H., Townsend and A. R., Vörösmarty, C. J. 2004. Nitrogen cycles: past, present and future. Biogeochemistry 70:153-226.
Galloway, James N., Townsend, Alan R., Willem Erisman, Jan, Bekunda, Mateete, Cai, Zucong, Freney, Johan R., Martinelli, Luiz A., Seitzinger, Sybil P. and Sutton, Mark A.
2008. Transformation of the Nitrogen Cycle: Recent Trends, Questions, and Potential Solutions. Science 320:889-892.
Gauslaa, Yngvar and Coxon, Darwyn. 2011. Interspecific and intraspecific variations in water storage in epiphytic old forest foliose lichens. Botany 89:787-798.
Gauslaa, Yngvar, Lie, Marit and Ohlson, Mikael. 2008. Epiphytic lichen biomass in a boreal Norway spruce forest. The Lichenologist 40(3):257-266.
Gauslaa, Yngvar, Palmqvist, Kristin, Solhaug, Knut A., Hilmo, Olga, Holien, Håkan, Nybakken, Line and Ohlson, Mikael. 2009. Size-dependent growth of two old-growth associated macrolichen species. New Phytologist 181:683-692.
Geiser, Linda H., Jovan, Sarah E., Glavich, Doug A. and Porter, Matthew K. 2010. Lichen- based critical loads for atmospheric nitrogen deposition in Western Oregon and Washington Forests, USA. Environmental Pollution 158:2412-2421.
Gärdefors, Ulf (ed.). 2010. Rödlistade arter i Sverige 2010 – The 2012 Red List of Swedish Species. Uppsala: Artdatabanken, SLU.
Hawksworth, David L. 1971. Lichens as litmus for air pollution: a historical review.
International Journal of Environmental Studies 1:281-296.
Hale, Mason E. 1974. The biology of lichens 2nd edition. London: Edward Arnold.
Hogan, E. J., Minnullina, L. J., Sheppard, L. J., Leith, I. D. and Crittenden, P. D. 2010.
Response of phosphomonoesterase activity in the lichen Cladonia portentosa to nitrogen and phosphorus enrichment in a field manipulation experiment. New Phytologist 186:926-933.
19
Honegger, Rosmarie. 1991. Functional aspects of the lichen symbiosis. Annual Review of Plant Physiology and Plant Molecular Biology 42:553-578.
Honegger, Rosmarie. 1998. The lichen symbiosis – what is so spectacular about it?
Lichenologist 30(3):193-212.
Johansson, Otilia, Nordin, Annika, Olofsson, Johan and Palmqvist, Kristin. 2010. Responses of epiphytic lichens to an experimental whole-tree nitrogen-deposition gradient. New Phytologist 188:1075-1084.
Johansson, Otilia, Olofsson, Johan, Giesler, Reiner and Palmqvist, Kristin. 2011. Lichen responses to nitrogen and phosphorus additions can be explained by the different symbiont responses. New Phytologist 191:795-805.
Johansson, Otilia, Palmqvist, Kristin and Olofsson, Johan. 2012. Nitrogen deposition drives lichen community changes through differential species responses. Global Change Biology 18:2626-2635.
Larsson, Per, Solhaug, Knut A. and Gauslaa, Yngvar. 2012. Seasonal partitioning of growth into biomass and area expansion in a cephalolichen and a cyanolichen of the old forest genus Lobaria. New Phytologist 194:991-1000.
Levine, Jonathan M., Brewer, J. Stephen and Bertness, Mark D. 1998. Nutrients, competition and plant zonation in a New England salt marsh. Journal of Ecology 86:285-292.
McClean, Colin J., Van den Berg, Leon J. L., Ashmore, Mike R. and Preston, Chris D. 2011.
Atmospheric nitrogen deposition explains patterns of plant species loss. Global Change Biology 17:2882-2892.
McCune, Bruce. 1993. Gradients in Epiphyte Biomass in Three Pseudotsuga- Tsuga Forests of Different Ages in Western Oregon and Washington. The Bryologist 96(3): 405-411.
McCune, Bruce and Geiser, Linda. 2009. Macrolichens of the Pacific Northwest. Corvallis:
Oregon State University Press.
Money, Nicholas P. 2008. Insights on the mechanics of hyphal growth. Fungal Biology Reviews 22:71-76.
Nash III, Thomas H. 2008 a. Introduktion. In Lichen Biology 2nd edition Thomas H. Nash III (ed.), 1-8. New York: Cambrudge University Press.
Nash III, Thomas H. 2008 b. Lichen sensitivity to air pollution. In Lichen Biology 2nd edition Thomas H. Nash III (ed.), 299-314. New York: Cambrudge University Press.
Neff, Jason. C., Holland, Elisabeth. A., Dentener, Frank. J., McDowell, William H. and Russell, Kristina. M. 2002. The origin, composition and rates of organic nitrogen deposition: A missing piece of the nitrogen cycle? Biogeochemistry 57/58:99-136.
Nilsson, Jan and Grennfelt, Peringe. 1988. Critical loads for sulphur and nitrogen – Miljørapport 1988:15. Copenhagen: Nordic Council of Ministers.
Palmqvist, Kristin and Dahlman, Lena. 2006. Responses of the green algal foliose lichen Platismatia glauca to increased nitrogen supply. New Phytologist 171:343-356.
Palmqvist, Kristin, Dahlman, Lena, Jonsson, Anna and Nash III, Thomas H. 2008. The carbon economy of lichens. In Lichen Biology 2nd edition Thomas H. Nash III (ed.), 182- 215. New York: Cambrudge University Press.
Palmqvist, Kristin, Dahlman, Lena, Valladares, Fernando, Tehler, Anders, Sancho, Leopoldo G. and Mattsson, Jan-Eric. 2002. CO2 exchange and thallus nitrogen across 75 contrasting lichen associations from different climate zones. Oecologia 133:295-306.
Petterson, Roger B., Ball, John P., Renhorn, Karl-Erik, Esseen, Per-Anders and Sjöberg, Kjell.
1995. Invertebrate communities in boreal forest canopies as influenced by forestry and lichens with implications for passerine birds. Biological Conservation 74:57-63.
Pihl Karsson, Gunilla, Blomgren, Håkan, Petterson, Kjell, Svensson, Annika och Sjöberg, Karin. 2003. Nationell miljöövervakning av luft- och nederbördskemi 2002. IVL Svenska Miljöinstitutet AB.
Rajaniemi, Tara K. 2002. Why does fertilization reduce plant species diversity? Testing three competition-based hypotheses. Journal of Ecology 90:316-324.
Rai, A. N. 2002. Cyanolichens: Nitrogen metabolism. In Cyanobacteria in Symbiosis 97-116.
Dordrecht: Kluwer Academic Publishers.
20
R Core Team (2012). R: A language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. ISBN 3-900051-07-0, URL http://www.R- project.org/.
Sala, Osvaldo E., Chapin III, F. Stuart, Armesto, Juan J., Berlow, Eric, Bloomfield, Janine, Dirzo, Rodolfo, Huber-Sanwald, Elisabeth, Huenneke, Laura F., Jackson, Robert B., Kinzig, Ann, Leemans, Rik, Lodge, David M., Mooney, Harold A., Oesterheld, Martín, Poff, N. LeRoy, Sykes, Martin T., Walker, Brian H., Walker, Marilyn and Wall, Diana H. 2000.
Global Biodiversity Scenarios for the Year 2100. Science 287:1770-1774.
Scott, G. D. 1960. Studies of the lichen symbiosis. I. The relationship between nutrition and moisture content in the maintenance of the symbiotic state. New Phytologist 59:374-381.
Seaward, Mark R. D. 2008. Environmental role of lichens. In Lichen Biology 2nd edition Thomas H. Nash III (ed.), 274-198. New York: Cambrudge University Press.
SMHI, Swedish Meteorological and Hydrological Institute. 2011. Nationell kartläggning av atmosfärskemiska data för Sveriges miljöövervakning, database.
http://www.smhi.se/klimatdata/miljo/atmosfarskemi/atmosfarskemi-1.9347.
28.05.2013.
SMHI, Swedish Meteorological and Hydrological Institute. 2013. Nederbörd, database.
http://www.smhi.se/klimatdata/meteorologi/nederbord. 21.05.2013.
Smith, Anthony J. E. 1982. Bryophyte ecology. London: Chapman and Hall.
Socolow, Robert H. 1999. Nitrogen management and the future of food: Lessons from the management of energy and carbon. Proceedings of the National Academy of Sciences of the United States of America 96:6001-6008.
Strenbom, Joachim, Nordin, Annika, Näsholm, Torgny and Ericson, Lars. 2002. Parasitic fungus mediates change in nitrogen-exposed boreal forest vegetation. Journal of Ecology 90:61-67.
Van Dobben, H. F. and Ter Braak, C. J. F. 1998. Effects of atmospheric NH3 on epiphytic lichens in the Netherlands: the pitfalls of biological monitoring. Atmospheric Environment 32(3):551-557.
Van Dobben, H. F., Wolterbeek, H. Th., Wamelink, G. W. W. and Ter Braak, C. J. F. 2001.
Relationship between epiphytic lichens, trace elements and gaseous atmospheric pollutants. Environmental Pollution 112:163-169.
Vitousek, Peter M., Aber, John D., Howarth, Robert W., Likens, Gene E., Matson, Pamela A., Schindler, David W., Schlesinger, William H. and Tilman, David G. 1997. Human alteration of the global nitrogen cycle: sources and consequences. Ecological Applications 7(3):737-750.
Vitousek, Peter M., Hättenschwiler, Stephan, Olander, Lydia and Allison, Steven. 2002.
Nitrogen and Nature. Ambio 31(2):97-101.
Welch, A. R., Gillman, M. P. and John, E. A. 2006. Effect of nutrient application on growth rate and competitive ability of three foliose lichen species. The Lichenologist 38(2):177- 186.
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