2 Materials and Methods
3.3 Discussion
3.3.1 DMSP concentrations in Spartina anglica and sediment analysis
Overall, the DMSP concentrations measured in this study were in the same range as those determined in previous studies, but it is shown here for the first time, that
Spartina anglica plants (leaves, stems and roots) are rich in DMSP over an entire annual cycle. However, DMSP was either not detectable or only detectable in trace amounts in leaf wash and rhizosphere sediment samples. This could be ex- plained by the fact that DMSP is stored intracellularly in Spartina anglica
(Husband et al. 2012, Otte et al. 2004, van Diggelen et al. 1986).
In this study DMSP concentrations of up to ~20 µmol g-1 fresh weight have been measured in Spartina anglica tissue samples. Van Diggelen et al. (1986) me- asured concentrations between 4 and 50 µmol g-1 fresh weight in greenhouse- grown Spartina anglica leaves. However, van Diggelen et al. (1986) did not carry out background measurements to determine endogenous DMS release from the leaf samples of Spartina anglica. Furthermore, van Diggelen et al. (1986) incu- bated samples for only 30 minutes and it has been shown that short incubation times lead to an overestimation of the DMSP concentrations, due to the fact that DMS formation is faster from plant samples than from synthetic DMSP in stan- dards during the first 3 hours (Otte and Morris 1994). Therefore, the DMSP con- centrations measured by van Diggelen et al. (1986) could be an overestimation. Mulholland and Otte (2000) measured DMSP concentrations in Spartina anglica
leaves, stems and roots ranging from 8 to 30 µmol g-1 FW in leaves, 4 to 12 µmol g-1 FW in stems and 4 to 10 µmol g-1 FW in roots. Mulholland and Otte (2002) determined DMSP concentrations in Spartina anglica leaf samples ranging from 4 to 50 µmol g-1 FW.
Environmental factors including sediment properties have previously been shown to affect the DMSP content in different tissues of Spartina plants. However, all previous studies have only addressed this issue in greenhouse or laboratory grown
Spartina. In this study it was shown that different sediment properties can lead to different chemical properties and hence to different intracellular DMSP concentra-
tions in Spartina and it was shown for the first time that these trends seem to be holding true in Spartina samples obtained from the natural environment.
The sediment analysis revealed that Spartina anglica plants 1 to 3 grew in sand and plant 4 to 6 in sandy loam. Sand is generally low in organic matter and le- aching of nutrients takes place (Gove et al. 2001). Furthermore, sand has relative- ly large-sized particles and therefore no capability to interact with charged par- ticles. Sandy loam has a higher percentage of silt and clay compared to sand and can therefore hold nitrogen and other compounds for longer than sand as demon- strated in studies by e.g. Christensen (1985). In Table 3.2 it can be seen that total nitrogen and total carbon concentrations, organic matter, etc. are higher in the sandy loam compared to the sand. These results go along with the general knowledge that the soil type and texture is an important factor that influences the distribution of these compounds including nitrogen (Bechtold and Naiman 2006, Powell Gaines and Gaines 1994).
Elevated nitrogen concentrations in the sediment resulted in decreasing DMSP concentrations in Spartina leaves as shown in the present and in previous studies (Colmer et al. 1996, Mulholland and Otte 2000, Otte and Morris 1994). In the studies by Otte and Morris (1994) and Mulholland and Otte (2000) Spartina
plants were grown in greenhouses under varying nitrogen concentrations (0 – 2 mM). In the study by Colmer et al. (1996) Spartina was grown under laboratory conditions with increasing NH3- and NH4+ concentrations. Although, the men-
tioned studies came to the same result – decreasing DMSP concentrations in
Spartina leaves with increasing nitrogen concentrations, only the present study was able to confirm these results in the field and to show a negative correlation between the internal DMSP concentration in Spartina leaves and the nitrogen concentration in the surrounding sediment. Therefore, the varying DMSP concen- trations in the Spartina leaves (Figure 3.2) can be explained by varying nitrogen availability in the surrounding sediment of the Spartina plants. However, it could also be concluded that different sediment types (sand versus sandy loam) show different chemical properties (Table 3.2) and therefore different internal DMSP concentrations especially in leaves (Figure 3.2).
Mulholland and Otte (2000) showed in their study that although the DMSP con- centration in Spartina leaves decreased with increasing nitrogen concentrations the whole plant DMSP content increased. This result cannot be confirmed in the present study. Although the DMSP concentrations in Spartina stem samples in- creased with increasing nitrogen concentrations the DMSP concentrations in leaf and root samples decreased and therefore the overall DMSP concentration in the whole plant decreased with increasing nitrogen concentrations. The experiments by Mulholland and Otte (2000) were carried out with greenhouse-grown Spartina
plants whereas the results in this study are based on naturally salt marsh grown
Spartina plants. Spartina plants grown in greenhouses might behave differently to
Spartina plants grown in their natural environment and could therefore result in contrasting results. Furthermore, Mulholland and Otte (2000) suggested that the DMSP is translocated from the Spartina leaves to the stems upon increasing ni- trogen concentrations (Otte et al. 2004). In this study the DMSP concentrations decreased in leaves with increasing nitrogen concentrations and the DMSP con- centration in stems increased. However, if a translocation of DMSP from the leaves to the stems is the cause for the increase in DMSP in the stems could not been proven during this study.
DMSP is not the only compatible solute in Spartina plants. The quaternary am- monium compound glycine betaine and the α-amino acid proline are two nitrogen- based compatible solutes in Spartina (Mulholland and Otte 2002). Glycine beta- ine, proline and DMSP act as osmotically active substances in Spartina and the biosynthesis pathways of DMSP and glycine betaine are linked and therefore a physiological relationship between those two compounds exists (Mulholland and Otte 2002). DMSP itself is not a nitrogen compound and in studies by Mulholland and Otte (2002) intracellular DMSP concentrations decreased and glycine betaine concentrations increased with elevated nitrogen concentrations. With low nitrogen concentrations intracellular DMSP concentrations increased and glycine betaine concentrations decreased in Spartina plants. Therefore, it could also be hypothe- sised that with elevated nitrogen concentrations the nitrogen compound glycine betaine is produced as an osmolyte in Spartina plants and with low nitrogen con- centrations the nitrogen independent osmotically active substance DMSP is syn- thesized.
3.3.2 DMS degradation and potential DMS degradation rates
An aspect that has not been investigated previously is to what extent the Spartina phyllosphere is a habitat for DMS degrading bacteria. It was shown here for the first time using DMS degradation experiments that Spartina anglica is colonized by DMS-degrading microorganisms in the phyllosphere and rhizosphere. These experiments indicated that the most active microorganisms are located on the leaf surface (leaf wash) in, or on, the roots and in the rhizosphere sediment. Although Schäfer et al. (2010) speculated that DMS-degrading bacteria might be present in the phyllosphere of DMSP-producing plants, until now it has never been proven. This is the first study to confirm that DMS degradation not only takes place in salt marsh sediments (Ansede et al. 2001, Kiene 1988) but also in the phyllosphere and rhizosphere of the salt marsh plant Spartina anglica, and therefore provides an entirely new aspect to our understanding of the role of microbial communities in affecting fluxes of DMS from salt marshes to the atmosphere.
Complete degradation of the initially added 1 mM DMS took place only in leaf wash, root and rhizosphere sediment samples. A complete degradation of the DMS could not be measured in leaf and stem samples. This might be due to the fact that the overall DMS concentration in the incubation vials was much higher than 1 mM due to the release of endogenous DMS from the leaf and stem tissue. DMS concentrations above 2.5 mM are considered to be toxic for bacterial cul- tures (De Zwart et al. 1996a). The results in Figure 3.4 are already corrected for endogenous DMS release.
The calculated potential DMS degradation rates (between ~ 0.07 and 0.83 µmol DMS g-1 FW d-1) show that the microorganisms in the phyllosphere and rhizo- sphere of Spartina anglica have potentially a great potential for DMS degradation. However, the incubation times and the initial DMS concentrations used in this study are high and therefore these potential rates cannot be applied to the envi- ronment. Simó and Pedrós-Alió (1999) measured DMS uptake rates of 0 – 6 nM d-1 in the surface sea water of the subpolar North Atlantic during a bloom of coc- colithophores using 8 h for incubation. Lomans et al. (1999) determined DMS uptake rates of 7.68 – 118.8 nmol ml-1 d-1 in peatland sediment slurries using ini-