External mycorrhizal mycelia – the importance of quantification in natural ecosystems
The external ectomycorrhizal mycelia (EEM) that grow out into the soil from ectomycorrhizal (EM) roots have a fundamental role in soil ecosystems. Their importance for the nutrient uptake of trees is well known (Smith & Read, 1997), and they also serve as an important food source for soil animals and in this way fuel the soil ecosystem with energy derived from the photosynthetic activity of trees. The large proportion of photosynthetic activity that is allocated to EM fungi (10–50%; reviewed by Simard et al., 2002) indicates a high potential for EEM to influence carbon sequestration in the soil. EM fungi have also been suggested to reduce decomposition rates in the soil, which would further enhance carbon sequestration rates (Langley & Hungate, 2003). Furthermore, the large contributions of EEM to soil respiration and soil microbial biomass which have recently been demonstrated by girdling of mature forest trees (Högberg & Högberg, 2002) emphasize the important role of EEM in many ecosystems.
It has been difficult to quantify the production of EEM in natural systems, mainly because biochemical markers for fungi cannot distinguish between saprotrophs (SAP) and EM fungi. Although molecular techniques for identifying EM fungi in the soil have made substantial progress in the last decade (e.g. Genney et al., 2006), it is still difficult to perform quantitative analyses of EEM production with these methods. We developed a mesh bag method to quantitatively estimate the production of EEM in forest ecosystems by using sand-filled in-growth mesh bags. These mesh bags are mainly colonized by EM fungi, and analysis of the carbon isotopic composition of mycelia extracted from the mesh bags can separate the contributions from EM and SAP fungi (Wallander et al., 2001). Furthermore, Kjöller (2006) recently investigated the composition of the EM community in sand-filled mesh bags incubated in a beech (Fagus sylvatica) forest, and found that the majority (83%) of the sequences obtained from the mesh bags originated from EM fungi. Boletoid EM fungi and Tomentella species that produce abundant EEM were especially common in the mesh bags. The in-growth mesh bag method together with fungal biomass estimates of soil samples has now successfully been used in arctic ecosystems to estimate potential changes in EEM growth in response to experimental treatments such as fertilization and warming (Clemmensen et al., in this issue, pp. 391–404).
‘The doubling of external ectomycorrhizal mycelial growth after fertilization could thus have significant potential to counterbalance the increased release of carbon dioxide from arctic ecosystems in response to long-term fertilization’
Opposing effects of fertilization in arctic and boreal ecosystems
Clemmensen et al. (2006) found that cycling of carbon and nitrogen (N) through EM fungi in arctic ecosystems has the potential to increase in the future, and this has important implications for future changes in carbon sequestration in these ecosystems. It is especially interesting to note that the effects of increased nutrient availability on EM growth appear to be different in arctic ecosystems compared with coniferous forests in the boreal region. Clemmensen et al. (2006) found significant stimulation of EM fungi by fertilization, while many earlier studies found reduced growth of EM fungi after N additions (reviewed by Wallenda & Kottke, 1998), and recent studies using the in-growth mesh bag method have also demonstrated reductions in EEM growth in response to elevated N availability in coniferous forests (Nilsson & Wallander, 2003; Nilsson et al., 2005; Hendricks et al., 2006). The reason for the contrasting effects of N addition on EM growth in arctic ecosystems compared with coniferous forests could be stronger N limitation in arctic ecosystems. Arctic ecosystems can be extremely N constrained (Schmidt et al., 2002) and N limitation of EEM growth is therefore not unlikely. In the heath tundra studied by Clemmensen et al. (2006), fertilization improved EEM growth without influencing growth of EM host plants, which may suggest N limitation of EEM growth at this site. In a recent paper, Hendricks et al. (2006) presented data suggesting N limitation of EEM growth in a coniferous forest where EEM growth was improved threefold by using more nutrient-rich natural soil rather than acid-washed sand in the mesh bags. On the basis of these results, Hendricks et al. (2006) promoted the use of natural soil rather than sand in mesh bags when estimating EEM growth, and this would clearly be a more realistic approach, although it requires that the background fungal biomass in the introduced soil is low. An alternative way to make the mesh bags more similar to the surrounding soil could be to introduce ion exchange resin into the mesh bags and load them with a simulated soil solution before introducing them into the soil.
The mechanisms for improved EEM growth in the tussock tundra, as reported in the study by Clemmensen et al. (2006), appeared to be different from the mechanisms in the heath tundra, as fertilization increased the biomass of the EM plants (Betula nana) eightfold at this site. A positive effect on the growth of the host plant is likely to have a positive influence on EEM growth, because more photosynthetic tissue will become available for carbon assimilation. This has been demonstrated in laboratory experiments where N addition has produced a positive growth response of both host plants and EEM (Ekblad et al., 1995) in N-limited systems, while addition of excess N to systems with balanced nutrition resulted in reduced growth of EEM and had no effect on the growth of the host plant (Wallander & Nylund, 1992). This shift towards more EM-dependent vegetation at arctic sites after fertilization may have important consequences for carbon sequestration of these ecosystems.
In a recent paper, Godbold et al. (2006) found that growth of EEM was the dominant pathway through which carbon entered the soil organic matter (SOM) pool in a deciduous forest ecosystem in southern Europe. By using C3 trees (Populus sp.) and C4 soil, they were able to estimate that approximately twice as much of the carbon that entered the SOM pool over a 3-year period originated from EEM compared with the amount originating from fine roots. The doubling of EEM growth found by Clemmensen et al. (2006) after fertilization could thus have significant potential to counterbalance the increased release of carbon dioxide from arctic ecosystems in response to long-term fertilization which has been reported by Mack et al. (2004).
Are ericoid mycorrhizal fungi important for carbon flux in arctic ecosystems?
Ericoid mycorrhizal (ErM) plants can dominate many arctic ecosystems, but the contribution of ErM fungi to the carbon flux, the production of external mycorrhizal mycelia and the total fungal biomass has not been conclusively determined. A recent study by Olsrud et al. (2004) demonstrated that ErM colonization of hair roots was enhanced in response to warming and fertilization in a heath tundra close to the site investigated by Clemmensen et al. (2006). ErM fungi are unlikely to colonize in-growth mesh bags as they do not extend far from the roots, but they can be expected to make up a significant proportion of the total fungal biomass in many arctic soils. Data from Nilsson et al. (2005) suggest that ErM fungi may contribute more than EM fungi to total fungal biomass in nutrient-poor coniferous forests with a field layer dominated by ericaceous plants. Methods to distinguish the biomass of ErM, EM and other fungi in the soil are highly desirable in order to elucidate the importance of these fungi for carbon sequestration in various ecosystems. The recent finding that some ErM fungi also can colonize EM hosts (Villareal-Ruiz et al., 2004) makes this an even more interesting research area.
Methodological problems to consider in future research
The use of in-growth mesh bags to measure EEM growth is especially useful for relative comparisons, for example to evaluate the effects of fertilization and temperature on EM growth, as in the study by Clemmensen et al. (2006). However, estimates of the absolute carbon flux through mycorrhizal systems are more problematic as production of EEM in in-growth mesh bags is unlikely to be the same as production in the soil. The substrate in the mesh bags is different from the soil, and the mesh bags represent an empty space in the soil that is likely to favour fast growers and early colonizers.
Quantification of fungal biomass in the soil using various biochemical markers is also problematic because conversion factors for converting biochemical markers such as ergosterol or phospholipid fatty acids to fungal biomass are necessary (discussed in Klamer & Bååth, 2004). Different fungal species contain different amounts of these biomarkers, which will cause problems, especially if the dominating fungal species in the sample has a divergent concentration of the biomarker. Usually this problem is less acute when large numbers of fungal species are included in the sample. Another problem with fungal biomass estimates based on ergosterol measurements is that these usually give unrealistically high values of fungal biomass when compared with other estimates of microbial biomass. The reason for this is not clear, but recent studies of ergosterol in the soil suggest a slower turnover rate than expected, which may indicate that some of the ergosterol in the soil may not be associated with living fungal biomass (Zhao et al., 2005).
The study by Clemmensen et al. (2006) emphasizes the importance of studying arctic ecosystems in order to understand future changes in the global carbon cycle. More detailed studies of carbon flux through mycorrhizal mycelia (both EM and ErM) and turnover of these mycelia are important future research fields in arctic as well as in boreal ecosystems.
I would like to thank Erland Bååth and Lars Ola Nilsson for valuable comments on the manuscript.