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The Hyphal Network of Arbuscular Mycorrhizal Fungi in the Soil: A Potential Key to a Better Understanding of Ecosystems and the Management of Horticultural Soils.

Further development of techniques for the extraction and analysis of soil-grown arbuscular mycorrhiza fungal hyphae

A sample of extraradical mycelium extracted from the soil	In the past, a system which allows for the extraction of arbuscular mycorrhiza (AM) mycelium was established at the IGZ. This method will be further developed to allow for the intact extraction of finely branched hyphae from the soil. Digital image processing will be adopted to assess AM mycelium architecture.
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Development of compartmented planting systems to measure the cost/benefit ratio of endomycorrhizal symbioses by isotope labelling

A planting pot with lateral compartments to which only fungal hyphae have access

Compartmented planting pots allow for the quantification of the contribution of a part of the AM fungal extraradical mycelium to plant nutrient uptake. Within this activity, 32/33P and 14C will be applied to quantify the contribution of the AM fungal mycelium to plant phosphorus uptake and the demand of the symbiont for plant carbohydrates.  The short-lived radioactive isotope 11C will be used to track photoassimilate partitioning between plant and AM fungal tissues at a very high spatial and temporal resolution. Suitable compartmented planting systems allowing for the application of radioactive P an C isotopes will be developed and constructed within this activity.

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Identification of molecular markers for the functioning of the arbuscular mycorrhiza fungal symbiosis

Extraradical AM fungal hyphae stained for alkaline phosphatase activity (dark dots within the hyphae), a potential marker for AM efficiency in P transport.

The presence and activity of phosphate transporters and proton pumps in the AM fungal extraradical mycelium will be investigated in order to identify potential markers for the current contribution of the AM symbiosis to plant phosphorus uptake. The expression of hexose transporters, proton pumps and enzymes of the energy metabolism in the intraradical mycelium could be an indicator for the supply of the AM fungus with plant photoassimilates. Genes with specific functions will be identified from cDNA libraries (complementary expression in yeast). The expression of these genes in the AM extraradical mycelium under different environmental conditions will then be investigated by in-situ hybridization and RT-PCR.

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Assessment of the cost/benefit-ratio of the AM symbiosis and the extraradical mycelium architecture in different developmental stages of the plant-fungal association.

Hypothesized changes in the cost/benefit-ratio of the AM symbiosis in different developmental stages

When mycorrhizae establish from resting propagules in the soil, the development of AM fungal root colonisation follows a sigmoidal curve with (I) a lag-phase, (II) a log-phase, (III) a plateau phase and in some cases a fourth phase where the extent of AM fungal root colonisation decreases. Architecture of the extraradical mycelium and the cost/benefit-ratio of the symbiosis will be investigated in different stages of AM development within this activity. It will also be tested, whether combinations of plant and AM fungal species with synchronized life-cycles lead to symbioses with higher functional compatibility compared with combinations of symbiotic partners differing in their life-cycle strategy.

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Investigation of the effect of the soil microfauna on the development of the AM extraradical mycelium and the cost/benefit-ratio of the AM symbiosis.

Do AM hyphae and spores (Picture shows bottles with mycelium samples) serve as food for soil animals?

The mycelium of AM fungi interacts with other soil microorganisms and the soil microfauna. Collembola and nematodes, which are very common in terrestrial soils, are frequently feeding on fungal tissue. Grazing of AM hyphae could lead to a decrease in the amount of AM mycelium in the soil and thus negatively affect mycorrhiza functioning. In contrast, soil animals feeding on saprophytic fungi could improve AM development and functioning by eliminating AM competitors for mineral elements in the soil. This activity will apply phospholipid fatty acid (PFLA) analysis to  investigate the interaction of the extraradical part of AM fungi with saprophytic fungi and different functional groups of  collembola and nematodes.

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A study on the influence of the presence of noncolonized host plant roots on AM extraradical mycelium architecture and the contribution of the AM symbiosis to plant growth and plant nutrient uptake.

Three compartment split root system to investigate  mycorrhiza in roots of two plants sharing the same soil volume

Arbuscular mycorrhizal fungi form morphologically different types of extraradical hyphae in the soil: Coarse, thick-walled and finely branched, thin hyphae. It has been hypothesized that coarse hyphae may function mainly in the spread of infection, whereas finely branched hyphae may be a key site of AM nutrient uptake from the soil. Within this activity, it will be investigated whether the presence of noncolonized host plant roots triggers preferential formation of coarse AM hyphae functioning in the spread of infection. It will also be investigated whether hyphae formed in presence of a noncolonized host plant root differ in their cost/benefit-ratio.

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Studies on the extraradical development of the AM symbiosis and its contribution to plant phosphate uptake in response to different types of phosphorus resources in the soil.

A fertilization experiment: Fungal compartments prepared with different types of  fertilizer are inserted into a planting pot

Nutrient resources are often not homogeneously distributed within the soil. To date it is not completely clear, whether and how mycorrhizal and nonmycorrhizal roots differ in their ability to exploit nutrient rich microsites in the soil. Within this activity, AM fungal strains from different field sites will be compared for their extraradical development in response to heterogeneously or homogeneously distributed sources of phosphorus in the soil. The contribution of different mycorrhizae to phosphorus uptake from different types of fertilizer (e.g. mineral vs organic form) will also be assessed.

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Investigation of the extraradical mycelium of AM fungi in the field

Insertion of fungal compartments into field soil (field experiments at the IBDF)

Development of  AM extraradical mycelium will be assessed in the long-term field experiments at the IBDF. Fungal compartments filled with minerally or organically fertilized substrate will be inserted into the field soil to obtain mycelium of the fungi from the field. Compartments filled with 15N laballed substrate will be used to assess nitrogen uptake via endomycorrhizal fungi by plants of different fertilization treatments. In addition, communities of soil microorganisms and microfauna in the different field plots will be characterized by PLFA analysis.

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