2.3 ECM identification and classification
Root samples prepared in 5% glycerin were gently washed in running water and soil particles adhering to root tips were removed with fine forceps under a stereoscopic microscope. When roots were covered by fungal mantles, they were classified as ectomycorrhiza. The morphology of ECM was determined under a photographic stereo microscope (LEICA, M205FA, Germany), the macroscopic and microscopic characteristics of the mycorrhizae were identified based on Agerer (1987-2006) (i.e., ECM system, color, mantle surface structure, cystidia, emanating hyphae, rhizomorphs, etc.). The living and dead root tips were distinguished by discerning the freshness or elasticity of the root tips during the microscope observation, and the tip numbers of living and dead root tips in each soil block were counted and the ratio of living to dead root tips (Root-tipsratio) were calculated. For representative ECM root samples of each morphotype in each soil block, three root tips were used for diameter (d , mm) and length (l , mm) measurements with the photographic stereo microscope. The total root tip number in each soil block was counted and identified by ECM morphotypes. The morphology diversity of ECM root tips (MDI) was measured by Simpson‘s index of diversity as in Lande (1996) and Matsuda & Hijii (2004); the ECM colonization ratio (Cratio) was measured as the percentage of the infected root tips over the total root tips. The ECM root tips per unit root biomass (ECMtips) were also measured. The superficial area of ECM root tips (SA) was measured for all ECM root tips in each soil block, with root tips determined as a combination of cylinder and hemisphere by:
SA (m2 m-3) =\(\sum_{i=1}^{N}[\)(2π (\(\frac{di}{2}\))2 +π di (li -di ))ⅹni ]ⅹ103
where di represents the average diameter of the ECM root tips of morphotype i ; lirepresents the average length of the ECM root tips of morphotypei ; ni represents the number of ECM root tips of morphotype i ; and N represents the total number of ECM morphology types.
We used the classification of Agerer (2001) and the Information System for Characterization and Determination of Ectomycorrhizae (DEEMY) database (http://www.deemy.de/) to assess the nutrient uptake strategies of ECM roots through exploration types. The ECM exploration types associated with A. faxonianawere categorized into contact (CE), short-distance (SDE) and medium-distance exploration (MDE) by the morphology types of ECM roots photographed with a stereo microscope. The CE type is described by the ECM roots with a smooth mantle and only a few emanating hyphae of negligible length, SDE by the ECM roots with a voluminous envelope of emanating hyphae of 0 – 1 mm in length but no rhizomorphs, and MDE by the ECM roots formed with rhizomorphs of 0.1 – 1 cm emanates. The frequency of ECM occurrence in each exploration type was calculated as the number of root tips of the specific type over the total root tip number.