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.