Light microscopy was used to select material with fungal coils.
Transversal sections were cut from the middle part of each
root sample by hand using a razor blade. Sections were
stained by Methyl blue 0.05 % solution (C. I. 42780, Merck) in
lactic acid for 10 min on microscopic slides. The samples
were examined in fresh lactic acid at 100- to 1000-fold magni-
fication (Leitz SM-LUX or Zeiss Axioskop 2).
Root pieces of 1 cm length of all the samples displaying
high frequency of vital looking hyphal coils, 56 in total and
at least ten of each species, were fixed in 2.5 % glutaraldehyde-formaldehyde
in Sørensen buffer (Karnovsky 1965),
post-fixed in 1 % osmium tetroxide for 1 h, dehydrated in an
acetone series and flat embedded in Spurr’s resin low viscosity,
longer pot-life formulation (Spurr 1969). Semithin sections
were cut from the embedded samples, stained with 0.6 % neofuchsin
crystal-violet, mounted in Entellan, and observed in
the light microscope. 20 samples with apparently vital
hyphae, originating from different plant individuals, were
selected for ultrathin cutting. Sections were mounted on Formvar-coated
copper grids and stained with 1 % uranyl acetate
(40 min) and lead citrate (12 min). Sections were examined using
transmission electron microscopes Zeiss TEM 902 or Zeiss
TEM109.