In the botanical expeditions throughout Greece, 15 samples of tulip roots were collected by the Institute of Plant Breeding and Genetic Resources, which were placed in containers with alcohol and were transferred for analysis to Soil Laboratory, School of Agriculture, Aristotle University of Thessaloniki, Greece.

Root samples were placed in Petri dishes to remove soil residue with tweezers and on absorbent paper to evaporate the alcohol. Up to 0.1 g of up to 10 individuals of the same species per region were used for DNA extraction. In the case of small quantities, complex samples were created. The remaining volume was placed in centrifuge tubes for staining and counting mycorrhizal colonization under a microscope.

As the number of individuals in which colonization was counted varies according to the amount of available root after weighing/storage for DNA extraction, complex samples were created on a case-by-case basis. Spores were extracted from the rhizosphere for counting and homogeneous groups of spores per morphotype were kept in slides for microscopic observation (see representative photos below). Also, the rhizosphere soil was applied as a vaccine in potters with 2 kg of substrate (sterile mixture 1: 1 sand: vermiculite) with alfalfa, pentaneur and corn as plant-traps for vaccine propagation and spore isolation.

Photos of isolated morphotypes of spores from the rhizospheres of Greek tulip species.
Photos of mycorrhizal colonization on roots of Greek tulip species: A: Cysts from sample T1, B: Abscess – point of entry from sample T21, Γ: Tufts from sample T22, Δ: very thin textures from sample T39.
Photos from non-mycorrhizal colonization or simultaneous colonization by mycorrhizal and non-mycorrhizal fungi on the roots of Greek tulip species. A: Black textures and tassels from sample T22. B: Sclerotia from sample T1, Γ: Sclerotia from sample T39, Δ and E: Black textures inside and outside the root and H-type ligaments from samples T2 and T22, ΣΤ and Z: Cysts without textures.