[show abstract][hide abstract] ABSTRACT: The xanthene dyes sulforhordamine G, phloxine B, rose Bengal, and 4,5,6,7-tetrachloroflorescein were used as fluorochromes for laser scanning confocal microscopy of LR-White resin-embedded mycorrhizae. Sulforhodamine G was the most effective dye, giving an even staining of cell components throughout the material, with minimal background fluorescence of LR-White resin. Confocal microscopy of stained blocks of tissue on a slide, viewed without the use of a coverslip, revealed the three-dimensional nature of various mycorrhizal structures; these structures included arbuscules, vesicles, and coiled hyphae in arbuscular mycorrhizae; coiled hyphae in orchid mycorrhizae; mantle and Hartig net hyphae in ectomycorrhizae; and intracellular hyphae in arbutoid mycorrhizae. Sections mounted on slides viewed with confocal microscopy provided exceptional clarity of fungal form and cytoplasmic contents and showed the relationship to the plant cells, also with negligible background fluorescence. Mounting and staining blocks of resin-embedded material provided a fast and effective technique for the visualization of a variety of plant and fungal tissues. Stain penetration in whole-mounted samples was sufficient to reconstruct clear three-dimensional images using confocal microscopy.Key words: mycorrhizae, xanthene dyes, confocal microscopy, resin embedding.
Canadian Journal of Botany 02/2011; 76(1):174-178. · 1.40 Impact Factor
[show abstract][hide abstract] ABSTRACT: Ectomycorrhizae synthesized between Alpova diplophloeus and Alnus rubra are of two morphological types: one with a mantle formed along the entire length of the lateral roots and the other, the clavate type, with the mantle confined to the apical portion of the laterals. The morphology of the mycorrhiza is dependent on the stage of lateral root elongation at the time of colonization by fungal hyphae. Clavate mycorrhizae form on lateral roots that have already elongated at the time of fungal colonization. Fungal hyphae interact with root hairs at the base of clavate mycorrhizae. Mantles of both types are fairly compact with few extramatrical hyphae. Hartig net hyphae, which branch profusely primarily in the radial direction, are confined to the epidermis and midway along the radial walls of the outer layer of cortical cells. Second-order lateral root primordia are initiated in the mature Hartig net zone. Cells in the outer layer of the cortex of mycorrhizal roots collapse during fixation, indicating the possible presence of a barrier in the cell wall blocking the ingress of fixative.
Canadian Journal of Botany 02/2011; 67(1):191-200. · 1.40 Impact Factor
[show abstract][hide abstract] ABSTRACT: Cenococcum geophilum sclerotium morphology, germination, and formation in pure culture and growth pouches is described. On agar, germination of most field-collected sclerotia occurred within 30 days. In vitro attempts to initiate sclerotia succeeeded to varying degrees for 20 of the 22 isolates (collected in three main habitats in Oregon) in agar or peat – vermiculite but not in liquid culture. Sclerotium formation was initiated in 1-month-old cultures by branching, lateral growth, and subsequent septation of hyphae. Surface cells of compact hyphae of young sclerotia elongated, became irregularly arranged, and melanized. In growth pouches, sclerotium initials developed on the paper wick from a hyphal mat formed from convoluted hyphae. Mature sclerotia appeared as small spherical structures with linear hyphae radiating out from the surface. Sections revealed a well-differentiated rind and a medulla with pseudoparenchymatous organization. Numerous nuclei and vacuoles were distributed in both rind and medullary hyphae. Key words: Cenococcum, sclerotium, initiation, in vitro.
Canadian Journal of Botany 02/2011; 70(1):125-132. · 1.40 Impact Factor
[show abstract][hide abstract] ABSTRACT: All members of the Monotropoideae (Ericaceae), including the species, Allotropa virgata and Pleuricospora fimbriolata, are mycoheterotrophs dependent on associated symbiotic fungi and autotrophic plants for their carbon needs. Although the fungal symbionts have been identified for A. virgata and P. fimbriolata, structural details of the fungal-root interactions are lacking. The objective of this study was, therefore, to determine the structural features of these plant root-fungus associations. Root systems of these two species did not develop dense clusters of mycorrhizal roots typical of some monotropoid species, but rather, the underground system was composed of elongated rhizomes with first- and second-order mycorrhizal adventitious roots. Both species developed mantle features typical of monotropoid mycorrhizas, although for A. virgata, mantle development was intermittent along the length of each root. Hartig net hyphae were restricted to the host epidermal cell layer, and fungal pegs formed either along the tangential walls (P. fimbriolata) or radial walls (A. virgata) of epidermal cells. Plant-derived wall ingrowths were associated with each fungal peg, and these resembled transfer cells found in other systems. Although the diffuse nature of the roots of these two plants differs from some members in the Monotropoideae, the structural features place them along with other members of the Monotropoideae in the "monotropoid" category of mycorrhizas.
[show abstract][hide abstract] ABSTRACT: Most species in the family Cistaceae are found in the Mediterranean basin. Several hosts are of special interest, owing to their associations with truffle species, while many are important as pioneer plants in disturbed areas and in soil stabilization. For these reasons, understanding their root systems and their associated fungal symbionts is important. Most studies of the structure of mycorrhizas in this family involve two genera, Cistus and Helianthemum. The present study examines structural features of mycorrhizas in two North American species, Hudsonia ericoides L. and Hudsonia tomentosa Nutt. Root systems of both species are highly branched with most fine roots colonized by mycorrhizal fungi. Based on morphological features, several mycorrhizal fungi were identified; structural details also provided evidence of more than one fungal symbiont for each host species. All mycorrhizas had a multi-layered fungal mantle and Hartig net hyphae confined to radially elongated epidermal cells; no intracellular hyphae were observed. Although the Hartig net was confined to the epidermis, the outer row of cortical cell walls lacked suberin, a known barrier to fungal penetration. Mycorrhizas in H. ericoides and H. tomentosa differed from those of Cistus and Helianthemum species that have a Hartig net that extends into the root cortex, as well as frequently present intracellular hyphae.
[show abstract][hide abstract] ABSTRACT: Achlorophyllous plants that are dependent on an association with fungi linked to photosynthetic plants for their carbon source are known as mycoheterotrophs. Arachnitis uniflora Phil., a monotypic member of the monocotyledonous family Corsiaceae, fits this category, as it relies on a glomalean fungus belonging to Glomus Group A for carbon acquisi-tion. Although basic structural features of root colonization have been reported for A. uniflora, the nutrient exchange inter-face has not been studied. This is the first study to use confocal microscopy, transmission electron microscopy, and cytochemical procedures to study the interface between a glomalean fungus and the roots of a mycoheterotrophic species. Results showed that arbuscules are never formed, and that the ''vesicles in bundles'' reported earlier are unlike typical glo-malean vesicles, in that they form in clusters by the enlargement of hyphal branches and have a complex multilayered wall. The thick inner wall layer consists primarily of b-1,3-glucans (callose) and is surrounded by a thin outer layer of chi-tin. Each structure is surrounded by host cell wall material and a perifungal membrane, suggesting an involvement in nu-trient exchange. The cytoplasm contains a complex of small b-1,3-glucan-containing vacuoles, lipid bodies, endobacteria, and many nuclei. These structures enlarge to occupy most of the cortical cell volume and then degrade, releasing lipids and other materials into the host cell. We suggest that these structures should not be equated with typical glomalean vesicles but are unique structures that may be involved, along with the hyphal coils, in nutrient acquisition by the host. Résumé : On appelle mycohétérotrophes les plantes sans chlorophylle dépendant d'une association avec des champignons liés à des plantes photosynthétiques pour leur source de carbone. L'Arachnitis uniflora, un membre monotypique de la fa-mille monocotylée des Corsiaceae, correspond à cette catégorie et fait intervenir des champignons glomaléens appartenant aux Glomus du Groupe A pour l'acquisition du carbone. Bien qu'on ait rapporté des caractéristiques structurales de base de la colonisation racinaire chez l'A. uniflora, aucune étude n'a été conduite sur l'interface de l'échange des nutriments. Il s'agit du premier travail effectué à l'aide de la microscopie confocale, de la microscopie électronique à balayage et de mé-thodes cytochimiques, pour étudier l'interface entre un champignon glomaléen et les racines d'une espèce hétéromyco-trophe. Les résultats montrent que les arbuscules ne se forment jamais et les vésicules en faisceaux (vesicles in bundles) préalablement rapportées ne ressemblent pas à des vésicules glomaléennes typiques, en ce qu'elles se forment en amas par l'agrandissement de ramifications d'hyphes, et comportent un paroi complexe à plusieurs couches. La paroi interne épaisse est constituée surtout de b-1,3 glucans (callose) et est entourée par une mince couche de chitine externe. Du matériel parié-tal de la cellule hôte entoure chaque structure et l'on observe une membrane périfongique suggérant une implication dans l'échange de nutriments. Le cytoplasme montre un complexe de petites vacuoles contenant des b-1,3 glucans, des corps li-pidiques, des endobactéries ainsi que plusieurs noyaux. Ces structures s'agrandissent jusqu'à occuper la majeure partie du volume de cellules corticales, avant de se dégrader en relâchant des lipides et autres matériaux dans la cellule hôte. Les auteurs suggèrent que ces structures ne devraient pas être assimilées aux vésicules glomaléennes typiques, mais constitue-raient plutôt des structures uniques pouvant être impliquées, avec les pelotons mycéliens, dans l'acquisition des nutriments par l'hôte.
[show abstract][hide abstract] ABSTRACT: Arbuscular mycorrhizal fungi, obligate symbionts of most plant species, are able to accumulate heavy metals, thereby, protecting plants from metal toxicity. In this study, the ultrastructural localization of Zn, Cu, and Cd in the extraradical mycelium and spores of the arbuscular mycorrhizal fungus Glomus intraradices grown in monoxenic cultures was investigated. Zinc, Cu, or Cd was applied to the extraradical mycelium to final concentrations of 7.5, 5.0, or 0.45 mmol/L, respectively. Samples were collected at time 0, 8 h, and 7 days after metal application and were prepared for rapid freezing and freeze substitution. Metal content in different subcellular locations (wall, cytoplasm, and vacuoles), both in hyphae and spores, was determined by energy-dispersive X-ray spectroscopy. In all treatments and fungal structures analysed, heavy metals accumulated mainly in the fungal cell wall and in the vacuoles, while minor changes in metal concentrations were detected in the cytoplasm. Incorporation of Zn into the fungus occurred during the first 8 h after metal addition with no subsequent accumulation. On the other hand, Cu steadily accumulated in the spore vacuoles over time, whereas Cd steadily accumulated in the hyphal vacuoles. These results suggest that binding of metals to the cell walls and compartmentalization in vacuoles may be essential mechanisms for metal detoxification.
Canadian Journal of Microbiology 03/2008; 54(2):103-10. · 1.20 Impact Factor
[show abstract][hide abstract] ABSTRACT: Genera in the tribe Pyroleae (subfamily Monotropoideae, family Ericaceae) occur as understory plants in northern temperate zones where some form major components of ecosystems. Most have been poorly studied in terms of their association with symbiotic fungi. In this study, colonization patterns of mycorrhizal roots of five members of the Pyroleae (Pyrola asarifolia Michx., Pyrola chlorantha Sw., Orthilia secunda (L.) House, Chimaphila umbellata (L.) W. Bart., Moneses uniflora (L.) Gray) were explored. Root samples were processed for light, fluorescence, and laser scanning confocal, scanning electron, and transmission electron microscopy, as well as for immunocytochemistry. Roots of all species had enlarged epidermal cells containing hyphal complexes, Hartig nets confined to the epidermis, and mantles. Epidermal cells were penetrated by hyphae originating from the Hartig net at more than one site either along the inner tangential wall or radial walls. The outer tangential wall of epidermal cells of all species, except M. uniflora, was thicker than radial and inner tangential walls and consisted of two layers, the outer containing nonesterified pectins that were labeled with JIM 5 antibodies. Radial walls and inner tangential walls did not label, but cortical cell walls did. Intracellular hyphal complexes developed initially around centrally positioned, enlarged epidermal cell nuclei and, through branching, occupied most of the cell volume. Senescence and degradation of the complexes followed. The fungal species in these symbiotic associations may be important functionally in nutrient exchange, as well as in contributing to broader linkages with other hosts in these plant communities. Résumé : On retrouve les genres de la tribu des Pyroleae (sous-famille Monotropoideae, famille Ericaceae) comme plantes de sous-bois dans les zones tempérées boréales, où certaines forment une composante majeure des écosystèmes. Dans la plupart des cas, on a peu étudié ces plantes quant à leurs associations avec des champignons symbiotiques. Les auteurs ont étudié les patrons de colonisation des racines mycorhizées chez cinq membres des Pyrolae (Pyrola asarifolia Michx., Py-rola chlorantha Sw., Orthilia secunda (L.) House, Chimaphila umbellata (L.) W. Bart., Moneses uniflora (L.) Gray). Ils ont préparé les échantillons racinaires pour la microscopie photonique, en fluorescence, électronique par balayage et par transmission, et confocale par balayage au laser, ainsi que pour l'immunocytochimie. Les racines de toutes les espèces montrent des cellules épidermiques élargies contenant des complexes d'hyphes, des réseaux de Hartig confinés à l'épiderme, et des manteaux. Des hyphes provenant du réseau de Hartig pénètrent les cellules épidermiques à plus d'un site, le long des parois tangentielles internes ou radiales. Chez toutes les espèces, sauf chez le M. uniflora, la paroi tangentielle externe des cellules épidermiques est plus épaisse que les parois radiales et tangentielles internes et consiste de deux couches, la plus externe contenant des pectines non-estérifiées qui réagissent aux anticorps JIM 5. Les parois radiales et tangentielles internes ne réagissent pas au marquage, alors que les parois des cellules corticales le font. Les complexes d'hyphes intracellulaires se développent d'abord autour des noyaux en position centrale des cellules épidermiques élargies, et par ramification occupent ensuite la majeure partie du volume cellulaire. Par la suite, on observe la sénescence et la dégradation des complexes. Les espèces fongiques de ces associations symbiotiques peuvent jouer un rôle important dans les échanges nutritionnels ainsi que contribuer à des connexions plus étendues avec d'autres hôtes dans ces communautés végétales.
[show abstract][hide abstract] ABSTRACT: Pityopus californicus (Eastw.) H. F. Copel., a monotypic member of the Monotropoideae in the family Ericaceae, is a myco-heterotrophic species with distribution limited to the Pacific Northwest of the USA. Young embryos of P. californicus developed mycorrhizal associations in seed packets that had been buried for up to 681 days, suggesting that seeds of P. californicus may require the presence of a fungus to achieve germination. Samples of nongerminated seeds and early stages in embryo and root development were subsequently processed for light microscopy, histochemistry, and transmission electron microscopy (TEM). Nongerminated seeds possessed a thick testa, lacked a shoot and root meristem, and consisted of an embryo with large parenchymatous cells containing protein bodies and starch grains as storage reserves. In the earliest developmental stage (seed coat still attached), fungal hyphae were present on the testa surface and between the testa and embryo. This stage was followed by embryo elongation, the organization of a root apical meristem, and the development of a well-developed fungal mantle surrounding the elongated embryo. At least two morphotypes were identified based on structural characteristics of the mantle. One of these, with ascomycetous septa, had Cenococcum-like features. Late-stage embryo/early root development revealed a typical mantle and Hartig net, with fungal pegs penetrating the outer tangential walls of epidermal cells. Transfer cell-like deposits of wall material, similar to those described in Monotropa spp., enclosed fungal pegs. The development of a Hartig net and fungal pegs suggests that nutrient exchange interfaces are required for seedling development.
[show abstract][hide abstract] ABSTRACT: Arachnitis uniflora, a myco-heterotrophic plant species, has fleshy tuberous roots colonized by the arbuscular mycorrhizal fungal genus Glomus (Phylum Glomeromycota). These roots produce apical and lateral propagules, both reported here for the first time. The objective of the study was to characterize the ontogeny and structure of the propagules, and to determine their function. Scanning electron microscopy, laser scanning confocal microscopy and light microscopy were used to study the ontogeny and structure of the propagules. Propagules developed either from cortical parenchyma cells or from cells immediately beneath the root cap; they developed a shoot meristem and cells in the basal region which were colonized by various fungal structures including hyphae and vesicles. These propagules may detach from the roots, establishing new plants.
New Phytologist 02/2006; 169(1):191-7. · 6.74 Impact Factor
[show abstract][hide abstract] ABSTRACT: The extraradical mycelium of ectomycorrhizas is comprised of a network of hyphae that may initiate rhizomorphs, sclerotia and sexual reproductive structures. The development of these structures requires photosynthates produced by host trees. In this study, the initiation and early development of Laccaria bicolor (Maire) Orton fruitbodies (basidiocarps) were studied. Seedlings of Pinus resinosa Ait. and Pinus sylvestris L. were colonized by L. bicolor, a broad host epigeous ectomycorrhizal basidiomycetous fungal species, in growth pouches. Ectomycorrhizas with an extensive extraradical mycelium formed on short roots within 7-12 days after fungal inoculum was introduced. Numerous sites of hyphal aggregation, many of which subsequently developed into basidiocarp primordia, were initiated in the extraradical mycelium. Initial changes in aggregating hyphae included swelling and branching followed by growth of hyphae perpendicular to the paper wick in growth pouches. A stipe and a pileus primordium developed but none of these basidiocarp primordia matured. Distinct regions were evident in the stipe and pileus when sections were stained by various methods. Loose hyphae at the apex and periphery of the pileus were separated by mucilage. In Petri dishes with vermiculite as the substrate, basidiocarps of Laccaria laccata formed in ectomycorrhizal associations with Pinus sylvestris, developed a mature pileus with an hymenium and mature basidiospores.
[show abstract][hide abstract] ABSTRACT: The annual developmental cycle of tuberous roots of Ranunculus asiaticus was studied with respect to structure and content of their cells, to understand how these roots are adapted to desiccation, high temperature and rehydration. Light microscopy, histochemical analysis, and protein analyses by SDS-PAGE were employed at eight stages of annual root development. During growth and maturation of the roots, cortical cells increased in size and their cell walls accumulated pectin materials in a distinct layer to the inside of the primary walls, with pits between adjoining cells. The number of starch granules and protein bodies also increased within the cells. Several discrete proteins accumulated. Following quiescence and rehydration of the roots there was a loss of starch and proteins from the cells, and cell walls decreased in thickness. The resurrection geophyte R. asiaticus possesses desiccation-tolerant annual roots. They store carbon and nitrogen reserves within their cells, and pectin within the walls to support growth of the plant following summer quiescence and rehydration.
New Phytologist 05/2005; 166(1):193-204. · 6.74 Impact Factor
[show abstract][hide abstract] ABSTRACT: Species in the subfamily Monotropoideae (family Ericaceae) are achlorophyllous and myco-heterotrophic. They have become highly specialized in that each plant species is associated with a limited number of fungal species which in turn are linked to autotrophic plants. This study provides an updated and comprehensive examination of the anatomical features of two species that have recently received attention with respect to their host-fungal specificity. Root systems of Monotropa uniflora and Pterospora andromedea collected from the field were characterized by light microscopy and scanning electron microscopy. All roots of both species were associated with fungi, each root having a well-developed mantle, paraepidermal Hartig net, and intracellular "fungal pegs" within epidermal cells. The mantle of M. uniflora was multi-layered and numerous outer mantle hyphae developed into cystidia of two distinct morphologies. Large calcium oxalate crystals were present, primarily on the mantle surface. The outer mantle of P. andromedea was more loosely organized, lacked cystidia, and had smaller plate-like as well as cylindrical crystals on the surface and between outer mantle hyphae. Fungal pegs in M. uniflora originated from inner mantle hyphae that penetrated the outer tangential wall of epidermal cells; in P. andromedea, these structures were initiated either from inner mantle hyphae or Hartig net hyphae and penetrated radial walls of epidermal cells. With respect to function, fungal pegs occurred frequently in both host species and, although presumed to be the sites of active nutrient exchange, no direct evidence exists to support this. Differences between these two monotropoid hosts, resulting from the mycorrhizal fungi with which each associates, are discussed.
[show abstract][hide abstract] ABSTRACT: A combination of light microscopy (including differential interference contrast) and laser scanning confocal microscopy was used to document the colonization patterns of epidermal cells and details of intracellular hyphal complexes of five native ericaceous hosts: Vaccinium oxycoccos L. (bog cranberry), Ledum groenlandicum Oeder. (Labrador tea), Vaccinium myrtilloides L. (velvet-leaf blueberry), Kalmia angustifolia L. (sheep laurel), and Gaultheria procumbens L. (wintergreen). Colonization patterns, hyphal complex morphology, and the structure of thick-walled epidermal cells varied considerably among hosts. Multiple hyphal connections were observed between adjacent epidermal cells, indicating that one fungal entry point may result in the colonization of more than one epidermal cell. Further field observations combined with fungal isolations from field-collected plants, identification, and reinoculation studies of other species in the large Ericaceae family are required to determine the full range of structural details in ericoid mycorrhizas.
[show abstract][hide abstract] ABSTRACT: Needles of Pinus strobus (white pine) were cleared and stained to survey the occurrence and location of Lophodermium sp., a fungal endophyte. Cytoplasmically dense endophytic hyphae with a pronounced lobed morphology and containing lipid bodies were localized intercellularly between the epidermis and hypodermis. These fungal infections did not appear quiescent, but rather exhibited signs of continual slow growth. A few associated host cells exhibited a hypersensitive response. Material embedded in resin and examined by light microscopy and transmission electron microscopy confirmed the location of hyphae between epidermal and hypodermal cells, and the presence of lipid bodies within the hyphae. In senescing needles, aggressive colonization of needle tissues occurred. Thus, for Lophodermium in white pine, endophytic infection is active rather than quiescent, and displays an alternate hyphal strategy to that seen in the reproductive phase.
Mycological Research 01/2001; 105(8):991-997. · 2.81 Impact Factor
[show abstract][hide abstract] ABSTRACT: The morphology and anatomy of ectomycorrhizas of Rhizopogon parksii, Rhizopogon vinicolor and Rhizopogon subcaerulescens, and a hypogeous relative, Truncocolumella citrina, synthesized on Douglas fir in glasshouse conditions using spore slurries as inoculum, are described and compared. Mycorrhizas formed with R. parksii and R. vinicolor did not exhibit their characteristic subtuberculate morphology in these tests, but rather had a pinnate form. All species had diagnostic features of ectomycorrhizas: a well-developed Hartig net and a fungal mantle. In addition, several species exhibited crystal inclusions in the outer mantle, usually at the interface between the mantle and soil. Truncocolumella citrina had crystal-like inclusions within the mantle but external to fungal hyphae, a feature rarely described in ectomycorrhizas.
New Phytologist 07/2000; 147(2):389 - 400. · 6.74 Impact Factor
[show abstract][hide abstract] ABSTRACT: The morphology and anatomy of ectomycorrhizas of Rhizopogon arctostaphyli, R. ellenae, R. flavofibrillosus, R. occidentalis, R. rubescens, R. smithii, R. subcaerulescens and R. truncatus synthesized on Ponderosa pine (Pinus ponderosa) in glasshouse conditions using spore slurries, are described and compared. All species produced a well-developed Hartig net, and a well-developed fungal mantle. The mantles of R. arctostaphyli, R. smithii and R. subcaerulescens ectomycorrhizas were two-layered with outer mantle hyphae of wider diameter than inner mantle hyphae. The mantle of R. subcaerulescens ectomycorrhizas also had distinctive peg-like structures (cystidia) along peripheral hyphae. Rhizopogon truncatus ectomycorrhizas were tuberculate in morphology and had a rind-like mantle enclosing adjacent roots. In addition, several species exhibited crystal inclusions in the outer mantle, presumably at the interface between mantle and soil.
New Phytologist 04/1999; 142(2):355 - 370. · 6.74 Impact Factor
[show abstract][hide abstract] ABSTRACT: Mycorrhiza ontogeny and details of Hartig net and mantle structure were compared in ectomycorrhizas synthesized in growth
pouches between the broad host range fungus Paxillus involutus and the tree species European black alder (Alnus glutinosa) and red pine (Pinus resinosa). In Alnus glutinosa, a paraepidermal Hartig net was restricted to the proximal (basal) portion of first-order laterals; the hypodermal layer
appeared to be a barrier to fungal penetration. Phi-thickenings were present in some cortical cells but these were not related
to lack of fungal ingress into the cortex. The mantle was often present close to the root apex but in many roots it was loosely
organized and patchy. In several instances, the mantle formed around the root apex was only temporary; renewed root growth
occurred without the formation of a mantle. In Pinus resinosa, the Hartig net developed between cortical cell layers of monopodial and dichotomously branched first–order laterals. Fungal
hyphae in the Hartig net exhibited a complex labyrinthine mode of growth. The mantle had a pseudoparenchymatous structure
and covered the root, including apices of dichotomously branched roots. The Paxillus–Pinus resinosa interaction had all the characteristics of a compatible ectomycorrhizal association. The Paxillus–Alnus glutinosa interaction, however, showed only aspects of superficial ectomycorrhizas, including the presence of a minimal (sometimes
absent) and mostly proximal Hartig net and variable mantle development. Sclerotia were produced in the extraradical mycelium
of Paxillus involutus when associated with either Alnus glutinosa or Pinus resinosa.