Tuberculate mycorrhizas of Castanopsis borneensis King and Engelhardtia roxburghiana Wall

Transcription

1 New Phytol. (1991), 117, Tuberculate mycorrhizas of Castanopsis borneensis King and Engelhardtia roxburghiana Wall BY I. HAUG\ R. WEBER\ F. OBERWINKLERi ^ND J. TSCHEN^ ^Universitdt Tubingen, Spezielle Botanik, Mykologie; Auf der Morgenstelle 1, 7400 Tubingen, West Germany '^National Chung Hsing University, Department of Botany, Taichung 40227, Taiwan {Received 23 April 1990 ; accepted 25 September 1990) SUMMARY Tuberculate mycorrhizas of Castanopsis borneensis King (Fagaceae) and Engelhardtia roxburghiana Wall, (Juglandaceae) were studied by light and electron microscopy. The tubercles of both trees showed similar structure and morphology. The fungal partner is an unidentified basidiomycete. Beside normally developed mycorrhizas in each tubercle some heavily-infected roots were found. These roots are characterized by a very thick hyphal mantle around a dead root tip and a circle-like ingrowth of the fungus up to the vascular tissue - the strangulation zone. All cells distal to the strangulation zone were dead, but basipetal to it, normal cortical tissue with an epidermal Hartig net was present. The outer cortex in this region showed wall ingrowths along the inner tangential walls and an accumulation of ER and mitochondria. The formation and function of the heavily-infected roots is discussed. Key words: Castanopsis borneensis, Engelhardtia roxburghiana, tuberculate mycorrhizas, ultrastructure, Taiwan, INTRODUCTION Tuberculate mycorrhizas consist of several densely branched mycorrhizal systems enveloped in a common hyphal sheath, McDougall (1922) and Melin (1923) described tuberculate mycorrhizas of Pinus sp, and Masui (1926) those of Quercus pausidentata Fr, in Japan, More recently tuberculate mycorrhizas have been reported from Pseudotsuga sp. (Dominik, 1963 ; Dominik & Majchrowicz, 1967; Trappe, 1965 ; Zak, 1971), from Tsuga mertensiana (Bong.) Carr, (Zak, 1973), from Photinia glabra (Thurb,) Maxim (Grand, 1971) and from Eucalyptus pilularis Sm. (Dell, Malajczuk & Thomson, 1990). A peculiar feature of the 'form A tuberculate mycorrhiza' described in Quercus pausidentata by Masui (1926) was the occurrence of 'heavily-infected' roots beside normal mycorrhizas. The same type of tuberculate mycorrhiza and the same 'heavily-infected' roots have been found on Castanopsis borneensis King (Fagaceae) and Engelhardtia roxburghiana Wall. (Juglandaceae) in Taiwan, To supplement the light microscopic studies of Masui (1926) some results from electron microscopy are presented in this paper. MATERIALS AND METHODS Roots were sampled in November 1988 in the Hui Sun Forest District (Mid-Taiwan, about 50 km east of Taichung, c. 800 m above sea level). The mixed subtropical forest consists of approximately 300 species belonging to many difterent families, Tuberculate mycorrhizas were found on C, borneensis and E. roxburghiana. The mycorrhizas were cleaned with a paint-brush and fixed in 2 % glutaraldehyde in 0-2 M cacodylate buffer (ph 7-2) for 10 week, postfixed with osmium tetroxide (1 % OsO^ in 0-1 M arsenic buffer, 2 h) and en bloc contrasted with uranyl acetate (1%, 1 h), Spurr's (1969) ERL was used as the embedding medium. Four infiltration steps were carried out: ERL: acetone in the proportions 1:2, 1:1, 2:1 each for \ h, followed by ERL for 3 d at 23 C, Polymerization took place at 70 C for 3 d. Semi-thin longitudinal sections (0-5-2 /^m) were stained with neofuchsin-crystal violet, Ultrathin sections ( nm) were contrasted with lead citrate.

2 26 /. Haug, R. Weber, F. Oberwinkler and J. Tschen ' t, FT ' - " \ ' 1 V -T Figures 1 3. Morphology and structure of tubercles of Castanopsis borneensis.

3 A. tuberculate mycorrhiza of Castanopsis and Engelhardtia 27 RESULTS General morphology of the tubercles The tubercles found on C. borneensis and E. roxburghiana showed a very similar morphology and structure. Differences concerning the root morphology of the two trees are described below. The description of the tubercles is combined for both trees. The tubercles are spherical to ovoid bodies 4-10 mm in diameter attached to a mother root (Fig. 1). In young stages they are white, turning brownish with age. The tubercles are enclosed by a rind of fungal tissue, which has a thickness of about 60 fiva (Figs 2, 3). The brown rind disintegrates in older tubercles, when mycorrhizas are dead. Rhizomorphs radiate from the tubercle surface. Fig. 2 and Fig. 3 show the internal structure of a tubercle consisting of numerous entangled mycorrhizas. The hyphal mantles are of loose to compact prosenchyma /im thick (Fig. 4). Individual roots within a tubercle may possess a hyphal mantle in common (Fig. 5), or may be separated by a recognizable space (Fig. 6). In the C. borneensis mycorrhizas the Hartig net is established only between the radially elongated epidermal cells (Fig. 7), in contrast to the mycorrhizas of E. roxburghiana where the Hartig net extends 1-2 cortical cell layers deep (Figs 8, 9). The morphology and structure of normal mycorrhizas in the tubercle is identical to single mycorrhizas occurring on both trees. The hyphae have dolipores with perforated parenthesomes in both associations (Fig. 14). The heavily-infected roots In each tubercle some mycorrhizas have dead tips (Figs 15, 16) and are embedded in an unusually large amount of mycelium (Figs 10-13) so that the hyphal mantle is up to 130/^m in width (Fig. 10). Immediately behind the apex the cortical cells are lacking and are replaced by numerous centripetallygrowing tightly-packed hyphae (Figs 12, 13). This area of the heavily-infected roots is called the 'strangulation zone'. The hyphae of the strangulation zone contain several nuclei (Fig. 18), a lot of lipid-like droplets (Figs 18, 21), many mitochondria (Figs 18, 19) and multivesicular bodies (Figs 19, 20). The multivesicular bodies are accumulated at the tips of the hyphae bordering upon the vascular tissue (Fig. 19). Some pictures may be interpreted as if the vesicles are released to the hyphal wall (Fig. 20). With exception of the multivesicular bodies the hyphae of the strangulation zone show no difference to the Hartig net hyphae. In the strangulation zone a few cells of the outer vascular tissue are found with intracellular hyphae (Fig. 21), but nowhere else - even towards the root tip where all cells are dead - does penetration of the hyphae into the cells occur (Figs 11, 16). There was no evidence of host cell wall reaction. The hyphae form a dense layer next to the dead root tissues and possess dense cytoplasm and small vacuoles (Fig. 16). In an advanced stage the hyphae at the root tip have thick cell walls and no cytoplasmic contents (Fig. 25). There is a clear boundary between living and dead root tissue in the strangulation zone. The cells extending from the strangulation zone up to the tip are dead (Figs 12, 13, 15, 16), the vascular tissue and the cortical cells behind the strangulation zone are alive (Figs 12, 13, 23). Basipetally from the strangulation zone the cortical tissue is normally developed with an epidermal Hartig net (Figs 12, 23). The epidermal cells are highly vacuolated or dead while the Hartig net hyphae have dense cytoplasm (Figs 22, 23). In the Castanopsis mycorrhizas the outer cortex (= hypodermis) bordering on the epidermis with the Hartig net consists of tangentially elongated cells, which are fully plasmatic. Prominent wall ingrowths are found along the inner tangential walls of the outer cortical cells ofthe heavily-infected roots (Figs 23, 24). A large amount of ER extends in parallel layers between the protuberances (Figs 23, 24) and many mitochondria are found in this region (Figs 23, 24). Between the outer cortical cells and the endodermis is one layer of cortical cells with much thickened inner walls (Fig. 23). These structural features of heavily-infected roots are summarized in Fig. 26. In one section a possible intermediate stage of infection was found. The root on the left in Fig. 10 (next to a heavily-infected root) shows normally developed cortex on the left with an epidermal Hartig net, but on the right the cortex is colonized totally by the fungus (Fig. 10,*). At the interface between fungal tissue and cortical tissue electron microscopic studies revealed remains of some cor- Abbreviations Figures B, multivesicular body; C, cortical cell; D, lipid-like droplets; E, epidermis; DA, dead apex; DAT, dead apical tissue; ER, endoplasmic reticulum; FT, rind of fungal tissue; H, hyphae; HM, hyphal mantle; HN, Hartig net; HW, hyphal wall; IC, inner cortex; M, mitochondrium; MY, mycorrhiza; N, nucleus; OC, outer cortex; R, rhizomorph; S, strangulation zone; V, vascular tissue. Figure 1. External appearance of a tubercle attached to a mother root and with radiating rhizomorphs (scale 1 mm). Figure 2. Internal appearance of a tuhercle with numerous entangled mycorrhizas and a rind of fugal tissue (scale 0-5 mm). Figure 3. Detail of a tuhercle: rind of fungal tissue and several mycorrhizas (scale 100/im).

4 28 /. Haug, R. Weber, F. Oberwinkler andj. Tschen i:.:'- r-. * s.. ' ' r '. '1 Figures 4 9. Mycorrhizal structures of tuherculate ectomycorrhizas. Figure 4. Tangential section in the region of a hyphal mantle: loose to compact prosenchymatous tissue {Castanopsis borneensis, scale 20 /im). Figure 5. Densely packed mycorrhizas with common hyphal mantles ( C borneensis, scale 100/im). Figure 6, Two adjacent mycorrhizas with little space between hyphal mantles ( C borfieensis, scale 20/^m). Figure 7. Cross section of C borneensis mycorrhiza with a Hartig net only between epidermal cells (scale 20 /im). Figure 8. Cross section of Engelhardtia roxburghiana mycorrhiza with a Hartig net between several cortical cell layers (scale 100/im). Figure 9. Longitudinal section of E. roxburghiana mycorrhiza (scale 100 tical cells between the hyphae (Fig. 17) consistmr 1 ( ^u J 11 1 only of their compressed walls and containing traces of polyphenols. The hyphae do not penetrate the cortical cells. It is difficult to estimate the frequency of heavily-infected roots in the tubercles, because it is not possible to section the whole tubercle, but from the parts sectioned, there may be about 3-5 % heavily-infected mycorrhizas. DISCUSSION The root morphology of C. borneensis mycorrhizas is similar to that of other members of the Fagaceae (Brundrett, Murase & Kendrick, 1990). There is a Hartig net between elongated epidermal cells. The outer cortex ( = hypodermis) has thickened outer walls and the inner cortex thickened inner walls. In

5 A. tuberculate mycorrhiza of Castanopsis and Engelhardtia 29 v-\\\- > ; Figures Heavily-infected roots in cross sections in the region of the apex. Figure 10. Right; cross section of a dead root tip with a very thick hyphal mantle. Left; cross section of a root with a normal epidermal Hartig net and cortical tissue on the left side and an ingrowth of the fungus (*) on the right side {E. roxburghiana, scale 100/im). Figure 11. Hyphae around a dead root apex {Engelhardtia roxburghiana, scale 20/tm). the Juglandaceae the situation is not so uniform. There are genera with VA-mycorrhizas {Juglans, Brundrett et al. 1990) and genera with eetomycorrhizas - among others Carya (Brundrett et al., 1990) and Engelhardtia. Carya has an epidermal Hartig net, which is typical for most angiosperms with eetomycorrhizas (Alexander & Hogberg, 1986; Massicotte, Ackerley & Peterson, 1987; Brundrett, et al, 1990). The Hartig net of E. roxburghiana is up to 2 layers deep between irregularly arranged cortical cells and represents therefore a rarity among the angiosperms. A comparison between the structural features of the tubercles of C. borneensis and E. roxburghiana and the detailed description of the tubercles of Q. pausidentata by Masui (1926) reveals many similarities in general morphology. Both the external structure (colour, size, rhizomorphs) and the internal

6 30 /. Haug, R. Weber, F. Oberwinkler and J. Tschen ^.V:^-.^.^ ""T^/^f-^ Figures Heavily-infected roots in longitudinal section. Figure 12. Median longitudinal section of a heavily-infected root: dead apex, strangulation zone, vital root with Hartig net {Castanopsis borneensis, scale 100/tm). Figure 13. Strangulation zone (C borneensis, scale 100 Structures (rind of fungal tissue, numerous entangled mycorrhizas, heavily-infected roots) show no differences. Further the structure of the hyphal mantles and the diameter of the hyphae are comparable. All the details which are given by Masui (1926) for the heavily-infected roots were also observed in the roots investigated here. Thus it is possible that the same fungus is involved. From the ultrastructural data we can only conclude that the fungus belongs to the basidiomycetes. Eventually the fungus influences the tree roots in such a way that the roots branch abnormally to produce, in a small volume, considerable short root initiation. Another unusual feature of the fungus is the intensive growth around dead root tips and the ingrowth up to the vascular tissue such that the cortical cells are lacking. Unfortunately it was not possible to observe the development of these heavily-infected roots. Even in young tubercles fully developed heavily-infected roots w^ere present. The half infected root (Fig. 10)

7 A. tuber culate mycorrhiza of Castanopsis and Engelhardtia! ' - 31 :. ' 'r" - \»,V'-,' -v- - '» < - '.. < 1-/ - 5.' ' -...-it f'. 'V. Figures Ultrastructure. Figure 14. Dolipore of the mycorrhizal fungus {Castanopsis borneensis, scale 0-5 /im). Figure 15. Dead apical tissue (C. borneensis, scale 5 /^m). Figure 15. Hyphae bordering on the dead apex cells in cross section {Engelhardtia roxburghiana, scale 5 /im). gives the impression that the fungus is able to push the cortical tissue away. It remains unclear how the apex dies. Masui (1926) postulated that the hyphae 'suck off' substances directly from the vascular tissue and that - together with the tight constriction - the nutrient supply to the growing apex becomes insufficient and the cells lose their vitality. The crucial point of this hypothesis is that the morphology of the vascular tissue is not influenced by the fungus. None of the cells of the vascular tissue have a significantly reduced diameter. Thus it cannot be assumed that the transport to the apex is interrupted. Whether or not the hyphae are able to 'suck off' substances from the host tissue is unclear. The cells of the root apex were not killed by ingrowth of the fungus. Even when all host cells were dead no ingrowth of the fungus occurred. In other cases intracellular infection is a common feature of mycorrhizal fungi (Harley, 1963; Atkinson, 1975; Harley & Smith, 1983). The death of the cells could be caused by a reduced supply of nutrients, but toxic substances may also play a role.

8 32 /. Hang, R. Weber, F. Oberwinkler and J. Tschen [) D ' E Figures Ultrastructure of heavily-infected roots. Figure 17. Compressed cortical cell between the hyphae {Engelhardtia roxburghiana, scale 2/im), Figure 18. Several nuclei in the hyphae of the strangulation zone {Castanopsis borneensis, scale 2 /im). Figure 19. Hyphae bordering on the vascular tissue with a multivesicular body (C, borneensis, scale 2 /im). Figure 20. Possible release of vesicles to the hyphal wall (C, borneensis, scale 0-5 fim). Figure 21. Intracellular hyphae in the outermost layer of the vascular tissue at the strangulation zone (C, borneensis, scale 5 fim). Figure 22. Vital Hartig net hyphae between dead epidermal cells basipetal to the strangulation zone (C, borneensis, scale 5 /im).

9 A. tuber culate mycorrhiza of Castanopsis and Engelhardtia 33.- Figures Ultrastructure of heavily-infected roots. Figure 23. Epidermis with Hartig net, outer cortex with prominent wall ingrowths ( ) and inner cortex with thickened walls (*) basipetal to the strangulation zone {Castanopsis borneensis, scale 5 /^m). Figure 24. Detail of Eigure 23: Wall ingrowths ( ) at the inner tangential wall of the outer cortex with endoplasmic reticulum and mitochondria; inner cortex with thickened walls (*) (C. borneensis, scale 1 /tm). Figure 25. Dead thick walled hyphae (*) at the root apex (C. borneensis, scale 5 The occurrence of vv'all ingrowths and the vital Hartig net hyphae make it probable that the transfer of nutrients does not take place only in the strangulation zone ~ as was postulated by Masui (1926) - but also behind this zone vv'here perhaps an accumulation of nutrients occurs. The outer cortical cells of C. borneensis show thickened walls towards the epidermis and Vi^all ingrowths towards the vascular tissue. Wall thickenings in outer cortex cells are a common feature of angiosperms with ectomycorrhizas (Brundrett et al. 1990) and are m^ost probably responsible for preventing Hartig net hyphal growth past the epidermis (Brundrett et al., 1990). The wall ingrowths are similar to the transfer ANP 117

10 34 /. Haug, R. Weber, F. Oberwinkler andj. Tschen Hyphal mantle _ Epidermis with Hartig net Outer cortex ~" (hyperdermis) Inner cortex I Endodermis with polyphenols Vascular tissue -Endodermis Dead apex- -Strangulation zone-» Figure 26. Schematic drawing of a heavily-infected root in a median longitudinal section. cell structures described by Gunning & Pate (1969). According to the investigations of Gunning & Pate (1969) transfer cells may be found in a wide variety of anatomical structures where intensive, short distance transport occurs. The authors observed the wall ingrowths to be best developed on those faces of the cell presumed to be most active in solute transport. Numerous mitochondria and a conspicuous endoplasmic reticulum usually accompany this wall specialization. Since this wall specialization is missing in the ' normal' mycorrhizas of C. borneensis and in other members of the Fagaceae {Fagus, Quercus, Brundrett et al., 1990; Pasania, Cyclobalanopsis, Haug, unpublished) these structures may be connected with the strangulation and may give rise to enhanced symplastic transport. Symplastic transport is particularly important in the Fagaceae because, as Brundrett et al. (1990) have shown, apoplastic transport is blocked by thick inner wall sclerenchyma. Wall ingrowths have been observed within naturally grown mycorrhizas of Pisonia grandis R. Br. (Ashford & Allaway, 1982, 1985), of Monotropa hypopytis L. (Duddridge & Read, 1982), of Sarcodes sanguinea Torrey and Pterospora andromedea Nuttall (Robertson & Robertson, 1982). Monotropa, Sarcodes and Pterospora have monotropoid mycorrhizas, in which the transfer-cell wall ingrowths were only found on the fungal pegs. In all four plant species the Hartig net is restricted to the epidermis and thus the exchange area is rather restricted. Wall ingrowths were also found in several in-vitro synthesized mycorrhizas (Duddridge & Read, 1984; Kottke & Oberwinkler, 1986; Warmbrodt & Eschrich, 1985), but it is uncertain whether these structures represent a defence reaction on the part of the host against the fungus or if they are comparable to those wall ingrowths produced in transfer cells. Finally there remains the question: what is the explanation for tuberculate mycorrhiza, i.e. the inclusion of single, fully developed mycorrhizas in a compact body? It is unlikely that the uptake of nutrients is improved in this way because the numerous root tips have no direct contact with the surrounding soil. Only the rhizomorphs extend from the tubercles into the soil. Certainly rhizomorphs play an important role in water and nutrient transfer (Duddridge, Malibari & Read, 1980; Brownlee et al, 1983). Gobi (1967) found increased amounts of P and K in the tubercles of Pinus cembra L. relative to single mycorrhizas and therefore concluded that the tubercles have a storage function. A real understanding of tuberculate mycorrhizas and especially the function of heavily-infected roots must await further study. ACKNOWLEDGEMENTS We express our sincere thanks to Mrs Angela Dressel for technical assistance and for skilfully copying the photographs and Mrs Yang Chao-Rong for assistance in Hui Sun, Taiwan. These studies were supported by a grant from the Volkswagenstiftung. REEERENCES ALEXANDER, I. J. & HOGBERG, P. (1986). Ectomycorrhizae of tropical angiosperm trees. New Phytologist 102, ASHFORD, A. E. & ALLAWAY, W. G. (1982). A sheathing mycorrhiza on Pisonia grandis R. Br. (Nyctaginaceae) with development of transfer cells rather than a Hartig net. New Phytologist 90, ASHFORD, A. E. & ALLAWAY,.W. G. (1985). Transfer cells and Hartig net in the epidermis of the sheathing mycorrhiza of

11 A. tuber culate mycorrhiza of Castanopsis and Engelhardtia 35 Pisonia grandis R. Br. from Seychelles. New Phytologist ATKINSON, D. A. (1975). The fine structure of mycorrhizas. D.Phil. Thesis, Oxford. BROWNLEE, C, DUDDRIDGE, J. A., MALIBARI, A. & READ, D. J. (1983). The structure and function of mycorrhizal systems of ectomycorrhizal roots with special reference to their role in forming inter-plant connections and providing pathways for assimilate and water transport. Plant and Soil 71, BRUNDRETT, M., MURASE, G. & KENDRICK, B. (1990). Comparative anatomy of roots and mycorrhizae of common Ontario trees. Canadian Journal of Botany 68, DELL, B., MALAJCZUK, N. & THOMSON, G. T. (1990). Ectomycorrhiza formation in Eucalyptus. V. A tubercalate ectomycorrhiza of Eucalyptus pilularis. New Phytologist DoMiNiK, T. (1963). Occurrence of Douglas-fir (Pseudotsuga taxifolia Britton) in various Polish stands. Prace Instytutu Badawczego Le'snictwa 258, DOMINIK, T. & MAJCHROWICZ, I. (1967). Studies on the tuberculate mycorrhizae of Douglas fir (Pseudotsuga taxifolia Britton). Ekologia Polska Seria A, 15 (3), DUDDRIDGE, J. A., MALIBARI, A. & READ, D. J. (1980). Structure and function of mycorrhizal rhizomorphs with special reference to their role in water transport. Nature, London 1%1, DUDDRIDGE, J. A. & READ, D. J. (1982). An ultrastructural analysis of the development of mycorrhizas in Monotropa hypopitys L. Neio Phytologist 29, DUDDRIDGE, J. A. & READ, D. J. (1984). Modification ofthe hostfungus interface in mycorrhizas synthesized between Suillus bovinus (Fr.) O. Kuntz and Pinus sylvestris L. New Phytologist 96, GoBL, E. (1967). Mykorrhiza-Untersuchungen in subalpinen Waldern. Mitteilungen der Forstlichen Bundesversuchsansalt 75, GRAND, L. E. (1971). Tuberculate and Cenococcum mycorrhizae oi Photinia (Rosaceae). Mycologia 63, GUNNING, B. E. & PATE, J. S. (1969). 'Transfer cells'-plant cells with wall ingrowths specialized in relation to short distance transport of solutes their occurrence, structure and development. Protoplasma 68, HARLEY, J. L. (1936). Mycorrhiza of Fagus sylvatica D.Phil. Thesis, Oxford. H.ARLEY, J. L. & SMITH, S. E. (1983). Mycorrhizal Symbiosis. Academic Press, London, New York. KoTTKE, I. & OBERWINKLER, E. (1986). Root-fungus interactions observed on initial stages of mantle formation and Hartig net establishment in mycorrhizas of Amanita muscaria (L. ex Er.) Hooker on Picea abies (L.) Karst. in pure culture. Canadian Journal of Botany 64, MASSICOTTE H. B., ACKERLEY, C. A. & PETERSON, R. L. (1987). The root-fungus interface as an indicator of symbiont interaction in ectomycorrhizae. Canadian Journal of Forest Research 17, MASUI, K. (1926). The compound mycorrhiza of Quercus pausidentata Er. Memoirs of the College of Science, Kyoto Imperial University, Series B, Vol. 11, McDouGALL, W. B. (1922): Mycorrhizas of coniferous trees. Journal of Forestry XX, No. 3, MELIN, E. (1923). Experimentelle Untersuchungen liber die Konstitution und Okologie der Mykorrhizen von Pinus sylvestris L. und Picea abies (L.) Karst. Mykologische Untersuchungen und Berichte 1, ROBERTSON, D. C. & ROBERTSON, J. A. (1982). Ultrastructure of Pterospora andromedea Nuttall and Sarcodes sanguinea Torrey mycorrhizas. New Phytologist 92, SPURR, A. R. (1969). A low viscosity epoxy resin embedding medium for electron microscopy. Journal of Ultrastructure Research 26, 31^3. TRAPPE, J. M. (1965). Tuberculate mycorrhizae of Douglas-fir. Forest Science 11, WARMBRODT, R. D. & EscHRiCH, W. (1985). Studies on the mycorrhizas of Pinus sylvestris L. produced in vitro with the basidiomycete Suillus variegatus (Sw. ex Er.) O. Kuntze. I. Ultrastructure ofthe mycorrhizal rootlets. New Phytologist 100, ZAK, B. (1971). Characterization and classification of mycorrhizae of Douglas fir. II. Pseudotsuga menziesii + Rhizopogon vinicolor. Canadian Journal of Botany 49, ZAK, B. (1973). Classification of ectomycorrhizae. In: 'Ectomycorrhizae'. (Ed by G.C.Marks & T. T. Kozlowski), pp Academic Press, New York and London. 3-2

12