Wood degradation by the white-rot basidiomycete Meripilus giganteus (Pers.: Pers.) Karst. was studied in naturally infected and artificially inoculated wood of beech (Fagus sylvatica L.) and large-leaved lime (Tilia platyphyllos Scop.). Semi-thin sections revealed that the secondary walls of most fibres contained internal cavities. Three distinct types of cavity formation, which differed not only between hosts, but also between cell type and location in the annual ring, were identified.

Within discoloured wood of naturally infected beech, the structure of the cavities and their formation by the associated hyphae were reminiscent of a soft-rot. By contrast, cavity formation in artificially inoculated beech and large-leaved lime wood differed from a soft-rot mode of attack as extensive delignification always preceded cavity formation, and neither T-branching, L-bending, nor hyphal growth were found within cell walls. The formation of half-moon shaped cavities in beech wood was present only in tension-wood fibres. From large diameter hyphae, growing within the fibre lumen, numerous fine perforation hyphae extended transversely via helical cracks into the cell wall. Subsequent degradation of cellulose within concentric layers of the tension-wood fibres commenced from the apices of perforation hyphae.

Sections stained with ruthenium red and hydroxlamine-ferric chloride, revealed that M. giganteus preferentially degrades pectin-rich regions of the middle lamellae in xylary ray cells. In large-leaved lime, such regions were uniformly located in the middle lamellae of axial and ray parenchyma. In beech wood, degradation of pectin-rich middle lamellae regions commenced after the delignification of secondary walls and resulted in a conspicuous hollowing of multiseriate xylem rays. Plasticity in wood degradation modes by M. giganteus in large-leaved lime and beech wood reflects variations in cell wall structure and/or prevailing wood conditions.

Summary 37

I. INTRODUCTION 37

II. DIGESTION OF PLANTS IN THE RUMEN: OLD AND NEW CONCEPTS 39

III. RUMEN-INDUCED PLANT METABOLISM: CELL DEGRADATION AND DEATH 41

1. Summary of plant cell death processes 41

2. Anaerobic stress and flooding tolerance of plants 42

3. Plant cell responses to elevated temperatures 45

4. Wounding responses/pathogen attachment 45

5. Senescence in the rumen? 47

IV. FUTURE PROSPECTS 50

Acknowledgements 51

References 51

It is generally assumed that breakdown of plant material in the rumen is a process mediated by gut microorganisms. This view arose because of the identification of a pre-gastric fermentation in the rumen, brought about by a large and diverse microbial population. The extensive use of dried and ground feed particles in forage evaluation might have helped to promote this assumption. However, although the assumption might be correct in animals feeding on conserved forage (hay and silage) where the cells of ingested forage are dead, it is possible that with grazed (living) forage, the role played by plant enzymes in the rumen has been overlooked. In a grazing situation, plant cells that remain intact on entering the rumen are not inert, but will respond to the perceived stresses of the rumen environment for as long as they are metabolically viable. Metabolic adjustments could include anaerobic and heat-shock responses that could promote premature senescence, leading to remobilization of cell components, especially proteins. Moreover, contact of plant cells with colonizing microorganisms in the rumen might promote a type of hypersensitive response, in much the same way as it does outside the rumen. After fresh plant material enters the rumen and prior to extensive plant cell-wall degradation, there is often a phase of rapid proteolysis providing N in excess of that required to maintain the rumen microbial population. The inefficient use of this ingested N results in generation of ammonia and urea in exhaled breath and urine, which promotes welfare and environmental pollution concerns. Therefore an important research goal in livestock agriculture is to find ways of decreasing this initial rate of proteolysis in the rumen. This will benefit the farmer financially (through decreased use of feed supplements), but will also benefit the environment, as N pollution can adversely affect pasture diversity and ecology. This review considers the possible responses of plant metabolism to the rumen environment, and how such considerations could alter current thinking in ruminant agriculture.

The accumulation of fructo-oligosaccharides and the activities of fructosyltransferase (sucrose[ratio ]sucrose 1F-fructosyltransferase (SST), 1F-fructosyltransferase (1F-FT) and 6G-fructosyltransferase (6G-FT)) in the bulbs of three onion cultivars were investigated from June to September 1993. The total fructo-oligosaccharide content increased from June until August, then decreased in September, except in one cultivar. The levels of neokestose and its related tetrasaccharides (1F, 6G-di-β-D-fructofuranosyl sucrose and 6G (1-β-D-fructofuranosyl)2sucrose) were higher than those of 1-kestose and nystose throughout growth. The activities of 6G-FT, 1F -FT and SST were high in June and July, then decreased; SST activity was very low in September. The activity ratios of 6G-FT to 1F-FT varied between 1·86 and 2·65 over the growth period. Two trisaccharides, three tetrasaccharides and four pentasaccharides were identified, together with a mixture of hexa- and heptasaccharides, all of which were synthesized in vitro from 0·1 M sucrose by an enzyme preparation of onion bulbs harvested in August. Octa- and nonasaccharides other than the saccharides formed from sucrose were also synthesized from 0·1 M 1-kestose or 0·1 M neokestose. All the saccharides produced from sucrose, 1-kestose or neokestose by the crude enzyme prepared from onion bulbs were identical to the saccharides occurring naturally in onion bulbs.

The period of active growth for spring ephemeral plants coincides with the period of high light, water and nutrient availability between snow melt and canopy closure in the understorey of deciduous forests in eastern North America. However, low temperatures prevail during this period and might restrict the performance of these plants. Remarkably, this peculiar phenology is extremely rare among annual plants. To understand better the role of light and water availability and of temperature in the phenology of spring ephemeral plants, we investigated the effects of two temperature regimes (low: 16/7 °C and high: 21/14 °C), three water availability levels (saturated, control and drought), and three photosynthetically active photon flux densities: low (85–100 μmol m−2 s−1); intermediate (182–196 μmol m−2 s−1); high (437–454 μmol m−2 s−1) on the growth and reproduction of the annual Floerkea proserpinacoides. Total biomass, total leaf area and flower and seed production increased with increasing temperature, water availability and light intensity. Total leaf area and total biomass were reduced in plants that were stressed under drought. However, at high temperatures, this reduction was less pronounced when droughted plants were partially shaded. At low temperatures, plants began to senesce after approximately 9 wk, whereas at higher temperatures, signs of senescence appeared after only 7 wk of growth. Despite shorter longevity, total biomass was approximately 1.5 times higher in the control water treatment at higher than at lower temperatures as a result of greater above-ground growth, and plants allocated a significantly greater proportion of mass gain to seed production. Although F. proserpinacoides can tolerate low temperatures such as those typical of early spring, higher temperatures such as those of late spring/early summer are more favorable for growth and reproduction as long as water and light are not limiting. Spring ephemeral annuals might be rare because low temperatures reduce growth rate and extend the life cycle. An annual plant might not have time to reproduce before resource availability deteriorates with canopy closure unless reproduction begins early in the life cycle of the species.

Abuscular mycorrhizal fungi are thought to survive adverse environmental conditions primarily as spores. Extraradical mycelia of two Glomus species were produced in fine mesh pouches which excluded roots but not hyphae. The mycelia in these pouches were exposed to freezing conditions, either in the field or in a controlled-temperature chamber. Bioassay plants were grown directly in the pouches and mycorrhizal colonization was assessed after 1 month. The mycelia remained infective in frozen soil over winter. This survival was not dependent on either the presence of root pieces or on the connection of mycelia to roots. Spores were not an effective inoculum in these bioassays. Overwinter survival of mycelia would enable plants to become incorporated into functional mycorrhizal associations early in spring.

Physiological responses of two provenances of European beech (Fagus sylvatica) were studied in seedlings grown at two [CO2] in combination with four temperature treatments. For the local Danish provenance, the average effect of elevated [CO2] during growth was to increase light-saturated net photosynthesis (An) and instantaneous water-use efficiency or transpiration efficiency (ITE). These increases were strongly related to the temperature treatment. Stomatal conductance (gs) was reduced in seedlings in high [CO2], but there was no statistically significant effect of temperature treatment. Stomatal conductance was 13–26% lower at elevated [CO2] and ITE was 89–156% higher, depending on growth temperature. The effects of [CO2] on An were considerably larger than those shown for many other woody species, but similar to those in other studies on European beech. The absolute value of An for a Romanian provenance of beech was 5–18% lower than in the Danish provenance at low [CO2] and 14–26% lower at high [CO2]. There was no statistically significant interaction between the provenances and [CO2], or between provenance and temperature. A model of the response of An to [CO2] at different temperatures gave predictions close to the measured results, except at the lowest temperature treatment where the model over-predicted the effect of elevated [CO2]. This and measurements of An made at a common, low [CO2] indicated a down-regulation of photosynthesis in the lowest temperature treatment at high [CO2]. Root plus soil respiration on a whole-tree basis (Rtr) was increased by elevated [CO2] at all but the lowest temperature, but no effect was seen of [CO2] on root respiration per unit root d. wt. Mean Rtr on any given date was significantly correlated with An, except at the lowest temperature treatment. It is hypothesized that low temperature limited the ability of the roots to use photosynthates resulting in a feedback inhibition of An when elevated [CO2] was combined with low temperature.

A mutation of a gene (discovered in Festuca pratensis Huds. and designated sid ) confers indefinite greenness on senescing leaves. Via intergeneric hybrids with Lolium multiflorum L. and Lolium perenne L., the mutant gene (sidy) has been introgressed into a range of Lolium backgrounds. Using genomic in situ hybridization we have identified segments carrying sidy in recombinant chromosomes of Lolium–Festuca introgression lines. We also used L. perenne lines segregating 1[ratio ]1 for the staygreen character to tag the gene with molecular markers. In two mapping populations a total of 84 genotypes were screened with isoenzymes, RAPD primers, RFLP probes and AFLP primer pairs. Over 180 polymorphic loci were identified, representing 10 linkage groups spanning 600 cM. Two AFLP markers are linked to sid at 4·6 and 14·9 cM, close enough to be usable for marker-assisted selection. Introgression of sidy into Lolium temulentum L. resulted in the production of near-isogenic inbreeding lines suitable for comparative studies of gene expression. Using a variation of the method of representational difference analysis a very small number of cDNAs have been identified as promising candidates for sid, or genes directly regulated by sid.

The effects of ambient and elevated ozone levels on growth and photosynthesis of beech (Fagus sylvatica) were studied by exposing seedlings in open-top chambers for one growing season to three treatments: charcoal-filtered (CF), non-filtered (NF) and non-filtered air with addition of ozone (30 ppb ozone) on clear days for 8–10 h d−1 (NF +). Ambient levels were relatively low and accumulated to an AOT40 (accumulated exposure over a threshold of 40 ppb) of 4055 ppb h (for the period 23 Apr–30 Sept). The NF + chambers received an AOT40 of 8880 ppb h. Throughout the growing season we measured growth and photosynthetic properties. The treatments did not cause strong effects: measurements of gas exchange (light-saturated assimilation rate, CO2 and light-response curves) and chlorophyll fluorescence showed slight and mostly non-significant reductions of several parameters. No significant differences were found for growth, though in the NF + treatment (AOT40 8880 ppb h) the relative growth rate for diameter increment was at times reduced by 12% compared with the control treatment.

Roots of Lablab purpureus (L.) Sweet were treated with tri-iodobenzoic acid (TIBA), kinetin or with nodulation factors (Nod factors) purified from Rhizobium sp. NGR234 and grown in the presence of a mycorrhizal inoculum (Glomus mosseae (Nicol. & Gerd.) Gerdemann & Trappe. Colonization by the mycorrhizal fungus was increased from <30% to c. 65% of root length when roots were treated with these growth regulators. Moreover, treatment of mycorrhizal L. purpureus roots with Nod factors or TIBA strongly induced sporocarp formation of Glomus mosseae. In parallel, the pool size of the fungal disaccharide trehalose was significantly affected in roots treated with TIBA and Nod factors alone, and with TIBA combined with all effectors, and increased from 0·06 mg g−1 d. wt in control roots to up to 1·7 mg g−1 d. wt (TIBA+kinetin). Conversely, the sucrose pool decreased from 5% d. wt to less than a half in roots treated with Nod factors. Activities of trehalase were significantly enhanced in mycorrhizal roots by the treatment with Nod factors or TIBA. When Nod factors and TIBA were added in combination, these activities were strongly enhanced suggesting synergism between these growth regulators.

Interactive effects of elevated atmospheric CO2 and soil N availability on N2 fixation and biomass production were examined using Gliricidia sepium, a tropical leguminous tree species. Our objective was to determine if elevated CO2 alters the inhibitory effects of soil N on N2 fixation, and whether the response of Gliricidia to elevated CO2 was a function of N source originating from either substrate N fertilizer or N2 fixation. We hypothesized that CO2 enrichment would ameliorate the inhibitory effects of N fertilization on seedling nodulation and N2 fixation through increased C partitioning to nodules. Seedlings were grown from seed for 100 d in growth chambers at either 350 or 700 μmol mol−1 CO2. Seedlings were inoculated with Rhizobium spp. and grown either with 0, 1 or 10 mM N fertilizer. The δ15N isotope-dilution technique was used to determine N source partitioning between N2 fixation and inorganic N fertilizer uptake. The addition of 10 mM N fertilizer significantly reduced nodule number and mass, specific nitrogenase activity, the specific rate of N2 fixation, and the proportion of plant N derived from N2 fixation. Elevated CO2, however, strongly ameliorated the inhibitory effects of N fertilization, indicating that increased C availability for nodule activity may partially offset the inhibition of N2 fixation caused by substrate N, as nodule sugar concentrations were stimulated with CO2 enrichment. This study clearly shows that elevated CO2 enhanced plant productivity and net N content of Gliricidia tree seedlings by stimulating N2 fixation. In addition, seedling biomass production was greatly enhanced by elevated CO2, regardless of whether plant N was derived from the substrate or from the atmosphere. We conclude from this study that CO2 enrichment mitigates the inhibitory effects of substrate N on nodule initiation and development and specific N2 fixation, either through increased C allocation to nodule production and activity, or through increased N demand by the plant for biomass production. This experiment with Gliricidia provides evidence for a positive feedback between increased atmospheric CO2 concentrations, C allocation to symbiotic N2-fixing bacteria, and plant C and N accumulation that may occur when N2-fixing plants are grown under conditions where substrate N may typically inhibit N2 fixation.

This work presents a cooperative effort to integrate new molecular (isozyme and SSU analyses) characters into the morphological taxonomy of the genus Gigaspora (Glomales). Previous analyses of published Gigaspora SSU sequences indicated the presence of a few polymorphic nucleotides in the region delimited by primers NS71-SSU 1492′. In our study, the SSU of 24 isolates of arbuscular mycorrhizal (AM) fungi from the Gigasporaceae were amplified and the NS71-SSU 1492′ region was directly sequenced. The corresponding sequences of four more isolates of AM fungi from Gigasporaceae, already published, were also included in our analyses. Three Gigaspora groups were identified on the basis of a 6 nucleotide-long ‘molecular signature’: Gigaspora rosea group (G. rosea+G. albida), Gigaspora margarita group (G. margarita+G. decipiens) and Gigaspora gigantea, which constituted a group by itself. The isozyme profiles (malate dehydrogenase, MDH) of 12 of these 28 isolates, and seven other isolates not sequenced, were compared. The results obtained further supported the grouping of isolates provided by the SSU analysis. Both SSU and MDH analysis indicated that two out of the 35 isolates had been misidentified, which was confirmed when their morphology was reassessed. The use of the Gigaspora intrageneric molecular signature as a quick, unambiguous and objective method to recognize Gigaspora isolates under any (field or laboratory) experimental conditions is suggested.

Following a precultivation with pedospheric nitrogen nutrition, Ricinus plants were supplied with nitrogen solely by spraying nitrate or ammonium solution onto the leaves during the experimental period. The chemical composition of tissues, xylem and phloem exudates was determined and on the basis of the previously determined nitrogen flows (Peuke et al., New Phytologist (1998), 138, 657–687) the flows of potassium, sodium, magnesium, calcium, chloride and ABA were modelled. These data, which permit quantification of net-uptake, transport in xylem and phloem, and utilization in shoot and root, were compared with results obtained in plants with pedospherically-supplied nitrate or ammonium and data in the literature. Although the overall effects on the chemical composition of supplying ammonium to the leaves were not as pronounced as in pedospherically supplied plants, there were some typical responses of plants fed with ammonium (ammonium syndrome). In particular, in ammonium-sprayed plants uptake and transport of magnesium decreased and chloride uptake was increased compared with nitrate-sprayed plants. Furthermore, acropetal ABA transport in the xylem in ammonium-sprayed Ricinus was threefold higher than in nitrate-sprayed plants. Additionally, concentrations of anions were more or less increased in tissues, particularly in the roots, and transport fluids. The overall signal from ammonium-sprayed leaves without a direct effect of ammonium ions on uptake and transport systems in the root is discussed.

Basidiocarps of Pisolithus associated with indigenous (Afzelia quanzensis Welw.) and introduced (Pinus caribaea Mor. and Eucalyptus camaldulensis Dehnh.) hosts in the lowland forests of the Coast Province of Kenya are morphologically distinct. Genetic variability among 52 Pisolithus basidiocarps, collected beneath the various host plants, was examined based on sequence polymorphism within the internal transcribed spacer (ITS) and intergenic spacer (IGS1) of ribosomal DNA genes. Variability in ITS and IGS1 sequences indicated that the three host-associated morphotypes were genetically different. Consensus trees generated by bootstrap analysis of sequence data of Pisolithus isolates from Australia and Kenya are polyphyletic and strongly suggest that the three different morphotypes/genotypes present in Kenya represent separate biological species. In addition, our data indicate that little genetic exchange occurs in silva between these species.

Afrothismia winkleri develops fleshy rhizomes, densely covered with small root tubercles, narrowing to filiform roots with age. The exclusively intracellular mycorrhizal fungus has distinct morphologies in different tissues of the plant. In the filiform root the hyphae grow straight and vesicles are borne on short hyphal stalks. The straight hyphae are present in the epidermis of the root tubercles, but change to loosely coiled and swollen hyphae in the rhizome tissue. No penetration from epidermis to root cortex was found. From the rhizome, a separating cell layer permits only one or rarely two hyphal penetrations into the cortex of each root tubercle. The hyphae proceed apically within the root hypodermis in a spiral row of distinctively coiled hyphae, branches of which colonize the inner root cortex. In the inner root cortex the hyphal coils degenerate to amorphous clumps. In older roots the cortex itself also deteriorates, but epidermis, hypodermis, endodermis and central cylinder persist. The mycorrhizal pattern in A. winkleri is interpreted as an elaborate exploitation system whereby the fungus provides carbon and nutrients to the plant and, simultaneously but spatially distinct, its hyphae are used to translocate and store the matter within the plant. Several features indicate that the endophyte is an arbuscular mycorrhizal fungus.

Several DNA-based techniques, developed for identifying and differentiating fungi in the Gaeumannomyces–Phialophora complex associated with take-all diseases of cereals and grasses, were used to compare fungi from maize. Maize isolates obtained as G. graminis (Sacc.) Arx & H Olivier var. tritici Walker, from the UK, having been identified by ascospore morphology and in pathogenicity tests on wheat, were indistinguishable from isolates of the same variety obtained from wheat. Isolates of G. graminis (Sacc.) Arx & H Olivier var. maydis Yao et al., recently described as the maize take-all fungus from China, were identical in DNA tests to the anamorphic fungus Phialophora radicicola Cain and almost identical to Phialophora zeicola Deacon & Scott, whose description was originally based on isolates from South Africa and France. These species appear to represent the holomorph of the same fungus. The late wilt pathogen of maize, from India and Egypt, commonly known as Cephalosporium maydis Samra et al., but suggested as being the Phialophora anamorph of a Gaeumannomyces species, was closely related to other Gaeumannomyces species included in the tests.

Copper and zinc tolerances of 10 micropropagated birch (Betula pendula and B. pubescens) clones were studied in hydroponic culture. Tolerance indices were determined, based on the mean growth rate of the longest root in 1 wk. A seed-derived clone (142A), from a lead/Zn-contaminated site showed more tolerance to Cu and Zn than bud-derived clones (HA02 and HA18) from a Cu/nickel-contaminated site or an ozone-tolerant clone (KL-2-M) from an uncontaminated area. For Cu, the EC50 values were 30, 14, 8 and 11 μm in clones 142A, HA02, HA18 and KL-2-M, respectively. For Zn, the EC50s were 4000 and 350 μm in clones 142A and KL-2-M, respectively. The relative Cu and Zn tolerances of the other clones were estimated by growing the plants in 30 μm CuSO4, and in 2000 or 350 μm ZnSO4, respectively. It is of interest that the Zn-tolerant clone 142A was tolerant to Cu, although this metal was present at a very low concentration in the soil where the parent tree grows. Another clone (142B), from another seed of the same parent tree, was tolerant neither to Zn nor Cu. Compared with their own EC50s for root growth for Cu, 142A took up more Cu than KL-2-M, suggesting that the higher tolerance of the former clone is not explained by reduced uptake of Cu. The Zn uptake in clones 142A and KL-2-M was studied at 4000 μm and 800 μm Zn, respectively. Interestingly, the roots of both clones contained the same amount of Zn, even though clone 142A was exposed to a fivefold concentration of Zn.

Within the growth zone, salt-affected leaves of sorghum (Sorghum bicolor) had narrower protoxylem and metaxylem cells than controls. Leaf width and cross-sectional area were also reduced, so that the salt treatment had no effect on the area of protoxylem per area of leaf cross section. Dye uptake studies suggested that in controls most of the veins, but in salt-affected leaves only half of the veins, are functional in water transport. Volumetric water flow was greatly diminished in the salt-affected plants. The reduced flow rate was largely explained by the salt-induced decrease in leaf surface area. Some decreases in flow rates per unit leaf mass or area were also produced by salinity, particularly in late developmental stages. Not surprisingly, leaf conductance measured with a diffusion porometer did not appear to be correlated with the diameter of the protoxylem or metaxylem elements. By contrast, published values of water deposition rates are strongly related to the size of the protoxylem elements: the rates of water deposition into the growing leaf tissue are proportional to the square of the protoxylem radius. Thus environmentally produced change of the hydraulic architecture of monocot leaves may cause change in local growth rates.

Phialocephala fortinii is among the few identified hyphomycetes belonging to the Mycelium radicis atrovirens complex. This ‘dark-septate endophyte’ has a global distribution and colonizes a wide variety of host plants. In this study, the spatial distribution of discrete genets of P. fortinii on the forefront of a receding glacier was analysed using the randomly amplified polymorphic DNA (RAPD) technique to determine plants colonized and patterns of colonization. In two consecutive years of sampling, a total of 74 isolates of P. fortinii were obtained from nine plant species, typically ectomycorrhizal, ericoid mycorrhizal or non-mycorrhizal. The isolates showed substantial variation, sharing on average approx. half the RAPD markers. In the first year, three isolates belonging to a single genet were obtained from two plants separated by a distance of nearly 1.5 m. The continuity of this genet was assessed by a sampling the following year. No isolates similar to that, or any of the genets collected the year before were observed. Consequently, the identical isolates from the previous year were concluded to represent discontinuous ramets. Two additional large genets were observed during the second year of sampling, which inhabited roots of several plants representing three different species. These data suggest that the sharing of P. fortinii genets among plant species might play a fundamental role in adaptation and interaction within the whole plant community in a system undergoing primary succession.

Osmotic adjustment (OA) and increased cell-wall extensibility required for expansive leaf growth are well defined components of adaptation to water stress in dry soil, which might interact with soil phosphorus (P) concentration and defoliation frequency for intensively grazed white clover in legume-based pastures. Experiments were conducted with frequently and infrequently defoliated mini-swards of white clover growing in dry soil with low and high P concentrations. The higher yielding high-P plants were able to dry the soil to greater soil water suctions; their leaves had lower water potential values, yet they showed fewer water stress symptoms and underwent a more complete recovery from the water stress symptoms on rewatering, than the low-P plants. High- P plants had greater OA, proline concentration and leaf expansion rate. On the other hand, low-P plants showed an increased osmotic concentration when there was no change in the total solute content per unit of leaf d. wt, indicating more loss of water from the leaf tissue. The key measures that appeared to be directly associated with plant recovery over a short period following water stress were increased proline concentration and leaf expansion rate, probably resulting from increased cell-wall extensibility rather than increased production of cells for the high-P plants.

The use of biofuels has been proposed as one possible substitute for fossil fuels, which contribute substantially to the increase in [CO2] in the atmosphere. However, increased harvesting of forest residues for biofuel might affect the availability of base cations, P and N, as well as the development, community dynamics and function of ectomycorrhizas. This in turn might influence nutrient uptake and tree growth. In this study we investigated the effects of repeated forest residue harvesting on ectomycorrhizal species colonizing spruce roots in the humus layer of a 35-yr-old forest. Harvesting significantly decreased the thickness of the humus layer as well as decreasing the numbers of ectomycorrhizal root tips both per metre root length and per unit humus volume. Changes in mycorrhizal community structure were studied by ITS typing with the use of PCR–RFLP analysis. In total, 19 different ITS types were found on two different sampling occasions (autumn and spring); 11 of these were common to both samplings. Nine of the ITS types were identified to at least the genus level by comparison with RFLP patterns of identified fruiting bodies or axenic cultures. Five species, Cortinarius sp. 2, Thelephora terrestris (Ehrenb.) Fr., Lactarius theiogalus (Bull.[ratio ]Fr.) S. F. Gray s.st. Neuhoff, Tylospora fibrillosa Donk and Tö-96-12, occurred on over 5% of the total sampled root tips. Together these five types colonized 63% of the mycorrhizas screened. A similarity index assessment showed no shift in mycorrhizal community structure as a result of harvesting. Our findings suggest that the repeated removal of forest residues might have a strong effect on the quantity and development of ectomycorrhizal roots in the organic horizon, but little effect on the species composition of the community.