Species-specific impacts of collembola grazing on fungal foraging ecology

Fungi are primary agents of organic matter decomposition in forests. Although invertebrate grazing affects fungal biomass and morphology, the species-specific consequences of these interactions are little understood. Using three collembola species (Folsomia candida, Protaphorura armata, Proisotoma minuta) we employed a multi-trophic approach to investigate the individual effects of invertebrate grazing on four species of saprotrophic basidiomycete fungi growing in two species (one fungus: one collembola) soil microcosms. We studied these effects at three trophic levels: the rate of wood decay brought about by the fungi was assessed; fungal growth was characterized across multiple time points using a range of image analysis parameters (radial extension, hyphal coverage, fractal dimension); and collembola abundance was determined at the end of the experiment. Collembola species had different impacts both within and across fungal species; F. candida had the greatest effect on fungal mycelia whereas P. armata often had little impact. Fungal species varied in their resilience to grazing; all collembola species modified Phanerochaete velutina and Hypholoma fasciculare morphology, that of Resinicium bicolor was only markedly affected by F. candida, and effects on Phallus impudicus were negligible. In the case of H. fasciculare, these grazing effects translated into effects on the rate of fungus-induced wood decay: F. candida and P. armata, but not P. minuta, reduced wood decay rate compared to ungrazed controls. Rate of wood decay was unaffected in the other three fungal species. Changes in collembola population size were generally consistent across fungal species, with each species achieving a greater abundance on P. velutina than on H. fasciculare and P. impudicus. The collembola species did, however, respond differently to R. bicolor, with F. candida being more successful than P. armata. Our study suggests that a wide range of impacts can occur during fungus–collembola interactions, and that caution should be exercised when treating saprotrophic fungi and mycophagous collembola as uniform functional components.     

Decomposer animals (Lumbricidae, Collembola) and organic matter distribution affect the performance of Lolium perenne (Poaceae) and Trifolium repens (Fabaceae)

Decomposer animals stimulate plant growth by indirect effects such as increasing nutrient availability or by modifying microbial communities in the rhizosphere. In grasslands, the spatial distribution of organic matter (OM) rich in nutrients depends on agricultural practice and the bioturbation activities of large detritivores, such as earthworms. We hypothesized that plants of different functional groups with contrasting nutrient uptake and resource allocation strategies differentially benefit from sites in soil with OM accumulation and the presence of decomposer animals. In a greenhouse experiment we investigated effects of spatial distribution of ¹⁵N-labelled grass litter, earthworms and collembola on a simple grassland community consisting of Lolium perenne (grass) and Trifolium repens (legume). Litter aggregates (compared to homogeneous litter distribution) increased total shoot biomass, root biomass and ¹⁵N uptake by the plants. Earthworms and collembola did not affect total N uptake of T. repens; however, the presence of both increased ¹⁵N uptake by T. repens and L. perenne. Earthworms increased shoot biomass of T. repens 1.11-fold and that of L. perenne 2.50 fold. Biomass of L. perenne was at a maximum in the presence of earthworms, collembola and with litter concentrated in a single aggregate. Shoot biomass of T. repens increased in the presence of collembola, with L. perenne generally responding opposingly. The results indicate that the composition of the decomposer community and the distribution of OM in soil affect plant competition and therefore plant community composition.     

Invertebrate grazing determines enzyme production by basidiomycete fungi

Extracellular enzymes produced by heterotrophic microorganisms in the soil are responsible for the decomposition of organic compounds. Basidiomycete fungi are the primary decomposer agents in temperate wooded ecosystems and contribute extensively to extracellular enzyme activity and nutrient mineralisation within soils. Growth and development of basidiomycete mycelia is influenced by soil-dwelling invertebrate grazers with potential implications for fungal activity and ecosystem functioning. The impacts of four invertebrate taxa belonging to Isopoda, Myriapoda, Collembola and Nematoda on the production of eight hydrolytic enzymes by four saprotrophic basidiomycetes (Phanerochaete velutina, Resinicium bicolor and two strains of Hypholoma fasciculare) were compared in a factorial microcosm study. Grazing generally increased enzyme production but invertebrates had species-specific impacts on enzyme activity. The magnitude of grazing influenced enzyme activity; macrofauna (woodlice and millipedes) induced the greatest responses. Enzymatic responses varied markedly between fungi. Grazing enhanced enzyme activity in the exploitative mycelial networks of P. velutina and H. fasciculare, while the opposite effects were observed in the explorative R. bicolor networks. The impacts of soil fauna on nutrient mineralisation depend on fungal community composition. β-glucosidase, cellobiohydrolase, N-acetylglucosaminidase, acid phosphatase and phosphodiesterase activities were affected most frequently by grazing and invertebrate activity, and thus had direct consequences for carbon, nitrogen and phosphorous cycling. The results indicate that invertebrate diversity and community composition may influence the spatial distribution and activity of extracellular enzymes with direct implications for nutrient mineralisation and trunover in woodland soils.     

Species richness of ectomycorrhizal hyphal necromass increases soil CO2 efflux under laboratory conditions

The ectomycorrhizal mycelium is a large component of boreal and temperate forest soil microbial biomass and the resulting necromass is likely to be an important source of nutrients for saprotrophic microorganisms. Here we test the effects of species richness of ectomycorrhizal mycelial biomass on short-term CO₂ efflux by amending forest soil with necromass from 8 fungal species added separately and in mixtures of 2, 4 and 8 species. All additions of necromass rapidly increased soil CO₂ efflux compared to unamended controls but CO₂ efflux increased significantly with species richness. Efflux of CO₂ did not correlate with the carbon (C) or nitrogen (N) contents or the C:N ratio of the added necromass. The study demonstrates that species diversity of dead ectomycorrhizal fungal hyphae can have important consequences for soil CO₂ efflux, and suggests decomposition of hyphae is regulated by specific constituents of the nutrient pools in the necromass rather than the total quantities added.     

Saprotrophic fungi transform organic phosphorus from spruce needle litter

Fungal decomposition of and phosphorus transformation from spruce litter needles (Picea abies) were simulated in systems containing litter needles inoculated with individual saprotrophic fungal strains and their mixtures. Fungal strains of Setulipes androsaceus (L.) Antonín, Chalara longipes (Preus) Cooke, Ceuthospora pinastri (Fr.) Höhn., Mollisia minutella (Sacc.) Rehm, Scleroconidioma sphagnicola Tsuneda, Currah & Thormann and an unknown strain NK11 were used as representatives of autochthonous mycoflora. Systems were incubated for 5.5 months in laboratory conditions. Fungal colonization in systems and competition among strains were assessed using the reisolation of fungi from individual needles. After incubation, needles were extracted with NaOH and extracts were analysed using ³¹P nuclear magnetic resonance spectroscopy (NMR). Needle decomposition was determined based on the decrease in C:N ratio. Systems inoculated with the basidiomycete S. androsaceus revealed substantial decrease in C:N ratio (from 25.8 to 11.3) while the effect of ascomycetes on the C:N ratio was negligible. We suppose that tested strains of saprotrophic ascomycetes did not participate substantially in litter decomposition, but were directly involved in phosphorus transformation and together with S. androsaceus could transform orthophosphate monoesters and diesters from spruce litter needles into diphosphates, polyphosphates and phosphonates. These transformations seem to be typical for saprotrophic fungi involved in litter needle decomposition, although the proportion of individual phosphorus forms differed among studied fungal strains. Phosphonate presence in needles after fungal inoculation is of special interest because no previous investigation recorded phosphonate synthesis and accumulation by fungi. Our results confirmed that the ³¹P NMR spectroscopy is an excellent instrumental method for studying transformations of soil organic phosphorus during plant litter decomposition. We suggest that polyphosphate production by S. androsaceus may contribute to the phosphorus cycle in forest ecosystems because this fungus is a frequent litter colonizer that substantially participates in decomposition.     

Origin of the nitrogen assimilated by soil fauna living in decomposing beech litter

We investigated the nitrogen source for main taxa of soil fauna in two beech forests of contrasted humus type using ¹⁵N-labelled beech litter and ¹⁵N analysis of soil fauna. ¹⁵N-labelled beech litter was deposited on the topsoil in December 2000 in four stands of different ages at Leinefelde (Germany) with mull humus and in one mature stand at Sorø (Denmark) with moder humus. The fate of the tracer isotope was measured in litter and soil, as well as in the soil fauna, and for each taxa, we calculated the proportion of N in the animal derived from the labelled substrate. Of the original N contained in the litter, 20-41% was lost after 9 months at Leinefelde, and only 10% at Sorø. This loss was counterbalanced by the incorporation of 24-31% external N at Leinefelde, and 31% at Sorø, partly originating from fungal colonisation of the added litter. The proportion of N assimilated from the labelled litter by the different soil animals varied in relation to their mobility and feeding preferences. Large and mobile soil animals, especially predators, derived on average less ¹⁵N because they were also able to feed outside the labelled litter boxes. Detritivores assimilated at most 15% of their nitrogen content at Leinefelde and 11% at Sorø from the decomposing labelled litter. The most labelled taxa at Leinefelde were small fungivorous and coprophagous species, mainly isotomid Collembola such as Isotomiella and Folsomia. At Sorø, best labelled taxa were saprophagous species such as Enchytraeidae, Glomeridae and Phthiracaroidea. These low rates of ¹⁵N assimilation indicate that fresh litter is not directly the main N source for soil animals. The results obtained suggest that soil fauna fed preferentially upon microorganisms colonising the litter at Leinefelde (mull) and from litter itself at Sorø (moder).     

Collembolan grazing affects the growth strategy of the cord-forming fungus Hypholoma fasciculare

Mycelia of cord-forming fungi show remarkable patterns of reallocation of biomass and nutrients indicating an important role of these, often extensive, organisms in the spatial translocation of energy and nutrients in forest soils. Despite the rich tradition of interaction studies between soil microarthropods and fungi, the spatial implications of these interactions, due to the potential growth responses of the fungi and to the translocation of energy and nutrients within the mycelial network, have been largely ignored. In this paper we analyse fungal growth responses in two-dimensional model systems composed of compressed soil, the cord-forming fungus Hypholoma fasciculare and three fungivorous Collembolan species. We hypothesised that (i) the highly co-ordinated nature of cord-forming fungi would lead to growth responses following collembolan grazing, and that, (ii) such changes are dependent on grazing intensity, and (iii) changes are dependent on the species grazing. Mycelial extent and hyphal cover decreased with increasing grazing density; at highest grazing density also the fractal dimension of the mycelial border decreased, indicating a less branched foraging front due to the regression of fine hyphae and the development of mycelial cords. Effects differed greatly between collembolan species although they exerted comparable grazing pressure (the smaller species were added in larger numbers according to their allometric size-metabolic rate relationships): while grazing by Folsomia candida resulted in less mycelial extension and hyphal cover, these variables were not affected when Proisotoma minuta and Hypogastrura cf. tullbergi grazed. The effects of a species mix suggested an additive effect of the component species. This shows that fungal mycelia may suffer from damage caused by few but large collembolans, affecting extension as well as coverage of the mycelium, but that fungi may compensate for the biomass loss caused by more but slightly smaller collembolans. In about 20% of the model systems H. fasciculare switched from a growth pattern with a broad contiguous foraging front and uniform growth in all directions to a pattern with fast growing sectors while other sectors stopped growth completely. The switch occurred in grazed systems exclusively; thus we interpret this observation as a fugitive response and as a strategy for quickly escaping from places where grazing pressure is experienced.     

The effect of collembolan grazing on fungal activity in differently managed upland pastures: A microcosm study

Laboratory microcosms containing litter from three tussock grasslands were used to assess the impact of grazing by a collembolan, Onychiurus procampatus, on the abundance, nutrient release, and respiration of the saprotrophic fungus, Phoma exigua. The fungal biomass and respiration rate were significantly reduced only when Collembola were present in excess of mean field densities but perhaps more typical of spatial aggregations in the soil. A high efficiency of nutrient immobilization by P. exigua was demonstrated but nutrient release was not significantly affected by the fauna. Problems associated with the use of microcosms in the simulation of field conditions are discussed.     

Collembola grazing on arbuscular mycorrhiza fungi modulates nutrient allocation in plants

Arbuscular mycorrhiza (AM) mycelia networks are important for nutrient allocation in many plants, but fungivorous soil invertebrates such as Collembola can modulate the symbiosis by grazing on the extra-radical mycelium (ERM). This study employs a dual biomarker approach with stable isotopes and fatty acids to disentangle trophic interactions of Collembola in a plant-fungal soil system with maize (Zea mays) and the AM fungus Glomus mosseae. To separate ERM and root mediated effects, root (RC) and hyphal compartments (HC) were used, and the latter was spiked with labeled ¹⁵N substrate. The euedaphic Collembola species Protaphorura fimata was introduced as the fungal and root grazer. Generally, the presence of Collembola in RC fostered biomass and phosphorous uptake in roots colonized with AM. Nitrogen transport from HC to RC was not altered, indicating that Collembola did not disrupt the ERM network via grazing. Collembola–fungus interactions fostered AM hyphal proliferation in HC, whereas in RC it induced a change from fungal senescence with build-up of storage reserves, to an active foraging phase. A distinct diet switch by Collembola between HC and RC indicated different ERM palatability meditated by the presence or absence of the host plant. Overall, Collembola grazing increased ERM nutrient sequestration, particularly phosphorus, and in turn plant performance. Collembola modified fungal phenology, favoring fungal colonization over reproductive phases. These trophic interactions were strongly determined by fungal life stage, with the establishment of a functional mycorrhiza as a crucial factor.     

Endogeic earthworms alter carbon translocation by fungi at the soil-litter interface

The effect of endogeic earthworms (Octolasion tyrtaeum (Savigny)) on the translocation of litter-derived carbon into the upper layer of a mineral soil by fungi was investigated in a microcosm experiment. Arable soil with and without O. tyrtaeum was incubated with ¹³C/¹⁵N-labelled rye leaves placed on plastic rings with gaze (64 μm mesh size) to avoid incorporation of leaves by earthworms. The plastic rings were positioned either on or 3 cm above the soil surface, to distinguish between biotic and chemical/physical translocation of nutrients by fungi and leaching.Contact of leaves to the soil increased ¹³C translocation, whereas presence of O. tyrtaeum reduced the incorporation of ¹³C into the mineral soil in all treatments. Although biomass of O. tyrtaeum decreased during the experiment, more ¹³C and ¹⁵N was incorporated into earthworm tissue in treatments with contact of leaves to the soil. Contact of leaves to the soil and the presence of O. tyrtaeum increased cumulative ¹³CO₂-C production by 18.2% and 14.1%, respectively.The concentration of the fungal bio-indicator ergosterol in the soil tended to be increased and that of the fungal-specific phospholipid fatty acid 18:2ω6 was significantly increased in treatments with contact of leaves to the soil. Earthworms reduced the concentration of ergosterol and 18:2ω6 in the soil by 14.0% and 43.2%, respectively. Total bacterial PLFAs in soil were also reduced in presence of O. tyrtaeum, but did not respond to the addition of the rye leaves. In addition, the bacterial community in treatments with O. tyrtaeum differed from that without earthworms and shifted towards an increased dominance of Gram-negative bacteria.The results indicate that litter-decomposing fungi translocate litter-derived carbon via their mycelial network in to the upper mineral soil. Endogeic earthworms decrease fungal biomass by grazing and disruption of fungal hyphae thereby counteracting the fungal-mediated translocation of carbon in soils.     

Size matters: What have we learnt from microcosm studies of decomposer fungus–invertebrate interactions?

The ongoing research ‘boom’ in soil ecology has been advanced by a widespread use of laboratory experiments to investigate mechanisms that could not be unravelled with field observations alone. Interactions between soil fungi and invertebrates have received considerable attention due to their trophic and functional importance in belowground systems. Saprotrophic cord-forming basidiomycete fungi are major agents of primary decomposition in woodland ecosystems, where they are also an important source of nutrition for fungal-feeding soil invertebrates. A plethora of microcosm experiments, with their main benefit being that they enable most variables to be kept constant while just a few are manipulated, have provided detailed insights into the ecology of fungus–invertebrate interactions. This review identifies important trends from this body of work (including a meta-analysis of grazing effects on fungal growth and wood decomposition) and explores the extent to which these patterns are supported by the few related experiments conducted in more complex mesocosm and field systems. Grazing in microcosms reduced fungal growth and increased decomposition, but with interaction-specific magnitude, reflecting invertebrate feeding preferences for different fungi. Macro-invertebrates (woodlice and millipedes) had stronger effects than micro- (e.g. nematodes) and meso- (e.g. collembola) invertebrates. This greater grazing pressure generally increased enzyme activities beneath mycelia during interactions in which wood decay was increased. Top-down effects of fungal-feeding can be extrapolated to more complex systems, but only for macro-invertebrates, particularly woodlice. Soil enzyme activity was stimulated, in microcosms and more complex systems, by short-term or low intensity grazing, but reduced when large areas of mycelium were removed by high-intensity grazing. Effects of differential fungal palatability on invertebrate populations are evident in microcosm studies of collembola. These bottom-up effects can be extrapolated more broadly than top-down effects; fungal community dominance determined collembola abundance and diversity, in mesocosms, and woodlouse abundance in the field. Using, as a case study, a series of experiments conducted at a range of scales, mechanisms underlying potential climate change effects on grazing interactions and decomposition are also explored. Biotic effects on decomposer community functioning are heterogeneous, depending on fungal dominance and the density of key macro-invertebrate taxa.     

Fermentation of sugar beet waste by Aspergillus niger facilitates growth and P uptake of external mycelium of mixed populations of arbuscular mycorrhizal fungi

Sugar beet waste has potential value as a soil amendment and this work studied whether fermentation of the waste by Aspergillus niger would influence the growth and P uptake of arbuscular mycorrhizal (AM) fungi. Plants were grown in compartmentalised growth units, each with a root compartment (RC) and two lateral root-free compartments (RFC). One RFC contained untreated soil while the other RFC contained soil, which was uniformly mixed with sugar beet waste, either untreated (SB) or degraded by A. niger (ASB) in a rock phosphate (RP)-supplied medium. The soil in each pair of RFC was labelled with ³³P and ³²P in order to measure P uptake by the AM fungal mycelium, of which length density was also measured. Whole cell fatty acid (WCFA) signatures were used as biomarkers of the AM fungal mycelium and other soil microorganisms. The amount of biomarkers of saprotrophic fungi and both Gram-positive and Gram-negative bacteria was higher in SB than in ASB treatments. Whilst ASB increased growth and activity of AM mycelium, SB had the opposite effect. Moreover, shoot P content was increased by the addition of ASB, and by inoculation with AM fungi. Modification of soil microbial structure and production of exudates by A. niger, as a consequence of fermentation process of sugar beet waste, could possibly explain the increase of AM growth in ASB treatments. On the other hand, the highest P uptake was a result of the solubilisation of rock phosphate by A. niger during the fermentation.     

Effects of mycorrhizal roots and extraradical hyphae on N uptake from vineyard cover crop litter and the soil microbial community

The objectives of this study were to evaluate the contribution of arbuscular mycorrhizal (AM) fungal hyphae to ¹⁵N uptake from vineyard cover crop litter (Medicago polymorpha), and to examine the soil microbial community under the influence of mycorrhizal roots and extraradical hyphae. Mycorrhizal grapevines (Vitis vinifera) were grown in specially designed containers, within which a polyvinyl chloride (PVC) mesh core was inserted. Different sizes of mesh allowed mycorrhizal roots (mycorrhizosphere treatment) or extraradical hyphae (hyphosphere treatment) to access dual labeled ¹⁵N and ¹³C cover crop litter that was placed inside the cores after 4 months of grapevine growth. Mesh cores in the bulk soil treatment, which served as a negative control, had the same mesh size as the hyphosphere treatment, but frequent rotation prevented extraradical hyphae from accessing the litter. Grapevines and soils were harvested 0, 7, 14, and 28 days after addition of the cover crop litter and examined for the presence of ¹⁵N. Soil microbial biomass and the soil microbial community inside the mesh cores were examined using phospholipid fatty acid analysis. ¹⁵N concentrations in grapevines in the hyphosphere treatment were twice that of grapevines in the bulk soil treatment, suggesting that extraradical hyphae extending from mycorrhizal grapevine roots may have a role in nutrient utilization from decomposing vineyard cover crops in the field. Nonetheless, grapevines in the mycorrhizosphere treatment had the highest ¹⁵N concentrations, thus highlighting the importance of a healthy grapevine root system in nutrient uptake. We detected similar peaks in soil microbial biomass in the mycorrhizosphere and hyphosphere treatments after addition of the litter, despite significantly lower microbial biomass in the hyphosphere treatment initially. Our results suggest that although grapevine roots play a dominant role in the uptake of nutrients from a decomposing cover crop, AM hyphae may have a more important role in maintaining soil microbial communities associated with nutrient cycling.     

Isotopic fractionation by saprotrophic microfungi: Effects of species, temperature and the age of colonies

Saprotrophic fungi represent an important resource for a number of fungivorous and omnivorous soil animals, but little is known about the patterns of isotopic fractionation by soil fungi. We grew five common species of saprotrophic microfungi in laboratory cultures on simple artificial substrate based on carbohydrates derived either from C3 or C4 plants. Fungal cultures were kept at 15, 20 or 25 °C. Isotopic composition of carbon (¹³C/¹²C) and nitrogen (¹⁵N/¹⁴N) in bulk fungal tissue was determined after 11, 21 and 32 days. The fractionation of carbon and nitrogen stable isotopes was species-specific, but generally did not differ in C3- and C4-based growth media. The Zygomycete Mucor plumbeus did not differ in δ¹³C from the carbon source used, though Ascomycetes (Alternaria alternata, Cladosporium cladosporioides, Trichoderma harzianum and Ulocladium botrytis) were depleted in heavy carbon relative to the carbon source by 0.5-0.9‰. Three species were significantly depleted in ¹⁵N relative to the sodium nitrate that was used as a single source of nitrogen. In all species, δ¹⁵N but not δ¹³C tended to increase with the age of fungal colonies. The effect of temperature on δ¹⁵N was weak and inconsistent in different species. In contrast, all fungi except T. harzianum accumulated more ¹³С at 25 °C than at 15 °C. The overall variation in the isotopic signatures of saprotrophic fungi growing in identical conditions reached 8‰ for δ¹⁵N and 2.5‰ for δ¹³C due to species-specific differences in the isotopic fractionation and the age of individual fungal colonies. This variation should be incorporated into the interpretation of the isotopic composition of fungivorous soil animals.     

Respiration and growth of a fungus, Mortierella isabellina, in response to grazing by Onychiurus armatus (Collembola)

Colonies of the fungus Mortierella isabellina (Oudem) were grazed by the collembolan Onychiurus armatus (Tullb.) for short periods interrupted by incubation without grazing in laboratory experiments. The grazed fungus had a lower respiration than ungrazed when Collembola were present on the mycelium. Fungal respiration was significantly increased with grazing when Collembola were periodically removed from the mycelium. Metal contamination of the substrate increased fungal respiration, regardless of grazing. Length of fungal mycelium was significantly greater in a natural soil in the presence of Collembola but reduced in a metal-polluted soil. Factors that may determine the impact of grazing on fungi are discussed.     

Release of C and N from roots of peas and oats and their availability to soil microorganisms

Nutrient mobilisation in the rhizosphere is driven by soil microorganisms and controlled by the release of available C compounds from roots. It is not known how the quality of release influences this process in situ. Therefore, the present study was conducted to investigate the amount and turnover of rhizodeposition, in this study defined as root-derived C or N present in the soil after removal of roots and root fragments, released at different growth stages of peas (Pisum sativum L.) and oats (Avena sativa L.). Plants were grown in soil columns placed in a raised bed under outdoor conditions and simultaneously pulse labelled in situ with a ¹³C-glucose-¹⁵N-urea solution using a stem feeding method. After harvest, ¹³C and ¹⁵N was recovered in plant parts and soil pools, including the microbial biomass. Net rhizodeposition of C and N as a percentage of total plant C and N was higher in peas than in oats. Moreover, the C-to-N ratio of the rhizodeposits was lower in peas, and a higher proportion of the microbial biomass and inorganic N was derived from rhizodeposition. These results suggest a positive plant-soil feedback shaping nutrient mobilisation. This process is driven by the C and N supply of roots, which has a higher availability in peas than in oats.     

Natural abundance δN and δC of DNA extracted from soil

We report the first simultaneous measurements of δ¹⁵N and δ¹³C of DNA extracted from surface soils. The isotopic composition of DNA differed significantly among nine different soils. The δ¹³C and δ¹⁵N of DNA was correlated with δ¹³C and δ¹⁵N of soil, respectively, suggesting that the isotopic composition of DNA is strongly influenced by the isotopic composition of soil organic matter. However, in all samples DNA was enriched in ¹³C relative to soil, indicating microorganisms fractionated C during assimilation or preferentially used ¹³C enriched substrates. Enrichment of DNA in ¹⁵N relative to soil was not consistently observed, but there were significant differences between δ¹⁵N of DNA and δ¹⁵N of soil for three different sites, suggesting microorganisms are fractionating N or preferentially using N substrates at different rates across these contrasting ecosystems. There was a strong linear correlation between δ¹⁵N of DNA and δ¹⁵N of the microbial biomass, which indicated DNA was depleted in ¹⁵N relative to the microbial biomass by approximately 3.4‰. Our results show that accurate and precise isotopic measurements of C and N in DNA extracted from the soil are feasible, and that these analyses may provide powerful tools for elucidating C and N cycling processes through soil microorganisms.     

Spatial distribution of fungal communities in a coastal grassland soil

In grasslands, saprotrophic fungi, including basidiomycetes, are major decomposers of dead organic matter, although spatial distributions of their mycelial assemblages are little described. The aim of this study was to characterise the scale and distribution of saprotrophic fungal communities in a coastal grassland soil using terminal restriction fragment length polymorphism (T-RFLP).Soil fungi were sampled at Point Reyes, California, USA, by taking forty-five 26 mm diam. cores in a spatially defined manner. Within each sampled core, complete core sections at 1-2 cm and 14-15 cm depths were removed and sub-sampled for DNA extraction and amplification using the primer pairs ITS1F-FAM/ITS4 (general fungi) or ITS1F-FAM/ITS4B (basidiomycete-specific).Nonmetric Multidimensional Scaling showed that general fungal communities could be clearly separated by depth, although basidiomycete communities could not. There were no strong patterns of community similarity or dissimilarity for general or basidiomycete fungal communities at horizontal geographical distances from 25 cm to 96 m in the upper horizon. These results show considerable vertical, but little horizontal, variability in fungal community structure in a semi-natural grassland at the spatial scales measured here.     

CO2 and inorganic N supply modify competition for N between co-occurring grass plants, tree seedlings and soil microorganisms

Plant-plant and plant-soil interactions play a key role in determining plant community structure and ecosystem function. However, the effects of global change on the interplay between co-occurring plants and soil microbes in successional communities are poorly understood. In this study, we investigated competition for nitrogen (N) between soil microorganisms, grass plants and establishing tree seedlings under factorial carbon dioxide (CO₂) and N treatments. Fraxinus excelsior seedlings were germinated in the presence or absence of grass competition (Dactylis glomerata) at low (380 μmol mol⁻¹) or high (645 μmol mol⁻¹) CO₂ and at two levels of N nutrition in a mesocosm experiment. Pulse ¹⁵N labelling was used to examine N partitioning among plant and soil compartments. Dactylis exerted a strong negative effect on Fraxinus biomass, N capture and ¹⁵N recovery irrespective of N and CO₂ treatment. In contrast, the presence of Dactylis had a positive effect on the microbial N pool. Plant and soil responses to N treatment were of a greater magnitude compared with responses to elevated CO₂, but the pattern of Fraxinus- and microbial-N pool response to N and CO₂ varied depending on grass competition treatment. Within the Dactylis competition treatment, decreases in Fraxinus biomass in response to N were not mirrored by decreases in tree seedling N content, suggesting a shift from below- to above-ground competition. In the Dactylis-sown pots, ¹⁵N recovery could be ranked Dactylis > microbial pool > Fraxinus in all N and CO₂ treatment combinations. Inequalities between Fraxinus and soil microorganisms in terms of ¹⁵N recovery were exacerbated by N addition. Contrary to expectations, elevated CO₂ did not increase plant-microbe competition. Nevertheless, microbial ¹⁵N recovery showed a small positive increase in the high CO₂ treatment. Overall, elevated CO₂ and N supply did not interact on plant/soil N partitioning. Our data suggest that the competitive balance between establishing tree seedlings and grass plants in an undisturbed sward is relatively insensitive to CO₂ or N-induced modifications in N competition between plant and soil compartments.     

Carbon allocation in mycelia of arbuscular mycorrhizal fungi during colonisation of plant seedlings

The direction of carbon (C) allocation in mycorrhizal mycelia is of fundamental importance to coexistence of individual plants. We therefore investigated the transfer of C from established plants to plant seedlings through fungal mycelia. C allocation by the arbuscular mycorrhizal (AM) fungus Glomus intraradices, from ‘donor’ plants to mycelia in soil and two different species of introduced ‘receiver’ seedlings, was investigated in a pot experiment using ¹³C labelling and fatty acid analysis. After ¹³CO₂ application to the shoots of Trifolium subterraneum or Plantago lanceolata, used as donor plants, T. subterraneum and P. lanceolata receiver seedlings were introduced. Samples were collected 4-20 days after ¹³CO₂ application and analysed regarding ¹³C and the fatty acid 16:1ω5, the signature of AM fungi. ¹³C transfer from T. subterraneum donor plants was demonstrated by ¹³C enrichment of the roots of the receiver seedlings, but not from the P. lanceolata donor plants. ¹³C allocation to the neutral lipid fatty acid 16:1ω5 was only 1 ng in each receiver seedling, but 2 μg of the fatty acid in whole soil. The results indicate that C allocation through mycelial networks is influenced by the donor plant species, but is not directed towards receiver seedlings to any higher degree than towards other directions. The importance of the extraradical AM fungal mycelium as a C sink was demonstrated.