When the forest dies: the response of forest soil fungi to a bark beetle-induced tree dieback.

Coniferous forests cover extensive areas of the boreal and temperate zones. Owing to their primary production and C storage, they have an important role in the global carbon balance. Forest disturbances such as forest fires, windthrows or insect pest outbreaks have a substantial effect on the functioning of these ecosystems. Recent decades have seen an increase in the areas affected by disturbances in both North America and Europe, with indications that this increase is due to both local human activity and global climate change. Here we examine the structural and functional response of the litter and soil microbial community in a Picea abies forest to tree dieback following an invasion of the bark beetle Ips typographus, with a specific focus on the fungal community. The insect-induced disturbance rapidly and profoundly changed vegetation and nutrient availability by killing spruce trees so that the readily available root exudates were replaced by more recalcitrant, polymeric plant biomass components. Owing to the dramatic decrease in photosynthesis, the rate of decomposition processes in the ecosystem decreased as soon as the one-time litter input had been processed. The fungal community showed profound changes, including a decrease in biomass (2.5-fold in the litter and 12-fold in the soil) together with the disappearance of fungi symbiotic with tree roots and a relative increase in saprotrophic taxa. Within the latter group, successive changes reflected the changing availability of needle litter and woody debris. Bacterial biomass appeared to be either unaffected or increased after the disturbance, resulting in a substantial increase in the bacterial/fungal biomass ratio.

Laccase activity in soils: considerations for the measurement of enzyme activity.

Laccases (benzenediol: oxygen oxidoreductases, EC 1.10.3.2) are copper-containing enzymes that catalyze the oxidative conversion of a variety of chemicals, such as mono-, oligo-, and polyphenols and aromatic amines. Laccases have been proposed to participate in the transformation of organic matter and xenobiotics as well as microbial interactions. Several laccase assays have been proposed and used in soils. Here, we show that the optimal pH conditions for the laccase substrates 2,2'-azinobis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS, pH 3-5), 2,6-dimethoxyphenol (4-5.5), L-3,4-dihydroxyphenylalanine (DOPA; 4-6), guaiacol (3.5-5), 4-methylcatechol (3.5-5), and syringaldazine (5.5-7.0) are similar between purified laccases from Trametes versicolor and Pyricularia sp. and soil extracts; the substrate affinities of purified enzymes (K(M)) and soil extracts were also similar. The laccase assays showed specificity overlap with tyrosinase and ligninolytic peroxidases when hydrogen peroxide is present. The ABTS oxidation assay is able to reliably detect the presence of 13.5 pg mL(-1) or 0.199×10(-12) mol mL(-1) of T. versicolor laccase, which is three times more sensitive than the 2,6-dimethoxyphenol-based assay and more than 40 times more sensitive than any of the other assays. The low molecular mass soil-derived compounds and the isolated fulvic and humic acids influence the laccase assays and should be removed from the soil extracts before measurements of the enzyme activity are performed.

Active and total microbial communities in forest soil are largely different and highly stratified during decomposition.

Soils of coniferous forest ecosystems are important for the global carbon cycle, and the identification of active microbial decomposers is essential for understanding organic matter transformation in these ecosystems. By the independent analysis of DNA and RNA, whole communities of bacteria and fungi and its active members were compared in topsoil of a Picea abies forest during a period of organic matter decomposition. Fungi quantitatively dominate the microbial community in the litter horizon, while the organic horizon shows comparable amount of fungal and bacterial biomasses. Active microbial populations obtained by RNA analysis exhibit similar diversity as DNA-derived populations, but significantly differ in the composition of microbial taxa. Several highly active taxa, especially fungal ones, show low abundance or even absence in the DNA pool. Bacteria and especially fungi are often distinctly associated with a particular soil horizon. Fungal communities are less even than bacterial ones and show higher relative abundances of dominant species. While dominant bacterial species are distributed across the studied ecosystem, distribution of dominant fungi is often spatially restricted as they are only recovered at some locations. The sequences of cbhI gene encoding for cellobiohydrolase (exocellulase), an essential enzyme for cellulose decomposition, were compared in soil metagenome and metatranscriptome and assigned to their producers. Litter horizon exhibits higher diversity and higher proportion of expressed sequences than organic horizon. Cellulose decomposition is mediated by highly diverse fungal populations largely distinct between soil horizons. The results indicate that low-abundance species make an important contribution to decomposition processes in soils.

Saprotrophic basidiomycete mycelia and their interspecific interactions affect the spatial distribution of extracellular enzymes in soil.

Saprotrophic cord-forming basidiomycetes are important decomposers of lignocellulosic substrates in soil. The production of extracellular hydrolytic enzymes was studied during the growth of two saprotrophic basidiomycetes, Hypholoma fasciculare and Phanerochaete velutina, across the surface of nonsterile soil microcosms, along with the effects of these basidiomycetes on fungi and bacteria within the soil. Higher activities of α-glucosidase, ß-glucosidase, cellobiohydrolase, ß-xylosidase, phosphomonoesterase and phosphodiesterase, but not of arylsulphatase, were recorded beneath the mycelia. Despite the fact that H. fasciculare, with exploitative hyphal growth, produced much denser hyphal cover on the soil surface than P. velutina, with explorative growth, both fungi produced similar amounts of extracellular enzymes. In the areas where the mycelia of H. fasciculare and P. velutina interacted, the activities of N-acetylglucosaminidase, α-glucosidase and phosphomonoesterase, the enzymes potentially involved in hyphal cell wall damage, and the utilization of compounds released from damaged hyphae of interacting fungi, were particularly increased. No significant differences in fungal biomass were observed between basidiomycete-colonized and noncolonized soil, but bacterial biomass was reduced in soil with H. fasciculare. The increases in the activities of ß-xylosidase, ß-glucosidase, phosphomonoesterase and cellobiohydrolase with increasing fungal:bacterial biomass ratio indicate the positive effects of fungal enzymes on nutrient release and bacterial abundance, which is reflected in the positive correlation of bacterial and fungal biomass content.

Transformation of Quercus petraea litter: successive changes in litter chemistry are reflected in differential enzyme activity and changes in the microbial community composition.

The links among the changes in litter chemistry, the activity of extracellular enzymes and the microbial community composition were observed in Quercus petraea litter. Three phases of decomposition could be distinguished. In the early 4-month stage, with high activities of ß-glucosidase, ß-xylosidase and cellobiohydrolase, 16.4% of litter was decomposed. Hemicelluloses were rapidly removed while cellulose and lignin degradation was slow. In months 4-12, with high endocellulase and endoxylanase activities, decomposition of cellulose prevailed and 31.8% of litter mass was lost. After the third phase of decomposition until month 24 with high activity of ligninolytic enzymes, the litter mass loss reached 67.9%. After 2 years of decay, cellulose decomposition was almost complete and most of the remaining polysaccharides were in the form of hemicelluloses. Fungi largely dominated over bacteria as leaf endophytes and also in the litter immediately before contact with soil, and this fungal dominance lasted until month 4. Bacterial biomass (measured as phospholipid fatty acid content) in litter increased with time but also changed qualitatively, showing an increasing number of Actinobacteria. This paper shows that the dynamics of decomposition of individual litter components changes with time in accordance with the changes in the microbial community composition and its production of extracellular enzymes.

Ecology of coarse wood decomposition by the saprotrophic fungus Fomes fomentarius.

Saprotrophic wood-inhabiting basidiomycetes are the most important decomposers of lignin and cellulose in dead wood and as such they attracted considerable attention. The aims of this work were to quantify the activity and spatial distribution of extracellular enzymes in coarse wood colonised by the white-rot basidiomycete Fomes fomentarius and in adjacent fruitbodies of the fungus and to analyse the diversity of the fungal and bacterial community in a fungus-colonised wood and its potential effect on enzyme production by F. fomentarius. Fungus-colonised wood and fruitbodies were collected in low management intensity forests in the Czech Republic. There were significant differences in enzyme production by F. fomentarius between Betula pendula and Fagus sylvatica wood, the activity of cellulose and xylan-degrading enzymes was significantly higher in beech wood than in birch wood. Spatial analysis of a sample B. pendula log segment proved that F. fomentarius was the single fungal representative found in the log. There was a high level of spatial variability in the amount of fungal biomass detected, but no effects on enzyme activities were observed. Samples from the fruiting body showed high ß-glucosidase and chitinase activities compared to wood samples. Significantly higher levels of xylanase and cellobiohydrolase were found in samples located near the fruitbody (proximal), and higher laccase and Mn-peroxidase activities were found in the distal ones. The microbial community in wood was dominated by the fungus (fungal to bacterial DNA ratio of 62-111). Bacterial abundance composition was lower in proximal than distal parts of wood by a factor of 24. These results show a significant level of spatial heterogeneity in coarse wood. One of the explanations may be the successive colonization of wood by the fungus: due to differential enzyme production, the rates of biodegradation of coarse wood are also spatially inhomogeneous.

Differential degradation of oak (Quercus petraea) leaf litter by litter-decomposing basidiomycetes.

Due to production of lignocellulose-degrading enzymes, saprotrophic litter-decomposing basidiomycetes can significantly contribute to the turnover of soil organic matter. The production of lignin and polysaccharide-degrading enzymes and changes in the chemical composition of litter was studied with Marasmius quercophilus, Mycena inclinata and Pholiota lenta, three basidiomycete species typical of oak (Quercus petraea) forests. Within 12weeks of incubation, M. inclinata decomposed 33%, M. quercophilus 36% and P. lenta 48% of the substrate dry mass. All fungi produced laccase and Mn-peroxidase and none of them produced lignin peroxidase or Mn-independent peroxidases. M. inclinata and M. quercophilus produced considerable laccase activity, while production by P. lenta was low. M. quercophilus and P. lenta produced most Mn-peroxidase at the beginning of the experiment, while the production by M. inclinata was more stable in time. Endo-1,4-beta-xylanase exhibited the highest activity among endocleaving glycosyl hydrolases while 1,4-beta-glucosidase was the main exocleaving enzyme. All fungi decreased the C:N ratio of the litter from 27 to 13-17 and M. inclinata and M. quercophilus also decreased the lignin content. Analytical pyrolysis of decayed litter showed changes in litter composition similar to those caused by white-rot fungi during wood decay, e.g. a decrease in the syringyl/guaiacyl lignin ratio. These changes were more pronounced in M. inclinata and M. quercophilus. The results indicate that different litter-decomposing fungi can cause substantial litter transformation despite considerable differences in the production of lignocellulose-degrading enzymes.