Fungal community composition and genetic potential regulate fine root decay in northern temperate forests.

Understanding how genetic differences among soil microorganisms regulate spatial patterns in litter decay remains a persistent challenge in ecology. Despite fine root litter accounting for ~50% of total litter production in forest ecosystems, far less is known about the microbial decay of fine roots relative to aboveground litter. Here, we evaluated whether fine root decay occurred more rapidly where fungal communities have a greater genetic potential for litter decay. Additionally, we tested if linkages between decay and fungal genes can be adequately captured by delineating saprotrophic and ectomycorrhizal fungal functional groups based on whether they have genes encoding certain ligninolytic class II peroxidase enzymes, which oxidize lignin and polyphenolic compounds. To address these ideas, we used a litterbag study paired with fungal DNA barcoding to characterize fine root decay rates and fungal community composition at the landscape scale in northern temperate forests, and we estimated the genetic potential of fungal communities for litter decay using publicly available genomes. Fine root decay occurred more rapidly where fungal communities had a greater genetic potential for decay, especially of cellulose and hemicellulose. Fine root decay was positively correlated with ligninolytic saprotrophic fungi and negatively correlated with ECM fungi with ligninolytic peroxidases, likely because these saprotrophic and ectomycorrhizal functional groups had the highest and lowest genetic potentials for plant cell wall degradation, respectively. These fungal variables overwhelmed direct environmental controls, suggesting fungal community composition and genetic variation are primary controls over fine root decay in temperate forests at regional scales.

Decay by ectomycorrhizal fungi couples soil organic matter to nitrogen availability.

Interactions between soil nitrogen (N) availability, fungal community composition, and soil organic matter (SOM) regulate soil carbon (C) dynamics in many forest ecosystems, but context dependency in these relationships has precluded general predictive theory. We found that ectomycorrhizal (ECM) fungi with peroxidases decreased with increasing inorganic N availability across a natural inorganic N gradient in northern temperate forests, whereas ligninolytic fungal saprotrophs exhibited no response. Lignin-derived SOM and soil C were negatively correlated with ECM fungi with peroxidases and were positively correlated with inorganic N availability, suggesting decay of lignin-derived SOM by these ECM fungi reduced soil C storage. The correlations we observed link SOM decay in temperate forests to tradeoffs in tree N nutrition and ECM composition, and we propose SOM varies along a single continuum across temperate and boreal ecosystems depending upon how tree allocation to functionally distinct ECM taxa and environmental stress covary with soil N availability.

Anthropogenic N deposition alters soil organic matter biochemistry and microbial communities on decaying fine roots.

Fine root litter is a primary source of soil organic matter (SOM), which is a globally important pool of C that is responsive to climate change. We previously established that ~20 years of experimental nitrogen (N) deposition has slowed fine root decay and increased the storage of soil carbon (C; +18%) across a widespread northern hardwood forest ecosystem. However, the microbial mechanisms that have directly slowed fine root decay are unknown. Here, we show that experimental N deposition has decreased the relative abundance of Agaricales fungi (-31%) and increased that of partially ligninolytic Actinobacteria (+24%) on decaying fine roots. Moreover, experimental N deposition has increased the relative abundance of lignin-derived compounds residing in SOM (+53%), and this biochemical response is significantly related to shifts in both fungal and bacterial community composition. Specifically, the accumulation of lignin-derived compounds in SOM is negatively related to the relative abundance of ligninolytic Mycena and Kuehneromyces fungi, and positively related to Microbacteriaceae. Our findings suggest that by altering the composition of microbial communities on decaying fine roots such that their capacity for lignin degradation is reduced, experimental N deposition has slowed fine root litter decay, and increased the contribution of lignin-derived compounds from fine roots to SOM. The microbial responses we observed may explain widespread findings that anthropogenic N deposition increases soil C storage in terrestrial ecosystems. More broadly, our findings directly link composition to function in soil microbial communities, and implicate compositional shifts in mediating biogeochemical processes of global significance.

Anthropogenic N deposition, fungal gene expression, and an increasing soil carbon sink in the Northern Hemisphere.

Terrestrial ecosystems in the Northern Hemisphere are a globally important sink for anthropogenic CO2 in the Earth's atmosphere, slowing its accumulation as well as the pace of climate warming. With the use of a long-term field experiment (ca. 20 yr), we show that the expression of fungal class II peroxidase genes, which encode enzymes mediating the rate-limiting step of organic matter decay, are significantly downregulated (-60 to -80%) because of increases in anthropogenic N deposition; this response was consistent with a decline in extracellular peroxidase enzyme activity in soil, the slowing of organic-matter decay, and greater soil C storage. The reduction in peroxidase expression we document here occurred in the absence of a compositional shift in metabolically active fungi, indicating that an overall reduction in peroxidase expression underlies the slowing of decay and increases in soil C storage. This molecular mechanism has global implications for soil C storage and should be represented in coupled climate-biogeochemical models simulating the influence of enhanced terrestrial C storage on atmospheric CO2 and the future climate of an N-enriched Earth.

Soil microbial communities and elk foraging intensity: implications for soil biogeochemical cycling in the sagebrush steppe.

Foraging intensity of large herbivores may exert an indirect top-down ecological force on soil microbial communities via changes in plant litter inputs. We investigated the responses of the soil microbial community to elk (Cervus elaphus) winter range occupancy across a long-term foraging exclusion experiment in the sagebrush steppe of the North American Rocky Mountains, combining phylogenetic analysis of fungi and bacteria with shotgun metagenomics and extracellular enzyme assays. Winter foraging intensity was associated with reduced bacterial richness and increasingly distinct bacterial communities. Although fungal communities did not respond linearly to foraging intensity, a greater ß-diversity response to winter foraging exclusion was observed. Furthermore, winter foraging exclusion increased soil cellulolytic and hemicellulolytic enzyme potential and higher foraging intensity reduced chitinolytic gene abundance. Thus, future changes in winter range occupancy may shape biogeochemical processes via shifts in microbial communities and subsequent changes to their physiological capacities to cycle soil C and N.

Anthropogenic N deposition increases soil organic matter accumulation without altering its biochemical composition.

Accumulating evidence indicates that future rates of atmospheric N deposition have the potential to increase soil C storage by reducing the decay of plant litter and soil organic matter (SOM). Although the microbial mechanism underlying this response is not well understood, a decline in decay could alter the amount, as well as biochemical composition of SOM. Here, we used size-density fractionation and solid-state 13 C-NMR spectroscopy to explore the extent to which declines in microbial decay in a long-term (ca. 20 yrs.) N deposition experiment have altered the biochemical composition of forest floor, bulk mineral soil, as well as free and occluded particulate organic matter. Significant amounts of organic matter have accumulated in occluded particulate organic matter (~20%; oPOM); however, experimental N deposition had not altered the abundance of carboxyl, aryl, alkyl, or O/N-alkyl C in forest floor, bulk mineral soil, or any soil fraction. These observations suggest that biochemically equivalent organic matter has accumulated in oPOM at a greater rate under experimental N deposition, relative to the ambient treatment. Although we do not understand the process by which experimental N deposition has fostered the occlusion of organic matter by mineral soil particles, our results highlight the importance of interactions among the products of microbial decay and the chemical and physical properties of silt and clay particles that occlude organic matter from microbial attack. Because oPOM can reside in soils for decades to centuries, organic matter accumulating under future rates of anthropogenic N deposition could remain in soil for long periods of time. If temperate forest soils in the Northern Hemisphere respond like those in our experiment, then unabated deposition of anthropogenic N from the atmosphere has the potential to foster greater soil C storage, especially in fine-texture forest soils.

Microbial Potential for Ecosystem N Loss Is Increased by Experimental N Deposition.

Fossil fuel combustion and fertilizer use has increased the amount of biologically available N entering terrestrial ecosystems. Nonetheless, our understanding of how anthropogenic N may alter the physiological mechanisms by which soil microorganisms cycle N in soil is still developing. Here, we applied shotgun metagenomics to a replicated long-term field experiment to determine how two decades of experimental N deposition, at a rate expected by mid-century, has affected the genetic potential of the soil microbial community to cycle N in soils. Experimental N deposition lead to a significant and persistent increase in functional assemblages mediating N cycle transformations associated with ecosystem N loss (i.e., denitrification and nitrification), whereas functional assemblages associated with N input and retention (i.e., N fixation and microbial N assimilation) were less positively affected. Furthermore, the abundance and composition of microbial taxa, as well as functional assemblages involved in housekeeping functions (i.e., DNA replication) were unaffected by experimental N deposition. Taken together, our results suggest that functional genes and gene pathways associated with ecosystem N loss have been favored by experimental N deposition, which may represent a genetic mechanism fostering increased N loss as anthropogenic N deposition increases in the future.

Active microorganisms in forest soils differ from the total community yet are shaped by the same environmental factors: the influence of pH and soil moisture.

Predicting the impact of environmental change on soil microbial functions requires an understanding of how environmental factors shape microbial composition. Here, we investigated the influence of environmental factors on bacterial and fungal communities across an expanse of northern hardwood forest in Michigan, USA, which spans a 500-km regional climate gradient. We quantified soil microbial community composition using high-throughput DNA sequencing on coextracted rDNA (i.e. total community) and rRNA (i.e. active community). Within both bacteria and fungi, total and active communities were compositionally distinct from one another across the regional gradient (bacteria P = 0.01; fungi P < 0.01). Taxonomically, the active community was a subset of the total community. Compositional differences between total and active communities reflected changes in the relative abundance of dominant taxa. The composition of both the total and active microbial communities varied by site across the gradient (P < 0.01) and was shaped by differences in soil moisture, pH, SOM carboxyl content, as well as C and N concentration. Our study highlights the importance of distinguishing between metabolically active microorganisms and the total community, and emphasizes that the same environmental factors shape the total and active communities of bacteria and fungi in this ecosystem.

Anthropogenic N Deposition Slows Decay by Favoring Bacterial Metabolism: Insights from Metagenomic Analyses.

Litter decomposition is an enzymatically-complex process that is mediated by a diverse assemblage of saprophytic microorganisms. It is a globally important biogeochemical process that can be suppressed by anthropogenic N deposition. In a northern hardwood forest ecosystem located in Michigan, USA, 20 years of experimentally increased atmospheric N deposition has reduced forest floor decay and increased soil C storage. Here, we paired extracellular enzyme assays with shotgun metagenomics to assess if anthropogenic N deposition has altered the functional potential of microbial communities inhabiting decaying forest floor. Experimental N deposition significantly reduced the activity of extracellular enzymes mediating plant cell wall decay, which occurred concurrently with changes in the relative abundance of metagenomic functional gene pathways mediating the metabolism of carbohydrates, aromatic compounds, as well as microbial respiration. Moreover, experimental N deposition increased the relative abundance of 50 of the 60 gene pathways, the majority of which were associated with saprotrophic bacteria. Conversely, the relative abundance and composition of fungal genes mediating the metabolism of plant litter was not affected by experimental N deposition. Future rates of atmospheric N deposition have favored saprotrophic soil bacteria, whereas the metabolic potential of saprotrophic fungi appears resilient to this agent of environmental change. Results presented here provide evidence that changes in the functional capacity of saprotrophic soil microorganisms mediate how anthropogenic N deposition increases C storage in soil.

Dispersal limitation and the assembly of soil Actinobacteria communities in a long-term chronosequence.

It is uncertain whether the same ecological forces that structure plant and animal communities also shape microbial communities, especially those residing in soil. We sought to uncover the relative importance of present-day environmental characteristics, climatic variation, and historical contingencies in shaping soil actinobacterial communities in a long-term chronosequence. Actinobacteria communities were characterized in surface soil samples from four replicate forest stands with nearly identical edaphic and ecological properties, which range from 9500 to 14,000 years following glacial retreat in Michigan. Terminal restriction fragment length polymorphism (TRFLP) profiles and clone libraries of the actinobacterial 16S rRNA gene were constructed in each site for phenetic and phylogenetic analysis to determine whether dispersal limitation occurred following glacial retreat, or if community composition was determined by environmental heterogeneity. At every level of examination, actinobacterial community composition most closely correlated with distance, a surrogate for time, than with biogeochemical, plant community, or climatic characteristics. Despite correlation with leaf litter C:N and annual temperature, the significant and consistent relationship of biological communities with time since glacial retreat provides evidence that dispersal limitation is an ecological force structuring actinobacterial communities in soil over long periods of time.

Isolation of fungal cellobiohydrolase I genes from sporocarps and forest soils by PCR.

Cellulose is the major component of plant biomass, and microbial cellulose utilization is a key step in the decomposition of plant detritus. Despite this, little is known about the diversity of cellulolytic microbial communities in soil. Fungi are well known for their cellulolytic activity and mediate key functions during the decomposition of plant detritus in terrestrial ecosystems. We developed new oligonucleotide primers for fungal exocellulase genes (cellobiohydrolase, cbhI) and used these to isolate distinct cbhI homologues from four species of litter-decomposing basidiomycete fungi (Clitocybe nuda, Clitocybe gibba, Clitopilus prunulus, and Chlorophyllum molybdites) and two species of ascomycete fungi (Xylaria polymorpha and Sarcoscypha occidentalis). Evidence for cbhI gene families was found in three of the four basidiomycete species. Additionally, we isolated and cloned cbhI genes from the forest floor and mineral soil of two upland forests in northern lower Michigan, one dominated by oak (Quercus velutina, Q. alba) and the other dominated by sugar maple (Acer saccharum) and American basswood (Tilia americana). Phylogenetic analysis demonstrated that cellobiohydrolase genes recovered from the floor of both forests tended to cluster with Xylaria or in one of two unidentified groups, whereas cellobiohydrolase genes recovered from soil tended to cluster with Trichoderma, Alternaria, Eurotiales, and basidiomycete sequences. The ability to amplify a key fungal gene involved in plant litter decomposition has the potential to unlock the identity and dynamics of the cellulolytic fungal community in situ.