François Maillard

Mycelial residues, also known as fungal necromass, represent a substantial fraction of soil organic matter (SOM) pools in terrestrial ecosystems worldwide. Although microbial decomposers are increasingly recognised as key drivers of fungal necromass carbon stock formation, the diversity and composition of their microbial predators—and the roles these predators play in mediating fungal necromass decomposition—have not been explored to date. To address this gap, we produced fungal necromass of varying biochemical quality from Hyaloscypha bicolor and decomposed it in forest topsoil in Minnesota, USA, to investigate how microbial decomposer (bacteria and fungi) and predator (protists and nematodes) communities differ between soil and necromass. We also examined whether microbial predators influence the abundance of fungal necromass decomposers and affect necromass decomposition rates. Over two sampling times (4 and 12 weeks), necromass exhibited rapid early mass loss followed by reduced decay, with a higher stabilised mass in high melanin necromass. Microbial abundances were higher in necromass than in surrounding soil, especially in low melanin necromass. Community composition of both decomposers and their predators differed between soil and necromass and shifted markedly with necromass quality and decomposition stage. Predator community composition was linked to bacterial and fungal abundances at both early and late stages of decay and was marginally associated with decomposition rates. We conclude that fungal necromass acts as a microbial ‘hotspot’ not only for decomposers but also for their predators. These findings highlight the importance of microbial predator–decomposer interactions to better understand the formation of fungal‐derived SOM.

Changes in temperature and rainfall regimes will have significant yet potentially contrasting impacts on rates of soil organic matter (SOM) decomposition. To assess how a combined stress treatment of warming and drought impacts the decomposition of fungal necromass—a fast-cycling soil organic matter (SOM) pool—we incubated Hyaloscypha bicolor necromass under both ambient and altered conditions (air and soil warming +3.3°C and ∼40% reduced rainfall) at the B4Warmed experiment in Minnesota, USA. We conducted two multi-week incubations, one assessing mass loss and microbial community composition on decaying necromass after 1, 2, 7, and 14 weeks and the second characterizing the substrate utilization capacities of necromass- associated microbial communities after weeks 1 and 7. Warming and reduced rainfall significantly accelerated the initial rate of necromass decay by ∼20%, but overall mass loss was not different between treatments at the end of the 14-week incubation. The accelerated initial rate of decay paralleled shifts in microbial community composition and activity in the altered plots, demonstrating a higher metabolic capability to utilize C and N substrates early in decomposition but a lower capability later in decay. These findings highlight the dynamic, stage-dependent response of fungal necromass decomposition to altered climate regimes, underscoring the importance of considering both temporal dynamics and the functional capacity of microbial communities when assessing the impacts of climate change on soil carbon and nutrient cycling in forest ecosystems.

The keystone species concept holds that certain members of an ecological community, despite their low abundance, exert disproportionately large effects on species diversity and composition. In microbial ecology, experimental validation of this concept has been limited because targeted removal of individual species remains technically challenging. Here, we developed a procedure to test the keystone species concept within a soil microbial food web by selectively suppressing a protist predator at the microscale via phototoxicity in a microfluidic soil chip system. We targeted a hypotrich ciliate (subclass Hypotrichia), and combined microscopy with high-throughput amplicon sequencing of microbial taxonomic markers to assess, across multiple trophic levels, how its suppression affected microbial community abundance, diversity, and composition. Over the 20-day incubation, the chip system supported complex communities of bacteria, fungi, and protists. Following Hypotrichia suppression, two distinct ecological responses were observed: first, an increase in the relative abundance of flagellates, consistent with mesopredator release, accompanied by a significant rise in overall protist diversity; second, a convergence in protist community composition, indicative of biotic homogenization. Bacterial community abundance, richness, and composition remained unchanged, likely due to compensatory predation from a relative increase in bacterivorous flagellates. In contrast, fungal diversity decreased, presumably because the altered protist community favored facultative fungal consumers. Collectively, these findings provide direct experimental evidence that low-abundance microbial predators can function as keystone species, modulating predator community composition and diversity, and exerting cascading effects on lower trophic levels within microbial brown food webs.

Despite fungal necromass being increasingly recognized a major source of carbon (C) and nitrogen (N) in soils, understanding of how soil mineral and microbial properties interact with initial necromass chemistry to influence its accumulation and persistence remains limited. To track the fate of fungal C and N into mineral associated organic matter (MAOM), we generated dual-labeled ¹³C and ¹⁵N fungal necromass of high and low melanin phenotypes of Hyaloscypha bicolor. Necromass was incubated in live and sterile soils from six forests in Indiana, USA that differed in initial microbial and physiochemical characteristics. At the end of the 38-day incubation, considerably more initial fungal necromass N was incorporated into MAOM relative to necromass C and low melanin necromass contributed more to MAOM-C formation. Though soil iron oxide (FeOx) and clay content were positively correlated with MAOM-C formation from low melanin necromass, initial microbial community composition was the best predictor of MAOM-N formation for both high and low melanin necromass additions. In particular, soils with higher relative abundances of bacterial copiotrophs at the start of the study formed less fungal necromass-derived MAOM-N. We also used synthesized FeOx minerals to investigate how initial necromass chemistry impacts mineral sorption in vitro. We found higher adsorption of amide functionalities for low melanin necromass on goethite minerals after 24 hours. However, 30% less initial necromass-N was converted into MAOM-N in sterile soils regardless of necromass type. When considered together, our results indicate that the microbial synthesis pathway drives fungal necromass-derived MAOM-N formation and that microbial processing limits fungal necromass-derived MAOM-C formation, highlighting the importance of microbial controls on the preservation of fungal-derived SOM in soils.

Summary: Ectomycorrhizal (EcM) fungi play a crucial role in the mineral nitrogen (N) nutrition of their host trees. While it has been proposed that several EcM species also mobilize organic N, studies reporting the EcM ability to degrade N‐containing polymers, such as chitin, remain scarce. Here, we assessed the capacity of a representative collection of 16 EcM species to acquire ¹⁵N from ¹⁵N‐chitin. In addition, we combined genomics and transcriptomics to identify pathways involved in exogenous chitin degradation between these fungal strains. Boletus edulis, Imleria badia, Suillus luteus, and Hebeloma cylindrosporum efficiently mobilized N from exogenous chitin. EcM genomes primarily contained genes encoding for the direct hydrolysis of chitin. Further, we found a significant relationship between the capacity of EcM fungi to assimilate organic N from chitin and their genomic and transcriptomic potentials for chitin degradation. These findings demonstrate that certain EcM fungal species depolymerize chitin using hydrolytic mechanisms and that endochitinases, but not exochitinases, represent the enzymatic bottleneck of chitin degradation. Finally, this study shows that the degradation of exogenous chitin by EcM fungi might be a key functional trait of nutrient cycling in forests dominated by EcM fungi.