Aeration and mineral composition of soil mediate microbial CUE

Abstract. In ecosystem studies, microbial carbon use efficiency (CUE) is often used to estimate the proportion of organic substrate (glucose) consumed by microbial biomass that is not released from soil as CO2. While most studies assume aerobic conditions, with CO2 and microbial biomass as the predominant products of organic substrate processing, anoxic microniches are common inside soil aggregates. Microorganisms in these microniches perform fermentation and anaerobic respiration using alternative electron acceptors: processes connected with the release of extracellular intermediates. These extracellular intermediates and other compounds are not traditionally accounted for but may represent a significant C flux when compared to microbial biomass formation. Climate change may modulate soil microbial activity by altering soil aeration status on a local level. Therefore, CUE as an intrinsic parameter that is used in ecosystem studies and modelling should be defined for more realistic assumptions regarding soil aeration. This study focused on the effect of oxygen and Fe availability on C mineralisation in forest soils and quantified C distribution between biomass and different extracellular metabolites. Forest soils from two Bohemian Forest (Czechia) sites, with low and high Fe content, were incubated under oxic and anoxic conditions. A solution of 13C-labelled glucose was used to track C incorporation into the biomass, respired CO2, and extracellular metabolites. We estimated CUE based on measured cumulative microbial respiration, residual glucose, biomass, and extracellular metabolites concentration. RNA-SIP was used to identify the active bacteria under each treatment. Under oxic conditions, glucose was rapidly consumed and largely converted to CO2, with greater microbial biomass and CUE observed in the low-Fe soil compared to the high-Fe soil. In contrast, under anoxic conditions, glucose consumption was slower, leading to the accumulation of fermentation products, especially in the high-Fe soil, and higher carbon storage efficiency. Microbial growth and turnover were generally lower under anoxic conditions. A large and diverse portion of the microbial community rapidly incorporated 13C-labelled glucose under oxic conditions, with over 300 active amplicon sequence variants (ASVs) identified – primarily from dominant phyla like Proteobacteria, Actinomycetota, and Bacteroidota. In contrast, anoxic conditions led to much slower and more limited labelling, with only a few ASVs (mainly Firmicutes) incorporating 13C. Our findings confirm that anoxia in soils enhances short-term C preservation and suggest that excluding exudates in mass flux calculations would underestimate C retention in the soil, especially under anoxic conditions.