Consumption of easily-available carbon does not alter microbial carbon use efficiency in soils

Microorganisms metabolize soil organic carbon (C) as a source of energy and biosynthetic precursors. Conventional metabolic flux analysis (MFA), coupled to 13C-labelling, can reconstruct C allocation through central metabolic pathways, but only reflects mass flow and not the thermodynamics of metabolism. We coupled metabolic energetics (13C), mass flow (18O) and calorespirometry in soil using an optimal set of isotopomer tracers. Fifteen position-specific or uniformly 13C-labelled isotopomers - four for alanine, seven glucose, and four glutamic acid – were added to a Luvisol, and substrate-derived 13CO2 fluxes along with microbial use efficiencies (CUE and SUE) were quantified as well as heat dissipation via isothermal microcalorimetry. Our results demonstrate that the temporal dynamics of catabolic CO2 release resemble that of heat dissipation, with both peaking approximately 18 h after substrate addition, irrespective of whether the tracer enters the central metabolic pathway at the monosaccharide level (glucose), at the pyruvate level (alanine) or the citric acid cycle (glutamic acid). This indicates that heat dissipation during the growth phase was strongly dominated by the microbial metabolic processes. Heat dissipation declined disproportionally compared to C mineralization after multiplicative growth, resulting in a lower calorespirometric ratio. Substrate-derived microbial biomass C (13C-MBC) pools showed that amino acids were incorporated, and retained in the biomass with intensive recycling, whereas glucose gets taken up, incorporated but ongoingly consumed, which leads to the peak biomass. This suggests that sugar may be a good tracer for metabolism. Glucose isotopomer utilization indicated dominance of the pentose phosphate and Entner Douderoff pathways over glycolysis, suggesting high activity of fast-growing organisms with considerable C allocation to anabolism. While calorespirometric ratio declined stepwise from 2426 kJ mol-1 , SUE and EUE were close to 100% during initial stage after the addition and declined when substance respiration started. In contrast, neither 18O-water- nor 13C-MFA-based CUE were altered by substrate supply, indicating that exogeneous substrate did not alter the microbial utilization and microbial quick regulation. Therefore, substrate mixtures do not induce a major shift in metabolic pathways during growing on them, leaving overall CUE largely unaffected. This study shows that the heat dissipation of growing microbial communities under high C supply is closely linked to their catabolic CO2 release. Consumption of easily-available carbon does not alter CUE (i.e. metabolic and physiological state of the soil microbiome), but strongly reduces SUE and EUE during ongoing substrate use. We furthermore demonstrated that coupled MFA and calorespirometry provides a powerful tool to understand in-situ microbial C and energy use in soils.