Long lasting non-cellular reactions in sterile soils recapitulate most of the intermediates of the Krebs cycle

Over the past decade, chemical evidence has emerged that non-enzymatic metabolic pathways, such as the Krebs cycle, may have existed before cellular life on the primitive Earth. However, the question of whether non-cellular reactions analogous to cell respiration metabolism are still “active” in today’s biosphere and whether they contribute to CO ₂ emissions in contemporary ecosystems remains open. In the present study, we investigated the long-term fate (> 6 months) of organic substrates supplied in sterilised soils in which cell life was not detectable. Through a series of analytical studies performed on the water-extractable fraction of soil exometabolites using chromatography, mass spectrometry and isotope labelling, we demonstrate that sterile soil matrix incubated with [ ¹³ C ₆ ]-glucose and [ ¹³ C ₆ ]-citrate can spontaneously generate intermediates of the Krebs cycle, alongside by-products such as acetate and formate. These findings support the hypothesis that extracellular metabolisms (EXOMETs) form a network of non-cellular reactions resembling metabolic pathways involved in aerobic respiration and anaerobic fermentation of cells. This research not only provides insights into the chemical continuity between chemistry and biochemistry but also raises questions about the implications of non-cellular pathways for soil ecosystem functioning and carbon fluxes in the contemporary biosphere.