Autor:innen:
Prof. Dr. Michaela A. Dippold | University of Tuebingen | Germany
Prof. Dr. Paul Dijkstra | Northern Arizona University | United States
Pro Bruce Hungate | Northern Arizona University | United States
Dr. Lingling Shi | University of Tuebingen | Germany
Prof. Dr. Yakov Kuzyakov | University of Goettingen | Germany
Dr. Weichao Wu | Shanghai Ocean University | China
The central carbon (C) metabolic network harvests energy to power the cell and feed biosynthesis for growth. In pure cultures, bacteria use some but not all of the network’s major pathways, such as glycolysis, pentose phosphate and Entner-Doudoroff pathways. However, how these pathways are used in microorganisms in intact soil communities is unknown. Methodological challenges arise, when multiple metabolic pathways lead to the same product. Then, compound-specific isotope analysis may not provide enough information to quantify the activities of the contributing pathways. Instead, identification of where in the molecule the 13C is incorporated is required. Here we show how knowledge of position-specific 13C incorporation from glucose isotopomers into phospholipid fatty acids (PLFA) and PLFA fragments can be used to quantitatively estimate the fluxes through the central C metabolic network. We developed a method to measure 13C enrichment of PLFA and PLFA fragments (ethanoate, propionate) using electron impact gas chromatography-mass spectrometry and tested this with fatty acids extracted from two pure cultures (Bacillus licheniformis and Pseudomonas fluorescence) after position-specific glucose labeling. Metabolic flux modeling based on the 13C enrichment of ethanoate and propionate fragments showed that B. licheniformis used Embden-Meyerhof-Parnas and pentose phosphate pathway (66% and 30%, respectively), whereas P. fluorescens utilized Entner-Doudoroff and pentose phosphate pathway (72% and 27%). Applying this method to disentangle central C metabolic network activities of co-occurring microbial groups in soils allowed to distinguish at least three groups of PLFA on the basis of their metabolic organization. We showed that the two groups of Gram-positive and the Gram-negative bacteria utilize different pathways to metabolize glucose in an intact agricultural soil and disentangled glucose C allocation to cata- and anabolism. These groups also differ in C Use Efficiency (CUE), the efficiency with which a substrate is used for biosynthesis. We demonstrated that CUE is mainly driven by their citric acid cycle activity. Our results provide experimental evidence for diversity among microbes in the organization of their central carbon metabolic network and CUE under in situ conditions in soils. These results involve important implications for how community composition and their metabolic organization affects soil C cycling and organic matter formation.