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International audience; Predicting mixing processes, especially transverse mixing, downstream of river confluences, is necessary for assessing and modeling the fate of pollutants transported in river networks, but it is still challenging. Typically, there is a lack of transverse mixing solutions implemented in 1-D hydrodynamical models widely used in river engineering applications. To investigate the mixing processes developing downstream of a medium-sized river confluence, three high-resolution in situ surveys are conducted at the Rhône-Saône confluence in France, based on geolocated specific conductivity and hydroacoustic measurements. Contrasting mixing situations are observed depending on hydrological conditions. In some cases, the two flows mix slowly due to turbulent shear at their vertical interface. This can be modeled by an analytical solution of the advection-diffusion equation. In other cases, the waters from one of the two tributaries move under the waters of the other tributary. The induced local circulation enhances transverse mixing but not vertical mixing and the flow remains stratified vertically, which may be missed when surface or satellite images are analyzed qualitatively. Stratification may be predicted by comparing the time scales for shear and density-driven adjustment. Shear-dominated transverse mixing of depth-averaged concentrations can be predicted analytically and implemented in 1-D hydrodynamical models. However, the initiation of apparently rapid transverse mixing due to density-driven circulation remains to be better understood and quantified. Plain Language Summary Predicting how waters mix downstream of river confluences is necessary for assessing and modeling the fate of pollutants transported in river networks, but it is still challenging. Typically, there is a lack of transverse mixing solutions implemented in models widely used in river engineering applications. To investigate the mixing processes developing downstream of a medium-sized river confluence, three high-resolution in situ surveys are conducted at the Rhône-Saône confluence in France. Contrasting slow or rapid mixing situations are observed depending on hydrological conditions. The transverse mixing of depth-averaged concentrations can be predicted analytically and implemented in 1-D hydrodynamical models. However, the initiation of rapid transverse mixing due to difference in fluid density remains to be better understood and quantified.

International audience; Nuclear magnetic resonance (NMR)-based fluxomics seeks to measure the incorporation of isotope labels in selected metabolites to follow kinetically the synthesis of the latter. It can however equally be used to understand the biosynthetic origin of the same metabolites. We investigate here different NMR approaches to optimize such experiments in terms of resolution and time requirement. Using the isoleucine biosynthesis as an example, we explore the use of different field strengths ranging from 500 MHz to 1.1 GHz. Because of the different field dependence of chemical shift and heteronuclear J couplings, the spectra change at different field strengths. We equally explore the approach to silence the leucine/valine methyl signals through the use of a suitable deuterated precursor, thereby allowing selective observation of the Ile C-13 labeling pattern. Combining both approaches, we arrive at an efficient procedure for the NMR-based exploration of Ile biosynthesis.

Prebiotics are increasingly used as food supplements, especially in infant formulas, to modify the functioning and composition of the microbiota. However, little is currently known about the mechanisms of prebiotic recognition and transport by gut bacteria, while these steps are crucial in their metabolism. In this study, we established a new strategy to profile the specificity of oligosaccharide transporters, combining microbiomics, genetic locus and strain engineering, and state-of-the art metabolomics. We revisited the transporter classification database and proposed a new way to classify these membrane proteins based on their structural and mechanistic similarities. Based on these developments, we identified and characterized, at the molecular level, a fructooligosaccharide transporting phosphotransferase system, which constitutes a biomarker of diet and gut pathology. The deciphering of this prebiotic metabolization mechanism by a nonbeneficial bacterium highlights the controversial use of prebiotics, especially in the context of chronic gut diseases., Prebiotic oligosaccharides, such as fructooligosaccharides, are increasingly being used to modulate the composition and activity of the gut microbiota. However, carbohydrate utilization analyses and metagenomic studies recently revealed the ability of deleterious and uncultured human gut bacterial species to metabolize these functional foods. Moreover, because of the difficulties of functionally profiling transmembrane proteins, only a few prebiotic transporters have been biochemically characterized to date, while carbohydrate binding and transport are the first and thus crucial steps in their metabolization. Here, we describe the molecular mechanism of a phosphotransferase system, highlighted as a dietary and pathology biomarker in the human gut microbiome. This transporter is encoded by a metagenomic locus that is highly conserved in several human gut Firmicutes, including Dorea species. We developed a generic strategy to deeply analyze, in vitro and in cellulo, the specificity and functionality of recombinant transporters in Escherichia coli, combining carbohydrate utilization locus and host genome engineering and quantification of the binding, transport, and growth rates with analysis of phosphorylated carbohydrates by mass spectrometry. We demonstrated that the Dorea fructooligosaccharide transporter is specific for kestose, whether for binding, transport, or phosphorylation. This constitutes the biochemical proof of effective phosphorylation of glycosides with a degree of polymerization of more than 2, extending the known functional diversity of phosphotransferase systems. Based on these new findings, we revisited the classification of these carbohydrate transporters. IMPORTANCE Prebiotics are increasingly used as food supplements, especially in infant formulas, to modify the functioning and composition of the microbiota. However, little is currently known about the mechanisms of prebiotic recognition and transport by gut bacteria, while these steps are crucial in their metabolism. In this study, we established a new strategy to profile the specificity of oligosaccharide transporters, combining microbiomics, genetic locus and strain engineering, and state-of-the art metabolomics. We revisited the transporter classification database and proposed a new way to classify these membrane proteins based on their structural and mechanistic similarities. Based on these developments, we identified and characterized, at the molecular level, a fructooligosaccharide transporting phosphotransferase system, which constitutes a biomarker of diet and gut pathology. The deciphering of this prebiotic metabolization mechanism by a nonbeneficial bacterium highlights the controversial use of prebiotics, especially in the context of chronic gut diseases.