Metatranscriptomics and metabolic modeling to identify bacterial metabolic interactions during the manufacture of a model pressed cheese.
Résumé
In cheese production, lactic acid bacteria (LAB) and propionic bacteria are key players in the production of metabolites that confer nutritional and organoleptic qualities. However, the contribution of each species to the final quality of cheeses is not fully elucidated. The objective was to determine which species contribute to acidify and produce flavour compounds, by which metabolic pathways, according to which temporality and also to investigate the bacterial metabolic interactions contributing to the functioning of the cheese ecosystem.
We sequenced and annotated: Lactococcus lactis subsp. lactis biovar diacetylactis CIRM-BIA1206 (LL), Lactiplantibacillus plantarum CIRM-BIA465 (LP), Propionibacterium freudenreichii CIRM-BIA122 (PF). We reconstructed the metabolic pathways and developed a community metabolic model. Four semi-hard cheeses were made with LL, LP and PF and were analyzed throughout manufacturing. Bacteria, sugars, organic acids and flavour compounds were quantified and RNA sequenced.
The analysis of differentially expressed genes showed at which moment of the manufacturing process the genes involved in the catabolism of lactose and citrate and in the synthesis pathways of different flavour compounds: lactic, acetic, propionic, isovaleric (old cheese flavour), acetoin (cheesy flavour) acids were induced. Lactose catabolism was induced in LP and PF (Leloir pathway) during acidification and then in LL (tagatose pathway) during ripening. Acetoin production was induced in LL, LP and PF from the beginning of the manufacturing process to the beginning of ripening. The synthesis of propionic and isovaleric acids were attributed to PF and the corresponding metabolisms were induced from the beginning of the manufacturing then stably expressed during the first month of the ripening. Since the mixed fermentation pathways were induced during ripening, acetic acid was likely produced by LL, LP and PF at this stage. Implementation of the metabolic community model in the Smetana tool revealed the molecular basis of the previously discussed commensalism between LAB and PF. LAB produce lactic acid and potentially ribose, succinate, glycerol, serine and phenylalanine for the benefit of PF. These interactions, identified in silico, remain to be validated in vitro.
All these results make metatranscriptomics associated with metabolic models, tools of choice to better understand and control the metabolisms and interactions governing the functioning of cheese ecosystems.
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