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Demonstration of improved dairy products through the application of starter strains of lactic acid bacteria with engineered fermentation pathways

Objectif



Lactic acid bacteria have long been used for the preparation of a number of food products from vegetal and animal origins. Their role in food processing is two-fold: to ensure transformation and conservation through the production of lactic acid, and to bring flavour and taste either indirectly through the secretion or release of hydrolytic enzymes such as proteases, peptidases or lipases, or directly through the production of aromatic compounds such as acetaldehyde or diacetyl. For a long time, lactic fermentation has been achieved through artisanal methods relying exclusively on natural blends of strains propagated from batch to batch in the farmhouse. But the vast majority of dairy products put on the market today in Europe are produced in large, highly industrialized plants with a limited number of starter strains themselves produced industrially and marketed following sophisticated processing such as lyophilization. As a consequence, the dairy industry today relies on a limited number of acidifier starter strains with quite restricted organoleptic profiles, in contrast with the immense diversity of strains with rich taste and flavour potentials available from natural niches. Yet, the current trend is towards mild, sweet and aromatic dairy products.
Our objective is to exploit metabolic engineering technology for the construction of improved dairy starters. Indeed, many starters are too "lactic" in the sense that their fermentative metabolism is primarily channeled towards the production of lactate, and the resulting dairy products are too acid and lack taste. Metabolic engineering will allow re-routing of the carbon flux towards the production of secondary metabolises enhancing the organoleptic qualities of the product. Directed genetic modification offers the advantage over conventional selection or random mutagenesis strategies that it is targeted at a given gene or pathway, leaving intact the whole genetic and physiological backgrounds of the selected starter. This project will be performed in an integrated way by five partners from four European countries: one University laboratory (UCL), two Industry-funded Research Foundations (NIZO and ITG), one large Multinational industry (NRC) and one Small-Medium sized Enterprise (CSK). Demonstration will be in three stages:
1. Metabolically engineered prototype starters will be constructed using state of the art food-grading methods. Three species with distinct dairy applications will be engineered: Lactococcus lactis (Gouda cheese and fermented dairy drinks), Lactobacillus helveticus (Emmental cheese) and Streptococcus thermophilus (yoghurt and Emmental). Engineering will involve modulation of lactate production through ldh genes disruption, production of the natural sweetener alanine through expression of the gene encoding alanine dehydrogenase either constitutively through swapping with lactate dehydrogenase genes or inducibly through nisin-driven expression. Food-grade overexpression of the NADH oxidase gene in an acetolactate decarboxylase mutant will also allow to enhance production of the flavour compound diacetyl.
2. These prototypes will be validated under real industrial conditions to show that they can be safely and reliably used for the production of dairy products with enhanced flavour and taste: buttery flavour (buttermilk), mild and/or sweet taste (yoghurt, Gouda and Emmental cheeses). Products will be manufactured at small and pilot scales and their physico-chemical and organoleptic characteristics will be thoroughly analysed. Validation will also include the demonstration that the properties of the engineered prototypes are maintained following pilot scale production.
3. Since producing and putting GMO on the market is a very sensitive matter for both producers and consumers, we will address an extended audience of representatives of the food industry and consumers associations through workshops, newsletters and website. We see this demonstration project as a test case for improving the public acceptance of properly engineered lactic acid bacteria.

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Coordinateur

Université Catholique de Louvain
Contribution de l’UE
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Adresse
5,Croix de Sud
1348 Louvain-la-Neuve
Belgique

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