The fast milk acidifying phenotype of Streptococcus thermophilus can be acquired by natural transformation of the genomic island encoding the cell-envelope proteinase PrtS

In industrial fermentation processes, the rate of milk acidification by Streptococcus thermophilus is of major technological importance. The cell-envelope proteinase PrtS was previously shown to be a key determinant of the milk acidification activity in this species. The PrtS enzyme is tightly anchored to the cell wall via a mechanism involving the typical sortase A (SrtA) and initiates the breakdown of milk casein into small oligopeptides. The presence or absence of PrtS divides the S. thermophilus strains into two phenotypic groups i.e. the slow and the fast acidifying strains. The aim of this study was to improve the milk acidification rate of slow S. thermophilus strains, and hence optimise the fermentation process of dairy products

CRISPR provides acquired resistance against viruses in prokaryotes

Clustered regularly interspaced short palindromic repeats (CRISPR) are a distinctive feature of the genomes of most Bacteria and Archaea and are thought to be involved in resistance to bacteriophage. We found that following viral challenge, bacteria integrated new spacers derived from phage genomic sequences. Removal or addition of particular spacers modified the phage-resistance phenotype of the cell. Thus, CRISPR, together with associated cas genes, provided resistance against phages, whereby specificity is determined by spacer/phage sequence similarity

The Complete Genome of Propionibacterium freudenreichii CIRM-BIA1T, a Hardy Actinobacterium with Food and Probiotic Applications

Background: Propionibacterium freudenreichii is essential as a ripening culture in Swiss-type cheeses and is also considered for its probiotic use [1]. This species exhibits slow growth, low nutritional requirements, and hardiness in many habitats. It belongs to the taxonomic group of dairy propionibacteria, in contrast to the cutaneous species P. acnes. The genome of the type strain, P. freudenreichii subsp. shermanii CIRM-BIA1 (CIP 103027T), was sequenced with an 11-fold coverage. Methodology/Principal Findings: The circular chromosome of 2.7 Mb of the CIRM-BIA1 strain has a GC-content of 67% and contains 22 different insertion sequences (3.5% of the genome in base pairs). Using a proteomic approach, 490 of the 2439 predicted proteins were confirmed. The annotation revealed the genetic basis for the hardiness of P. freudenreichii, as the bacterium possesses a complete enzymatic arsenal for de novo biosynthesis of aminoacids and vitamins (except panthotenate and biotin) as well as sequences involved in metabolism of various carbon sources, immunity against phages, duplicated chaperone genes and, interestingly, genes involved in the management of polyphosphate, glycogen and trehalose storage. The complete biosynthesis pathway for a bifidogenic compound is described, as well as a high number of surface proteins involved in interactions with the host and present in other probiotic bacteria. By comparative genomics, no pathogenicity factors found in P. acnes or in other pathogenic microbial species were identified in P. freudenreichii, which is consistent with the Generally Recognized As Safe and Qualified Presumption of Safety status of P. freudenreichii. Various pathways for formation of cheese flavor compounds were identified: the Wood-Werkman cycle for propionic acid formation, amino acid degradation pathways resulting in the formation of volatile branched chain fatty acids, and esterases involved in the formation of free fatty acids and esters. Conclusions/Significance: With the exception of its ability to degrade lactose, P. freudenreichii seems poorly adapted to dairy niches. This genome annotation opens up new prospects for the understanding of the P. freudenreichii probiotic activity

Genie microbiologique et technologie a membrane.

*INRA, C.R. de Rennes, Technologie Laitiere, 35042 Rennes Diffusion du document : INRA, C.R. de Rennes, Technologie Laitiere, 35042 RennesNational audienc

Couplage fermentation-ultrafiltration et production de biomasse

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Two-Dimensional Electrophoresis Study of Lactobacillus delbrueckii subsp. bulgaricus Thermotolerance

The response of Lactobacillus delbrueckii subsp. bulgaricus cells to heat stress was studied by use of a chemically defined medium. Two-dimensional electrophoresis (2-DE) analysis was used to correlate the kinetics of heat shock protein (HSP) induction with cell recovery from heat injury. We demonstrated that enhanced viability, observed after 10 min at 65°C, resulted from the overexpression of HSP and from mechanisms not linked to protein synthesis. In order to analyze the thermoadaptation mechanisms involved, thermoresistant variants were selected. These variants showed enhanced constitutive tolerance toward heat shock. However, contrary to the wild-type strain, these variants were poorly protected after osmotic or heat pretreatments. This result suggests that above a certain threshold, cells reach a maximum level of protection that cannot be easily exceeded. A comparison of protein patterns showed that the variants were able to induce more rapidly their adaptive mechanisms than the original strain. In particular, the variants were able to express constitutively more HSP, leading to the higher level of thermoprotection observed. This is the first report of the study by 2-DE of the heat stress response in L. delbrueckii subsp. bulgaricus

Le séchage des microorganismes par atomisation

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Lactobacillus delbrueckii ssp. bulgaricus thermotolerance

International audienceLactobacillus delbrueckii ssp. bulgaricus is a lactic acid bacterium widely used in the dairy food industry. Since the industrial processes are a succession of constraints, it is essential to understand the behaviour of L. bulgaricus when facing usual stresses. The influence of heat stress was investigated on the viability of L. bulgaricus cells grown in a chemically defined medium. The susceptibility of cells to heat-shock was obvious only above 55 $^{\circ}$C. We investigated the acquisition of thermotolerance as a result of exposure to a moderate heat-shock, and the acquisition of a cross-stress-tolerance by exposure to a mild osmotic stress. When cells were submitted, before lethal temperature challenge (65 $^{\circ}$C), to a heat pre-treatment at 50 $^{\circ}$C or to a hyper-osmotic pre-treatment, the viability of cells increased. For the industrial strain RD 546, the addition of glycine betaine (GB) in 0.4 mol.L$^{-1}$ NaCl during the pre-treatment decreased the acquired thermotolerance, while GB alone enhanced cell viability. The thermotolerance of the type strain was not influenced by GB. We demonstrated that the stress tolerance induced by a moderate heat-shock was dependent on protein synthesis, while the effect of GB on RD 546 thermotolerance was independent of such biosynthesis. Thermotolerance acquired in presence of GB depends on a strain-dependant mechanism that differs from the mechanism involved after a moderate heat-shock.Stress thermique et thermotolérance chez Lactobacillus delbrueckii ssp. bulgaricus . Lactobacillus delbrueckii ssp. bulgaricus est une bactérie lactique largement utilisée en industrie alimentaire. Les procédés industriels étant une succession de contraintes, il est essentiel de connaître le comportement de L. bulgaricus face aux stress rencontrés. L'influence du stress thermique sur la viabilité de 2 souches de L. bulgaricus, cultivées en milieu chimiquement défini, a été étudiée. La viabilité des cellules n'est affectée qu'au-delà de 55 $^{\circ}$C. Les cellules acquièrent une thermotolérance vis-à-vis d'un choc thermique à 65 $^{\circ}$C durant 10 min, c'est-à-dire une viabilité accrue, après exposition à un prétraitement thermique modéré à 50 $^{\circ}$C ou un prétraitement hyperosmotique. Pour la souche industrielle RD 546, l'addition de glycine bétaïne (GB) dans le milieu de prétraitement contenant 0,4 mol.L$^{-1}$ NaCl diminue la thermotolérance acquise, tandis que la glycine bétaïne seule en tant que prétraitement avant le choc létal augmente la viabilité cellulaire. La thermotolérance induite par un prétraitement thermique modéré est dépendante de la synthèse protéique, tandis que l'effet de la GB sur la thermotolérance est indépendant de biosynthèse. La thermotolérance acquise en présence de GB nécessite un mécanisme souche-dépendant distinct par rapport à celui impliqué lors d'un prétraitement thermique

Reconstruction in silico de voies métaboliques (application aux voies glycolitiques de Propionibacterium freudenreichii subsp. shermanii)

Depuis la dernière décennie, la recherche en microbiologie fait face à un véritable changement qualitatif et quantitatif avec l émergence de la génomique (qui concerne les génomes et leur analyse) et le développement de la biologie à haut débit (tous les ʺomiquesʺ). Ce changement permet d aborder l étude du fonctionnement cellulaire (voies de signalisations, voies métaboliques) dans sa globalité, et non plus par l étude individuelle de ses acteurs biologiques. Dans le cadre de cette thèse, nous nous intéressons à la reconstruction des voies métaboliques d une bactérie d intérêt agro-alimentaire, Propionibacterium freudenreichii subsp. Shermanii (P. shermanii), à partir de l analyse de son génome. La problématique de la reconstruction de voies métaboliques est double. D une part, il faut pouvoir identifier les fonctions enzymatiques présentes dans l organisme, ce qui se traduit par l analyse du génome de P. shermanii. D autre part, il s agit de reconstruire le réseau des voies métaboliques à partir des fonctions enzymatiques précédemment identifiées. Après avoir mis en place des méthodologies et des outils bioinformatiques permettant d analyser les données génomiques de P. shermanii, nous en avons reconstruit manuellement et automatiquement les voies glycolytiques (glycose et pentose phosphates).RENNES-Agrocampus-CRD (352382323) / SudocSudocFranceF