Caractérisation de bactéries lactiques psychrotrophes en vue de leur utilisation dans la biopréservation des aliments (étude physiologique et moléculaire des mécanismes d'adaptation au froid)

Des souches de bactéries lactiques ont été isolées à partir de produits de la mer et regroupées en fonction de leur pouvoir inhibiteur contre 14 bactéries appartenant à la flore d'altération ou pathogène de ces produits, dans le but de développer des applications en biopréservation. Elles ont été identifiées majoritairement comme leuconostoc gelidum et Lactococcus piscium. L'identification approfondie d'une souche de Lc. piscium montre qu'il s'agit d'une nouvelle espèce de Lactococcus. Des analyses sensorielles ont permis de montrer que 2 souches appartenant à l'espèce Leuconostoc gelidum permettait de prolonger la durée de vie de crevettes cuites emballées sous vide et qu'une souche de Lactococcus sp. avait le même effet sur du saumon fumé. Un test d'inhibition de bactéries pathogènes (Listeria monocytogenes, Staphylococcus aureus et Vibrio cholerae) a été effectué sur du saumon fumé. La souche de Lc.sp.EU2241 (non productrice de bactériocine) s'est révélée plus efficace que la souche de Ln. Gelidum EU 2247 (productrice de bactériocine) pour inhiber ces pathogènes, avec une inhibition atteignant 2 log par rapport au temoin non biopréservé. Les caractéristiques de croissance de la souche Lc sp. EU2241 à différentes températures ont montré un optimum de croissance à 26C et aucune croissance à 29C ou au dessus. L'analyse du protéome a révélé la surproduction d'une protéine de choc froid lors de la croissance à 5C, et le gène correspondant a pu être identifié et séquencé. Il s'agit de la première mise en évidence de la surproduction d'une protéine de choc froid lors de la croissance à température suboptimale pour une bactérie lactique.Lactic acid bacteria strains were isolated from seafood products and clustered according to their inhibiting capacities against 14 food borne spoiling and pathogenic strains in order to develop applications in biopreservation. Most of the strains were identified as Leuconostoc gelidum and Lactococcus piscium. Detailed identification of one strain of the Lactococcus piscium strains suggests that it is a new specie. According to sensory analysis results, 2 strains belonging to the Leuconostoc gelidum specie were the most efficient for the enhancement of the shelf life of vacuum packed peeled and cooked shrimps. Similar results were obtained in cold smoked salmon with one strains of Lactococcus sp. A challenge test against three pathogenic strains (Listeria monocytogenes, Staphylococcus aureus and Vibrio cholerae) was performed in cold smoked salmon. The Lc. sp. EU2241 strain (non bacteriocinogenic) enabled a more efficient inhibition of the pathogens than the Ln. gelidum EU2247 strain (bacteriocinogenic). The maximum recorded inhibition was 2 logs compared to the uninoculated samples. Growth characteristics of strain Lc. sp. EU2241 were determined at different temperatures showed an optimal growth at 26C and no growth at 29C or above. Proteomic analysis showed a cold shock protein overproduction during growth at 5C. The gene coding for this protein was identified and sequenced. Overproduction of a cold shock protein during growth at suboptimal temperature has never been described in a lactic acid bacterium before.NANTES-BU Sciences (441092104) / SudocSudocFranceF

Etudes des interactions entre une bactérie bioprotectrice, Lactococcus piscium CNCM I-4031, et Brochotrix thermosphacta et Listeria monocytogenes dans la crevette tropicale

Le potentiel bioprotecteur de Lactococcus piscium CNCM I-4031 vis à vis d une bactérie altérante, Brochothrix thermosphacta, a été testé sur de la crevette cuite décortiquée conservée sous atmosphère protectrice. En co-culture, L. piscium permet d inhiber la croissance de B. thermosphacta de 4 log ufc/g, tout en améliorant la qualité sensorielle du produit pendant 31 jours. L inhibition n est pas due à la production d acide lactique ni à une compétition pour les acides aminés libres. La croissance de L. piscium et B. thermosphacta a été modélisée en fonction du pH, de la température et du taux de NaCl. Les valeurs optimales de croissance sont proches mais B. thermosphacta présente une tolérance au sel plus élevée et L. piscium un caractère psychrotolérant plus marqué. L effet du niveau d inoculation par L. piscium, en combinaison avec la température et le sel, sur l interaction avec B. thermosphacta a été modélisé. Un inoculum fort en L. piscium (6-7 log ufc/g) est nécessaire pour maintenir le nombre de B. thermosphacta inférieur à 4 log ufc/g, cependant une amélioration sensorielle de la crevette est obtenue même avec un inoculum assez bas. L inhibition de Listeria monocytogenes par L. piscium a ensuite été testée sur la crevette cuite, montrant une inhibition de 4 log ufc/g. Les mécanismes d action impliqués dans cette activité ont été étudiés sur un milieu chimiquement défini (MSMA) reproduisant l inhibition. Celle-ci n est pas liée à la consommation du glucose, la production d acide lactique ou la compétition pour les acides aminés, les bases ou les vitamines. En revanche le contact cellulaire entre les deux bactéries est nécessaire pour obtenir l inhibition.The bioprotective potential of Lactococcus piscium CNCM I-4031 towards a specific spoilage organism, Brochothrix thermosphacta, was studied on tropical cooked peeled shrimp packed under modified atmosphere. In co-culture, the growth of this spoilage bacterium was inhibited by 4 log ufc/g by L. piscium and the sensory quality of the product was improved during 31 days. The inhibition could not be attributed to lactic acid or nutritional competition for amino acids. The growth modelling of L. piscium and B. thermosphacta was performed in different conditions of temperature, pH and NaCl concentrations. The optimal growth values of these two bacteria are quite similar, however L. piscium seemed to be more psychrotolerant than B. thermosphacta but twice less tolerant to salt. The effect of the inoculation level of L. piscium in combination with temperature and salt on the interaction with B. thermosphacta was modelled. A high concentration (6-7 log ufc/g) was necessary to maintain the number of B. thermosphacta lower than 4 log ufc/g. However sensory improvement of shrimp was obtained with lower inoculation level. Lastly, the interaction between L. piscium and Listeria monocytogenes was studied on cooked peeled shrimp showing a 4 log cfu/g inhibition. The mechanisms involved in this inhibition were studied on a chemically defined medium (MSMA) allowing to obtain a similar inhibition. The inhibitory activity was not linked to glucose consumption, lactic acid production or competition for amino acids, bases or vitamins. However, cellular contact is necessary to obtain the inhibition.NANTES-BU Sciences (441092104) / SudocSudocFranceF

Influence of temperature, pH and NaCl concentration on the maximal growth rate of Brochothrix thermosphacta and a bioprotective bacteria Lactococcus piscium CNCM I-4031

International audienceThe maximum specific growth rate (mu(max)) of Brochothrix thermosphacta, a spoilage bacteria of cooked peeled shrimp, and Lactococcus piscium CNCM I-4031, a bioprotective strain, was investigated under different conditions of temperature, NaCl concentrations and pH. The basic modelling approach used was the Gamma concept (gamma-concept) and the model developed was then adapted to shrimp. Cardinal growth parameters were quite similar for the two strains, except for NaCl. No NaCl was required for growth and the NaClmax was three-times higher for B. thermosphacta than for L. piscium (62 and 23 g l(-1) respectively). However, tolerance to NaCl was higher in seafood than in liquid broth, possibly due to presence of osmoltically active molecules. L. piscium and B. thermosphacta were psychrotolerant, with T-min = -4.8 and -3.4 degrees C, T-opt = 23.4 and 27.0 degrees C and T-max = 27.2 and 30.8 degrees C respectively. The optimal pH was neutral and growth possible till pH = 4.8 for the two strains, assuming possible applications of the bioprotective strain in lightly marinated seafood. The mu(max) of B. thermosphacta in shrimp was a little higher than in L piscium whatever the environmental conditions. Validation of the model showed that the gamma-concept was suitable for predicting mu(max) of B. therm osphacta in shrimp. Data generated in this study can be used to adapt the model to other foods with few additional experiments and the effect of different parameters may be added in the future. The model was less accurate for the bioprotective strain and the effect of NaCl must be studied in more detail directly in the matrix. (C) 2012 Elsevier Ltd. All rights reserved

Pulsed light system as a novel food decontamination technology: a review

International audienceIn response to consumer preferences for high quality foods that are as close as possible to fresh products, athermal technologies are being developed to obtain products with high levels of organoleptic and nutritional quality but free of any health risks. Pulsed light is a novel technology that rapidly inactivates pathogenic and food spoilage microorganisms. It appears to constitute a good alternative or a complement to conventional thermal or chemical decontamination processes. This food preservation method involves the use of intense, short-duration pulses of broad-spectrum light. The germicidal effect appears to be due to both photochemical and photothermal effects. Several high intensity flashes of broad spectrum light pulsed per second can inactivate microbes rapidly and effectively. However, the efficacy of pulsed light may be limited by its low degree of penetration, as microorganisms are only inactivated on the surface of foods or in transparent media such as water. Examples of applications to foods are presented, including microbial inactivation and effects on food matrices

Odnos porabnikov do akcijskih nakupov kot oblike pospeševanja prodaje

Optimization of Carnobacterium divergens V41 growth and bacteriocin activity in a culture medium deprived of animal protein, needs for food bioprotection, was performed by using a statistical approach. In a screening experiment, 12 factors (pH, temperature, carbohydrates, NaCl, yeast extract, soy peptone, sodium acetate, ammonium citrate, magnesium sulfate, manganese sulfate, ascorbic acid and thiamine) were tested for their influence on the maximal growth and bacteriocin activity using a two-level incomplete factorial design with 192 experiments performed in microtiter plate wells. Based on results, a basic medium was developed and three variables (pH, temperature and carbohydrates concentration) were selected for a scale-up study in bioreactor. A 23 complete factorial design was performed, allowing the estimation of linear effects of factors and all the first order interactions. The best conditions for the cell production were obtained with a temperature of 15°C and a carbohydrates concentration of 20 g/l whatever the pH (in the range 6.5–8), and the best conditions for bacteriocin activity were obtained at 15°C and pH 6.5 whatever the carbohydrates concentration (in the range 2–20 g/l). The predicted final count of C. divergens V41 and the bacteriocin activity under the optimized conditions (15°C, pH 6.5, 20 g/l carbohydrates) were 2.4 × 1010 CFU/ml and 819200 AU/ml respectively. C. divergens V41 cells cultivated in the optimized conditions were able to grow in cold-smoked salmon and totally inhibited the growth of Listeria monocytogenes (< 50 CFU g−1) during 5 weeks of vacuum storage at 4 and 8°C

Adaptation to Cold and Proteomic Responses of the Psychrotrophic Biopreservative Lactococcus piscium Strain CNCM I-4031

International audienceThere is considerable interest in the use of psychrotrophic bacteria for food biopreservation and in the understanding of cold adaptation mechanisms. The psychrotrophic biopreservative Lactococcus piscium strain CNCM I-4031 was studied for its growth behavior and proteomic responses after cold shock and during cold acclimation. Growth kinetics highlighted the absence of growth latency after cold shock, suggesting a very high promptness in cold adaptation, a behavior that has never been described before for lactic acid bacteria (LAB). A comparative proteomic analysis was applied with two-dimensional gel electrophoresis (2-DE), and upregulated proteins were identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Both cold shock and cold acclimation triggered the upregulation of proteins involved in general and oxidative stress responses and fatty acid and energetic metabolism. However, 2-DE profiles and upregulated proteins were different under both conditions, suggesting a sequence of steps in cold adaptation. In addition, the major 7-kDa Csp protein was identified in the L. piscium CNCM I-4031 genome but was not cold regulated. The implication of the identified cold shock proteins and cold acclimation proteins in efficient cold adaptation, the possible regulation of a histidyl phosphocarrier protein, and the roles of a constitutive major 7-kDa Csp are discussed

Uptake of choline from salmon flesh and its conversion to glycine betaine in response to salt stress in Shewanella putrefaciens

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Lactic acid bacteria selection for biopreservation as a part of hurdle technology approach applied on seafood

As fragile food commodities, microbial and organoleptic qualities of fishery and seafood can quickly deteriorate. In this context, microbial quality and security improvement during the whole food processing chain (from catch to plate), using hurdle technology, a combination of mild preserving technologies such as biopreservation, modified atmosphere packaging and superchilling, are of great interest. Thirty-five lactic acid bacteria (LAB) belonging to the species Carnobacterium maltaromaticum, Carnobacterium inhibens, Lactococcus piscium, Leuconostoc gelidum, Vagococcus fluvialis, Vagococcus penaei and Aerococcus viridans, known to possess interesting antimicrobial activity, were chosen for their potential application as bioprotective agents as a part of hurdle technology applied to fishery products. The selection approach was based on 7 criteria including antimicrobial activity, alteration potential, resistance to chitosan coating and superchilling process, cross inhibition, biogenic amines production (histamine, tyramine) and antibiotics resistance. Antimicrobial activity was assessed against 6 common spoiling bacteria in fishery products (Shewanella baltica, Photobacterium phosphoreum, Brochothrix thermosphacta, Lactobacillus sakei, Hafnia alvei, Serratia proteamaculans) and 1 pathogenic bacteria (Listeria monocytogenes) in co-culture inhibitory assays miniaturized in 96-well microtiter plates. Antimicrobial activity and spoilage evaluation, both performed in cod and salmon juice, highlighted the existence of sensory signatures and inhibition profiles, which seem to be species related. Finally 6 LAB with no unusual antibiotics resistance profile nor histamine production ability were selected as bioprotective agents for further in situ inhibitory assays in cod and salmon based products, alone or in combination with other hurdles (chitosan, modified atmosphere packing and superchilling)

Bioprotective Effect of Lactococcus piscium CNCM I-4031 Against Listeria monocytogenes Growth and Virulence

Listeria monocytogenes is a Gram-positive pathogen occurring in many refrigerated ready-to-eat foods. It is responsible for foodborne listeriosis, a rare but severe disease with a high mortality rate (20–30%). Lactococcus piscium CNCM I-4031 has the capacity to prevent the growth of L. monocytogenes in contaminated peeled and cooked shrimp and in a chemically defined medium using a cell-to-cell contact-dependent mechanism. To characterize this inhibition further, the effect of L. piscium was tested on a collection of 42 L. monocytogenes strains. All strains were inhibited but had different sensitivities. The effect of the initial concentration of the protective and the target bacteria revealed that the inhibition always occurred when L. piscium had reached its maximum population density, whatever the initial concentration of the protective bacteria. Viewed by scanning electron microscopy, L. monocytogenes cell shape and surface appeared modified in co-culture with L. piscium CNCM I-4031. Lastly, L. monocytogenes virulence, evaluated by a plaque-forming assay on the HT-29 cell line, was reduced after cell pre-treatment by the protective bacteria. In conclusion, the bioprotective effect of L. piscium toward L. monocytogenes growth and virulence was demonstrated, and a hypothesis for the inhibition mechanism is put forward

Validity of Soprolife camera and Calcivis device in caries lesion activity assessment

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