Environnement des bactéries et sensibilité aux biocides : mise au point d'une technique rapide pour déterminer in situ l'efficacité bactéricide d'agents antimicrobiens

Controlling the hygiene status of surfaces remains a constant concern in different areas. Indeed, the biofouling of surfaces can be a source of dramatic public health problems in food industries (food toxi-infections) or in hospitals (nosocomial infections). The efficiency of biocides remains variable from one application to the others (especially on adherent micro-organisms), it is necessary to improve the disinfectant formula and/or the associated procedures of disinfection. This improvement involves new test methods that allow rapid assessment of the antibacterial activity of marketed or not products, in conditions closed to the reality. So, through this work, we defined a protocol to determine lethal activity of biocides on sessile bacteria, protocol made up of four principal steps: i) selection of micro-organisms and standardisation of their storage and growth conditions, ii) setting a reproductible adhesion protocol of bacteria on soiled or not surfaces, iii) disinfection test on sessile micro-organisms and iv) optimisation of this protocol for in situ observations and reducing the time needed to obtain results by employing fluorescent markers which give indications on cellular viability.La maîtrise de l'hygiène des surfaces demeure une préoccupation constante dans de nombreux secteurs d'activité. En effet, la contamination microbiologique des surfaces peut être à l'origine de problèmes de santé publique plus ou moins sévères dans les industries agro-alimentaires (toxi-infections alimentaires) ou le milieu médical (infections nosocomiales). L'efficacité des agents désinfectants restant variable d'une application à l'autre (notamment sur des micro-organismes adhérant), il s'avère nécessaire d'améliorer les formulations désinfectantes et/ou les procédures de désinfection qui y sont associées. Cette amiélioration passe par la mise en place de méthodes tests permettant d'évaluer rapidement l'activité antimicrobienne de produits commercialisés ou non, dans des conditions proches de la réalité. Ainsi, au cours de ce travail, nous avons défini un protocole pour déterminer l'activité létale des produits désinfectants sur cellules adhérentes, protocole composé de quatre étapes principales : i) choix des micro-organismes et standardisation de leurs conditions de conservation et de croissance, ii) mise en place d'un protocole d'adhésion reproductible sur supports conditionnés ou non, iii) test de désinfection sur cellules adhérentes et iv) optimisation de ce protocole dans le caractère in situ et le délai d'obtention des résultats par l'utilisation de marqueurs fluorescents, indicateurs de la viabilité cellulaire

ENVIRONNEMENT DES BACTERIES ET SENSIBILITE AUX BIOCIDES (MISE AU POINT D'UNE TECHNIQUE RAPIDE POUR DETERMINER IN SITU L'EFFICACAITE BACTERICIDE D'AGENTS ANTIMICROBIENS)

MASSY-AgroParisTech (913772301) / SudocSudocFranceF

Material influence on biocontamination level and adhering cell physiology Influence des matériaux sur le niveau de biocontamination et la physiologie des cellules adhérentes

Dans la plupart des environnements, les microorganismes vivent préférentiellement au sein de biofilms. De nombreux facteurs influencent leur formation i.e. les nutriments, la température, le régime du fluide environnent, la microflore et les matériaux. Dans le mécanisme de biocontamination, décrit en quatre étapes successives, l'adhésion initiale est un élément clé de la bioadhésion et les matériaux un élément majeur pour son contrôle. L'acier inoxydable est très utilisé dans de nombreux secteurs d'activité pour sa bonne nettoyabilité et son excellente résistance à la corrosion. Pour se différencier, certains matériaux mettent en avant d'autres propriétés. Ainsi, la sélection du matériau le mieux adapté à un problème donné nécessite de connaitre son aptitude à la biocontamination ainsi que son impact sur la physiologie des microorganismes. Pour tous les matériaux testés, la mortalité des bactéries adhérentes est inférieure à 55 %. Les résultats obtenus ont montré qu'un matériau dit antimicrobien n'induit pas plus de cellules endommagées comparativement aux autres matériaux. In most environments, association with a surface in a structure known as a biofilm is the prevailing microbial lifestyle. Several factors may influence the biofilm formation e.g. nutrients, temperature, flow velocity, initial microflora and the nature of materials. Considering the biocontamination mechanism described in four steps, the initial adhesion is a key element in the biocontamination phenomenon and the substratum is of major concern in controlling bacterial adhesion. Stainless steel is well used in numerous markets because of its high cleanability and corrosion resistance properties. However, other materials are put forward by focusing on properties which differentiate them from those of stainless steel. Thereby, to select the material best suited to the problem, there should have data on their aptitude for biocontamination as well as adhesion impact on cell physiology. For all materials, the ratio of dead adhering cells is lower than 55%. The results obtained show that cell injury is not higher on material known to be bactericidal than on other ones

Material influence on biocontamination level and adhering cell physiology

In most environments, association with a surface in a structure known as a biofilm is the prevailing microbial lifestyle. Several factors may influence the biofilm formation e.g. nutrients, temperature, flow velocity, initial microflora and the nature of materials. Considering the biocontamination mechanism described in four steps, the initial adhesion is a key element in the biocontamination phenomenon and the substratum is of major concern in controlling bacterial adhesion. Stainless steel is well used in numerous markets because of its high cleanability and corrosion resistance properties. However, other materials are put forward by focusing on properties which differentiate them from those of stainless steel. Thereby, to select the material best suited to the problem, there should have data on their aptitude for biocontamination as well as adhesion impact on cell physiology. For all materials, the ratio of dead adhering cells is lower than 55%. The results obtained show that cell injury is not higher on material known to be bactericidal than on other ones.Dans la plupart des environnements, les microorganismes vivent pr ́ ef ́ erentiellement au sein de biofilms. De nombreux facteurs influencent leur formation i.e. les nutriments, la temp ́ erature, le r ́ egime du fluide environnent, la microflore et les mat ́ eriaux. Dans le m ́ ecanisme de biocontamination, d ́ ecrit en quatre ́ etapes successives, l’adh ́ esion initiale est un ́ el ́ ement cl ́ e de la bioadh ́ esion et les mat ́ eriaux un ́ el ́ ement majeur pour son contr ˆ ole. L’acier inoxydable est tr ` es utilis ́ e dans de nombreux secteurs d’activit ́ e pour sa bonne nettoyabilit ́ e et son excellente r ́ esistance ` a la corrosion. Pour se diff ́ erencier, certains mat ́ eriaux mettent en avant d’autres propri ́ et ́ es.Ainsi,las ́ election du mat ́ eriau le mieux adapt ́ e ` a un probl ` eme donn ́ en ́ ecessite de connaitre son aptitude ` a la biocontamination ainsi que son impact sur la physiologie des microorganismes. Pour tous les mat ́ eriaux test ́ es, la mortalit ́ e des bact ́ eries adh ́ erentes est inf ́ erieure ` a55%.Lesr ́ esultats obtenus ont montr ́ e qu’un mat ́ eriau dit antimicrobien n’induit pas plus de cellules endommag ́ ees comparativement aux autres mat ́ eriaux

Hydrophobicity of abiotic surfaces governs droplets deposition and evaporation patterns

International audienc

Antifouling stainless steel surface: Competition between roughness and surface energy

7. International Conference on Processing and Manufacturing of Advanced Materials Quebec City, Canada : AUG 01-05, 2011 Parts: 1-4International audienceTo increase the shelf-life qualities of dairy products, a heat treatment is usually done. However, heat treatments induce physico-chemical modifications of the products. Some of them lead to the expected product but an unwanted consequence of this process is the formation of a fouling deposit on the surfaces in contact with the processed fluid. To eliminate fouling, cleaning processes have to be done once a day. It increases the processing and maintenance costs. To control and to decrease the fouling are the main problems in food industries and an active research is carried out on efficient antifouling surface treatments. In the present study, a 316L 2B stainless steel was submitted to different surface treatments (Flame and plasma pre-treatments, Plasma Enhanced Chemical Vapour Deposition, hydrophobic coatings, mechanical polishing ...) to try to establish correlations between different surface parameters (roughness, hydrophobicity, nanostructuration, surface energy, ...) onto the fouling in heat exchangers. All the treated plates were then submitted to a fouling test using an aqueous solution of beta-lactoglobulin at 1% (p/p) with a final calcium concentration of 910 mg/L and compared to a bare steel plate. The results obtained imply different influences of each parameter depending on the surface roughness: the effect of a non organized micrometric roughness is preponderant compared to the surface energy: the fouling comes from a mechanical effect mainly due to rubbing. However, when the surface is nanostructured, fouling decreases. When the roughness reaches the nanometer scale (between 100 and 400 nm), it is the surface energy and the polar/apolar components which become preponderant compared to the roughness. Fouling is this time mainly due to the hydrophilicity of the surface and to the adsorption of the beta-lactoglobulin on acido-basic sites. Finally, when the roughness reaches less than 50 nm, polar/apolar components show no effect anymore, the preponderant parameter is the hydrophobicity of the surface

Durabilité des aciers inoxydables et développement de biofilms dans les réseaux d’eau potable : étude comparative avec d’autres matériaux couramment utilisés

De nombreux matériaux sont couramment utilisés dans les systèmes de distribution d’eau potable ; le choix du matériau dépend de sa disponibilité, de sa facilité de mise en œuvre et des coûts d’investissement et de maintenance. L’expérience montre que certains matériaux sont particulièrement sensibles à l’encrassement et au développement d’un biofilm alors que les autres y sont moins sujets mais au prix d’une corrosion généralisée de la surface entraînant un relargage d’ions non négligeable. En raison de leurs propriétés physico-chimiques de surface et de leur excellente résistance à la corrosion, les aciers inoxydables apparaissent comme de bons candidats pour transporter l’eau potable dans les meilleures conditions d’hygiène et de durabilité. Cet article présente les résultats d’une étude menée sur des boucles alimentées en eau potable naturelle pendant deux ans. Des nuances d’aciers inoxydables ferritique 444 (EN 1.4521), austénitiques 304L (EN 1.4307), 316L (EN 1.4404), 316LN (EN 1.4429) et duplex 2304 (EN 1.4362), 2205 (EN 1.4462) ont été évaluées. Le comportement de ces alliages vis-à-vis de la corrosion, du développement d’un biofilm, de l’entartrage et de la biocontamination a été comparé à celui du cuivre, de l’acier galvanisé, du polyéthylène et du ciment. Les résultats montrent (i) des taux d’encrassement et d’entartrage maximum pour l’acier galvanisé et le ciment ; (ii) une minéralisation des biofilms pour tous les matériaux, couplée à un phénomène de stratification plus marqué dans le cas des aciers inoxydables ; (iii) l’initiation d’un endommagement par corrosion localisée uniquement visible sur l’acier galvanisé ; et (iv) des vitesses moyennes de corrosion uniforme très faibles pour tous les aciers inoxydables avec pour conséquence des concentrations attendues en cations métalliques dans l’eau bien en deçà des normes internationales en vigueur

Plasma-surface engineering for biofilm prevention: evaluation of anti-adhesive and antimicrobial properties of a silver-nanocomposite thin film

International audienceBiofilms can be generally defined as a complex and dynamic ecosystem, constituted by a community of microorganisms adherent to a substrate and often embedded within a selfproduced extracellular polymeric matrix. Biofilm formation is a well-recognized phenomenon, especially in medical and food industries and often leads to undesirable effects (nosocomial infections, energy losses, accelerated corrosion, food spoilage, and spread of foodborne diseases…). In this framework, surface engineering for preventing biofilm formation is a challenging question, which has fuelled an explosion of research in surface science for the development of antimicrobial and/or anti-adhesive materials by physical or chemical modifications. The surface treatment can prevent biofilm formation by limiting the initial microbial adhesion and/or by killing microorganisms as they come in close contact with the solid surface. Among the different approaches considered, a growing interest is focused on thin silver coatings, like silver-based composite materials, due to their extended time-release properties. In the present work, plasma-mediated thin films (~170 nm), containing silver nanoparticles embedded in an organosilicon matrix, were deposited onto stainless steel. The process originality relies on a dual strategy, associating silver sputtering and simultaneous Plasma Enhanced Chemical Vapour Deposition, in an argon-hexamethyldisiloxane plasma, using an asymmetrical radiofrequency discharge at 13.56 MHz. SEM demonstrated the nanoparticle-based morphology of the deposited layer. X-ray photoelectron spectroscopy confirmed the presence of metallic silver nanoparticles embeddedin the organosilicon matrix. The film anti-adhesive potentialities were evaluated in vitro towards the model yeast Saccharomyces cerevisiae by performing shear-flow induced detachment experiments, under well-controlled hydrodynamic and physico-chemical conditions. The maximal effect was achieved for the organosilicon matrix alone. When silver nanoparticles were incorporated, yeast detachment was less pronounced, probably due to the strong affinity of embedded silver for biological groups of the cell wall surface. The presence of methyl groups in the matrix network could also promote enhanced hydrophobic cell/coating interactions. An antifungal action of released silver (Ag+ ions and/ornanoparticles) at the immediate vicinity of the coating surface occurred, since a 1.4 log reduction in viable counts was observed, compared to control conditions with bare stainless steel. TEM observations of the yeast ultrastructure demonstrated morphological and structural damages. The presence of electron-dense silver clusters was also detected not only on the cell surface but also within the cell. In parallel, the coating antimicrobial properties against bacteria were assessed (reduction in viable counts of 1.5 and 2.4 log and for Escherichia coli and Staphylococcus aureus, respectively)

Plasma-deposited nanocomposite polymer-silver coating against Escherichia coli and Staphylococcus aureus: antibacterial properties and ageing

International audienceA plasma-deposited coating, containing silver nanoparticles embedded in an organosilicon matrix, was synthesized using AISI 316L stainless steel as the underlying substrate. The coating antibacterial property was evaluated on the Gram-negative Escherichia coli K12 MG1655 and the Gram-positive Staphylococcus aureus MW2 strains, by combining indirect (plate counting) and in situ (fluorescent bacteria labelling) methods. Both approaches were shown to be highly complementary and converged on a maximal antibacterial efficacy against E. coli, as plate counts showed a decrease of 6 and 1 Log and dead bacteria represented 25% and 2% of the total adhering bacteria for E. coli and S. aureus, respectively. The coating antibacterial potential was then determined over time on E. coli biofilm. Whatever the biofilm age, growth inhibition was observed due to silver-mediated bacteriostatic effect. The coating bactericidal activity was initially strong. However, differences between coated and bare stainless steel surfaces tended to collapse above a 2-day contact time. A thorough characterization of the film properties after ageing in biological suspension or saline solution (short and longer-term exposures) revealed an oxidation of both the organosilicon matrix and the silver nanoparticles, accompanied by silver release at the extreme surface. However, a silver reservoir was still present and potentially active in the deep layers of the coating

CHASE-Containing Histidine Kinase Receptors in Apple Tree: From a Common Receptor Structure to Divergent Cytokinin Binding Properties and Specific Functions

Cytokinin signaling is a key regulatory pathway of many aspects in plant development and environmental stresses. Herein, we initiated the identification and functional characterization of the five CHASE-containing histidine kinases (CHK) in the economically important Malus domestica species. These cytokinin receptors named MdCHK2, MdCHK3a/MdCHK3b, and MdCHK4a/MdCHK4b by homology with Arabidopsis AHK clearly displayed three distinct profiles. The three groups exhibited architectural variations, especially in the N-terminal part including the cytokinin sensing domain. Using a yeast complementation assay, we showed that MdCHK2 perceives a broad spectrum of cytokinins with a substantial sensitivity whereas both MdCHK4 homologs exhibit a narrow spectrum. Both MdCHK3 homologs perceived some cytokinins but surprisingly they exhibited a basal constitutive activity. Interaction studies revealed that MdCHK2, MdCHK4a, and MdCHK4b homodimerized whereas MdCHK3a and MdCHK3b did not. Finally, qPCR analysis and bioinformatics approach pointed out contrasted expression patterns among the three MdCHK groups as well as distinct sets of co-expressed genes. Our study characterized for the first time the five cytokinin receptors in apple tree and provided a framework for their further functional studies