Exploiting the Richness of Environmental Waterborne Bacterial Species to Find Natural Legionella pneumophila Competitors

Legionella pneumophila is one of the most tracked waterborne pathogens and remains an important threat to human health. Despite the use of biocides, L. pneumophila is able to persist in engineered water systems with the help of multispecies biofilms and phagocytic protists. For few years now, high-throughput sequencing methods have enabled a better understanding of microbial communities in freshwater environments. Those unexplored and complex communities compete for nutrients using antagonistic molecules as war weapons. Up to now, few of these molecules were characterized in regards of L. pneumophila sensitivity. In this context, we established, from five freshwater environments, a vast collection of culturable bacteria and investigated their ability to inhibit the growth of L. pneumophila. All bacterial isolates were classified within 4 phyla, namely Proteobacteria (179/273), Bacteroidetes (48/273), Firmicutes (43/273), and Actinobacteria (3/273) according to 16S rRNA coding sequences. Aeromonas, Bacillus, Flavobacterium, and Pseudomonas were the most abundant genera (154/273). Among the 273 isolates, 178 (65.2%) were shown to be active against L. pneumophila including 137 isolates of the four previously cited main genera. Additionally, other less represented genera depicted anti-Legionella activity such as Acinetobacter, Kluyvera, Rahnella, or Sphingobacterium. Furthermore, various inhibition diameters were observed among active isolates, ranging from 0.4 to 9 cm. Such variability suggests the presence of numerous and diverse natural compounds in the microenvironment of L. pneumophila. These molecules include both diffusible secreted compounds and volatile organic compounds, the latter being mainly produced by Pseudomonas strains. Altogether, this work sheds light on unexplored freshwater bacterial communities that could be relevant for the biological control of L. pneumophila in manmade water systems

Bacterial-based additives for the production of artificial snow: What are the risks to human health?

International audienceFor around two decades, artificial snow has been used by numerous winter sports resorts to ensure good snow cover at low altitude areas or more generally, to lengthen the skiing season. Biological additives derived from certain bacteria are regularly used to make artificial snow. However, the use of these additives has raised doubts concerning the potential impact on human health and the environment. In this context, the French health authorities have requested the French Agency for Environmental and Occupational Health Safety (Afsset) to assess the health risks resulting from the use of such additives. The health risk assessment was based on a review of the scientific literature, supplemented by professional consultations and expertise. Biological or chemical hazards from additives derived from the ice nucleation active bacterium Pseudomonas syringae were characterised. Potential health hazards to humans were considered in terms of infectious, toxic and allergenic capacities with respect to human populations liable to be exposed and the means of possible exposure. Taking into account these data, a qualitative risk assessment was carried out, according to four exposure scenarios, involving the different populations exposed, and the conditions and routes of exposure. It was concluded that certain health risks can exist for specific categories of professional workers (mainly snowmakers during additive mixing and dilution tank cleaning steps, with risks estimated to be negligible to low if workers comply with safety precautions). P. syringae does not present any pathogenic capacity to humans and that the level of its endotoxins found in artificial snow do not represent a danger beyond that of exposure to P. syringae endotoxins naturally present in snow. However, the risk of possible allergy in some particularly sensitive individuals cannot be excluded. Another important conclusion of this study concerns use of poor microbiological water quality to make artificial snow

Etude de Dac gbI: allergene glycoproteique majeur du pollen de dactyle (Dactylis glomerata). Application de la spectrometrie de masse a l'analyse structurale de glycanes

SIGLEAvailable from INIST (FR), Document Supply Service, under shelf-number : T 81959 / INIST-CNRS - Institut de l'Information Scientifique et TechniqueFRFranc

Dessalement des eaux de mer, autopsie et nettoyage de membranes d'osmose inverse isolées d'usines de production et d'unités pilotes

POITIERS-BU Sciences (861942102) / SudocSudocFranceF

Prévention de la formation de biofilms, élaboration de surfaces antimicrobiennes

La résistance des biofilms aux antibiotiques classiques rend leur éradication et/ou leur inhibition difficile et pose un problème de santé publique, environnemental et industriel. La première partie de ce travail a consisté à évaluer l'activité antimicrobienne et anti-biofilm de surfaces modifiées par greffage des peptides antimicrobiens (Magainine I, Gramicidine A) par voie chimique. Les tests réalisés contre Listeria ivanovii, Staphylococcus aureus, Enterococcus faecalis, Escherichia coli et Candida albicans après 3 h d'incubation à 37C montrent une inhibition du nombre de cellules microbiennes adhérées sur les supports avec peptides. De plus, ces surfaces réduisent la formation du biofilm bactérien pendant au moins 24 h. Les résultats de microscopie confocale suggèrent une action inhibitrice se faisant par un effet plutôt bactériostatique et non par inhibition de l'adhérence. La seconde partie de ce travail a consisté au développement de films d hydrogels de k- carraghéane antibactériens soit par immersion des films préformés dans une solution de nisine (système immergé), soit par incorporation d'une solution de nisine dans la solution de carraghéane en surfusion (système inclus), soit en combinant les deux systèmes incorporation puis immersion (système hybride). Les films obtenus pour le système immergé et hybride présentent une activité antibactérienne dirigée contre L.ivanovii, S.aureus et E.faecalis. Par contre, le système inclus n'a d effet inhibiteur que sur L.ivanovii pour les films contenant une concentration de 4000 UI/mL en nisine. Les résultats de microscopie confocale des bactéries GFP montrent un effet bactéricide et antiadhésif de la nisine.The resistance of the biofilms to traditional antibiotics returns difficult their eradication and/or their inhibition and poses public health problems, environmental and industrial. The first part of this work consisted in evaluating the antimicrobial and anti-biofilm activity of modified surfaces by grafting of antimicrobial peptides (Magainin I, Gramicidin A) by chemical way. The tests carried out against Listeria ivanovii, Staphylococcus aureus, Enterococcus faecalis, Escherichia coli and Candida albicans after 3h of incubation to 37C show an inhibition amongst microbial cells adhered on the supports with peptides. Moreover, these surfaces reduce the formation of the bacterial biofilm during at least 24 h. The results of confocal microscopy suggest an inhibiting action being done by an effect rather bacteriostatic and not by inhibition of adherence. The second part of this work consisted with the development of antibacterial hydrogel films of k-carrageenan either by immersion of films preformed in a solution of nisin (immersed system) or by combining the two systems incorporation then immersion (hybrid system). The films obtained for the immersed and hybrid systems present an activity antibacterial directed against L.ivanovii, S.aureus et E.faecalis. On the other hand, the included system has inhibiting effect only on L.ivanovii for films containing a concentration of 4000 UI/mL in nisin. The results of microscopy confocale of bacteria GFP show a bactericidal or anti-adhesive effect of the nisinPOITIERS-BU Sciences (861942102) / SudocSudocFranceF

Implication de toxines fongiques dans l'esca de la vigne, mise au point d'un test de diagnostic et recherche d'un traitement contre la maladie

L esca est une maladie vasculaire très dommageable pour la vigne puisqu elle provoque la mort du cep à plus ou moins long terme. Cette maladie, très complexe, met en jeu plusieurs champignons pathogènes parmi lesquels Phaeomoniella chlamydospora et Phaeoacremonium aleophilum sont considérés comme agents pionniers. Ces champignons sécrètent des molécules toxiques dans leur milieu de culture. Appliquées sur cellules de vigne, ces molécules agissent sur des étapes clés des réponses de la plante aux agents pathoge nes : modifications du potentiel de membrane, des flux de protons, de l absorption des nutriments et induction de la mort cellulaire. Ces modifications ont été reliées à des variations d activité d enzymes régulant ces divers processus : H+-ATPase de la membrane plasmique, NADPH oxydase et phénylalanine ammonia-lyase. La nature polypeptidique de composés sécrétés respectivement par P. chlamydospora et P. aleophilum a permis d obtenir des anticorps polyclonaux dirigés contre ces molécules. Un test immunologique réalisé à partir d empreintes de sarments sur membrane de nitrocellulose, méthode simple et peu destructive, a permis de détecter la présence ou non des champignons dans la plante. Ce test a été validé en serre sur des boutures de vigne inoculées sélectivement par chacun des champignons, puis utilisé au vignoble sur des plantes présentant ou non des symptômes de la maladie. Un traitement contre l esca a été mis au point en utilisant un mélange de molécules présentant des propriétés antifongiques ou élicitrices de réactions de défense. Après détermination de l activité antifongique du mélange sur P. chlamydospora et P. aleophilum en culture in vitro, un traitement a été réalisé en serre par pulvérisation foliaire de boutures de vigne inoculées sélectivement. L efficacité du traitement a été contrôlée en utilisant le test de détection immunologique.Esca is a vascular disease very harmful to the vine since it causes long term stock death. This complex disease involves several pathogenic fungi among which Phaeomoniella chlamydospora and Phaeoacremonium aleophilum are viewed as the first invader fungi. The above mentioned fungi secreted several toxic molecules into their culture medium. Applied to grapevine cells, these molecules act on key stages of plant responses to pathogenic agents namely: modifications of membrane potential, proton fluxes, nutrient absorption and induction of cell death. These modifications have been linked with changes in enzyme activities controlling the following processes: H+-ATPase of the plasma membrane, NADPH oxidase and phenylalanine ammonia-lyase. The polypeptidic nature of certain compounds secreted by P. chlamydospora and P. aleophilum enabled us to obtain polyclonal antibodies directed against these polypeptides which were subsequently used for the optimisation and validation of a dot-blot method. This simple test could distinguish between infected and non-infected plants. This test was initially validated on grapevine cuttings grown in a greenhouse and which were inoculated selectively by each fungus. The test was then used in a vineyard on plants in the presence or absence of foliar esca symptoms. A treatment against esca was developed by either mixing molecules having antifungal action or inducing plant defence reactions. After verification of the antifungal activity of the association on P. chlamydospora and P. aleophilum on an in vitro culture, a treatment was evaluated in a greenhouse by foliar pulverization on grapevine cuttings inoculated selectively. The efficiency of the treatment was monitored by using the dot-blot methodPOITIERS-BU Sciences (861942102) / SudocSudocFranceF

Protection des matrices alimentaires par antagonisme bactérien

Y105, dans les conditions précitées, a mis en évidencParmi les bactériocines de bactéries lactiques, peptides anti-bactériens, celles de la sous-classe IIa présentent un motif consensus YGNGV et une activité anti-Listeria. Une nouvelle technique permettant de visualiser les antagonismes bactériens a été développée. Cette technique, utilisant la bioluminescence, a permis de rechercher des bactéries lactiques ayant une activité inhibitrice des bactéries Listeria et entérocoque. La méthode a permis de détecter une nouvelle bactériocine, la sakacine G. Bactériocine dite "courte", de 37 acides aminés, elle possède deux ponts disulfures, et son gène de structure est dupliqué ce qui en fait une bactériocine à part. L'étude de l'impact des facteurs physico-chimiques (pH, température, salinité) sur la production de certaines bactériocines, dont la mésentéricine Y105, a mis en évidence des effets variés selon les bactéries. L'analyse du gène de structure de la mésentéricine e une régulation de son expression.Among lactic acid bacteria bacteriocins, anti-microbial peptides, subclass IIa bacteriocins share a YGNGV consensus motif and are active against Listeria. A new method for the detection of antibacterial activities was developed. These technique allowed us to detect the antagonistic activity of lactic acid bacteria against bioluminescent Listeria and Enterococcus. A new bacteriocin, sakacin G was purified and characterized. The peptide is short, 37 amino acids, but has two disulfide bonds and its genetic locus exhibits a specific features, namely the duplication of the sakacin G gene. This is the first example of a bacteriocin gene "duo" in the same operon. Physico-chemicals factors impact (pH, temperature, salt) on the production of some bacteriocins, as well as mesentericin Y105, from their producing strains, showed varied effects according to the strains. Studies on the structural gene of mesentericin Y105 showed a regulation of its expression.POITIERS-BU Sciences (861942102) / SudocSudocFranceF

Etude du transport et de la régulation de la mésentéricine Y105

POITIERS-BU Sciences (861942102) / SudocSudocFranceF

Développement de nouvelles approches in vitro pour l'inhibition des biofilms de candida (recherche de molécules fongiques inhibitrices et mise au point d'un modèle de traitement verrou)

POITIERS-BU Médecine pharmacie (861942103) / SudocSudocFranceF

Développement de matériaux à activité anti-biofilm de Candida albicans

POITIERS-BU Médecine pharmacie (861942103) / SudocSudocFranceF