Inhibition of attachment of some fouling diatoms and settlement of Ulva lactuca zoospores by film-forming bacterium and their extracellular products isolated from biofouled substrata in Northern Chile

The biofouling of surfaces submerged in the marine environment includes primary colonization of the substrate by microorganisms including bacteria, microalgae, and microscopic reproductive propagules of macroorganisms such as algal zoospores. The present study reports the evaluation of the inhibitory potential of biofilms and extracellular products (EP) of the indigenous bacterium Alteromonas sp strain Ni1-LEM on the settlement of marine biofouling such as: (i) eight marine benthic diatoms and (ii) zoospores of the alga Ulva lactuca , as well as the germination of these zoospores and was compared with reference strains with proven antifouling properties, Halomonas marina (ATCC 25374) and Pseudoalteromonas tunicata . Highest antifouling activity was found for the indigenous strain. In attempts to better define the chemical nature of the antifouling substance in the EP of the Alteromonas sp strain Ni1-LEM, the culture filtrates were tested for activity after heat treatment, enzymatic treatments, dialysis through semipermeable membranes, and separation into polar (aqueous) and non-polar (organic) fractions. The results suggested that the antifouling substance in the culture filtrates to be protein or peptide in nature, thermostable, hydrophilic, and equal to or greater than 3500 daltons in molecular size. Antifouling substances from bacteria may lead to the development of novel antifouling agents in the future

Transcriptomics and cellular response of red abalone Haliotis rufescens grown in industrial hatchery to the metal and pathogens stress : role of probiotics in the survival of organisms

Dans les écosystèmes marins côtiers, l‘activité anthropique et les paramètres naturels induisent chez lesorganismes aquatiques des situations de «multistress». Parmi ces paramètres, deux d‘entre eux ont été étudiés:La contamination métallique et l’infection bactérienne. Dans ce contexte la compréhension des réponsesmoléculaires et physiologiques de l’ormeau rouge Haliotis rufescens a été étudiée, en particulier, face l’impactmétallique (cuivre), et au pathogène (Vibrio parahaemolyticus) en interaction avec des bactéries probiotiquespendant le stade juvénile de ce mollusque en conditions contrôlées. Dans un première temps, une évaluation surl’effet d’un consortium probiotique formé de trois souches bactériennes: Vibrio sp C21, Agarivorans albus F1et Vibrio sp F15 a été testée et les paramètres de survie et de croissance des juvéniles de l’ormeau ont été suivis.Les résultats montrent que les probiotiques augmentent les performances de ces deux paramètres de trait de viecomparés aux individus témoins sans probiotiques. Dans un second temps, une approche génomique parbanques soustractives a été réalisée afin d’identifier les transcrits régulés par l’impact métallique. Cetteméthode a permis d’identifier 108 gènes potentiellement impliqués dans la réponse cellulaire et physiologiquedes organismes face au stress. La cinétique de l’expression génique a été suivie à l’aide de 14 transcrits et leurvariation temporelle pendant la phase critique révèle des régulations parfois complexes et différentielles chezles organismes. Ensuite, les réponses immunitaires des hémocytes d’ormeau face l'exposition au cuivre ont étérecherchées. Les résultats montrent que l’exposition au métal provoque des disfonctionnement importants desactivités hémocytaires. Enfin, l’interaction entre les bactéries pathogènes et probiotiques ont étés étudiées etleurs effets sur le mollusque ont été également recherchés. La présence des bactéries pathogènes régulel’expression génique des transcrits impliqués dans des fonctions physiologiques clés des organismes (lesdéfenses immunitaires et l’état énergétique). De plus, les résultats de cette interaction nous renseignent sur lerôle capital accompli par les bactéries probiotiques sur les performances en termes d’amélioration de la survieet de la croissance des jeunes ormeaux en condition de stress.In coastal ecosystems, anthropogenic activity and natural factors induce “multi-stress” situations. Inthis study, we focused on two such stress-inducing factors: metal contamination and bacterial infection.Specifically, under controlled conditions, we examined the molecular and physiological responses of redabalone Haliotis rufescens toward the impact of a metal (copper) and a pathogen (Vibrio parahaemolyticus)with respect to this mollusk’s interaction with probiotic bacteria during the juvenile stage. First, we assessed theeffect of a consortium of three probiotic bacterial strains—Vibrio sp. C21, Agarivorans albus F1, and Vibrio sp.F15—on two abalone lifecycle-related parameters, namely, survival and growth. The results showed that,compared to the control treatment (no probiotics), the probiotics improved the performance of these twolifecycle parameters. In a second step, the molecular mechanisms underlying the juvenile abalone response tocopper stress were studied under experimental conditions, using a suppression subtractive hybridizationmethod. This method identified 108 partial sequences of genes involved in the cellular and physiologicalresponses to stress. The kinetics of gene expression were followed using 14 of these transcripts, and theirtemporal variations during the critical phase showed complex and differential regulation exerted by copper inthese organisms. We then investigated the immune response of abalone hemocytes to copper exposure. Theresults showed that exposure to the metal causes a significant dysfunction of hemocyte activities. Finally, theinteraction between probiotics and pathogenic bacteria (transcriptomic and cellular aspects) was studied and theeffects of these bacteria on mollusk performance were also investigated. The presence of pathogenic bacteriawas found to regulate gene expression of transcripts involved in key physiological functions, including thosethat regulate immune responses and energy metabolism. Examination of these interactions thus indicates therole of probiotic bacteria in performance in terms of the improved survival and growth of juvenile abaloneunder stress conditions

Réponse transcriptomique et cellulaire de l'ormeau rouge Haliotis rufescens, cultivé en écloserie industrielle face aux stress métalliques et aux pathogènes : rôle des probiotiques dans la survie des organismes

In coastal ecosystems, anthropogenic activity and natural factors induce “multi-stress” situations. Inthis study, we focused on two such stress-inducing factors: metal contamination and bacterial infection.Specifically, under controlled conditions, we examined the molecular and physiological responses of redabalone Haliotis rufescens toward the impact of a metal (copper) and a pathogen (Vibrio parahaemolyticus)with respect to this mollusk’s interaction with probiotic bacteria during the juvenile stage. First, we assessed theeffect of a consortium of three probiotic bacterial strains—Vibrio sp. C21, Agarivorans albus F1, and Vibrio sp.F15—on two abalone lifecycle-related parameters, namely, survival and growth. The results showed that,compared to the control treatment (no probiotics), the probiotics improved the performance of these twolifecycle parameters. In a second step, the molecular mechanisms underlying the juvenile abalone response tocopper stress were studied under experimental conditions, using a suppression subtractive hybridizationmethod. This method identified 108 partial sequences of genes involved in the cellular and physiologicalresponses to stress. The kinetics of gene expression were followed using 14 of these transcripts, and theirtemporal variations during the critical phase showed complex and differential regulation exerted by copper inthese organisms. We then investigated the immune response of abalone hemocytes to copper exposure. Theresults showed that exposure to the metal causes a significant dysfunction of hemocyte activities. Finally, theinteraction between probiotics and pathogenic bacteria (transcriptomic and cellular aspects) was studied and theeffects of these bacteria on mollusk performance were also investigated. The presence of pathogenic bacteriawas found to regulate gene expression of transcripts involved in key physiological functions, including thosethat regulate immune responses and energy metabolism. Examination of these interactions thus indicates therole of probiotic bacteria in performance in terms of the improved survival and growth of juvenile abaloneunder stress conditions.Dans les écosystèmes marins côtiers, l‘activité anthropique et les paramètres naturels induisent chez lesorganismes aquatiques des situations de «multistress». Parmi ces paramètres, deux d‘entre eux ont été étudiés:La contamination métallique et l’infection bactérienne. Dans ce contexte la compréhension des réponsesmoléculaires et physiologiques de l’ormeau rouge Haliotis rufescens a été étudiée, en particulier, face l’impactmétallique (cuivre), et au pathogène (Vibrio parahaemolyticus) en interaction avec des bactéries probiotiquespendant le stade juvénile de ce mollusque en conditions contrôlées. Dans un première temps, une évaluation surl’effet d’un consortium probiotique formé de trois souches bactériennes: Vibrio sp C21, Agarivorans albus F1et Vibrio sp F15 a été testée et les paramètres de survie et de croissance des juvéniles de l’ormeau ont été suivis.Les résultats montrent que les probiotiques augmentent les performances de ces deux paramètres de trait de viecomparés aux individus témoins sans probiotiques. Dans un second temps, une approche génomique parbanques soustractives a été réalisée afin d’identifier les transcrits régulés par l’impact métallique. Cetteméthode a permis d’identifier 108 gènes potentiellement impliqués dans la réponse cellulaire et physiologiquedes organismes face au stress. La cinétique de l’expression génique a été suivie à l’aide de 14 transcrits et leurvariation temporelle pendant la phase critique révèle des régulations parfois complexes et différentielles chezles organismes. Ensuite, les réponses immunitaires des hémocytes d’ormeau face l'exposition au cuivre ont étérecherchées. Les résultats montrent que l’exposition au métal provoque des disfonctionnement importants desactivités hémocytaires. Enfin, l’interaction entre les bactéries pathogènes et probiotiques ont étés étudiées etleurs effets sur le mollusque ont été également recherchés. La présence des bactéries pathogènes régulel’expression génique des transcrits impliqués dans des fonctions physiologiques clés des organismes (lesdéfenses immunitaires et l’état énergétique). De plus, les résultats de cette interaction nous renseignent sur lerôle capital accompli par les bactéries probiotiques sur les performances en termes d’amélioration de la survieet de la croissance des jeunes ormeaux en condition de stress

Inhibition of attachment of some fouling diatoms and settlement of Ulva lactuca zoospores by film-forming bacterium and their extracellular products isolated from biofouled substrata in Northern Chile

The biofouling of surfaces submerged in the marine environment includes primary colonization of the substrate by microorganisms including bacteria, microalgae, and microscopic reproductive propagules of macroorganisms such as algal zoospores. The present study reports the evaluation of the inhibitory potential of biofilms and extracellular products (EP) of the indigenous bacterium Alteromonas sp strain Ni1-LEM on the settlement of marine biofouling such as: (i) eight marine benthic diatoms and (ii) zoospores of the alga Ulva lactuca , as well as the germination of these zoospores and was compared with reference strains with proven antifouling properties, Halomonas marina (ATCC 25374) and Pseudoalteromonas tunicata . Highest antifouling activity was found for the indigenous strain. In attempts to better define the chemical nature of the antifouling substance in the EP of the Alteromonas sp strain Ni1-LEM, the culture filtrates were tested for activity after heat treatment, enzymatic treatments, dialysis through semipermeable membranes, and separation into polar (aqueous) and non-polar (organic) fractions. The results suggested that the antifouling substance in the culture filtrates to be protein or peptide in nature, thermostable, hydrophilic, and equal to or greater than 3500 daltons in molecular size. Antifouling substances from bacteria may lead to the development of novel antifouling agents in the future

Utilización de una biopelícula microalgal multiespecífica para optimizar la fijación larval y el crecimiento de abalón (Haliotis rufescens) en un criadero comercial

This study evaluates the use of a microalgal biofilm composed by a mixture of benthic diatoms such as Amphora cf. capitellata (NV), Nitzschia ovalis arnott (NC), Cylindrotheca closterium (CC) and Navicula cryptocephala (NAV) to improve larval settlement and growth of H. rufescens postlarvae in commercial hatcheries in the north of Chile. Preliminarily the optimal growth dynamic conditions of selected diatoms were evaluated in relation light intensity by the use of polycarbonate carboys containing PVC filaments for supporting the growth of diatoms. The results showed that the different diatom strains reached the highest cell densities in terms of cell coverage, at 50 ìmol· m-2· s-1 of light intensity after 120 h for NC, CC and NV strains and 144 h for NAV strain. These results guarantee the massive productions of the selected diatoms for their use as commercial scale. The determination of the multi-especific effect of selected diatoms on the settlement and growth of differents postlarvae cohorts, after 60 and 120 days from the larval metamorphosis, showed significant improvements on settlement and growth of abalone seeds. These results suggest that the utilization of benthic diatom mixtures for the culture of abalone postlarvae at commercial scale improve the disponibility of food for the abalone Chilean industry.Este estudio evalua el uso de una biopelicula microalgal compuesta por una mezcla de diatomeas bentonicas como Amphora cf. capitellata (NV), Nitzschia ovalis arnott (NC), Cylindrotheca closterium (CC) y Navicula cryptocephala (NAV), para mejorar el asentamiento y crecimiento de postlarvas de H. rufescens en criaderos en el norte de Chile. En forma preliminar se evaluaron las condiciones optimas de crecimiento de las diatomeas seleccionadas, en relacion a la dinamica de crecimiento y la intensidad luminica optima, mediante la utilizacion de botellones de policarbonato con filamentos de PVC como sustrato para el crecimiento bentonico. Los resultados mostraron que las distintas cepas alcanzaron los mejores crecimientos, en terminos de cobertura celular, a 50 �Êmol m-2 s-1 de intensidad luminica, transcurridas 120 h de cultivo para las cepas NC, CC y NV y 144 h para la cepa NAV, garantizando la produccion masiva de estas especies para su uso a nivel comercial. La determinacion de esta mezcla multiespecifica de diatomeas sobre el asentamiento y crecimiento de post larvas para distintas cohortes, posterior a 60 y 120 dias desde la metamorfosis larval, mostro mejoras significativas sobre el asentamiento y el crecimiento de las semillas de abalon. Estos resultados sugieren que el uso de mezclas de diatomeas bentonicas para el cultivo de postlarvas de abalon a nivel masivo, mejora la oferta alimentaria para la industria abalonera en Chile

Transcriptomic and cellular response to bacterial challenge (pathogenic Vibrio parahaemolyticus) in farmed juvenile Haliotis rufescens fed with or without probiotic diet.

International audienceThe abalone production in Chile has increased considerably in recent years with no sign of tapering off. Open and semi-closed circuits in the marine water zones in the north and south of Chile are the preferred areas of culture. Coastal ecosystems are subjected to a wide variety of contaminants that generate stress that affects populations via their impacts to individuals at both physiological and genetic levels. This work investigated the genomic and cellular response of post-weaning juvenile Haliotis rufescens abalone under hatchery conditions, fed with probiotic diets, and subsequently challenged with Vibrio parahaemolyticus. The expression patterns of 16 selected genes associated with different metabolic pathways were analyzed using Real-Time PCR. Gene expression was then compared to immunological response parameters in the abalone and quantification of V. parahaemolyticus during the experimental period. Both transcriptomic and immunological analyses indicated significant alteration of physiological processes in H. rufescens correlated to exposure to the pathogenic bacteria, as well as to probiotic nutrition

Identification of genes expressed in juvenile Haliotis rufescens in response to different copper concentrations in the north of Chile under controlled conditions

International audienceThis study reports molecular markers potentially associated with resistance or sensitivity to the impact of copper in juvenile red abalone, Haliotis rufescens, in the north of Chile under experimental conditions. Genomic analysis was made applying subtractive hybridization libraries (SSH) to identify genes up-and down regulated during cooper exposure in abalone over periods of 12 and 168 h exposed to 2.5 and 10 μg/L of Cu+2. Results obtained from the SSH library revealed 368 different sequences regulated by copper, that correspond to eight major physiological functions. The validation of these sequences obtained by SSH as well as their expression kinetics were made by PCR in real time on 14 potential genes regulated by metal stress. This study provides information for the characterization of potential genomic markers that may be used in future environmental monitoring and to investigate new mechanisms of stress to copper in this commercially important marine species

Wave-mixed and orbitally shaken single-use photobioreactors for diatom algae propagation

Although vertical column, flat-plate, tubular, bubble column and airlift photobioreactors are widely used for the cultivation of diatoms, it has been shown that bubble-induced hydrodynamic stress created in these types of bioreactor can damage cells. Therefore, three single-use surface-aerated bag bioreactors, known for their outstanding results in cultivating shear-sensitive mammalian cells, were for the first time investigated for their suitability in growing the model microalgae Phaeodactylum tricornutum. All of the systems, which were additionally aerated with CO2 and equipped with illumination systems providing different light qualities, guaranteed a 22- to 43-fold increase in cell density within seven days, without any addition of cell protection agents or changes in cell morphology. Maximum cell density and dry biomass were achieved in the orbitally shaken 2D-bag by using cool-white fluorescent tubes