Les communautés bactériennes d'un holobionte méditerranéen, la gorgone rouge Paramuricea clavata : diversité, stabilité et spécificité

Coralligenous communities dominated by gorgonian species have been severely affected by diseases and mass mortality events linked to the current warming trends reported for the Mediterranean Sea. The characterization of the natural microbial compartment of this temperate gorgonian species becomes a crucial step in the evaluation of the bacterial contribution to health and functioning of the Paramuricea clavata holobiont.Under these circumstances, the global aim of this PhD work was to describe the interactions existing between the red gorgonian P. clavata and its associated bacteria in the Northwestern Mediterranean basin. The culture-independent analyses based on the bacterial 16S ribosomal DNA included (i) the characterization of spatiotemporal variation of the bacterial communities, (ii) the localization of the bacteria within host tissues, (iii) the evaluation of the stability of gorgonian-bacterial associations under stress conditions and (iv) the determination of the host-specificity of dominant bacteria in different sympatric gorgonian species (Eunicella singularis, Eunicella cavolini and Corallium rubrum).The results of this study highlighted that P. clavata and its microbiota form a holobiont in which host and bacteria live in close association. This association is spatiotemporally stable and maintained under stress conditions. Associated bacterial communities are mostly endosymbiotic and dominated by a bacterial ribotype belonging to a new genus within the Hahellaceae family that seems to be host-specific. These results suggest a particular role of this bacterial genus in the gorgonian holobionts. The present work enabled to initiate the understanding of the bacterial compartment in the 4 most abundant gorgonian species dwelling in the Mediterranean coastal habitats. This knowledge and the different tools that have been developed can be incorporated to new researches on the role of symbiotic associations in health and future trajectories of gorgonian populations in the current climate change context.Les communautés du coralligène dominées par des gorgonaires ont été sévèrement affectées par des évènements de mortalités massives liés au réchauffement de la Méditerranée. Pour évaluer la contribution des bactéries associées à l’holobionte Paramuricea clavata à son fonctionnement et sa santé, il est apparu primordial de caractériser le compartiment microbien naturel de ce gorgonaire tempéré.Dans ce contexte, l’objectif général de cette thèse était de décrire les interactions existant entre la gorgone rouge P. clavata et ses bactéries associées en Méditerranée nord-occidentale. Les analyses entreprises par des techniques culture-indépendantes basées sur l’analyse des ADN ribosomiques 16S bactériens ont inclus (i) la caractérisation de la variation spatio-temporelle des communautés bactériennes, (ii) la localisation des bactéries dans les tissus de l’hôte, (iii) l’évaluation de la stabilité des associations gorgones-bactéries en conditions de stress et (iv) la détermination de la spécificité d’hôte des bactéries dominantes entre différentes espèces de gorgonaires sympatriques (Eunicella singularis, Eunicella cavolini et Corallium rubrum). Les résultats obtenus ont établi que P. clavata et son microbiote forment un holobionte au sein duquel hôte et bactéries vivent en étroite association, stable dans le temps et l’espace ou en conditions de stress. Les communautés bactériennes associées sont principalement endosymbiotiques et dominées par un ribotype bactérien appartenant à un genre nouveau de la famille des Hahellaceae qui semble présenter une forte spécificité d’hôte. Ces résultats suggèrent un rôle particulier de ce genre bactérien chez les holobiontes gorgonaires. Ces travaux ont permis d’initier la connaissance du compartiment bactérien des quatre espèces de gorgonaires les plus abondantes dans les habitats côtiers. Les éléments acquis et les différents outils mis au point pourront être intégrés à de nouvelles recherches sur le rôle des associations symbiotiques dans la santé et le devenir des populations de gorgonaires face aux changements environnementaux en cours

Opportunities for topical antimicrobial therapy: permeation of canine skin by fusidic acid

BACKGROUND: Staphylococcal infection of the canine epidermis and hair follicle is amongst the commonest reasons for antimicrobial prescribing in small animal veterinary practice. Topical therapy with fusidic acid (FA) is an attractive alternative to systemic therapy based on low minimum inhibitory concentrations (MICs, commonly <0.03 mg/l) documented in canine pathogenic staphylococci, including strains of MRSA and MRSP (methicillin-resistant Staphylococcus aureus and S. pseudintermedius). However, permeation of canine skin by FA has not been evaluated in detail. This study aimed to define the degree and extent of FA permeation in canine skin in vitro from two sites with different hair follicle density following application of a licensed ophthalmic formulation that shares the same vehicle as an FA-betamethasone combination product approved for dermal application in dogs. Topical FA application was modelled using skin held in Franz-type diffusion cells. Concentrations of FA in surface swabs, receptor fluid, and transverse skin sections of defined anatomical depth were determined using high-performance liquid chromatography and ultraviolet (HPLC-UV) analysis. RESULTS: The majority of FA was recovered by surface swabs after 24 h, as expected (mean ± SEM: 76.0 ± 17.0%). FA was detected within 424/470 (90%) groups of serial sections of transversely cryotomed skin containing follicular infundibula, but never in 48/48 (100%) groups of sections containing only deeper follicular structures, nor in receptor fluid, suggesting that FA does not permeate beyond the infundibulum. The FA concentration (mean ± SEM) in the most superficial 240 μm of skin was 2000 ± 815 μg/g. CONCLUSIONS: Topically applied FA can greatly exceed MICs for canine pathogenic staphylococci at the most common sites of infection. Topical FA therapy should now be evaluated using available formulations in vivo as an alternative to systemic therapy for canine superficial bacterial folliculitis.Peer reviewedFinal Published versio

Genetic landscape of a large cohort of Primary Ovarian Insufficiency : New genes and pathways and implications for personalized medicine

Background Primary Ovarian Insufficiency (POI), a public health problem, affects 1-3.7% of women under 40 yield-ing infertility and a shorter lifespan. Most causes are unknown. Recently, genetic causes were identified, mostly in single families. We studied an unprecedented large cohort of POI to unravel its molecular pathophysiology.Methods 375 patients with 70 families were studied using targeted (88 genes) or whole exome sequencing with pathogenic/likely-pathogenic variant selection. Mitomycin-induced chromosome breakages were studied in patients' lymphocytes if necessary. Findings A high-yield of 29.3% supports a clinical genetic diagnosis of POI. In addition, we found strong evidence of pathogenicity for nine genes not previously related to a Mendelian phenotype or POI: ELAVL2, NLRP11, CENPE, SPATA33, CCDC150, CCDC185, including DNA repair genes: C17orf53(HROB), HELQ, SWI5 yielding high chromo-somal fragility. We confirmed the causal role of BRCA2, FANCM, BNC1, ERCC6, MSH4, BMPR1A, BMPR1B, BMPR2, ESR2, CAV1, SPIDR, RCBTB1 and ATG7 previously reported in isolated patients/families. In 8.5% of cases, POI is the only symptom of a multi-organ genetic disease. New pathways were identified: NF-kB, post-translational regulation, and mitophagy (mitochondrial autophagy), providing future therapeutic targets. Three new genes have been shown to affect the age of natural menopause supporting a genetic link.Interpretation We have developed high-performance genetic diagnostic of POI, dissecting the molecular pathogene-sis of POI and enabling personalized medicine to i) prevent/cure comorbidities for tumour/cancer susceptibility genes that could affect life-expectancy (37.4% of cases), or for genetically-revealed syndromic POI (8.5% of cases), ii) predict residual ovarian reserve (60.5% of cases). Genetic diagnosis could help to identify patients who may benefit from the promising in vitro activation-IVA technique in the near future, greatly improving its success in treating infertility.Funding Universite? Paris Saclay, Agence Nationale de Biome?decine.Copyright (c) 2022 The Author(s). Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/)Peer reviewe

Bacterial communities associated with a Mediterranean holobiont, the red gorgonian Paramuricea clavata : diversity, stability and specificity

Les communautés du coralligène dominées par des gorgonaires ont été sévèrement affectées par des évènements de mortalités massives liés au réchauffement de la Méditerranée. Pour évaluer la contribution des bactéries associées à l’holobionte Paramuricea clavata à son fonctionnement et sa santé, il est apparu primordial de caractériser le compartiment microbien naturel de ce gorgonaire tempéré.Dans ce contexte, l’objectif général de cette thèse était de décrire les interactions existant entre la gorgone rouge P. clavata et ses bactéries associées en Méditerranée nord-occidentale. Les analyses entreprises par des techniques culture-indépendantes basées sur l’analyse des ADN ribosomiques 16S bactériens ont inclus (i) la caractérisation de la variation spatio-temporelle des communautés bactériennes, (ii) la localisation des bactéries dans les tissus de l’hôte, (iii) l’évaluation de la stabilité des associations gorgones-bactéries en conditions de stress et (iv) la détermination de la spécificité d’hôte des bactéries dominantes entre différentes espèces de gorgonaires sympatriques (Eunicella singularis, Eunicella cavolini et Corallium rubrum). Les résultats obtenus ont établi que P. clavata et son microbiote forment un holobionte au sein duquel hôte et bactéries vivent en étroite association, stable dans le temps et l’espace ou en conditions de stress. Les communautés bactériennes associées sont principalement endosymbiotiques et dominées par un ribotype bactérien appartenant à un genre nouveau de la famille des Hahellaceae qui semble présenter une forte spécificité d’hôte. Ces résultats suggèrent un rôle particulier de ce genre bactérien chez les holobiontes gorgonaires.Coralligenous communities dominated by gorgonian species have been severely affected by diseases and mass mortality events linked to the current warming trends reported for the Mediterranean Sea. The characterization of the natural microbial compartment of this temperate gorgonian species becomes a crucial step in the evaluation of the bacterial contribution to health and functioning of the Paramuricea clavata holobiont.Under these circumstances, the global aim of this PhD work was to describe the interactions existing between the red gorgonian P. clavata and its associated bacteria in the Northwestern Mediterranean basin. The culture-independent analyses based on the bacterial 16S ribosomal DNA included (i) the characterization of spatiotemporal variation of the bacterial communities, (ii) the localization of the bacteria within host tissues, (iii) the evaluation of the stability of gorgonian-bacterial associations under stress conditions and (iv) the determination of the host-specificity of dominant bacteria in different sympatric gorgonian species (Eunicella singularis, Eunicella cavolini and Corallium rubrum).The results of this study highlighted that P. clavata and its microbiota form a holobiont in which host and bacteria live in close association. This association is spatiotemporally stable and maintained under stress conditions. Associated bacterial communities are mostly endosymbiotic and dominated by a bacterial ribotype belonging to a new genus within the Hahellaceae family that seems to be host-specific. These results suggest a particular role of this bacterial genus in the gorgonian holobionts

Les communautés bactériennes d'un holobionte méditerranéen, la gorgone rouge Paramuricea clavata : diversité, stabilité et spécificité

Coralligenous communities dominated by gorgonian species have been severely affected by diseases and mass mortality events linked to the current warming trends reported for the Mediterranean Sea. The characterization of the natural microbial compartment of this temperate gorgonian species becomes a crucial step in the evaluation of the bacterial contribution to health and functioning of the Paramuricea clavata holobiont.Under these circumstances, the global aim of this PhD work was to describe the interactions existing between the red gorgonian P. clavata and its associated bacteria in the Northwestern Mediterranean basin. The culture-independent analyses based on the bacterial 16S ribosomal DNA included (i) the characterization of spatiotemporal variation of the bacterial communities, (ii) the localization of the bacteria within host tissues, (iii) the evaluation of the stability of gorgonian-bacterial associations under stress conditions and (iv) the determination of the host-specificity of dominant bacteria in different sympatric gorgonian species (Eunicella singularis, Eunicella cavolini and Corallium rubrum).The results of this study highlighted that P. clavata and its microbiota form a holobiont in which host and bacteria live in close association. This association is spatiotemporally stable and maintained under stress conditions. Associated bacterial communities are mostly endosymbiotic and dominated by a bacterial ribotype belonging to a new genus within the Hahellaceae family that seems to be host-specific. These results suggest a particular role of this bacterial genus in the gorgonian holobionts. The present work enabled to initiate the understanding of the bacterial compartment in the 4 most abundant gorgonian species dwelling in the Mediterranean coastal habitats. This knowledge and the different tools that have been developed can be incorporated to new researches on the role of symbiotic associations in health and future trajectories of gorgonian populations in the current climate change context.Les communautés du coralligène dominées par des gorgonaires ont été sévèrement affectées par des évènements de mortalités massives liés au réchauffement de la Méditerranée. Pour évaluer la contribution des bactéries associées à l’holobionte Paramuricea clavata à son fonctionnement et sa santé, il est apparu primordial de caractériser le compartiment microbien naturel de ce gorgonaire tempéré.Dans ce contexte, l’objectif général de cette thèse était de décrire les interactions existant entre la gorgone rouge P. clavata et ses bactéries associées en Méditerranée nord-occidentale. Les analyses entreprises par des techniques culture-indépendantes basées sur l’analyse des ADN ribosomiques 16S bactériens ont inclus (i) la caractérisation de la variation spatio-temporelle des communautés bactériennes, (ii) la localisation des bactéries dans les tissus de l’hôte, (iii) l’évaluation de la stabilité des associations gorgones-bactéries en conditions de stress et (iv) la détermination de la spécificité d’hôte des bactéries dominantes entre différentes espèces de gorgonaires sympatriques (Eunicella singularis, Eunicella cavolini et Corallium rubrum). Les résultats obtenus ont établi que P. clavata et son microbiote forment un holobionte au sein duquel hôte et bactéries vivent en étroite association, stable dans le temps et l’espace ou en conditions de stress. Les communautés bactériennes associées sont principalement endosymbiotiques et dominées par un ribotype bactérien appartenant à un genre nouveau de la famille des Hahellaceae qui semble présenter une forte spécificité d’hôte. Ces résultats suggèrent un rôle particulier de ce genre bactérien chez les holobiontes gorgonaires. Ces travaux ont permis d’initier la connaissance du compartiment bactérien des quatre espèces de gorgonaires les plus abondantes dans les habitats côtiers. Les éléments acquis et les différents outils mis au point pourront être intégrés à de nouvelles recherches sur le rôle des associations symbiotiques dans la santé et le devenir des populations de gorgonaires face aux changements environnementaux en cours

Transient Shifts in Bacterial Communities Associated with the Temperate Gorgonian Paramuricea clavata in the Northwestern Mediterranean Sea

International audienceBackground Bacterial communities that are associated with tropical reef-forming corals are being increasingly recognized for their role in host physiology and health. However, little is known about the microbial diversity of the communities associated with temperate gorgonian corals, even though these communities are key structural components of the ecosystem. In the Northwestern Mediterranean Sea, gorgonians undergo recurrent mass mortalities, but the potential relationship between these events and the structure of the associated bacterial communities remains unexplored. Because microbial assemblages may contribute to the overall health and disease resistance of their host, a detailed baseline of the associated bacterial diversity is required to better understand the functioning of the gorgonian holobiont.Methodology/Principal Findings The bacterial diversity associated with the gorgonian Paramuricea clavata was determined using denaturing gradient gel electrophoresis, terminal-restriction fragment length polymorphism and the construction of clone libraries of the bacterial 16S ribosomal DNA. Three study sites were monitored for 4 years to assess the variability of communities associated with healthy colonies. Bacterial assemblages were highly dominated by one Hahellaceae-related ribotype and exhibited low diversity. While this pattern was mostly conserved through space and time, in summer 2007, a deep shift in microbiota structure toward increased bacterial diversity and the transient disappearance of Hahellaceae was observed.Conclusion/Significance This is the first spatiotemporal study to investigate the bacterial diversity associated with a temperate shallow gorgonian. Our data revealed an established relationship between P. clavata and a specific bacterial group within the Oceanospirillales. These results suggest a potential symbiotic role of Hahellaceae in the host-microbe association, as recently suggested for tropical corals. However, a transient imbalance in bacterial associations can be tolerated by the holobiont without apparent symptoms of disease. The subsequent restoration of the Hahellaceae-dominated community is indicative of the specificity and resilience of the bacteria associated with the gorgonian host

BACTERIAL COMMUNITIES ASSOCIATED WITH THE MEDITERRANEAN GORGONIAN PARAMURICEA CLAVATA

International audienceLarge-scale disease outbreaks in Paramuricea clavata and other gorgonian species have recently occurred in the northwestern Mediterranean. The environmental driver of these events was an increase in seawater temperature during climatic anomalies in summer. From diseased P. clavata colonies, we previously isolated a Vibrio strain that showed thermodependent virulence, demonstrating that pathogenic microbes are associated to gorgonian mortalities. The composition and dynamics of the natural microbial communities living in association with P. clavata are unknown. The aim of the present study is to establish a baseline for diversity and abundance of associated bacteria. These data will be used to investigate changes in the normal microbiota at elevated temperature and to test specific hypothesis about factors leading to pathogen development and disease during anomalies. Preliminary data obtained by culture-independent techniques indicate that bacterial communities from geographically remote populations of P. clavata are similar, while observed seasonal changes in bacterial assemblages suggest that environmental factors could influence their composition. As part of the multidisciplinary Medchange program, this work will contribute to provide a better understanding of marine ecological responses to climate change

Spatial and temporal analysis of microbial diversity associated with the Mediterranean gorgonian Paramuricea clavata

International audienceIn Nortwestern Mediterranean Sea, recurrent mass mortalities have been observed affecting benthic macroinvertebrate species during climatic anomalies in summer. One of the most affected species was the red gorgonian Paramuricea clavata, and the infectious pathogen Vibrio coralliilyticus was implicated in disease in this organism. As the composition and dynamics of the natural microbial communities living in association with temperate Mediterranean gorgonians are unknown, we attempted to establish a baseline for diversity and abundance of the bacterial community of P. clavata. In this study, we used both terminal-restriction fragment length polymorphism (T-RFLP) and clone library analysis to assess the spatio-temporal changes of bacterial assemblages associated with the gorgonian. We found a clear difference between the bacterial communities during summer and winter, while T- RFLP profiles were highly similar between P. clavata populations separated by hundreds of kilometers. Sequencing data from 16S rDNA clone libraries demonstrated the presence of distinct phylogenetic taxa during summer and winter, with an increased diversity in warm season. Monitoring of bacterial assemblage variability during a longer period of time is currently in process to assess if the seasonal pattern observed between summer and winter is repeated during successive years. In the context of recurrent gorgonian diseases and mortalities, this picture of the bacterial communities structure will be helpful to monitor the effects of thermal anomalies on the resident microbial flora

Bacteria associated with the Mediterreanean gorgonian Paramuricea clavata: a spatio-temporal characterization

International audienceIn Nortwestern Mediterranean Sea, recurrent mass mortalities have been observed affecting benthic macroinvertebrate species during climatic anomalies in summer. One of the most affected species was the red gorgonian Paramuricea clavata, and the infectious pathogen Vibrio coralliilyticus was implicated in disease in this organism. As the composition and dynamics of the natural microbial communities living in association with temperate Mediterranean gorgonians are unknown, we attempted to establish a baseline for diversity and abundance of the bacterial community of P. clavata. In this study, we used both terminal-restriction fragment length polymorphism (T-RFLP) and clone library analysis to assess the spatio-temporal changes of bacterial assemblages associated with the gorgonian. We found a clear difference between the bacterial communities during summer and winter, while T-RFLP profiles were highly similar between P. clavata populations separated by hundreds of kilometers. Sequencing data from 16S rDNA clone libraries demonstrated the presence of distinct phylogenetic taxa during summer and winter, with an increased diversity in warm season. Monitoring of bacterial assemblage variability during a longer period of time is currently in process to assess if the seasonal pattern observed between summer and winter is repeated during successive years. In the context of recurrent gorgonian diseases and mortalities, this picture of the bacterial communities structure will be helpful to monitor the effects of thermal anomalies on the resident microbial flora

Spatial and temporal analysis of bacterial diversity associated with the Mediterranean gorgonian Paramuricea clavata

International audienceWe combined terminal-restriction fragment length polymorphism (T-RFLP) and clone library analyses to investigate the diversity and the spatio-temporal changes of bacterial assemblages associated with the gorgonian Paramuricea clavata. We found a clear difference between the bacterial communities during winter and summer while T-RFLP profiles were highly similar between P. clavata populations separated by hundreds of kilometres. Sequencing data from 16S rDNA clone libraries demonstrated the presence of distinct phylogenetic taxa during summer and winter, with an increased diversity in the warm season. In the context of recurrent gorgonian diseases and mortalities, this picture of the structure of bacterial communities will be helpful to monitor the effects of thermal anomalies on the resident microbial flora