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[hal-04662155] T7 DNA polymerase treatment improves quantitative sequencing of both double-stranded and single-stranded DNA viruses
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ano.nymous@ccsd.cnrs.fr.invalid (Maud Billaud) 25 Jul 2024
https://hal.inrae.fr/hal-04662155
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[hal-04662284] Escherichia coli CRISPR arrays from early life fecal samples preferentially target prophages
Abstract CRISPR-Cas systems are defense mechanisms against phages and other nucleic acids that invade bacteria and archaea. In Escherichia coli, it is generally accepted that CRISPR-Cas systems are inactive in laboratory conditions due to a transcriptional repressor. In natural isolates, it has been shown that CRISPR arrays remain stable over the years and that most spacer targets (protospacers) remain unknown. Here, we re-examine CRISPR arrays in natural E. coli isolates and investigate viral and bacterial genomes for spacer targets using a bioinformatics approach coupled to a unique biological dataset. We first sequenced the CRISPR1 array of 1769 E. coli isolates from the fecal samples of 639 children obtained during their first year of life. We built a network with edges between isolates that reflect the number of shared spacers. The isolates grouped into 34 modules. A search for matching spacers in bacterial genomes showed that E. coli spacers almost exclusively target prophages. While we found instances of self-targeting spacers, those involving a prophage and a spacer within the same bacterial genome were rare. The extensive search for matching spacers also expanded the library of known E. coli protospacers to 60%. Altogether, these results suggest that E. coli’s CRISPR-Cas is an anti-prophage system and highlight the importance of reconsidering the criteria use to deem CRISPR-Cas systems active.
ano.nymous@ccsd.cnrs.fr.invalid (Moïra Dion) 25 Jul 2024
https://hal.inrae.fr/hal-04662284
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[hal-03981886] Impact d'un transfert de microbiotes maternels et de leurs métabolites sur l'expression de marqueurs entéroendocrines chez le raton
Le microbiote intestinal est un acteur majeur de la physiologie de l’hôte notamment via ses interactions avec l’épithélium intestinal. Initialement transmis de la mère à l’enfant, le métabolisme des bactéries primocolonisatrices peut avoir un impact immédiat et futur sur la santé. Ainsi, le transfert néonatal de microbiotes différents issus de rates résistantes (obese-resistant, OR) ou prédisposées (obese-prone, OP) à l’obésité a des effets à court et long terme sur le comportement alimentaire de la descendance (Pocheron et al. Frontiers in Microbiology, 2021), en faveur d’un risque de surpoids. La régulation de la prise alimentaire par l’axe intestin-cerveau fait intervenir les cellules entéroendocrines capables de détecter les métabolites bactériens. Nous avons recherché des relations potentielles entre la composition et l’activité des microbiotes transférés et la fonction endocrine intestinale. Les microbiotes (vaginaux, fécaux et issus du lait) de mères OP et OR, dont la composition taxonomique différait, ont été inoculés à des ratons nés de mères conventionnelles Fischer F344 de la naissance au 15ème jour de vie, constituant ainsi trois groupes : F-OP, F-OR, et F-Sham. A 21 jours, l’analyse métagénomique des contenus caecoliques révélait une composition taxonomique et un potentiel fonctionnel significativement différents entre les groupes traités et le groupe F-Sham, qui ne se traduisait que par des différences mineures dans l’abondance des métabolites (RMN) dans les surnageants caecocoliques. De même, l’analyse transcriptomique in vivo des iléons et côlons (qPCR) et in vitro sur la lignée STC-1 incubée en présence des surnageants a révélé des différences d’expression de certains marqueurs endocrines uniquement entre les groupes traités et F-Sham. Le transfert néonatal de microbiote impacterait donc les cellules entéroendocrines indépendamment des différences taxonomiques (OP/OR). L’identification des marqueurs bactériens impliqués est en cours via la recherche de corrélations statistiques.
ano.nymous@ccsd.cnrs.fr.invalid (Gwenola Le Dréan) 10 Feb 2023
https://hal.inrae.fr/hal-03981886
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[hal-04659886] Identification of <i>V. parvula</i> and <i>S. gordonii</i> adhesins mediating co-aggregation and its impact on physiology and mixed biofilm structure
The dental plaque is a polymicrobial community where biofilm formation and co-aggregation, the ability to bind to other bacteria, play a major role in the construction of an organized consortium. One of its prominent members is the anaerobic diderm Veillonella parvula, considered as a bridging species, which growth depends on lactate produced by oral Streptococci . Understanding how V. parvula co-aggregates and the impact of aggregation has long been hampered due to the lack of appropriate genetic tools. Here we studied co-aggregation of the naturally competent strain V. parvula SKV38 with various oral bacteria and its effect on cell physiology. We show that V. parvula requires different trimeric autotransporters of the type V secretion system to adhere to oral Streptococci and Actinomyces . In addition, we describe a novel adhesin of Streptococcus gordonii, VisA (SGO_2004), as the protein responsible for co-aggregation with V. parvula . Finally, we show that co-aggregation does not impact cell-cell communication, which is mainly driven by environmental sensing, but plays an important role in the architecture and species distribution within the biofilm.
ano.nymous@ccsd.cnrs.fr.invalid (Louis Dorison) 23 Jul 2024
https://hal.inrae.fr/hal-04659886
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[hal-04395352] Serine protease inhibitors and human wellbeing interplay: new insights for old friends
Serine Protease Inhibitors (Serpins) control tightly regulated physiological processes and their dysfunction is associated to various diseases. Thus, increasing interest is given to these proteins as new therapeutic targets. Several studies provided functional and structural data about human serpins. By comparison, only little knowledge regarding bacterial serpins exists. Through the emergence of metagenomic studies, many bacterial serpins were identified from numerous ecological niches including the human gut microbiota. The origin, distribution and function of these proteins remain to be established. In this report, we shed light on the key role of human and bacterial serpins in health and disease. Moreover, we analyze their function, phylogeny and ecological distribution. This review highlights the potential use of bacterial serpins to set out new therapeutic approaches.
ano.nymous@ccsd.cnrs.fr.invalid (Héla Mkaouar) 15 Jan 2024
https://hal.science/hal-04395352
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[hal-02941029] Sulfiredoxin Protects Mice from Lipopolysaccharide-Induced Endotoxic Shock
Peroxiredoxins constitute a major family of cysteine-based peroxide-scavenging enzymes. They carry an intriguing redox switch by undergoing substrate-mediated inactivation via overoxidation of their catalytic cysteine to the sulfinic acid form that is reverted by reduction catalyzed by the sulfinic acid reductase sulfiredoxin (Srx). The biological significance of such inactivation is not understood, nor is the function of Srx1. To address this question, we generated a mouse line with a null deletion of the Srx1-encoding Srxn1 gene. We show here that Srxn1(-/-) mice are perfectly viable and do not suffer from any apparent defects under laboratory conditions, but have an abnormal response to lipopolysaccharide that manifests by increased mortality during endotoxic shock. Microarray-based mRNA profiles show that although the response of Srxn1(-/-) mice to lipopolysaccharide is typical, spanning all spectrum and all pathways of innate immunity, it is delayed by several hours and remains intense when the response of Srxn1(+/+) mice has already dissipated. These data indicate that Srx1 activity protects mice from the lethality of endotoxic shock, adding this enzyme to other host factors, as NRF2 and peroxiredoxin 2, which by regulating cellular reactive oxygen species levels act as important modifiers in the pathogenesis of sepsis.
ano.nymous@ccsd.cnrs.fr.invalid (Anne-Gaëlle Planson) 16 Sep 2020
https://hal.inrae.fr/hal-02941029
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[pasteur-04145613] Extracellular succinate induces spatially organized biofilm formation in Clostridioides difficile
Clostridioides difficile infection associated to gut microbiome dysbiosis is the leading cause for nosocomial diarrhea. The ability of C. difficile to form biofilms has been progressively linked to its pathogenesis as well as its persistence in the gut. Although C. difficile has been reported to form biofilms in an increasing number of conditions, little is known about how these biofilms are formed in the gut and what factors may trigger their formation. Here we report that succinate, a metabolite abundantly produced by the dysbiotic gut microbiota, induces in vitro biofilm formation of C. difficile strains. We characterized the morphology and spatial composition of succinate-induced biofilms, and compared to non-induced or deoxycholate (DCA) induced biofilms. Biofilms induced by succinate are significantly thicker, structurally more complex, and poorer in proteins and exopolysaccharides (EPS). We then applied transcriptomics and genetics to characterize the early stages of succinate-induced biofilm formation and we showed that succinate-induced biofilm results from major metabolic shifts and cell-wall composition changes. Similar to DCA-induced biofilms, biofilms induced by succinate depend on the presence of a rapidly metabolized sugar. Finally, although succinate can be consumed by the bacteria, we found that the extracellular succinate is in fact responsible for the induction of biofilm formation through complex regulation involving global metabolic regulators and the osmotic stress response. Thus, our work suggests that as a gut signal, succinate may drive biofilm formation and help persistence of C. difficile in the gut, increasing the risk of relapse.
ano.nymous@ccsd.cnrs.fr.invalid (Emile Auria) 29 Jun 2023
https://pasteur.hal.science/pasteur-04145613
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[hal-04659883] Effect of surface treatment and shear flow on biofilm formation over materials employed in space water storage and distribution systems
The prolonged duration of future manned space missions conceals potential threats associated with microbial contamination. Such closed environments are susceptible to formation of complex biofilm communities, where microorganisms can thrive and further evolve. The objective of this study was to evaluate the impact of surface type, surface treatment and shear stress on biofilm formation in water facilities. To that aim, the ability of Pseudomonas fluorescens SBW25 to adhere on three space applications related materials, including passivated (SS) and both passivated and electropolished (SSEP) stainless steel, as well as Ti-6Al-4V (Ti) alloy was studied under stagnant and shear stress conditions after 24 h of exposure. Results indicated that surface type strongly affects bacterial adhesion under the same conditions. Surface coverage during static experiments was in the following order: SS &gt; Ti &gt; SSEP, while SS exhibited a fourfold surface coverage compared to SSEP highlighting the significance of surface treatment. Moreover, SS and Ti stimulate the formation of several microcolonies and their growth. On the other hand, the application of shear stress diminished bacterial attachment to the studied materials, the degree of which relied on the material type. In this case, bacterial settlement on SS and Ti was dependent on the surface texture, implying that surface roughness may also play an important role in cell adhesion under shear conditions. Furthermore, the metallic surfaces did not hinder bacterial attachment when silver ions were previously deposited on their surface. The deposition that occurs on metallic surfaces when in contact with water disinfected with silver ions, for example, during space missions, highlights its impact on the loss of disinfection capacity of silver ions.
ano.nymous@ccsd.cnrs.fr.invalid (Dimitrios I Avgoulas) 23 Jul 2024
https://hal.inrae.fr/hal-04659883
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[hal-04659516] In vitro modelling of oral microbial invasion in the human colon
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ano.nymous@ccsd.cnrs.fr.invalid (Lucie Etienne-Mesmin) 23 Jul 2024
https://hal.inrae.fr/hal-04659516
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[hal-04660777] Neolectins, towards new tools for selective sugar binding
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ano.nymous@ccsd.cnrs.fr.invalid (Nastassja Burrini) 24 Jul 2024
https://univ-orleans.hal.science/hal-04660777
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[hal-04013541] Anti-inflammatory Streptococcus thermophilus CNRZ160 limits sarcopenia induced by low-grade inflammation in older adult rats
Background and aims Aging is characterized, at the systemic level, by the development of low-grade inflammation, which has been identified as determining sarcopenia by blunting postprandial muscle anabolism. The causes of this “inflammageing” is still not clearly defined. An increased intestinal permeability, a microbiota dysbiosis and subsequent generation of intestinal then generalized inflammation have been hypothesized. The objective of this study was to test in vivo during aging if (1) a chronic low-grade intestinal inflammation can lead to anabolic resistance and muscle loss and (2) if a bacterial strain presenting anti-inflammatory properties could prevent these adverse effects. Methods Young adult (6 m) and elderly rats (18 m) received Dextran Sodium Sulfate (DSS) for 28 days to generate low-grade intestinal inflammation, and received (PB1 or PB2 groups) or not (DSS group) one of the two S. Thermophilus strains (5 × 10 9 CFU/day) previously shown to present an anti-inflammatory potential in vitro . They were compared to pair fed control (PF). Muscle and colon weights and protein synthesis (using 13 C Valine) were measured at slaughter. Muscle proteolysis, gut permeability and inflammatory markers were assessed only in old animals by RT-PCR or proteins quantifications (ELISA). Results In both adult and old rats, DSS reduced absolute protein synthesis (ASR) in gastrocnemius muscle [−12.4% (PB1) and −9.5% (PB2) vs. PF, P &lt; 0.05] and increased ASR in colon (+86% and +30.5%, respectively vs. PF, P &lt; 0.05). PB1 (CNRZ160 strain) but not PB2 resulted in a higher muscle ASR as compared to DSS in adults (+18%, P &lt; 0.05), a trend also observed for PB1 in old animals (+12%, P = 0.10). This was associated with a blunted increase in colon ASR. In old rats, PB1 also significantly decreased expression of markers of autophagy and ubiquitin-proteasome pathways vs. DSS groups and improved gut permeability (assessed by Occludin, Zonula Occludens 1 and Claudin 1 expression, P &lt; 0.05) and alleviated systemic inflammation (A2M: −48% vs. DSS, P &lt; 0.05). Conclusion The loss of muscle anabolism associated with low-grade intestinal inflammation can be prevented by supplementation with anti-inflammatory CNRZ160 strain. We propose that the moderated gut inflammation by CNRZ160 may result in curtailed amino acids (AA) utilization by the gut, and subsequent restored AA systemic availability to support muscle protein accretion. Therefore, CNRZ160 could be considered as an efficient probiotic to modulate muscle mass loss and limit sarcopenia during aging.
ano.nymous@ccsd.cnrs.fr.invalid (Isabelle Savary-Auzeloux) 03 Mar 2023
https://hal.inrae.fr/hal-04013541
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[hal-04395377] Mucosal-associated invariant T cells promote inflammation and intestinal dysbiosis leading to metabolic dysfunction during obesity
Obesity is associated with low-grade chronic inflammation promoting insulin-resistance and diabetes. Gut microbiota dysbiosis is a consequence as well as a driver of obesity and diabetes. Mucosal-associated invariant T cells (MAIT) are innate-like T cells expressing a semi-invariant T cell receptor restricted to the non-classical MHC class I molecule MR1 presenting bacterial ligands. Here we show that during obesity MAIT cells promote inflammation in both adipose tissue and ileum, leading to insulin resistance and impaired glucose and lipid metabolism. MAIT cells act in adipose tissue by inducing M1 macrophage polarization in an MR1-dependent manner and in the gut by inducing microbiota dysbiosis and loss of gut integrity. Both MAIT cell-induced tissue alterations contribute to metabolic dysfunction. Treatment with MAIT cell inhibitory ligand demonstrates its potential as a strategy against inflammation, dysbiosis and metabolic disorders.
ano.nymous@ccsd.cnrs.fr.invalid (Amine Toubal) 15 Jan 2024
https://hal.science/hal-04395377
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[hal-04623171] A mechanistic modelling approach of the host–microbiota interactions to investigate beneficial symbiotic resilience in the human gut
The health and well-being of a host are deeply influenced by the interactions with its gut microbiota. Contrasted environmental conditions, such as diseases or dietary habits, play a pivotal role in modulating these interactions, impacting microbiota composition and functionality. Such conditions can also lead to transitions from beneficial to detrimental symbiosis, viewed as alternative stable states of the host–microbiota dialogue. This article introduces a novel mathematical model exploring host–microbiota interactions, integrating dynamics of the colonic epithelial crypt, microbial metabolic functions, inflammation sensitivity and colon flows in a transverse section. The model considers metabolic shifts in epithelial cells based on butyrate and hydrogen sulfide concentrations, innate immune pattern recognition receptor activation, microbial oxygen tolerance and the impact of antimicrobial peptides on the microbiota. Using the model, we demonstrated that a high-protein, low-fibre diet exacerbates detrimental interactions and compromises beneficial symbiotic resilience, underscoring a destabilizing effect towards an unhealthy state. Moreover, the proposed model provides essential insights into oxygen levels, fibre and protein breakdown, and basic mechanisms of innate immunity in the colon and offers a crucial understanding of factors influencing the colon environment.
ano.nymous@ccsd.cnrs.fr.invalid (Marie Haghebaert) 25 Jun 2024
https://hal.inrae.fr/hal-04623171
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[hal-03918193] Four functional profiles for fibre and mucin metabolism in the human gut microbiome
Background With the emergence of metagenomic data, multiple links between the gut microbiome and the host health have been shown. Deciphering these complex interactions require evolved analysis methods focusing on the microbial ecosystem functions. Despite the fact that host or diet-derived fibres are the most abundant nutrients available in the gut, the presence of distinct functional traits regarding fibre and mucin hydrolysis, fermentation and hydrogenotrophic processes has never been investigated. Results After manually selecting 91 KEGG orthologies and 33 glycoside hydrolases further aggregated in 101 functional descriptors representative of fibre and mucin degradation pathways in the gut microbiome, we used non-negative matrix factorization to mine metagenomic datasets. Four distinct metabolic profiles were further identified on a training set of 1153 samples and thoroughly validated on a large database of 2571 unseen samples from 5 external metagenomic cohorts. Profiles 1 and 2 are the main contributors to the fibre-degradation-related metagenome: they present contrasted involvement in fibre degradation and sugar metabolism and are differentially linked to dysbiosis, metabolic disease and inflammation. Profile 1 takes over Profile 2 inhealthy samples, and unbalance of these profiles characterize dysbiotic samples. Furthermore, high fibre diet favours a healthy balance between Profiles 1 and Profile 2. Profile 3 takes over Profile 2 during Crohn’s disease, inducing functional reorientations towards unusual metabolism such as fucose and H2S degradation or propionate, acetone and butanediol production. Profile 4 gathers under-represented functions, like methanogenesis. Two taxonomic makes up of the profiles were investigated, using either the covariation of 203 prevalent genomes or metagenomic species, both providing consistent results in line with their functional characteristics. This taxonomic characterization showed that Profiles 1 and 2 were respectively mainly composed of bacteria from the phyla Bacteroidetes and Firmicutes while Profile 3 is representative of Proteobacteria and Profile 4 of methanogens. Conclusions Integrating anaerobic microbiology knowledge with statistical learning can narrow down the metagenomic analysis to investigate functional profiles. Applying this approach to fibre degradation in the gut ended with 4 distinct functional profiles that can be easily monitored as markers of diet, dysbiosis, inflammation and disease.
ano.nymous@ccsd.cnrs.fr.invalid (Simon Labarthe) 20 Oct 2023
https://hal.inrae.fr/hal-03918193v2
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[hal-02319042] Biopréservation et hautes pressions : des outils pour la maîtrise des dangers microbiologiques dans les aliments
L’utilisation d’additifs conservateurs est souvent nécessaire pour assurer la sécurité microbiologique des produits carnés réfrigérés faiblement acides. Le projet ANR BLac HP (2014-2019) a étudié une nouvelle stratégie de stabilisation des produits carnés réfrigérés pour assurer le contrôle des flores indésirables à la fois végétatives et sporulées. Grâce à une approche pluridisciplinaire, les travaux ont montré que la combinaison de la biopréservation par des bactéries lactiques et d’un traitement hautes pressions permettait d’assurer la qualité microbiologique de dés de jambon cuits à teneur réduite en nitrite pendant toute leur durée de vie. Le traitement permet de plus une qualité sensorielle optimale sans impact environnemental supplémentaire par rapport au procédé conventionnel. Mots-clés: spores, jambon, nitrites.
ano.nymous@ccsd.cnrs.fr.invalid (Hélène Simonin) 17 Oct 2019
https://institut-agro-dijon.hal.science/hal-02319042
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[hal-03271317] Maternal Microbiota Transfer Programs Offspring Eating Behavior
Understanding the link between mother’s obesity and regulation of the child’s appetite is a prerequisite for the design of successful preventive strategies. Beyond the possible contributions of genetic heritage, family culture, and hormonal and metabolic environment during pregnancy, we investigate in the present paper the causal role of the transmission of the maternal microbiotas in obesity as microbiotas differ between lean and obese mothers, maternal microbiotas are the main determinants of a baby’s gut colonization, and the intestinal microbiota resulting from the early colonization could impact the feeding behavior of the offspring with short- and long-term consequences on body weight. We thus investigated the potential role of vertical transfers of maternal microbiotas in programming the eating behavior of the offspring. Selectively bred obese-prone (OP)/obese-resistant (OR) Sprague-Dawley dams were used since differences in the cecal microbiota have been evidenced from males of that strain. Microbiota collected from vagina (at the end of gestation), feces, and milk (at postnatal days 1, 5, 10, and 15) of OP/OR dams were orally inoculated to conventional Fischer F344 recipient pups from birth to 15 days of age to create three groups of pups: F-OP, F-OR, and F-Sham group (that received the vehicle). We first checked microbiotal differences between inoculas. We then assessed the impact of transfer (from birth to adulthood) onto the intestinal microbiota of recipients rats, their growth, and their eating behavior by measuring their caloric intake, their anticipatory food reward responses, their preference for sweet and fat tastes in solutions, and the sensations that extend after food ingestion. Finally, we searched for correlation between microbiota composition and food intake parameters. We found that maternal transfer of microbiota differing in composition led to alterations in pups’ gut microbiota composition that did not last until adulthood but were associated with specific eating behavior characteristics that were predisposing F-OP rats to higher risk of over consuming at subsequent periods of their life. These findings support the view that neonatal gut microbiotal transfer can program eating behavior, even without a significant long-lasting impact on adulthood microbiota composition.
ano.nymous@ccsd.cnrs.fr.invalid (Anne-Lise Pocheron) 25 Jun 2021
https://hal.inrae.fr/hal-03271317
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[hal-03973492] DOHaD22-ABS-1830 NEONATAL TRANSFER OF MATERNAL MICROBIOTA HAS A LASTING EFFECT ON THE FEEDING BEHAVIOR OF THE OFFSPRING
Introduction: The microbiome is known to impact nearly every aspect of host physiology in health and disease, as it has a substantial effect on metabolic function. Vertical transmission from mother to child can affect the physiology from one generation to the next when changes in the composition of the microbiota have occurred due to maternal diet or obesity. Objectives: Our objective was to study whether obesity or thinness during gestation and lactation would impact different maternal microbiota and whether the transfer of microbiota to the newborn would modify feeding behavior, independently of the metabolic alterations of the mother. We thus characterized the potential role of vertical transfers of maternal microbiota in the programming of offspring feeding behavior and studied the potential mechanisms underlying the programming. Methods: Selectively obesity-prone (OP)/obesity-resistant (OR) Sprague Dawley dams were used because differences in cultures of cecal microbiota and milk microbiota were demonstrated. Microbiota collected from the vagina, feces and milk were orally inoculated into conventional Fischer F344 recipient puppies from birth to 15 days to create 3 groups of puppies: F-OP, FOR and F-Sham. Results: Early transfer of maternal microbiota was associated with specific feeding behavior traits that predisposed F-OP rats to a higher risk of overconsumption in later periods of life. The metagenomic analysis allowed us to identify a few species and the corresponding metagenomic functions positively or negatively associated with the alteration of food intake parameters and the cerebral functional pathway. DGE seq analysis of brain structure, neuroanatomy features, and NMR analysis of plasma and intestinal contents of transferred animals were also studied to search for a potential mechanistic relationship between microbiome activity and brain development. Conclusion: These results support the idea that neonatal transfer of gut microbiota can program feeding behavior, probably by acting on early phases of neurodevelopment.
ano.nymous@ccsd.cnrs.fr.invalid (Patricia Parnet) 04 Feb 2023
https://hal.inrae.fr/hal-03973492
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[hal-03973514] MATERNAL MICROBIOTAS TRANSFER IMPACTS MICROBIOTA-GUT-BRAIN AXIS IN OFFSPRING
Content: Introduction. The intestinal microbiota is a major player in the physiology of the host, particularly through its interactions with the intestinal epithelium. Initially transmitted from mother to child, the metabolism of primocolonizing bacteria can have an immediate and future impact on health. We previously demonstrated that neonatal transfer of different microbiota from obese-resistant (OR) or obese-prone (OP) dams has short-and long-term effects on the feeding behavior of offspring, supporting a risk of overweight. Regulation of food intake by the gut-brain axis involves enteroendocrine cells capable of detecting bacterial metabolites. Objectives. We looked for potential relationships between the composition and activity of transferred microbiota and gut endocrine function as a hypothesis for an early impact of microbiota-gut-brain on neurodevelopment and further eating behavior. Method. Microbiota (vaginal, fecal, and milk-derived) from OP and OR dams, which differed in taxonomic composition, were inoculated into pups born to conventional Fischer F344 dams from birth to day 15 of life, constituting three groups: F-OP, FOR , and F-Sham. Results. At 21 days of age, principal component analysis of caecocolic contents showed a discriminant separation of the 3 groups by metagenomic species and associated functions, which was reflected by only minor differences in metabolite abundance (NMR) in caecocolic supernatants. In vivo transcriptomic analysis of ileum and colon (RT-qPCR) and in vitro on the enteroendocrine STC-1 line incubated with the supernatants revealed differences in expression of some endocrine markers between the treated and F-Sham groups. The identification of the bacterial markers involved (short-chain fatty acids, neurotransmitters, bacterial peptides) is in progress via the search for statistical correlations. Conclusion. Maternal microbiotas tranfserred at birth altered the microbiota-gut-brain axis at leat in part via the enteroendocrine function.
ano.nymous@ccsd.cnrs.fr.invalid (Gwenola Le Dréan) 04 Feb 2023
https://hal.inrae.fr/hal-03973514
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[hal-02735602] Maternal obesogenic microbiota modified central neural circuits leading to eating behavior modulations in transplanted rat pups
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ano.nymous@ccsd.cnrs.fr.invalid (Anne-Lise Pocheron) 02 Jun 2020
https://hal.inrae.fr/hal-02735602
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[hal-04557632] The virtue of training: extending phage host spectra against vancomycin-resistant Enterococcus faecium strains using the Appelmans method
Phage therapy has (re)emerged as a serious possibility for combating multidrug-resistant bacterial infections, including those caused by vancomycin-resistant Enterococcus faecium strains. These opportunistic pathogens belong to a specific clonal complex 17, against which relatively few phages have been screened. We isolated a collection of 21 virulent phages growing on these vancomycin-resistant isolates. Each of these phages harbored a typical narrow plaquing host range, lysing at most 5 strains and covering together 10 strains of our panel of 14 clinical isolates. To enlarge the host spectrum of our phages, the Appelmans protocol was used. We mixed four out of our most complementary phages in a cocktail that we iteratively grew on eight naive strains from our panel, of which six were initially refractory to at least three of the combined phages. Fifteen successive passages permitted to significantly improve the lytic activity of the cocktail, from which phages with extended host ranges within the E. faecium species could be isolated. A single evolved phage able to kill up to 10 of the 14 initial E. faecium strains was obtained, and it barely infected nearby species. All evolved phages had acquired point mutations or a recombination event in the tail fiber genetic region, suggesting these genes might have driven phage evolution by contributing to their extended host spectra.
ano.nymous@ccsd.cnrs.fr.invalid (Julien Lossouarn) 24 May 2024
https://hal.inrae.fr/hal-04557632
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[hal-01204418] Automated classification of tailed bacteriophages according to their neck organization
The genetic diversity observed among bacteriophages remains a major obstacle for the identification of homologs and the comparison of their functional modules. In the structural module, although several classes of homologous proteins contributing to the head and tail structure can be detected, proteins of the head-to-tail connection (or neck) are generally more divergent. Yet, molecular analyses of a few tailed phages belonging to different morphological classes suggested that only a limited number of structural solutions are used in order to produce a functional virion. To challenge this hypothesis and analyze proteins diversity at the virion neck, we developed a specific computational strategy to cope with sequence divergence in phage proteins. We searched for homologs of a set of proteins encoded in the structural module using a phage learning database. Results: We show that using a combination of iterative profile-profile comparison and gene context analyses, we can identify a set of head, neck and tail proteins in most tailed bacteriophages of our database. Classification of phages based on neck protein sequences delineates 4 Types corresponding to known morphological subfamilies. Further analysis of the most abundant Type 1 yields 10 Clusters characterized by consistent sets of head, neck and tail proteins. We developed Virfam, a webserver that automatically identifies proteins of the phage head-neck-tail module and assign phages to the most closely related cluster of phages. This server was tested against 624 new phages from the NCBI database. 93% of the tailed and unclassified phages could be assigned to our head-neck-tail based categories, thus highlighting the large representativeness of the identified virion architectures. Types and Clusters delineate consistent subgroups of Caudovirales, which correlate with several virion properties. Conclusions: Our method and webserver have the capacity to automatically classify most tailed phages, detect their structural module, assign a function to a set of their head, neck and tail genes, provide their morphologic subtype and localize these phages within a "head-neck-tail" based classification. It should enable analysis of large sets of phage genomes. In particular, it should contribute to the classification of the abundant unknown viruses found on assembled contigs of metagenomic samples.
ano.nymous@ccsd.cnrs.fr.invalid (Anne Lopes) 28 May 2020
https://hal.science/hal-01204418
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[hal-01134590] Temperate phages acquire DNA from defective prophages by relaxed homologous recombination: the role of rad52-like recombinases.
Bacteriophages (or phages) dominate the biosphere both numerically and in terms of genetic diversity. In particular, genomic comparisons suggest a remarkable level of horizontal gene transfer among temperate phages, favoring a high evolution rate. Molecular mechanisms of this pervasive mosaicism are mostly unknown. One hypothesis is that phage encoded recombinases are key players in these horizontal transfers, thanks to their high efficiency and low fidelity. Here, we associate two complementary in vivo assays and a bioinformatics analysis to address the role of phage encoded recombinases in genomic mosaicism. The first assay allowed determining the genetic determinants of mosaic formation between lambdoid phages and Escherichia coli prophage remnants. In the second assay, recombination was monitored between sequences on phage λ, and allowed to compare the performance of three different Rad52-like recombinases on the same substrate. We also addressed the importance of homologous recombination in phage evolution by a genomic comparison of 84 E. coli virulent and temperate phages or prophages. We demonstrate that mosaics are mainly generated by homology-driven mechanisms that tolerate high substrate divergence. We show that phage encoded Rad52-like recombinases act independently of RecA, and that they are relatively more efficient when the exchanged fragments are divergent. We also show that accessory phage genes orf and rap contribute to mosaicism. A bioinformatics analysis strengthens our experimental results by showing that homologous recombination left traces in temperate phage genomes at the borders of recently exchanged fragments. We found no evidence of exchanges between virulent and temperate phages of E. coli. Altogether, our results demonstrate that Rad52-like recombinases promote gene shuffling among temperate phages, accelerating their evolution. This mechanism may prove to be more general, as other mobile genetic elements such as ICE encode Rad52-like functions, and play an important role in bacterial evolution itself.
ano.nymous@ccsd.cnrs.fr.invalid (Marianne de Paepe) 31 Mar 2015
https://hal.science/hal-01134590
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[hal-04616642] Dynamics of the viral community on the surface of a French smear-ripened cheese during maturation and persistence across production years
The surface of smear-ripened cheeses constitutes a dynamic microbial ecosystem resulting from the successive development of different microbial groups such as lactic acid bacteria, fungi, and ripening bacteria. Recent studies indicate that a viral community, mainly composed of bacteriophages, also represents a common and substantial part of the cheese microbiome. However, the composition of this community, its temporal variations, and associations between bacteriophages and their hosts remain poorly characterized. Here, we studied a French smear-ripened cheese by both viral metagenomics and 16S metabarcoding approaches to assess both the succession of phages and bacterial communities on the cheese surface during cheese ripening and their temporal variations in ready-to-eat cheeses over the years of production. We observed a clear transition of the phage community structure during ripening with a decreased relative abundance of viral species (vOTUs) associated with Lactococcus phages, which were replaced by vOTUs associated with phages infecting ripening bacteria such as Brevibacterium, Glutamicibacter, Pseudoalteromonas, and Vibrio . The dynamics of the phage community was strongly associated with bacterial successions observed on the cheese surface. Finally, while some variations in the distribution of phages were observed in ready-to-eat cheeses produced at different dates spanning more than 4 years of production, the most abundant phages were detected throughout. This result revealed the long-term persistence of the dominant phages in the cheese production environment. Together, these findings offer novel perspectives on the ecology of bacteriophages in smear-ripened cheese and emphasize the significance of incorporating bacteriophages in the microbial ecology studies of fermented foods. IMPORTANCE The succession of diverse microbial populations is critical for ensuring the production of high-quality cheese. We observed a temporal succession of phages on the surface of a smear-ripened cheese, with new phage communities showing up when ripening bacteria start covering this surface. Interestingly, the final phage community of this cheese is also consistent over large periods of time, as the same bacteriophages were found in cheese products from the same manufacturer made over 4 years. This research highlights the importance of considering these bacteriophages when studying the microbial life of fermented foods like cheese.
ano.nymous@ccsd.cnrs.fr.invalid (Thomas Paillet) 19 Jun 2024
https://hal.inrae.fr/hal-04616642
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[hal-04396027] Indole, a Signaling Molecule Produced by the Gut Microbiota, Negatively Impacts Emotional Behaviors in Rats
[...]
ano.nymous@ccsd.cnrs.fr.invalid (Mathilde Jaglin) 15 Jan 2024
https://hal.science/hal-04396027
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[hal-04630522] Gut Microbiome Integration in Drug Discovery and Development of Small Molecules
The ongoing discussion regarding the use of mixed or pure cultures of hydrogenotrophic methanogenic archaea in Power-to-Methane (P2M) bioprocess applications persists, with each option presenting its own advantages and disadvantages. To address this issue, a comparison of methane (CH4) yield between a novel methanogenic archaeon belonging to the species Methanothermobacter marburgensis (strain Clermont) isolated from a biological methanation column, and the community from which it originated, was conducted. This comparison included the type strain M. marburgensis str. Marburg. The evaluation also examined how exposure to oxygen (O2) for up to 240 min impacted the CH4 yield across these cultures. While both Methanothermobacter strains exhibit comparable CH4 yield, slightly higher than that of the mixed adapted culture under non-O2-exposed conditions, strain Clermont does not display the lag time observed for strain Marburg.
ano.nymous@ccsd.cnrs.fr.invalid (Patrick Jimonet) 01 Jul 2024
https://hal.inrae.fr/hal-04630522
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[hal-01197434] COnnecting LOcal to GLObal star formation via MIni-starburst
To connect local to global star formation, we compare the mass, size, line width, and star formation rate (SFR) density of cloud structures ranging from Galactic clouds ($\sim$1--10~pc in size) to galaxies (with $\sim$1--10~kpc sizes).Our focus is on molecular cloud structures, called massive complexes, which have intermediate sizes of $\sim$100~pc and masses larger than $10^6~\msun$.We propose using a starburstiness quantity $\zeta$ to quantify the star formation intensity of a molecular cloud complex, or how much its SFR deviates from the star formation relations such as Kennicutt and Lada ones. For molecular cloud complex that has $\zeta>3$ , we call it mini-starburst complex, otherwise normal or main-sequence complex. This $\zeta>3$ threshold leads to a SFR threshold. Most complexes can be classified as ministarburst as having $\zeta>3$. We propose that mini-starburst events represent an enhancement of the star formation activity, following dynamic events such as compression, colliding flows, or agglomeration which compress materials within a cloud complex.
ano.nymous@ccsd.cnrs.fr.invalid (Quang, Nguyen Luong) 11 Sep 2015
https://hal.science/hal-01197434
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[hal-01191221] Goat alphas1-casein genotype affects milk fat globule physicochemical properties and the composition of the milk fat globule membrane
Milk fat secretion is a complex process that initiates in the endoplasmic reticulum of the mammary epithelialcell by the budding of lipid droplets. Lipid droplets are finally released as fat globules in milk envelopedby the apical plasma membrane of the mammary epithelial cell. The milk fat globule membrane (MFGM)thus comprises membrane-specific proteins and polar lipids (glycerophospholipids and sphingolipids) surroundinga core of neutral lipids (mainly triacylglycerols and cholesterol esters). We have recently describedmajor proteins of the MFGM in the goat and we have highlighted prominent differences between goats andbovine species, especially regarding lactadherin, a major MFGM protein. Here, we show that, in the goatspecies, the well-documented genetic polymorphism at the αs1-casein (CSN1S1) locus affects both structureand composition of milk fat globules. We first evidenced that both milk fat globule size and ζ-potentialare related to the αs1-casein genotype. At midlactation, goats displaying strong genotypes for αs1-casein (A/Agoats) produce larger fat globules than goats with a null genotype at the CSN1S1 locus (O/O goats). Alinear relationship (R2 = 0.75) between fat content (g/kg) in the milk and diameter of fat globules (μm) wasestablished. Moreover, we found significant differences with regard to MFGM composition (including bothpolar lipids and MFGM proteins) from goats with extremegenotype at the CSN1S1 locus. At midlactation, the amount of polar lipids is significantly higher in theMFGM from goats with null genotypes for αs1-casein (O/O goats; 5.97 ± 0.11 mg/g of fat; mean ± standarddeviation) than in the MFGM from goats with strong genotypes for αs1-casein (A/A goats; 3.96 ± 0.12 mg/gof fat; mean ± standard deviation). Two MFGM-associated proteins, namely lactadherin and stomatin, arealso significantly upregulated in the MFGM from goats with null genotype for αs1-casein at early lactation. Ourfindings are discussed with regard to techno-functional properties and nutritional value of goat milk. In addition,the genetic polymorphism in the goat species appears to be a tool to provide clues to the lipid secretionpathways in the mammary epithelial cell.
ano.nymous@ccsd.cnrs.fr.invalid (Christelle Cebo) 29 May 2020
https://hal.science/hal-01191221
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[anses-03889040] Avis de l'Anses relatif aux modalités de maîtrise du risque lié à la présence de dangers microbiologiques dans les fromages et autres produits laitiers fabriqués à partir de lait cru
Les produits laitiers au lait cru et les fromages fabriqués à partir de lait cru font régulièrement l’objet de procédures de retrait-rappel et peuvent être notamment impliqués dans l’apparition de cas groupés de salmonelloses et de syndromes hémolytiques et urémiques (SHU). Les dernières alertes épidémiologiques interrogent sur l’état des connaissances et l’efficacité des mesures de maîtrise pouvant être mises en œuvre dans la filière des fromages et autres produits laitiers fabriqués à partir de lait cru ainsi que sur l’évaluation de leur impact sur la réduction du risque pour le consommateur.
ano.nymous@ccsd.cnrs.fr.invalid (Florence Dubois-Brissonnet) 30 Jan 2023
https://anses.hal.science/anses-03889040
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[hal-01190027] A polyphasic approach to study the dynamics of microbial population of an organic wheat sourdough during its conversion to gluten-free sourdough
To develop a method for organic gluten-free (GF) sourdough bread production, a long-term and original wheat sourdough was refreshed with GF flours. The dynamics of the sourdough microbiota during five months of back-slopping were analyzed by classical enumeration and molecular methods, including PCR-temporal temperature gel electrophoresis (PCR-TTGE), multiplex PCR, and pulsed field gel electrophoresis (PFGE). The results showed that the yeast counts remained constant, although Saccharomyces cerevisiae, present in the initial wheat sourdough, was no longer detected in the GF sourdough, while lactic acid bacteria (LAB) counts increased consistently. In the first phase, which was aimed at obtaining a GF sourdough from wheat sourdough, Lactobacillus sanfranciscensis, L. plantarum, and L. spicheri were the main LAB species detected. During the second phase, aimed at maintaining the GF sourdough, the L. plantarum and L. spicheri populations decreased whereas L. sanfranciscensis persisted and L. sakei became the predominant species. Multiplex PCRs also revealed the presence of several L. sakei strains in the GF sourdough. In a search for the origin of the LAB species, PCR-TTGE was performed on the flour samples but only L. sanfranciscensis was detected, suggesting a flour origin for this typical sourdough species. Thus, while replacement of the wheat flour by GF flour influenced the sourdough microbiota, some of the original sourdough LAB and yeast species remained in the GF sourdough. [Int Microbiol 2014; 17(1):1-9]
ano.nymous@ccsd.cnrs.fr.invalid (Emilie Lhomme) 01 Sep 2015
https://hal.science/hal-01190027
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[hal-04417180] A Streamlined Method to Obtain Biologically Active TcdA and TcdB Toxins from Clostridioides difficile
The major virulence factors of Clostridioides difficile (C. difficile) are enterotoxins A (TcdA) and B (TcdB). The study of toxins is a crucial step in exploring the virulence of this pathogen. Currently, the toxin purification process is either laborious and time-consuming in C. difficile or performed in heterologous hosts. Therefore, we propose a streamlined method to obtain functional toxins in C. difficile. Two C. difficile strains were generated, each harboring a sequence encoding a His-tag at the 3′ end of C. difficile 630∆erm tcdA or tcdB genes. Each toxin gene is expressed using the Ptet promoter, which is inducible by anhydro-tetracycline. The obtained purification yields were 0.28 mg and 0.1 mg per liter for rTcdA and rTcdB, respectively. In this study, we successfully developed a simple routine method that allows the production and purification of biologically active rTcdA and rTcdB toxins with similar activities compared to native toxins.
ano.nymous@ccsd.cnrs.fr.invalid (Diane Sapa) 18 Jul 2024
https://hal.inrae.fr/hal-04417180
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[hal-03745296] Role of adherent and invasive Escherichia coli in Crohn’s disease: lessons from the postoperative recurrence model
Objective We used the postoperative recurrence model to better understand the role of adherent and invasive Escherichia coli (AIEC) bacteria in Crohn’s disease (CD), taking advantage of a well-characterised postoperative cohort. Design From a prospective, multicentre cohort of operated patients with CD, AIEC identification was performed within the surgical specimen (M0) (N=181 patients) and the neoterminal ileum (n=119 patients/181) during colonoscopy performed 6 months after surgery (M6). Endoscopic postoperative recurrence was graded using Rutgeerts’ index. The mucosa-associated microbiota was analysed by 16S sequencing at M0 and M6. Relative risks or ORs were adjusted on potential confounders. Results AIEC prevalence was twofold higher within the neoterminal ileum at M6 (30.3%) than within the surgical specimen (14.9%) (p<0.001). AIEC within the neoterminal ileum at M6 was associated with higher rate of early ileal lesions (i1) (41.6% vs 17.1%; aRR 3.49 (95% CI 1.01 to 12.04), p=0.048) or ileal lesions (i2b+i3) (38.2% vs 17.1%; aRR 3.45 (95% CI 1.06 to 11.30), p=0.040) compared with no lesion (i0). AIEC within the surgical specimen was predictive of higher risk of i2b-endoscopic postoperative recurrence (POR) (aOR 2.54 (95% CI 1.01 to 6.44), p=0.049) and severe endoscopic POR (aOR 3.36 (95% CI 1.25 to 9.06), p=0.017). While only 5.0% (6/119) of the patients were AIEC-positive at both M0 and M6, 43.7% (52/119), patients with history of positive test for AIEC (M0 or M6) had higher risk of ileal endoscopic POR (aOR 2.32 (95% CI 1.01 to 5.39), p=0.048)), i2b-endoscopic postoperative recurrence (aOR 2.41 (95% CI 1.01 to 5.74); p=0.048) and severe endoscopic postoperative (aOR=3.84 (95% CI 1.32 to 11.18), p=0.013). AIEC colonisation was associated with a specific microbiota signature including increased abundance of Ruminococcus gnavus . Conclusion Based on the postoperative recurrence model, our data support the idea that AIEC are involved in the early steps of ileal CD. Trial registration number NCT03458195 .
ano.nymous@ccsd.cnrs.fr.invalid (Anthony Buisson) 04 Aug 2022
https://hal.inrae.fr/hal-03745296
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[hal-04659049] On behalf on the Medicen microbiome drug metabolism working group. Gut microbiome integration in drug discovery and development of small molecules opportunities and recommendations
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ano.nymous@ccsd.cnrs.fr.invalid (Patrick Jimonet) 22 Jul 2024
https://hal.inrae.fr/hal-04659049
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[hal-03356343] Transcriptome architecture and regulation at environmental transitions in flavobacteria: the case of an important fish pathogen
The family Flavobacteriaceae (phylum Bacteroidetes ) is a major component of soil, marine and freshwater ecosystems. In this understudied family, Flavobacterium psychrophilum is a freshwater pathogen that infects salmonid fish worldwide, with critical environmental and economic impact. Here, we report an extensive transcriptome analysis that established the genome map of transcription start sites and transcribed regions, predicted alternative sigma factor regulons and regulatory RNAs, and documented gene expression profiles across 32 biological conditions mimicking the pathogen life cycle. The results link genes to environmental conditions and phenotypic traits and provide insights into gene regulation, highlighting similarities with better known bacteria and original characteristics linked to the phylogenetic position and the ecological niche of the bacterium. In particular, osmolarity appears as a signal for transition between free-living and within-host programs and expression patterns of secreted proteins shed light on probable virulence factors. Further investigations showed that a newly discovered sRNA widely conserved in the genus, Rfp18, is required for precise expression of proteases. By pointing proteins and regulatory elements probably involved in host–pathogen interactions, metabolic pathways, and molecular machineries, the results suggest many directions for future research; a website is made available to facilitate their use to fill knowledge gaps on flavobacteria.
ano.nymous@ccsd.cnrs.fr.invalid (Cyprien Guérin) 28 Sep 2021
https://hal.inrae.fr/hal-03356343
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[hal-04611367] Hundreds of antimicrobial peptides create a selective barrier for insect gut symbionts
The spatial organization of gut microbiota is crucial for the functioning of the gut ecosystem, although the mechanisms that organize gut bacterial communities in microhabitats are only partially understood. The gut of the insect Riptortus pedestris has a characteristic microbiota biogeography with a multispecies community in the anterior midgut and a monospecific bacterial population in the posterior midgut. We show that the posterior midgut region produces massively hundreds of specific antimicrobial peptides (AMPs), the Crypt-specific Cysteine-Rich peptides (CCRs) that have membrane-damaging antimicrobial activity against diverse bacteria but posterior midgut symbionts have elevated resistance. We determined by transposon-sequencing the genetic repertoire in the symbiont Caballeronia insecticola to manage CCR stress, identifying different independent pathways, including AMP-resistance pathways unrelated to known membrane homeostasis functions as well as cell envelope functions. Mutants in the corresponding genes have reduced capacity to colonize the posterior midgut, demonstrating that CCRs create a selective barrier and resistance is crucial in gut symbionts. Moreover, once established in the gut, the bacteria differentiate into a CCR-sensitive state, suggesting a second function of the CCR peptide arsenal in protecting the gut epithelia or mediating metabolic exchanges between the host and the gut symbionts. Our study highlights the evolution of an extreme diverse AMP family that likely contributes to establish and control the gut microbiota.
ano.nymous@ccsd.cnrs.fr.invalid (Joy Lachat) 13 Jun 2024
https://hal.science/hal-04611367
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[hal-04279297] Responsible Research and Innovation Exploration of the transcriptional landscapes of culturable and nonculturable bacteria using in vitro transcription emulators
We aim to develop an in vitro platform to express and study gene expression in both culturable and non-culturable bacteria, with a specific focus on the human gut microbiome. The gut microbiome is largely composed of non-culturable bacteria that play a crucial role in human health but are poorly understood, as we have only recently become aware of their presence, mainly through sequencing. Our proposed platform will utilize established cell-free systems and a library of rationally designed plasmids to emulate the transcription machineries of non-model organisms. By employing this approach, we can overcome the limitations of standard protocols for gene expression in non-model organisms, allowing for a systematic and controlled study of their genomes. Through transcriptome analysis, we can identify and map novel transcription units and regulatory elements, as well as understand the dynamics of gene expression. This extensive dataset will not only enhance the functional annotation of non-culturable bacteria but also improve our ability to predict their functional properties. The project focuses on a better understanding of organism diversity as a key to a more profound knowledge of life. It will also pave the way for the identification of promising pathways involved in the production of valuable bio-based chemicals.
ano.nymous@ccsd.cnrs.fr.invalid (Olivier Borkowski) 10 Nov 2023
https://hal.science/hal-04279297
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[hal-01506467] Interactions between genotype and environment drive the metabolic phenotype within Escherichia coli isolates
To gain insights into the adaptation of the Escherichia coli species to different environments, we monitored protein abundances using quantitative proteomics and measurements of enzymatic activities of central metabolism in a set of five representative strains grown in four contrasted culture media including human urine. Two hundred and thirty seven proteins representative of the genome-scale metabolic network were identified and classified into pathway categories. We found that nutrient resources shape the general orientation of metabolism through coordinated changes in the average abundances of proteins and in enzymatic activities that all belong to the same pathway category. For example, each culture medium induces a specific oxidative response whatever the strain. On the contrary, differences between strains concern isolated proteins and enzymes within pathway categories in single environments. Our study confirms the predominance of genotype by environment interactions at the proteomic and enzyme activity levels. The buffering of genetic variation when considering life-history traits suggests a multiplicity of evolutionary strategies. For instance, the uropathogenic isolate CFT073 shows a deregulation of iron demand and increased oxidative stress response.
ano.nymous@ccsd.cnrs.fr.invalid (Victor Sabarly) 12 Apr 2017
https://hal.science/hal-01506467
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[inserm-01779144] Desert Hedgehog Promotes Ischemia-Induced Angiogenesis by Ensuring Peripheral Nerve Survival
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ano.nymous@ccsd.cnrs.fr.invalid (M. Renault) 26 Apr 2018
https://inserm.hal.science/inserm-01779144
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[hal-04540116] Design of a proteolytic module for improved metabolic modeling of Bacteroides caccae
The gut microbiota plays a crucial role in health and is significantly modulated by human diets. In addition to Western diets which are rich in proteins, high-protein diets are used for specific populations or indications, mainly weight loss. In this study, we investigated the effect of protein supplementation on Bacteroides caccae , a Gram-negative gut symbiont. The supplementation with whey proteins led to a significant increase in growth rate, final biomass, and short-chain fatty acids production. A comprehensive genomic analysis revealed that B. caccae possesses a set of 156 proteases with putative intracellular and extracellular localization and allowed to identify amino acid transporters and metabolic pathways. We developed a fully curated genome-scale metabolic model of B. caccae that incorporated its proteolytic activity and simulated its growth and production of fermentation-related metabolites in response to the different growth media. We validated the model by comparing the predicted phenotype to experimental data. The model accurately predicted B. caccae ’s growth and metabolite production ( R 2 = 0.92 for the training set and R 2 = 0.89 for the validation set). We found that accounting for both ATP consumption related to proteolysis, and whey protein accessibility is necessary for accurate predictions of metabolites production. These results provide insights into B. caccae ’s adaptation to a high-protein diet and its ability to utilize proteins as a source of nutrition. The proposed model provides a useful tool for understanding the feeding mechanism of B. caccae in the gut microbiome. IMPORTANCE Microbial proteolysis is understudied despite the availability of dietary proteins for the gut microbiota. Here, the proteolytic potential of the gut symbiont Bacteroides caccae was analyzed for the first time using pan-genomics. This sketches a well-equipped bacteria for protein breakdown, capable of producing 156 different proteases with a broad spectrum of cleavage targets. This functional potential was confirmed by the enhancement of growth and metabolic activities at high protein levels. Proteolysis was included in a B. caccae metabolic model which was fitted with the experiments and validated on external data. This model pinpoints the links between protein availability and short-chain fatty acids production, and the importance for B. caccae to gain access to glutamate and asparagine to promote growth. This integrated approach can be generalized to other symbionts and upscaled to complex microbiota to get insights into the ecological impact of proteins on the gut microbiota.
ano.nymous@ccsd.cnrs.fr.invalid (Amandine Paulay) 10 Apr 2024
https://hal.inrae.fr/hal-04540116
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[hal-02180951] Multiple links connect central carbon metabolism to DNA replication initiation and elongation in Bacillus subtilis
DNA replication is coupled to growth by an unknown mechanism. Here, we investigated this coupling by analyzing growth and replication in 15 mutants of central carbon metabolism (CCM) cultivated in three rich media. In about one-fourth of the condition tested, defects in replication resulting from changes in initiation or elongation were detected. This uncovered 11 CCM genes important for replication and showed that some of these genes have an effect in one, two or three media. Additional results presented here and elsewhere (Janniere, L., Canceill, D., Suski, C., et al. (2007), PLoS One, 2, e447.) showed that, in the LB medium, the CCM genes important for DNA elongation (gapA and ackA) are genetically linked to the lagging strand polymerase DnaE while those important for initiation (pgk and pykA) are genetically linked to the replication enzymes DnaC (helicase), DnaG (primase) and DnaE. Our work thus shows that the coupling between growth and replication involves multiple, medium-dependent links between CCM and replication. They also suggest that changes in CCM may affect initiation by altering the functional recruitment of DnaC, DnaG and DnaE at the chromosomal origin, and may affect elongation by altering the activity of DnaE at the replication fork. The underlying mechanism is discussed.
ano.nymous@ccsd.cnrs.fr.invalid (Hamid Nouri) 26 May 2020
https://hal.science/hal-02180951
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[hal-02356215] Immunome differences between porcine ileal and jejunal Peyer’s patches revealed by global transcriptome sequencing of gut-associated lymphoid tissues
The epithelium of the intestinal mucosa and the gut-associated lymphoid tissues (GALT) constitute an essential physical and immunological barrier against pathogens. In order to study the specificities of the GALT transcriptome in pigs, we compared the transcriptome profiles of jejunal and ileal Peyer's patches (PPs), mesenteric lymph nodes (MLNs) and peripheral blood (PB) of four male piglets by RNA-Seq. We identified 1,103 differentially expressed (DE) genes between ileal PPs (IPPs) and jejunal PPs (JPPs), and six times more DE genes between PPs and MLNs. The master regulator genes FOXP3, GATA3, STAT4, TBX21 and RORC were less expressed in IPPs compared to JPPs, whereas the transcription factor BCL6 was found more expressed in IPPs. In comparison between IPPs and JPPs, our analyses revealed predominant differential expression related to the differentiation of T cells into Th1, Th2, Th17 and iTreg in JPPs. Our results were consistent with previous reports regarding a higher T/B cells ratio in JPPs compared to IPPs. We found antisense transcription for respectively 24%, 22% and 14% of the transcripts detected in MLNs, PPs and PB, and significant positive correlations between PB and GALT transcriptomes. Allele-specific expression analyses revealed both shared and tissue-specific cis-genetic control of gene expression.
ano.nymous@ccsd.cnrs.fr.invalid (T. Maroilley) 26 May 2020
https://hal.science/hal-02356215
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[hal-00933734] Bacterial protein signals are associated with Crohn's disease.
OBJECTIVE: No Crohn's disease (CD) molecular maker has advanced to clinical use, and independent lines of evidence support a central role of the gut microbial community in CD. Here we explore the feasibility of extracting bacterial protein signals relevant to CD, by interrogating myriads of intestinal bacterial proteomes from a small number of patients and healthy controls. DESIGN: We first developed and validated a workflow-including extraction of microbial communities, two-dimensional difference gel electrophoresis (2D-DIGE), and LC-MS/MS-to discover protein signals from CD-associated gut microbial communities. Then we used selected reaction monitoring (SRM) to confirm a set of candidates. In parallel, we used 16S rRNA gene sequencing for an integrated analysis of gut ecosystem structure and functions. RESULTS: Our 2D-DIGE-based discovery approach revealed an imbalance of intestinal bacterial functions in CD. Many proteins, largely derived from Bacteroides species, were over-represented, while under-represented proteins were mostly from Firmicutes and some Prevotella members. Most overabundant proteins could be confirmed using SRM. They correspond to functions allowing opportunistic pathogens to colonise the mucus layers, breach the host barriers and invade the mucosae, which could still be aggravated by decreased host-derived pancreatic zymogen granule membrane protein GP2 in CD patients. Moreover, although the abundance of most protein groups reflected that of related bacterial populations, we found a specific independent regulation of bacteria-derived cell envelope proteins. CONCLUSIONS: This study provides the first evidence that quantifiable bacterial protein signals are associated with CD, which can have a profound impact on future molecular diagnosis.
ano.nymous@ccsd.cnrs.fr.invalid (Catherine Juste) 28 May 2020
https://hal.science/hal-00933734
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[hal-01204441] Following Pathogen Development and Gene Expression in a Food Ecosystem: the Case of a Staphylococcus aureus Isolate in Cheese
Human intoxication or infection due to bacterial food contamination constitutes an economic challenge and a public health problem. Information on the in situ distribution and expression of pathogens responsible for this risk is to date lacking, largely because of technical bottlenecks in detecting signals from minority bacterial populations within a complex microbial and physicochemical ecosystem. We simulated the contamination of a real high-risk cheese with a natural food isolate of Staphylococcus aureus, an enterotoxin-producing pathogen responsible for food poisoning. To overcome the problem of a detection limit in a solid matrix, we chose to work with a fluorescent reporter (superfolder green fluorescent protein) that would allow spatiotemporal monitoring of S. aureus populations and targeted gene expression. The combination of complementary techniques revealed that S. aureus localizes preferentially on the cheese surface during ripening. Immunochemistry and confocal laser scanning microscopy enabled us to visualize, in a single image, dairy bacteria and pathogen populations, virulence gene expression, and the toxin produced. This procedure is readily applicable to other genes of interest, other bacteria, and different types of food matrices.
ano.nymous@ccsd.cnrs.fr.invalid (Isabelle Fleurot) 28 May 2020
https://hal.science/hal-01204441
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[hal-03164832] Analysis of Polycerate Mutants Reveals the Evolutionary Co-option of HOXD1 for Horn Patterning in Bovidae
In the course of evolution, pecorans (i.e. higher ruminants) developed a remarkable diversity of osseous cranial appendages, collectively referred to as 'headgear', which likely share the same origin and genetic basis. However, the nature and function of the genetic determinants underlying their number and position remain elusive. Jacob and other rare populations of sheep and goats are characterized by polyceraty, the presence of more than two horns. Here, we characterize distinct POLYCERATE alleles in each species, both associated with defective HOXD1 function. We show that haploinsufficiency at this locus results in the splitting of horn bud primordia, likely following the abnormal extension of an initial morphogenetic field. These results highlight the key role played by this gene in headgear patterning and illustrate the evolutionary co-option of a gene involved in the early development of bilateria to properly fix the position and number of these distinctive organs of Bovidae.
ano.nymous@ccsd.cnrs.fr.invalid (Aurélie Allais-Bonnet) 28 Oct 2021
https://hal.science/hal-03164832
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[hal-02433421] Human IgA binds a diverse array of commensal bacteria
In humans, several grams of IgA are secreted every day in the intestinal lumen. While only one IgA isotype exists in mice, humans secrete IgA1 and IgA2, whose respective relations with the microbiota remain elusive. We compared the binding patterns of both polyclonal IgA subclasses to commensals and glycan arrays and determined the reactivity profile of native human monoclonal IgA antibodies. While most commensals are dually targeted by IgA1 and IgA2 in the small intestine, IgA1+IgA2+ and IgA1-IgA2+ bacteria coexist in the colon lumen, where Bacteroidetes is preferentially targeted by IgA2. We also observed that galactose-α terminated glycans are almost exclusively recognized by IgA2. Although bearing signs of affinity maturation, gut-derived IgA monoclonal antibodies are cross-reactive in the sense that they bind to multiple bacterial targets. Private anticarbohydrate-binding patterns, observed at clonal level as well, could explain these apparently opposing features of IgA, being at the same time cross-reactive and selective in its interactions with the microbiota.
ano.nymous@ccsd.cnrs.fr.invalid (Delphine Sterlin) 09 Jan 2020
https://hal.sorbonne-universite.fr/hal-02433421
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[hal-02627949] Microbiota-gut brain axis involvement in neuropsychiatric disorders
Introduction: The microbiota-gut brain (MGB) axis is the bidirectional communication between the intestinal microbiota and the brain. An increasing body of preclinical and clinical evidence has revealed that the gut microbial ecosystem can affect neuropsychiatric health. However, there is still a need of further studies to elucidate the complex gene-environment interactions and the role of the MGB axis in neuropsychiatric diseases, with the aim of identifying biomarkers and new therapeutic targets, to allow early diagnosis and improving treatments. Areas covered: To review the role of MGB axis in neuropsychiatric disorders, prediction and prevention of disease through exploitation, integration, and combination of data from existing gut microbiome/microbiota projects and appropriate other International '-Omics' studies. The authors also evaluated the new technological advances to investigate and modulate, through nutritional and other interventions, the gut microbiota. Expert opinion: The clinical studies have documented an association between alterations in gut microbiota composition and/or function, whereas the preclinical studies support a role for the gut microbiota in impacting behaviors which are of relevance to psychiatry and other central nervous system (CNS) disorders. Targeting MGB axis could be an additional approach for treating CNS disorders and all conditions in which alterations of the gut microbiota are involved.
ano.nymous@ccsd.cnrs.fr.invalid (Luigi Francesco Iannone) 26 May 2020
https://hal.inrae.fr/hal-02627949
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[hal-03343060] Condition-Dependent Transcriptome Reveals High-Level Regulatory Architecture in Bacillus subtilis
Bacteriophage-encoded single strand annealing proteins (SSAPs) are recombinases which can substitute the classical, bacterial RecA and manage the DNA metabolism at different steps of phage propagation. SSAPs have been shown to efficiently promote recombination between short and rather divergent DNA sequences and were exploited for in vivo genetic engineering mainly in Gram-negative bacteria. In opposition to the conserved and almost universal bacterial RecA protein, SSAPs display great sequence diversity. The importance for SSAPs in phage biology and phage-bacteria evolution is underlined by their role as key players in events of horizontal gene transfer (HGT). All of the above provoke a constant interest for the identification and study of new phage recombinase proteins in vivo , in vitro as well as in silico . Despite this, a huge body of putative ssap genes escapes conventional classification, as they are not properly annotated. In this work, we performed a wide-scale identification, classification and analysis of SSAPs encoded by the Firmicutes bacteria and their phages. By using sequence similarity network and gene context analyses, we created a new high quality dataset of phage-related SSAPs, substantially increasing the number of annotated SSAPs. We classified the identified SSAPs into seven distinct families, namely RecA, Gp2.5, RecT/Redβ, Erf, Rad52/22, Sak3, and Sak4, organized into three superfamilies. Analysis of the relationships between the revealed protein clusters led us to recognize Sak3-like proteins as a new distinct SSAP family. Our analysis showed an irregular phylogenetic distribution of ssap genes among different bacterial phyla and specific phages, which can be explained by the high rates of ssap HGT. We propose that the evolution of phage recombinases could be tightly linked to the dissemination of bacterial phage-resistance mechanisms (e.g., abortive infection and CRISPR/Cas systems) targeting ssap genes and be a part of the constant phage-bacteria arms race.
ano.nymous@ccsd.cnrs.fr.invalid (P. Nicolas) 13 Sep 2021
https://hal.inrae.fr/hal-03343060
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[hal-01204316] Segmented Filamentous Bacterium Uses Secondary and Tertiary Lymphoid Tissues to Induce Gut IgA and Specific T Helper 17 Cell Responses
Segmented filamentous bacterium (SFB) is a symbiont that drives postnatal maturation of gut adaptive immune responses. In contrast to nonpathogenic E. coli, SFB stimulated vigorous development of Peyer's patches germinal centers but paradoxically induced only a low frequency of specific immunoglobulin A (IgA)-secreting cells with delayed accumulation of somatic mutations. Moreover, blocking Peyer's patch development abolished IgA responses to E. coli, but not to SFB. Indeed, SFB stimulated the postnatal development of isolated lymphoid follicles and tertiary lymphoid tissue, which substituted for Peyer's patches as inductive sites for intestinal IgA and SFB-specific T helper 17 (Th17) cell responses. Strikingly, in mice depleted of gut organized lymphoid tissue, SFB still induced a substantial but nonspecific intestinal Th17 cell response. These results demonstrate that SFB has the remarkable capacity to induce and stimulate multiple types of intestinal lymphoid tissues that cooperate to generate potent IgA and Th17 cell responses displaying only limited target specificity.
ano.nymous@ccsd.cnrs.fr.invalid (Emelyne Lecuyer) 23 Sep 2015
https://hal.science/hal-01204316
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[hal-03048771] The low-grade intestinal inflammation potentiates sarcopenia in Old Adults that can be restrained by a probiotic: Streptococcus Thermophilus CNRZ160
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ano.nymous@ccsd.cnrs.fr.invalid (Isabelle Savary-Auzeloux) 29 Nov 2021
https://hal.inrae.fr/hal-03048771
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[hal-03455954] Souche de Streptococcus thermophilus CNRZ160 pour le traitement et la prévention de l’inflammation intestinale et des désordres associés, chez un individu »: couvre la supplémentation avec S thermophilus pour traiter ou prévenir une inflammation intestinale ou un trouble associé
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ano.nymous@ccsd.cnrs.fr.invalid (Dominique Dardevet) 29 Nov 2021
https://hal.science/hal-03455954
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[hal-03048700] Streptococcus thermophilus CNRZ160 preserves muscle anabolism in old rats by targeting anti-inflamamtory mechanisms at the gut level: A probable gut-muscle crosstalk
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ano.nymous@ccsd.cnrs.fr.invalid (Isabelle Savary-Auzeloux) 29 Nov 2021
https://hal.inrae.fr/hal-03048700
