HAL ENVT (Ecole Nationale Vétérinaire de Toulouse)
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Real-time metabolomics on E. coli cell cultures
No full textInternational audienceFermentation monitoring is a powerful tool for bioprocess development, optimisation and control. Several parameters can be tracked including temperature, dissolved oxygen, pH, metabolites, etc... As metabolites represent the final step of biological regulation, they are key indicators. Metabolic changes can occur very rapidly, and traditional metabolomic analyses are time-consuming. Therefore, online high-resolution time course analysis is required. To achieve this, we adapt a real-time metabolomic workflow (Cortada-Garcia, et al., 2022). As a proof of concept, we monitored metabolites from an E.coli culture and the incorporation of 13C after a spike of 13C glucose. A set of modules, including culture pump, solvent pumps, 6 and 10 ports valves and filtration valve, links the bioreactor to the mass spectrometer. A workflow manager, developed in Python, was used to pilot fluidic system and launch analysis sequences. The E. coli culture was analysed every minute using flow injection mass spectrometry at resolution of 90 000 for m/z = 200 (Exploris 240, Thermo Scientific). An integrated solution for real-time data processing, enabling direct monitoring of the bioreactor is currently in progress. The system will be capable to measure, identify and quantify metabolites, visualize the results and modulate the fermentation process if needed. The next steps of this development involve improving metabolome coverage by MS, implementing metabolite quantification with internal standard or by benchtop NMR in series. This approach should be transposable to other types of culture, such as adherent cells
Hierarchical Long Short Term Memory Recurrent Neural Network for Goats Behaviour Prediction from Accelerometer Data
No full textInternational audienceGastrointestinal parasitism is a major challenge in small grazing ruminants, affecting animal welfare and farmers’income. In this regard, monitoring individual animal behaviour could help to develop new selection schemesthat favor animals with a lower risk of larval infestation, but also support the targeted use of anthelmintic byfocusing only on infested animals. Accelerometer sensors are widely used in combination with statistical modelsto predict the behaviour of grazing ruminants, but the lack of generalization of the models and the limited rangeof well-predicted behaviours are still challenging. In our study, we introduce an innovative methodology based onhierarchical long short term memory (LSTM) recurrent neural networks to predict the main behaviours of goatson pasture. For that purpose, we collected accelerometer data from the horns of 59 Creole goats and annotatedthe behaviour over 144 hours of data. We defined 13 moving features that are mathematical combinations ofthe raw data to get more information while preserving the temporal structure of the accelerometer time-series. Adata augmentation technique involving the addition of random noise was applied to sequences from the minoritybehaviour labels. A hierarchical LSTM model was then built to derive behaviours from a given accelerometersignal, by sequentially combining several models that first tackle simple classification tasks (e.g., grazing ornon-grazing segments), then increasingly complex ones (e.g., displacement or other activities), progressivelywithdrawing segments that have already been identified. The hierarchical LSTM model was validated using atesting set consisting of goats not seen during model training, and carefully selected to maximize behaviourallabels heterogeneity. Performance of the hierarchical LSTM model was also compared to those of a regular LSTMmodel which directly classified the raw signal into the 5 behaviours, used as the baseline. Highest performancewas obtained with the hierarchical LSTM model, reaching a Fscore of 87.84%, a precision of 89.44% and a recallof 86.3%. Best performance was obtained for grazing prediction (recall: 99.5%; precision: 99.4%), followingby resting (recall: 98.0%; precision: 98.4%) and ruminating (recall: 95.2%; precision: 89%). Most confusionsoccurred with the displacement (recall: 51.2%; precision: 67.8%), likely due to the low number of sequencesin the dataset (0.42% of the dataset). While other avenues remain to be explored for improving the predictionof such rare behaviours, our approach introduces key innovations that not only address the methodologicallimitations identified in the literature, but also facilitate further exploration of the role of goat behaviour inmanaging gastrointestinal parasitism on pasture
A consortium of seven commensal bacteria promotes gut microbiota recovery and strengthens ecological barrier against vancomycin-resistant enterococci
No full textInternational audienceVancomycin-resistant enterococci (VRE) often originate from the gastrointestinal tract, where their proliferation precedes dissemination into the bloodstream, and can lead to systemic infection. Uncovering the actors and mechanisms reducing the intestinal colonisation by VRE is essential to control infection. We aimed to identify commensal bacteria that interfere with VRE gut colonisation or act as an ecological barrier.ResultsWe performed a 3-week longitudinal analysis of the gut microbiota composition and VRE carriage levels during microbiota recovery in mice colonised with VRE after antibiotic-induced dysbiosis. By combining biological data and mathematical modelling, we identified 15 molecular species (OTUs) that negatively correlated with VRE overgrowth. Six strains representative of these OTUs were collected, cultivated and used in mixture with a seventh strain (Mix7) in two different mouse lines challenged with VRE. Of the seven strains, three belonged to Lachnospiraceae, one to Muribaculaceae, one to Ruminococcaceae and two to Lactobacillaceae. We found that Mix7 led to a better recovery of the gut microbiota composition and reduced VRE carriage. Differences in the effect of Mix7 were observed between responder and non-responder mice. These differences were associated with variations in the composition of the initial microbiota and during recovery and represent potential biomarkers for predicting response to Mix7. In a mouse model of alternative stable state of dysbiosis, response to Mix7 was associated with higher concentrations of short-chain fatty acids (acetate, propionate, butyrate) and a range of metabolites including bile acids, reflecting the recovery of the microbiota back to initial state. Furthermore, Muribaculum intestinale strain was required to obtain the Mix7 effect on VRE reduction in vivo, but the presence of at least one of the other six strains was needed. None of the supernatant of the seven strains, alone or in combination, inhibited VRE growth in vitro. Interestingly, five strains belong to species shared among humans and mice, and the other two have human functional equivalents.ConclusionsAn innovative approach based on mathematical modelling of the microbiota composition permitted to identify a mixture of commensal bacterial strains, which improves the ecological barrier effect against VRE. The mechanisms are dependent on the recovery and initial composition of the microbiota. Ultimately, this work will enable a move towards a personalised medicine by targeting predisposed patients presenting a risk of infection, such as neutropenic or bone-marrow transplant patients, and likely to respond to supplementation with commensal strains, providing new live biotherapeutic products and biomarkers to predict response to supplementation.--QgEhBLB8UrKv1oYceiTBVideo AbstractConclusionsAn innovative approach based on mathematical modelling of the microbiota composition permitted to identify a mixture of commensal bacterial strains, which improves the ecological barrier effect against VRE. The mechanisms are dependent on the recovery and initial composition of the microbiota. Ultimately, this work will enable a move towards a personalised medicine by targeting predisposed patients presenting a risk of infection, such as neutropenic or bone-marrow transplant patients, and likely to respond to supplementation with commensal strains, providing new live biotherapeutic products and biomarkers to predict response to supplementation
Low Mutation Rate and Atypical Mutation Spectrum in Prasinoderma coloniale : Insights From an Early Diverging Green Lineage
No full textInternational audienceMutations are the ultimate source of genetic diversity on which natural selection and genetic drift act, playing a crucial role in evolution and long-term adaptation. At the molecular level, the spontaneous mutation rate (µ), defined as the number of mutations per base per generation, thus determines the adaptive potential of a species. Through a mutation accumulation experiment, we estimate the mutation rate and spectrum in Prasinoderma coloniale, a phytoplankton species from an early-branching lineage within the Archaeplastida, characterized by an unusually high genomic guanine-cytosine (GC) content (69.8%). We find that P. coloniale has a very low total mutation rate of µ = 2.00 × 10−10. The insertion–deletion mutation rate is almost 5 times lesser than the single nucleotide mutation rate with µID = 3.40 × 10−11 and µSNM = 1.62 × 10−10. Prasinoderma coloniale also exhibits an atypical mutational spectrum: While essentially all other eukaryotes show a bias toward GC to AT mutations, no evidence of this AT-bias is observed in P. coloniale. Since cytosine methylation is known to be mutagenic, we hypothesized that this may result from an absence of C-methylation. Surprisingly, we found high levels of C-methylation (14% in 5mC, 25% in 5mCG contexts). Methylated cytosines did not show increased mutation rates compared with unmethylated ones, not supporting the prevailing notion that C-methylation universally leads to higher mutation rates. Overall, P. coloniale combines a GC-rich genome with a low mutation rate and original mutation spectrum, suggesting the almost universal AT-bias may not have been present in the ancestor of the green lineage
Implantation of Aspergillus Section Flavi in French Maize and Consequences on Aflatoxin Contamination of Maize at Harvest: Three-Year Survey
No full textInternational audienceThe worldwide distribution of aflatoxin B1, a carcinogenic mycotoxin, is changing due to climate change. This frequent contaminant of crops in tropical and subtropical regions is an emerging threat in Europe. Its first appearance in French maize was reported in 2015, a year with exceptional climatic conditions. But, from this year, such conditions (drought during spring and hot summers) occurred regularly, raising the question of a possible durable implantation of aflatoxigenic fungal species in French maize fields. To answer this question, 554 maize samples were collected during three consecutive years (2018–2020) throughout the French territory. They were subjected to mycological analysis and strains belonging to the Flavi section of the Aspergillus genus were identified, and their toxigenic potential was determined. This survey demonstrates that Aspergillus section Flavi are now frequent contaminants of maize since they were observed in more than 50% of samples in 2018 and 2019. This prevalence sharply increased in 2020 to reach 80% contamination. In parallel, the frequency of contamination with aflatoxins also increased. While it was very limited during the first two years of the study, despite favorable climatic conditions, contamination with aflatoxins was present in 16% of samples analyzed in 2020, exceeding E.U. regulation by 5%. Even if Aspergillus flavus is the dominant species, representing more than 90% of isolated strains, Aspergillus parasiticus seems to play a major role in grain contamination, as demonstrated by the presence of AFG in 50% of contaminated samples. These findings highlight the need to rethink the monitoring and management of aflatoxin risk in France for the future
Création et utilisation d’un jeu sérieux comme outil d’apprentissage de l’anatomie et de la physiologie de l’oreille des espèces d’intérêt vétérinaire
No full textPedagogy is constantly evolving, and the advent of new technologies offers a multitude of possibilities. The study of anatomy is no exception, and the development of serious games has been a viable option explored at the National Veterinary School of Toulouse for several years. The ear, a complex organ requiring in-depth anatomical knowledge, is particularly well-suited to this type of game, as it facilitates understanding. However, the contribution of these games to pedagogy remains insufficiently known and quantified. The implementation of specific evaluation criteria has made it possible to better assess students' satisfaction, their understanding, and their sense of immersion after participating in the game.La pédagogie est en constante évolution et l’arrivée des nouvelles technologies offrent une multitude de possibilités. L’apprentissage de l’anatomie ne fait pas exception et l’élaboration de jeux sérieux est une réelle option développée à l’École Nationale Vétérinaire de Toulouse depuis quelques années. L’oreille, organe complexe nécessitant un apprentissage anatomique approfondi, se prête parfaitement à la réalisation de ce type de jeu afin d’en faciliter la compréhension. Mais l’apport de ces jeux dans la pédagogie est encore trop peu connu et trop peu quantifié. La mise en place de certains critères d’évaluation ont permis de mieux quantifier la satisfaction des élèves ainsi que leur compréhension et leur sentiment d’immersion au sortir du jeu
Musique et animaux : effets d’un enrichissement auditif musical sur le bien-être de chiens utilisés à des fins scientifiques
No full textThis thesis aims to clarify the complex link between music and animal well-being, particularly its potential in the enrichment of the living environment of animals used for scientific purposes. The current challenge is to draw inspiration from bioacoustic research to create appropriate musical compositions that are specific to the behavioural objectives targeted. The goal of this thesis was to study the behavioural response of dogs exposed to musical auditory enrichment under controlled conditions. We worked with Beagle dogs from an experimental kennel, analysing their vocalisations and activity levels. In our experimental conditions, we found that the presence of music was associated with a significant reduction in vocalisations and overall activity, alongside an increase in time spent lying down. Introducing slow-tempo classical orchestral music would therefore promote calmness in dogs used for scientific purposes, thus representing a relevant form of enrichment, whether in research settings or for dogs living in group housingCette thèse a pour but d’expliciter le lien complexe qu’il existe entre musique et bien-être animal, et son intérêt dans le cadre de l’enrichissement du milieu de vie d’animaux utilisés à des fins scientifiques. L’enjeux actuel est donc de s’inspirer des travaux en bioacoustique afin d’utiliser une composition musicale adaptée, qui serait spécifique aux objectifs comportementaux visés. L’objectif de ce travail de thèse a été d’étudier la réponse comportementale de chiens soumis à un enrichissement auditif de type musical en condition contrôlées. Nous avons travaillé avec les chiens Beagles d’un chenil d’expérimentation, en analysant les vocalisations et leur activité. Dans nos conditions d’expérimentations, nous avons constaté que la présence de musique était liée une diminution significative des vocalisations ainsi que de l’activité générale des chiens, parallèlement à une augmentation du temps passé en position couchée. L'introduction de musique classique orchestrale à tempo lent favoriserait donc le calme chez les chiens utilisés à des fins scientifiques, représentant donc un enrichissement pertinent, que ce soit dans le cadre de la recherche ou pour des chiens vivant en collectivit
SPIDER Système Partagé d’Interopérabilité des Données d’Expérimentation et de Recherche: Une plateforme de convergence et d’interopérabilité des données animales
No full textInternational audienceSPIDER aims to make known and share all the information available on an animal or a list of animals, in INRAE's experimental facilities.It ensures interoperability with other data sources, such as CRB Anim, and enriches information with metadata (INRAE ontologies and thesauri)
Risques d'intoxication des animaux au pâturage suite à l'invasion des surfaces herbagères par le Séneçon du Cap Titre court : risques d'intoxication par le Séneçon du Cap
No full textInternational audienceThe invasion of European grasslands by Cape ragwort motivated to study its contents in alkaloids in comparison to European ragworts, and to evaluate the toxicological risks for herbivorous animals. Seneciphylline could not be quantified in Cape Ragwort but was measured in notable amounts in Common Ragwort. Contrarily, Cape ragwort showed high concentrations in senecionine and especially rétrorsine, clearly higher than those measured in Jacobaea vulgaris, Jacobaea eruciflora and Senecio vulgaris. It is essential that all stakeholders become aware of the risk linked to the spread of Cape ragwort for animals.Comparer les taux d'alcaloïdes du Séneçon du Cap (Senecio inaequidens), qui envahit les prairies d'Europe, avec ceux de 3 Séneçons indigènes (Jacobaea vulgaris, Jacobaea eruciflora et Senecio vulgaris), permet d'évaluer le risque toxicologique qu'ils présentent pour les herbivores. Dans le Séneçon du Cap, la sénéciphylline n'a pas pu être quantifiée, contrairement au Séneçon commun. Par contre, des concentrations élevées en sénécionine et en rétrorsine ont été mesurées dans le Séneçon du Cap, en comparaison aux 3 Séneçons européens. Il est indispensable que tous les acteurs prennent conscience du risque lié à la dissémination du séneçon du Cap pour les animaux