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The digestive physiology of the germ-free animal has a number of characteristics (cecal hypertrophy, slower small intestine cell renewal, slower gastric emptying and intestinal transit) which distinguish it from that of the conventional animal. If the germ-free model is to be used to determine the role of gastrointestinal microflora in the nutrition of the conventional animal, it is essential to complete the study of these characteristics by data on digestive enzymes in the germ-free. The present paper analyzes these data. There is little information on salivary amylase and none on gastric proteolytic enzymes and intestinal peptidases. More complete data on exocrine pancreas enzymes and intestinal disaccharidases show that the digestive equipment is similar in germ-free and conventional animals. Bile salts, not considered as digestive enzymes, are qualitatively and quantitatively different, depending on the digestive tract bacterial environment. In general, the germ-free animal has some characteristics which should permit better utilization of the diet ingested. Measurements of apparent digestibility do not confirm this hypothesis since results obtained in germ-free and conventional animals of the same species are contradictory.

Bile salt output and digestive utilization of a high-lipid diet were studied in pig bile collected from the common bile duct and infused into the duodenum then into the distal ileum of the animal. Five Large-White pigs of 50 kg body-weight were fitted with three permanent fistulae, one in the common bile duct for complete collection of bile secretion, and the other two in the duodenum and the ileum (at about 80 cm from the ileo-caecal valve) for returning the secretion to the animal. The diversion of the bile into the distal ileum led to a significant 82 p. 100 decrease in daily bile acid output and a 36 p. 100 decrease in apparent digestibility of dietary energy when compared with values obtained when bile was infused into the duodenum. The latter decrease was mainly due to a marked diminution ( - 60.4 p. 100) of the digestive utilization of the dietary lipids. The results show the role of the pig small intestine in maintaining an efficient enterohepatic circulation of bile acid as a result of optimal utilization of the dietary lipids.

Metaproteomics has matured into a powerful tool to assess functional interactions in microbial communities. While many metaproteomic workflows are available, the impact of method choice on results remains unclear. Here, we carry out a community-driven, multi-laboratory comparison in metaproteomics: the critical assessment of metaproteome investigation study (CAMPI). Based on well-established workflows, we evaluate the effect of sample preparation, mass spectrometry, and bioinformatic analysis using two samples: a simplified, laboratory-assembled human intestinal model and a human fecal sample. We observe that variability at the peptide level is predominantly due to sample processing workflows, with a smaller contribution of bioinformatic pipelines. These peptide-level differences largely disappear at the protein group level. While differences are observed for predicted community composition, similar functional profiles are obtained across workflows. CAMPI demonstrates the robustness of present-day metaproteomics research, serves as a template for multi-laboratory studies in metaproteomics, and provides publicly available data sets for benchmarking future developments., The authors present CAMPI, a large-scale multi-lab comparison of diverse metaproteomics workflows. CAMPI provides insights into the robustness of current methods, suggests further improvements to the field, and may pave the way for future community-driven metaproteomics projects.

Metaproteomics focuses on identifying and quantifying proteins in complex biological samples such as the human gut microbiota. The analysis of several hundred of samples is of interest given the growing recognition of this ecosystem as a health partner. However, the methods and protocols used so far in metaproteomics are not suitable for large-scale studies. We have therefore developed algorithms, evaluated and compared several identification approaches for peptides and proteins and proposed systematic evaluation criteria, with a particular interest in the replicability of identifications, in order to develop a pre-treatment pipeline suitable for wide-ranging studies. This work bring a methodological base so far missing in the field of the metaproteomics of the human gut microbiota. Quantification of peptides and proteins by XIC has never been performed on this type of data, we have also compared normalization methods and developed a methodology for imputing missing data to refine the abundance estimations obtained by the more classical method of SC. This thesis work has highlighted microbial biomarkers of potential interest for predicting the response to a slimming diet, or to characterize various phenotypes of IBD. We have also been able to analyse the metaproteome of more than 200 patients in the framework of the ProteoCardis ANR, which is ancillary to the European project MetaCardis, and which focuses on the potential link between gut microbiota and cardiovascular diseases. The search for proteins of interest among these data should allow us to discover protective or aggravating candidate biomarkers of cardiovascular diseases., La métaprotéomique s’attache à identifier et quantifier les protéines d’échantillons biologiques complexes comme le microbiote intestinal humain. L’analyse de plusieurs centaines d’échantillons revêt un intérêt évident compte tenu de la reconnaissance croissante de cet écosystème en tant que partenaire santé. Cependant, les méthodes et protocoles utilisés jusqu’à ce jour en métaprotéomique ne sont pas adaptés à des études de grande ampleur. Nous avons développé des algorithmes, évalué et comparé plusieurs approches d’identification des peptides et protéines et proposé des critères d’évaluation systématiques, avec un intérêt particulier porté sur la réplicabilité des identifications, afin de développer un pipeline de prétraitement adapté à des études d’envergure. Ce travail apporte un socle méthodologique jusqu’ici manquant dans le domaine de la métaprotéomique du microbiote intestinal humain. Nous avons également comparé des méthodes de normalisation des XIC et développé une méthodologie d’imputation des données manquantes permettant d’affiner les estimations d’abondances obtenues par la méthode de SC. Ce travail de thèse a permis de mettre en évidence des biomarqueurs microbiens potentiellement d’intérêt pour prédire la réponse à un régime amaigrissant ou pour caractériser différents phénotypes de MICI. Nous avons également analysé le métaprotéome de plus de 200 patients dans le cadre de l’ANR ProteoCardis adossée au projet MetaCardis, et s’intéressant au lien possible entre microbiote intestinal et maladies cardiovasculaires. La recherche de protéines d’intérêt parmi ces données devrait permettre de découvrir des candidats biomarqueurs de maladies cardiovasculaires.

Metaproteomics focuses on identifying and quantifying proteins in complex biological samples such as the human gut microbiota. The analysis of several hundred of samples is of interest given the growing recognition of this ecosystem as a health partner. However, the methods and protocols used so far in metaproteomics are not suitable for large-scale studies. We have therefore developed algorithms, evaluated and compared several identification approaches for peptides and proteins and proposed systematic evaluation criteria, with a particular interest in the replicability of identifications, in order to develop a pre-treatment pipeline suitable for wide-ranging studies. This work bring a methodological base so far missing in the field of the metaproteomics of the human gut microbiota. Quantification of peptides and proteins by XIC has never been performed on this type of data, we have also compared normalization methods and developed a methodology for imputing missing data to refine the abundance estimations obtained by the more classical method of SC. This thesis work has highlighted microbial biomarkers of potential interest for predicting the response to a slimming diet, or to characterize various phenotypes of IBD. We have also been able to analyse the metaproteome of more than 200 patients in the framework of the ProteoCardis ANR, which is ancillary to the European project MetaCardis, and which focuses on the potential link between gut microbiota and cardiovascular diseases. The search for proteins of interest among these data should allow us to discover protective or aggravating candidate biomarkers of cardiovascular diseases.; La métaprotéomique s’attache à identifier et quantifier les protéines d’échantillons biologiques complexes comme le microbiote intestinal humain. L’analyse de plusieurs centaines d’échantillons revêt un intérêt évident compte tenu de la reconnaissance croissante de cet écosystème en tant que partenaire santé. Cependant, les méthodes et protocoles utilisés jusqu’à ce jour en métaprotéomique ne sont pas adaptés à des études de grande ampleur. Nous avons développé des algorithmes, évalué et comparé plusieurs approches d’identification des peptides et protéines et proposé des critères d’évaluation systématiques, avec un intérêt particulier porté sur la réplicabilité des identifications, afin de développer un pipeline de prétraitement adapté à des études d’envergure. Ce travail apporte un socle méthodologique jusqu’ici manquant dans le domaine de la métaprotéomique du microbiote intestinal humain. Nous avons également comparé des méthodes de normalisation des XIC et développé une méthodologie d’imputation des données manquantes permettant d’affiner les estimations d’abondances obtenues par la méthode de SC. Ce travail de thèse a permis de mettre en évidence des biomarqueurs microbiens potentiellement d’intérêt pour prédire la réponse à un régime amaigrissant ou pour caractériser différents phénotypes de MICI. Nous avons également analysé le métaprotéome de plus de 200 patients dans le cadre de l’ANR ProteoCardis adossée au projet MetaCardis, et s’intéressant au lien possible entre microbiote intestinal et maladies cardiovasculaires. La recherche de protéines d’intérêt parmi ces données devrait permettre de découvrir des candidats biomarqueurs de maladies cardiovasculaires.