Clément Lozano

Effets des filtres UV organiques sur les bactéries marines hétérotrophes: Les filtres UV sont les principes actifs des produits de protection solaire et sont utilisés comme agents stabilisateurs dans les produits cosmétiques industriels. Plusieurs études ont signalé la présence de filtres UV dans les eaux côtières du monde entier et ont démontré leur bioaccumulation et leurs effets toxiques sur les organismes aquatiques. Les bactéries marines dominent la biomasse marine et sont d’une importance majeure pour les cycles biogéochimiques. En dépit de leurs rôles essentiels, la toxicité des filtres UV n'a jamais été évaluée sur les microorganismes. Cette thèse vise à explorer les effets des filtres UV organiques les plus courants, en tenant compte de la réponse physiologique, morphologique et moléculaire, sur des bactéries hétérotrophes environnementale, isolées de la baie de Banyuls (Méditerranée Nord-Ouest, France). Ce projet a démontré les effets bactéricides espèce et molécule dépendants des filtres UV organiques les plus communs sur les bactéries marines. Nos recherches ont contribué à une meilleure compréhension de la réponse cellulaire induite par l'exposition aux filtres UV. Nous avons montré, par le biais d'une étude morphologique et via des techniques de protéomique quantitative, que les bactéries présentaient une variété de réponses moléculaires liées au stress, corroborant que les bactéries sont des modèles d’étude écotoxicologiques pertinents. De futures recherches méta-omiques sur les communautés naturelles et artificielles nous permettront d'évaluer l'impact des apports anthropiques sur les fonctions des communautés microbiennes

Plasma proteomics is a rapid, noninvasive, and highly informative approach for identifying disease biomarkers. However, the wide dynamic range of protein concentration limits the depth of liquid chromatography–tandem mass spectrometry proteomics. To address this challenge, we evaluated, with an Orbitrap Astral instrument using DIA, the performance of several protein depleting and enriching technologies on platelet-free plasma. Specifically, we assessed: perchloric acid depletion, immunodepletion, ProteoMiner, MagNet-SAX, ENRICHplus, Proteonano and the combination of ProteoMiner and immunodepletion. All methods were assessed in terms of proteomic depths, protein quantification precision, and functional analysis. Proteonano exhibited the most effective enrichment for the lowest abundance proteins, confidently identifying 299 proteins with mapped blood concentrations below 10⁶ pg/L. Immunodepletion yielded the highest proteome coverage in the moderate abundance range (660 confident proteins). Also, ENRICHplus quantitative profile closely matched that of the neat plasma (93% correlation). Additionally, high repeatability (median coefficient of variation) was demonstrated by MagNet-SAX (13%), ProteoMiner (15%), and immunodepletion (16%). Combining ProteoMiner and immunodepletion reduced the plasma protein dynamic range, enabling deeper low abundance protein analysis but decreased repeatability. These results obtained on platelet-free plasma deviate from previously reported results on platelet-rich plasma, highlighting the crucial sample preparation stage for plasma proteomics.

Single-pot, solid-phase-enhanced sample preparation (SP3) is a rapid, automatable, and cost-effective protein cleanup technique facilitated by the aggregation and digestion of proteins on paramagnetic beads. In this study, we document how plasma protein-to-bead ratio influences the performance in terms of the number of identified peptides and proteins for two chemically modified paramagnetic SP3 beads, i.e., Carboxylate-Speedbeads and MagReSyn-Hydroxyl beads. The optimized protein-to-bead ratio enabled the identification of 17% more plasma protein groups than the nonoptimized condition. Furthermore, we evidenced differences in the quantification results of peptides upon aggregation on hydroxyl- and carboxylate-modified beads. By assessing the physicochemical properties of these peptides, significant differences were revealed in their pI values, charge, polarity, and aspartic and glutamic acid composition. Our results highlight that the choice of beads and protein-to-bead ratio are two important parameters that require optimization depending on the physicochemical properties of the targeted proteins.

The vast majority of marine microorganisms and their functions are yet to be explored. The considerable diversity they encompass is an endless source of knowledge and wealth that can be valued on an industrial scale, emphasizing the need to develop rapid and efficient identification and characterization techniques. In this study, we identified 26 microbial isolates from coastal water of the NW Mediterranean Sea, using phylopeptidomics, a cutting-edge tandem mass spectrometry proteotyping technique. Taxonomical identification at the species level was successfully conducted for all isolates. The presence of strains belonging to the newly described Balneolaeota phylum, yet uncharacterized at the proteomics scale, was noted. The very first proteomics-based investigation of a representative of the Balneolaeota phylum, Balneola vulgaris, is proposed, demonstrating the use of our proteotyping workflow for the rapid identification and in-depth molecular characterization, in a single MS/MS analytical run. Tandem mass spectrometry proteotyping is a valuable asset for culturomic programs as the methodology is able to quickly classify the most atypical isolates.