Examinando por Autor "Pacheco, Nicolas"

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    Exploiting the natural poly(3-hydroxyalkanoates) production capacity of Antarctic Pseudomonas strains: from unique phenotypes to novel biopolymers
    (Springer Verlag, 2019-08-01) Pacheco, Nicolas; Orellana-Saez, Matias; Pepczynska, Marzena; Enrione, Javier; Bassas-Galia, Monica; Borrero-de Acuña, Jose M.; Zacconi, Flavia C.; Marcoleta, Andrés E.; Poblete-Castro, Ignacio
    Extreme environments are a unique source of microorganisms encoding metabolic capacities that remain largely unexplored. In this work, we isolated two Antarctic bacterial strains able to produce poly(3-hydroxyalkanoates) (PHAs), which were classified after 16S rRNA analysis as Pseudomonas sp. MPC5 and MPC6. The MPC6 strain presented nearly the same specific growth rate whether subjected to a temperature of 4 °C 0.18 (1/h) or 30 °C 0.2 (1/h) on glycerol. Both Pseudomonas strains produced high levels of PHAs and exopolysaccharides from glycerol at 4 °C and 30 °C in batch cultures, an attribute that has not been previously described for bacteria of this genus. The MPC5 strain produced the distinctive medium-chain-length-PHA whereas Pseudomonas sp. MPC6 synthesized a novel polyoxoester composed of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate-co-3-hydroxyoctanoate-co-3-hydroxydecanoate-co-3-hydroxydodecanoate). Batch bioreactor production of PHAs in MPC6 resulted in a titer of 2.6 (g/L) and 1.3 (g/L), accumulating 47.3% and 34.5% of the cell dry mass as PHA, at 30 and 4 °C, respectively. This study paves the way for using Antarctic Pseudomonas strains for biosynthesizing novel PHAs from low-cost substrates such as glycerol and the possibility to carry out the bioconversion process for biopolymer synthesis without the need for temperature control.
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    In-depth genomic and phenotypic characterization of the antarctic psychrotolerant strain pseudomonas sp. MPC6 reveals unique metabolic features, plasticity, and biotechnological potential
    (Frontiers Media S.A., 2019-05) Orellana Saez, Matias; Pacheco, Nicolas; Costa, José I.; Mendez, Katterinne N.; Miossec, Matthieu J.; Meneses, Claudio; Castro Nallar, Eduardo; Marcoleta, Andrés E.; Poblete Castro, Ignacio
    We obtained the complete genome sequence of the psychrotolerant extremophile Pseudomonas sp. MPC6, a natural Polyhydroxyalkanoates (PHAs) producing bacterium able to rapidly grow at low temperatures. Genomic and phenotypic analyses allowed us to situate this isolate inside the Pseudomonas fluorescens phylogroup of pseudomonads as well as to reveal its metabolic versatility and plasticity. The isolate possesses the gene machinery for metabolizing a variety of toxic aromatic compounds such as toluene, phenol, chloroaromatics, and TNT. In addition, it can use both C6- and C5-carbon sugars like xylose and arabinose as carbon substrates, an uncommon feature for bacteria of this genus. Furthermore, Pseudomonas sp. MPC6 exhibits a high-copy number of genes encoding for enzymes involved in oxidative and cold-stress response that allows it to cope with high concentrations of heavy metals (As, Cd, Cu) and low temperatures, a finding that was further validated experimentally. We then assessed the growth performance of MPC6 on glycerol using a temperature range from 0 to 45C, the latter temperature corresponding to the limit at which this Antarctic isolate was no longer able to propagate. On the other hand, the MPC6 genome comprised considerably less virulence and drug resistance factors as compared to pathogenic Pseudomonas strains, thus supporting its safety. Unexpectedly, we found five PHA synthases within the genome of MPC6, one of which clustered separately from the other four. This PHA synthase shared only 40% sequence identity at the amino acid level against the only PHA polymerase described for Pseudomonas (63-1 strain) able to produce copolymers of short- and medium-chain length PHAs. Batch cultures for PHA synthesis in Pseudomonas sp. MPC6 using sugars, decanoate, ethylene glycol, and organic acids as carbon substrates result in biopolymers with different monomer compositions. This indicates that the PHA synthases play a critical role in defining not only the final chemical structure of the biosynthesized PHA, but also the employed biosynthetic pathways. Based on the results obtained, we conclude that Pseudomonas sp. MPC6 can be exploited as a bioremediator and biopolymer factory, as well as a model strain to unveil molecular mechanisms behind adaptation to cold and extreme environments. Copyright © 2019 Orellana-Saez, Pacheco, Costa, Mendez, Miossec, Meneses, Castro-Nallar, Marcoleta and Poblete-Castro.
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    The ArcB kinase sensor participates in the phagocyte-mediated stress response in Salmonella Typhimurium
    (Frontiers Media SA, 2025) Pardo-Esté, Coral; Urbina, Francisca; Aviles, Nicolas; Pacheco, Nicolas; Briones, Alan; Cabezas, Carolina; Rojas, Vicente; Pavez, Valentina; Sulbaran-Bracho, Yoelvis; Hidalgo, Alejandro A.; Castro-Severyn, Juan; Saavedra, Claudia P.
    The ArcAB two-component system includes a histidine kinase sensor (ArcB) and a regulator (ArcA) that respond to changes in cell oxygen availability. The ArcA transcription factor activates genes related to metabolism, membrane permeability, and virulence, and its presence is required for pathogenicity in Salmonella Typhimurium, which can be phosphorylated independently of its cognate sensor, ArcB. In this study, we aimed to characterize the transcriptional response to hypochlorous acid (HOCl) mediated by the presence of the ArcB sensor. HOCl is a powerful microbicide widely used for sanitization in industrial settings. We used wild-type S. Typhimurium and the mutant lacking the arcB gene exposed to NaOCl to describe the global transcriptional response. We also infected murine neutrophils to evaluate the expression levels of relevant genes related to the resistance and infection process while facing ROS-related stress. Our results indicate that the absence of the arcB gene significantly affects the ability of S. Typhimurium to grow under HOCl stress. Overall, 6.6% of Salmonella genes varied their expression in the mutant strains, while 8.6% changed in response to NaOCl. The transcriptional response associated with the presence of ArcB is associated with metabolism and virulence, suggesting a critical role in pathogenicity and fitness, especially under ROS-related stress. Our results show that ArcB influences the expression of genes associated with fatty acid degradation, protein secretion, cysteine and H2S biosynthesis, and translation, both in vitro and under conditions found within neutrophils. We found that protein carbonylation is significantly higher in the mutant strain than in the wild type, suggesting a critical function for ArcB in the response and repair processes. This study contributes to the understanding of the pathogenicity and adaptation mechanisms that Salmonella employs to establish a successful infection in its host.