Examinando por Autor "Pizarro-Guajardo, Marjorie"
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Ítem Early career interview: Marjorie Pizarro-Guajardo, Universidad Andrés Bello(Future Medicine Ltd., 2020-03) Pizarro-Guajardo, MarjorieI studied molecular biotechnology as an undergraduate at the University of Chile (Santiago, Chile), inspired by an interest that my high school biology professor, Nelly Parra, helped me to find. In 2011, I started to study the anaerobic pathogen Clostridium difficile, the pathogen responsible for nosocomial diarrhea, and how the spores contribute to the infection process. I felt a high level of passion for my work and after 1 year as Research Assistant I was accepted in the PhD program of Biotechnology at tge University Andr´es Bello (Santiago, Chile), where I focused my work on the development of a strategy to remove C. difficile spores from the host to make a therapy that can improve the clearance of the infection.Ítem Effect of microalgae on intestinal inflammation triggered by soybean meal and bacterial infection in zebrafish(PLoS ONE, 2017-11) Bravo-Tello, Karina; Ehrenfeld, Nicole; Solís, Camila J.; Ulloa, Pilar E.; Hedrera, Manuel; Pizarro-Guajardo, Marjorie; Paredes-Sabja, Daniel; Feijóo, Carmen G.Soybean meal has been used in many commercial diets for farm fish; despite this component inducing intestinal inflammation. On the other hand, microalgae have increasingly been used as dietary supplements in fish feed. Nevertheless, the vast quantity of microalgae species means that many remain under- or unstudied, thus limiting wide scale commercial application. In this work, we evaluated the effects to zebrafish (Danio rerio) of including Tetraselmis sp (Ts); Phaeodactylum tricornutum (Pt); Chlorella sp (Ch); Nannochloropsis oculata (No); or Nannochloropsis gaditana (Ng) as additives in a soybean meal-based diet on intestinal inflammation and survival after Edwardsiella tarda infection. In larvae fed a soybean meal diet supplemented with Ts, Pt, Ch, or Ng, the quantity of neutrophils present in the intestine drastically decreased as compared to larvae fed only the soybean meal diet. Likewise, Ts or Ch supplements in soybean meal or fishmeal increased zebrafish survival by more than 20% after being challenged. In the case of Ts, the observed effect correlated with an increased number of neutrophils present at the infection site. These results suggest that the inclusion of Ts or Ch in fish diets could allow the use of SBM and at the same time improve performance against pathogen. © 2017 Bravo-Tello et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.Ítem Identification of Clostridium difficile Immunoreactive Spore Proteins of the Epidemic Strain R20291(Wiley-VCH Verlag, 2018-09) Pizarro-Guajardo, Marjorie; Ravanal, María Cristina; Paez, Maria Daniela; Callegari, Eduardo; Paredes-Sabja, DanielPurpose: Clostridium difficile infections are the leading cause of diarrhea associated with the use of antibiotics. During infection, C. difficile initiates a sporulation cycle leading to the persistence of C. difficile spores in the host and disease dissemination. The development of vaccine and passive immunization therapies against C. difficile has focused on toxins A and B. In this study, an immunoproteome-based approach to identify immunogenic proteins located on the outer layers of C. difficile spores as potential candidates for the development of immunotherapy and/or diagnostic methods against this devastating infection is used. Experimental design: To identify potential immunogenic proteins on the surface of C. difficile R20291, spore coat/exosporium extracts are separated by 2D electrophoresis (2-DE) and analyzed for reactivity against C. difficile spore-specific goat sera. Finally, the selected spots are in-gel digested with chymotrypsin, peptides generated are separated by nanoUPLC followed by MS/MS using Quad-TOF-MS, corroborated by Ultimate 3000RS-nano-UHPLC coupled to Q-Exactive-Plus-Orbitrap MS. Results: The analysis identify five immunoreactive proteins: spore coat proteins CotE, CotA, and CotCB; exosporium protein CdeC; and a cytosolic methyltransferase. Conclusion: This data provides a list of spore surface protein candidates as antigens for vaccine development against C. difficile infections. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, WeinheimÍtem Induction of a Specific Humoral Immune Response by Nasal Delivery of Bcla2ctd of Clostridioides difficile(MDPI AG, 2020-02) Maia, Ana Raquel; Reyes-Ramírez, Rodrigo; Pizarro-Guajardo, Marjorie; Saggese, Anella; Castro-Córdova, Pablo; Isticato, Rachele; Ricca, Ezio; Paredes-Sabja, Daniel; Baccigalupi, LoredanaClostridioides difficile, formerly known as Clostridium difficile, is a spore-forming bacterium considered as the most common cause of nosocomial infections in developed countries. The spore of C. difficile is involved in the transmission of the pathogen and in its first interaction with the host; therefore, a therapeutic approach able to control C. difficile spores would improve the clearance of the infection. The C-terminal (CTD) end of BclA2, a spore surface protein of C. difficile responsible of the interaction with the host intestinal cells, was selected as a putative mucosal antigen. The BclA2 fragment, BclA2CTD, was purified and used to nasally immunize mice both as a free protein and after adsorption to the spore of Bacillus subtilis, a well-established mucosal delivery vehicle. While the adsorption to spores increased the in vitro stability of BclA2CTD, in vivo both free and spore-adsorbed BclA2CTD were able to induce a similar, specific humoral immune response in a murine model. Although in the experimental conditions utilized the immune response was not protective, the induction of specific IgG indicates that free or spore-bound BclA2CTD could act as a putative mucosal antigen targeting C. difficile spores. © 2020 by the authors. Licensee MDPI, Basel, Switzerland.Ítem Rol de la glicoproteína BclA3 y de la glicosiltransferasa SgtA en el ensamblaje de los apéndices polares y glicosilaciones en las esporas de Clostridioides difficile R20291(Universidad Andrés Bello, 2020) Ortega Lizárraga, César Daniel; Paredes-Sabja, Daniel; Pizarro-Guajardo, Marjorie; Facultad de Ciencias de la VidaClostridioides difficile es un importante patógeno formador de esporas, considerado el mayor agente causante en las enfermedades nosocomiales y de la diarrea asociada a antibióticos. Las esporas de C. difficile son clave para la transmisión, infección y persistencia en el ciclo infeccioso de C. difficile. La cepa de interés epidemiológico C. difficile R20291 presenta en la superficie de sus esporas estructuras filamentosas tipo vellosidades compuestas principalmente por las glicoproteínas tipo colágeno, BclA. En el presente estudio se buscó determinar cómo afectan la ausencia de la glicoproteína BclA3 y de sus glicosilaciones mediadas por SgtA en la frecuencia de los apéndices polares en las esporas y en la presencia de BclA3 en las esporas de C. difficile R20291. En el presente trabajo se identificó que la glicoproteína BclA3 está expuesta en la superficie de la espora y se localiza tanto en el cuerpo de la espora como en los apéndices polares, donde presentaría un rol en la formación de apéndices. Además, la mutación de la glicosiltransferasa SgtA afecta la presencia y/o la conformación de la glicoproteína BclA3 al menos en su dominio carboxilo terminal. Esta glicosiltransferasa aparentemente tiene un rol en el establecimiento de los apéndices al igual que BclA3. Así mismo, la presencia de SgtA afecta la estructura de BclA3. La caracterización de las principales glicoproteínas del exosporium es importante, ya que puede llevar al desarrollo de nuevas terapias basadas en la respuesta inmune para ofrecer herramientas alternativas en la prevención y tratamiento de las infecciones por C. difficile.Ítem Ultrastructural variability of the exosporium layer of Clostridium difficile spores(American Society for Microbiology, 2016-04) Pizarro-Guajardo, Marjorie; Calderón-Romero, Paulina; Castro-Córdova, Pablo; Mora-Uribe, Paola; Paredes-Sabja, DanielThe anaerobic sporeformer Clostridium difficile is the leading cause of nosocomial antibiotic-associated diarrhea in developed and developing countries. The metabolically dormant spore form is considered the transmission, infectious, and persistent morphotype, and the outermost exosporium layer is likely to play a major role in spore-host interactions during the first contact of C. difficile spores with the host and for spore persistence during recurrent episodes of infection. Although some studies on the biology of the exosporium have been conducted (J. Barra-Carrasco et al., J Bacteriol 195:3863–3875, 2013, https://doiorg.recursosbiblioteca.unab.cl/10.1128/JB.00369-13; J. Phetcharaburanin et al., Mol Microbiol 92:1025–1038, 2014, https://doi-org.recursosbiblioteca.unab.cl/10.1111/mmi.12611), there is a lack of information on the ultrastructural variability and stability of this layer. In this work, using transmission electron micrographs, we analyzed the variability of the spore's outermost layers in various strains and found distinctive variability in the ultrastructural morphotype of the exosporium within and between strains. Through transmission electron micrographs, we observed that although this layer was stable during spore purification, it was partially lost after 6 months of storage at room temperature. These observations were confirmed by indirect immunofluorescence microscopy, where a significant decrease in the levels of two exosporium markers, the N-terminal domain of BclA1 and CdeC, was observed. It is also noteworthy that the presence of the exosporium marker CdeC on spores obtained from C. difficile biofilms depended on the biofilm culture conditions and the strain used. Collectively, these results provide information on the heterogeneity and stability of the exosporium surface of C. difficile spores. These findings have direct implications and should be considered in the development of novel methods to diagnose and/or remove C. difficile spores by using exosporium proteins as targets.