Examinando por Autor "Tuttobene, Marisel Romina"
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Ítem Acceso Abierto Acinetobacter baumannii response to cefiderocol challenge in human urine(Springer Nature, 2022-05-24) Nishimura, Brent; Escalante, Jenny; Tuttobene, Marisel Romina; Subils, Tomás; Mezcord, Vyanka; Pimentel, Camila; Georgeos, Nardin; Pasteran, Fernando; Rodríguez, Cecilia; Sieira, Rodrigo; Actis, Luis A.; Tolmasky, Marcelo E.; Bonomo, Robert A.; Ramírez, María SoledadCefiderocol (CFDC) is a novel chlorocatechol-substituted siderophore antibiotic approved to treat complicated urinary tract infections (cUTI) and hospital-acquired and ventilator-acquired pneumonia (HAP/VAP). Previous work determined that albumin-rich human fluids increase the minimum inhibitory concentration (MICs) of Acinetobacter baumannii against CFDC and reduce the expression of genes related to iron uptake systems. This latter effect may contribute to the need for higher concentrations of CFDC to inhibit growth. The presence of human urine (HU), which contains low albumin concentrations, did not modify MIC values of two carbapenem-resistant A. baumannii. Levels of resistance to CFDC were not modified by HU in strain AMA40 but were reduced in strain AB5075. Expanding the studies to other carbapenem-resistant A. baumannii isolates showed that the presence of HU resulted in unmodified or reduced MIC of CDFC values. The expression of piuA, pirA, bauA, and bfnH determined by qRT-PCR was enhanced in A. baumannii AMA40 and AB5075 by the presence of HU in the culture medium. All four tested genes code for functions related to recognition and transport of ferric-siderophore complexes. The effect of HU on expression of pbp1, pbp3, blaOXA-51-like, blaADC, and blaNDM-1, genes associated with resistance to β-lactams, as well as genes coding for efflux pumps and porins was variable, showing dependence with the strain analyzed. We conclude that the lack of significant concentrations of albumin and free iron in HU makes this fluid behave differently from others we tested. Unlike other albumin rich fluids, the presence of HU does not impact the antibacterial activity of CFDC when tested against A. baumannii.Ítem Acceso Abierto Carbapenem‑resistant Acinetobacter baumannii (CRAB): metabolic adaptation and transcriptional response to human urine (HU)(Nature Research, 2024-08-19) Escalante, Jenny; Hamza, Mase; Nishimura, Brent; Melecio, Meghan; Davies Sala, Carol; Tuttobene, Marisel Romina; Subils, Tomás; Traglia, German M.; Pham, Chloe; Sieira, Rodrigo; Actis, Luis A.; Bonomo, Robert A.; Tolmasky, Marcelo E.; Ramírez, María SoledadCarbapenem-resistant Acinetobacter baumannii (CRAB) is a major human pathogen and a research priority for developing new antimicrobial agents. CRAB is a causative agent of a variety of infections in diferent body sites. One of the manifestations is catheter-associated urinary tract infection, which exposes the bacteria to the host’s urine, creating a particular environment. Exposure of two CRAB clinical isolates, AB5075 and AMA40, to human urine (HU) resulted in the diferential expression levels of 264 and 455 genes, respectively, of which 112 were common to both strains. Genes within this group play roles in metabolic pathways such as phenylacetic acid (PAA) catabolism, the Hut system, the tricarboxylic acid (TCA) cycle, and other processes like quorum sensing and bioflm formation. These results indicate that the presence of HU induces numerous adaptive changes in gene expression of the infecting bacteria. These changes presumably help bacteria establish and thrive in the hostile conditions in the urinary tract. These analyses advance our understanding of CRAB’s metabolic adaptations to human fuids, as well as expand knowledge on bacterial responses to distinct human fuids containing diferent concentrations of human serum albumin (HSA).Ítem Acceso Abierto Cerebrospinal fuid (CSF) augments metabolism and virulence expression factors in Acinetobacter baumannii(Nature Research, 2021-02-26) Martínez, Jasmine; Razo Gutiérrez, Chelsea; Razo Gutiérrez, Chelsea; Courville, Robert; Pimentel, Camila; Liu, Christine; Fung, Sammie E.; Tuttobene, Marisel Romina; Phan, Kimberly; Vila, Alejandro J.; Shahrestani, Parvin; Jimenez, Veronica; Tolmasky, Marcelo E.; Becka, Scott A.; Papp Wallace, Krisztina M.; Bonomo, Robert A.; Soler Bistue, Alfonso; Sieira, Rodrigo; Ramírez, María SoledadIn a recent report by the Centers for Disease Control and Prevention (CDC), multidrug resistant (MDR) Acinetobacter baumannii is a pathogen described as an “urgent threat.” Infection with this bacterium manifests as diferent diseases such as community and nosocomial pneumonia, bloodstream infections, endocarditis, infections of the urinary tract, wound infections, burn infections, skin and soft tissue infections, and meningitis. In particular, nosocomial meningitis, an unwelcome complication of neurosurgery caused by extensivelydrug resistant (XDR) A. baumannii, is extremely challenging to manage. Therefore, understanding how A. baumannii adapts to diferent host environments, such as cerebrospinal fuid (CSF) that may trigger changes in expression of virulence factors that are associated with the successful establishment and progress of this infection is necessary. The present invitro work describes, the genetic changes that occur during A. baumannii infltration into CSF and displays A. baumannii’s expansive versatility to persist in a nutrient limited environment while enhancing several virulence factors to survive and persist. While a hypervirulent A. baumannii strain did not show changes in its transcriptome when incubated in the presence of CSF, a lowvirulence isolate showed signifcant diferences in gene expression and phenotypic traits. Exposure to 4% CSF caused increased expression of virulence factors such as fmbriae, pilins, and iron chelators, and other virulence determinants that was confrmed in various model systems. Furthermore, although CSF’s presence did not enhance bacterial growth, an increase of expression of genes encoding transcription, translation, and the ATP synthesis machinery was observed. This work also explores A. baumannii’s response to an essential component, human serum albumin (HSA), within CSF to trigger the diferential expression of genes associated with its pathoadaptibility in this environment.Ítem Acceso Abierto Hetero-antagonism of avibactam and sulbactam with cefiderocol in carbapenem-resistant Acinetobacter spp(American Society for Microbiology, 2024-08-20) Wong, Olivia; Mezcord, Vyanka; Lopez, Christina; Traglia, German M.; Pasteran, Fernando; Tuttobene, Marisel Romina; Corso, Alejandra; Tolmasky, Marcelo E.; Bonomo, Robert A.; Ramírez, María Soledad; https://orcid.org/0000-0002-4780-5311; https://orcid.org/0000-0001-5840-5869; https://orcid.org/0000-0003-0234-4643; https://orcid.org/0000-0002-6298-7811; https://orcid.org/0000-0002-3299-894X; https://orcid.org/0000-0002-9904-7890Cefiderocol, a siderophore-cephalosporine conjugate antibiotic, shows promise as a therapeutic option for carbapenem-resistant (CR) Acinetobacter infections. While resistance has already been reported in A. baumannii, combination therapies with avibactam or sulbactam reduce MICs of cefiderocol, extending its efficacy. However, careful consideration is necessary when using these combinations. In our experiments, exposure of A. baumannii and A. lwoffii to cefiderocol and sulbactam or avibactam led to the selection of cefiderocol-resistant strains. Three of those were subjected to whole genome sequencing and transcriptomic analysis. The strains all possessed synonymous and non-synonymous substitutions and short deletions. The most significant mutations affected efflux pumps, transcriptional regulators, and iron homeostasis genes. Transcriptomics showed significant alterations in expression levels of outer membrane proteins, iron homeostasis, and β-lactamases, suggesting adaptive responses to selective pressure. This study underscores the importance of carefully assessing drug synergies, as they may inadvertently foster the selection of resistant variants and complicate the management of CR Acinetobacter infections.Ítem Acceso Abierto Histone-like nucleoid-structuring protein (H-NS) regulatory role in antibiotic resistance in Acinetobacter baumannii(Nature Research, 2021) Rodgers, Deja; Le, Casin; Pimentel, Camila; Tuttobene, Marisel Romina; Subils, Tomás; Escalante, Jenny; Nishimura, Brent; García Véscovi, Eleonora; Sieira, Rodrigo; Bonomo, Robert A.; Tolmasky, Marcelo E.; Ramírez, María SoledadÍtem Acceso Abierto Human serum albumin (HSA) regulates the expression of histone-like nucleoid structure protein (H-NS) in Acinetobacter baumannii(Nature Research, 2022-08-27) Escalante, Jenny; Nishimura, Brent; Tuttobene, Marisel Romina; Subils, Tomás; Pimentel, Camila; Georgeos, Nardin; Sieira, Rodrigo; Bonomo, Robert A.; Tolmasky, Marcelo E.; Ramírez, María SoledadÍtem Acceso Abierto Human Serum Proteins and Susceptibility of Acinetobacter baumannii to Cefiderocol: Role of Iron Transport(MDPI, 2022-03-03) Le, Casin; Pimentel, Camila; Pasteran, Fernando; Tuttobene, Marisel Romina; Subils, Tomás; Escalante, Jenny; Nishimura, Brent; Arriaga, Susana; Carranza, Aimee; Mezcord, Vyanka; Vila, Alejandro J.; Corso, Alejandra; Actis, Luis A.; Tolmasky, Marcelo E.; Bonomo, Robert A.; Ramírez, María SoledadÍtem Acceso Abierto Induced Heteroresistance in Carbapenem-Resistant Acinetobacter baumannii (CRAB) via Exposure to Human Pleural Fluid (HPF) and Its Impact on Cefiderocol Susceptibility(MDPI, 2023-07-21) Mezcord, Vyanka; Escalante, Jenny; Nishimura, Brent; Traglia, German M.; Sharma, Rajnikant; Vallé, Quentin; Tuttobene, Marisel Romina; Subils, Tomás; Marin, Ingrid; Pasteran, Fernando; Actis, Luis A.; Tolmasky, Marcelo E.; Bonomo, Robert A.; Rao, Gauri; Ramírez, María Soledad; https://orcid.org/0000-0003-1896-8450; https://orcid.org/0000-0002-4780-5311; https://orcid.org/0000-0001-5840-5869; https://orcid.org/0000-0001-9644-9088; https://orcid.org/0000-0002-6298-7811; https://orcid.org/0000-0002-9904-7890Infections caused by Carbapenem-resistant Acinetobacter baumannii (CRAB) isolates, such as hospital-acquired pneumonia (HAP), bacteremia, and skin and soft tissue infections, among others, are particularly challenging to treat. Cefiderocol, a chlorocatechol-substituted siderophore antibiotic, was approved by the U.S. Food and Drug Administration (FDA) in 2019 and prescribed for the treatment of CRAB infections. Despite the initial positive treatment outcomes with this antimicrobial, recent studies reported a higher-than-average all-cause mortality rate in patients treated with cefiderocol compared to the best available therapy. The cause(s) behind these outcomes remains unconfirmed. A plausible hypothesis is heteroresistance, a phenotype characterized by the survival of a small proportion of cells in a population that is seemingly isogenic. Recent results have demonstrated that the addition of human fluids to CRAB cultures leads to cefiderocol heteroresistance. Here, we describe the molecular and phenotypic analyses of CRAB heteroresistant bacterial subpopulations to better understand the nature of the less-than-expected successful outcomes after cefiderocol treatment. Isolation of heteroresistant variants of the CRAB strain AMA40 was carried out in cultures supplemented with cefiderocol and human pleural fluid (HPF). Two AMA40 variants, AMA40 IHC1 and IHC2, were resistant to cefiderocol. To identify mutations and gene expression changes associated with cefiderocol heteroresistance, we subjected these variants to whole genome sequencing and global transcriptional analysis. We then assessed the impact of these mutations on the pharmacodynamic activity of cefiderocol via susceptibility testing, EDTA and boronic acid inhibition analysis, biofilm formation, and static time-kill assays. Heteroresistant variants AMA40 IHC1 and AMA40 IHC2 have 53 chromosomal mutations, of which 40 are common to both strains. None of the mutations occurred in genes associated with high affinity iron-uptake systems or β-lactam resistance. However, transcriptional analyses demonstrated significant modifications in levels of expression of genes associated with iron-uptake systems or β-lactam resistance. The blaNDM-1 and blaADC-2, as well as various iron-uptake system genes, were expressed at higher levels than the parental strain. On the other hand, the carO and ompA genes’ expression was reduced. One of the mutations common to both heteroresistant strains was mapped within ppiA, a gene associated with iron homeostasis in other species. Static time-kill assays demonstrated that supplementing cation-adjusted Mueller–Hinton broth with human serum albumin (HAS), the main protein component of HPF, considerably reduced cefiderocol killing activity for all three strains tested. Notably, collateral resistance to amikacin was observed in both variants. We conclude that exposing CRAB to fluids with high HSA concentrations facilitates the rise of heteroresistance associated with point mutations and transcriptional upregulation of genes coding for β-lactamases and biofilm formation. The findings from this study hold significant implications for understanding the emergence of CRAB resistance mechanisms against cefiderocol treatment. This understanding is vital for the development of treatment guidelines that can effectively address the challenges posed by CRAB infections.Ítem Acceso Abierto Interaction of Acinetobacter baumannii with Human Serum Albumin: Does the host hetermine the outcome?(MDPI, 2021-07-08) Pimentel, Camila; Le, Casin; Tuttobene, Marisel Romina; Subils, Tomás; Bonomo, Robert A.; Tolmasky, Marcelo E.; Ramírez, María SoledadÍtem Desconocido Involvement of the histone-like nucleoid structuring protein (H-NS) in Acinetobacter baumannii’s natural transformation(MDPI, 2021-08-26) Le, Casin; Pimentel, Camila; Tuttobene, Marisel Romina; Subils, Tomás; Escalante, Jenny; Nishimura, Brent; Arriaga, Susana; Rodgers, Deja; Bonomo, Robert A.; Sieira, Rodrigo; Tolmasky, Marcelo E.; Ramírez, María Soledad; https://orcid.org/0000-0003-0234-4643; https://orcid.org/0000-0002-4495-563X; https://orcid.org/0000-0002-6298-7811; https://orcid.org/0000-0002-9904-7890Most Acinetobacter baumannii strains are naturally competent. Although some information is available about factors that enhance or reduce the frequency of the transformation of this bacterium, the regulatory elements and mechanisms are barely understood. In this article, we describe studies on the role of the histone-like nucleoid structuring protein, H-NS, in the regulation of the expression of genes related to natural competency and the ability to uptake foreign DNA. The expression levels of the natural transformation-related genes pilA, pilT, pilQ, comEA, comEC, comF, and drpA significantly increased in a ∆hns derivative of A. baumannii A118. The complementation of the mutant with a recombinant plasmid harboring hns restored the expression levels of six of these genes (pilT remained expressed at high levels) to those of the wild-type strain. The transformation frequency of the A. baumannii A118 ∆hns strain was significantly higher than that of the wild-type. Similar, albeit not identical, there were consequences when hns was deleted from the hypervirulent A. baumannii AB5075 strain. In the AB5075 complemented strain, the reduction in gene expression in a few cases was not so pronounced that it reached wild-type levels, and the expression of comEA was enhanced further. In conclusion, the expression of all seven transformation-related genes was enhanced after deleting hns in A. baumannii A118 and AB5075, and these modifications were accompanied by an increase in the cells’ transformability. The results highlight a role of H-NS in A. baumannii’s natural competence.Ítem Acceso Abierto Light modulates important pathogenic determinants and virulence in ESKAPE pathogens Acinetobacter baumannii, Pseudomonas aeruginosa, and Staphylococcus aureus(American Society for Microbiology, 2021-02-08) Tuttobene, Marisel Romina; Pérez, J. F.; Pavesi, Estefanía S.; Pérez Mora, Bárbara; Biancotti, Daiana; Cribb, Pamela; Altilio, Matías; Müller, Gabriela Leticia; Gramajo, Hugo Cesar; Tamagno, G.; Ramírez, María Soledad; Diacovich, Lautaro; Mussi, María Alejandra; https://orcid.org/0000-0002-9904-7890; https://orcid.org/0000-0002-3339-0100; https://orcid.org/0000-0002-4168-3624; Rabinovich, Gabriel: provide HaCaT cells; Voyich, Jovanka: provide the hla and complemented hla mutantLight sensing has been extensively characterized in the human pathogen Acinetobacter baumannii at environmental temperatures. However, the influence of light on the physiology and pathogenicity of human bacterial pathogens at temperatures found in warm-blooded hosts is still poorly understand. In this work, we show that Staphylococcus aureus, Acinetobacter baumannii, and Pseudomonas aeruginosa (ESKAPE) priority pathogens, which have been recognized by the WHO and the CDC as critical, can also sense and respond to light at temperatures found in human hosts. Most interestingly, in these pathogens, light modulates important pathogenicity determinants as well as virulence in an epithelial infection model, which could have implications in human infections. In fact, we found that alpha-toxin-dependent hemolysis, motility, and growth under iron-deprived conditions are modulated by light in S. aureus. Light also regulates persistence, metabolism, and the ability to kill competitors in some of these microorganisms. Finally, light exerts a profound effect on the virulence of these pathogens in an epithelial infection model, although the response is not the same in the different species; virulence was enhanced by light in A. baumannii and S. aureus, while in A. nosocomialis and P. aeruginosa it was reduced. Neither the BlsA photoreceptor nor the type VI secretion system (T6SS) is involved in virulence modulation by light in A. baumannii. Overall, this fundamental knowledge highlights the potential use of light to control pathogen virulence, either directly or by manipulating the light regulatory switch toward the lowest virulence/persistence configuration. IMPORTANCE: Pathogenic bacteria are microorganisms capable of producing disease. Dangerous bacterial pathogens, such as Staphylococcus aureus, Pseudomonas aeruginosa, and Acinetobacter baumannii, are responsible for serious intrahospital and community infections in humans. Therapeutics is often complicated due to resistance to multiple antibiotics, rendering them ineffective. In this work, we show that these pathogens sense natural light and respond to it by modulating aspects related to their ability to cause disease; in the presence of light, some of them become more aggressive, while others show an opposite response. Overall, we provide new understanding on the behavior of these pathogens, which could contribute to the control of infections caused by them. Since the response is distributed in diverse pathogens, this notion could prove a general concept.Ítem Embargo Modulación por luz en Acinetobacter baumannii y en otros patógenos bacterianos causantes de infecciones hospitalarias(2020) Tuttobene, Marisel Romina; Mussi, María Alejandra; Ramírez, María SoledadLa luz solar es un estímulo ambiental ubicuo para la gran mayoría de los organismos vivos en la Tierra; por lo tanto, es lógico esperar el desarrollo de "mecanismos de sensado de luz" que los lleven a adaptarse con éxito a nichos ecológicos particulares. Aunque estos mecanismos fueron inicialmente reconocidos en los organismos fotosintéticos, en la actualidad sabemos que la percepción de la luz en los organismos quimiotróficos también ocurre. En esta tesis se estudiaron los mecanismos de detección de luz azul en diferentes patógenos bacterianos relevantes para la clínica. Hemos caracterizado extensivamente la regulación por luz a temperaturas moderadas (23/24ºC) en Acinetobacter baumannii, mostrando que la luz ejerce un efecto global en su fisiología. Específicamente demostramos que la luz modula la captación de hierro, los niveles de catalasa, la formación de biofilms, el catabolismo del ácido fenilacético y acetoína, el quórum sensing, la expresión del sistema de secreción tipo VI, entre otros importantes procesos fisiológicos, muchos de los cuales están relacionados con la capacidad de este patógeno oportunista de persistir en entornos intrahospitalarios adversos. Pudimos dilucidar el mecanismo de transducción de la señal mostrando que muchos de estos procesos dependen del fotorreceptor de tipo BLUF, BlsA, el cual es un regulador global capaz de unirse y antagonizar el funcionamiento de diversos reguladores transcripcionales como Fur, el regulador global del metabolismo del hierro, y de AcoN, el represor del catabolismo de acetoína, de manera luz-dependiente. Así, por ejemplo, BlsA se une al represor Fur únicamente en oscuridad a temperaturas moderadas, posiblemente secuentrándolo e impidiendo así su unión al sitio blanco en el operador. De esta manera, se induce la expresión de los genes que codifican para el sideróforo acinetobactina y se ve estimulado el crecimiento sólo en esta condición de oscuridad, en ambos procesos de manera dependiente de BlsA. Por su parte, BlsA interacciona con AcoN sólo en presencia de luz, impidiendo posiblemente de esta manera su unión al operador del cluster de genes que codifican para enzimas del catabolismo de acetoína . Esto es consistente con la inducción de la expresión génica y el crecimiento en acetoína como única fuente de carbono solamente en esta condición de iluminación. En oscuridad, BlsA no interacciona con AcoN y la expresión génica se mantiene en estado basal. La regulación por luz ocurre así a nivel transcripcional, modulando numerosas vías y clusters génicos. De esta forma, se observa un comportamiento dual de BlsA pudiendo interaccionar en ambas condiciones, además de tratarse de un regulador global. Asimismo, mostramos que en otros patógenos como Staphylococcus aureus y Pseudomonas aeruginosa, que han sido reconocidos por la Organización Mundial de la Salud (OMS) y el Centro para el Control y la Prevención de Enfermedades de los Estados Unidos (CDC), junto a A. baumannii, como críticos, detectan y responden a la luz a temperaturas compatibles con los hosedadores de sangre caliente. Encontramos que en estos patógenos la luz modula importantes determinantes de patogenicidad, así como la virulencia frente a un modelo de infección epitelial, lo que podría tener implicaciones en las infecciones humanas. Tambien encontramos que la hemólisis dependiente de α toxina es modulada por la luz en S. aureus. A su vez, la luz modula el crecimiento en condiciones limitantes de hierro y el metabolismo en estos patógenos ESKAPE. La motilidad y la competición frente a C. albicans, son otras de las características que encontramos moduladas por luz a 37°C. Además de los avances en el conocimiento fundamental que plantean estos nuevos hallazgos, destacan el uso potencial de la luz para controlar la virulencia de estos patógenos, ya sea directamente o manipulando los reguladores del sensado de luz hacia el estado de menor virulencia / persistencia posible.Ítem Acceso Abierto Quorum and light signals modulate acetoin/butanediol catabolism in Acinetobacter spp(Frontiers Media, 2019-06-20) Tuttobene, Marisel Romina; Fernández-García, Laura; Blasco, Lucía; Cribb, Pamela; Ambroa, Anton; Müller, Gabriela Leticia; Fernández-Cuenca, Felipe; Bleriot, Inés; Rodríguez, Ramiro Esteban; Barbosa, Beatriz G. V.; Lopez-Rojas, Rafael; Trastoy, Rocío; López, María; Bou, Germán; Tomás, María; Mussi, María AlejandraÍtem Acceso Abierto The iron content of human serum albumin nodulates the susceptibility of Acinetobacter baumannii to Cefiderocol(MDPI, 2023-02-20) Escalante, Jenny; Nishimura, Brent; Tuttobene, Marisel Romina; Subils, Tomás; Mezcord, Vyanka; Actis, Luis A.; Tolmasky, Marcelo E.; Bonomo, Robert A.; Ramírez, María Soledad