Our research group focuses on theanalysisof mechanisms that govern bacterial virulence. Our aim is to elucidate gene expression regulatory networks and the mechanism of action of virulence factors that determine the infective process.
Screening and identification of new antibacterial compounds
The emergence of multi-resistant pathogenic bacteria threatensthe success in the treatment of most infectious illnesses, turning ineffective the antibiotic drugs currently administered. This makes urgent the development of new antibacterial agents directed against alternative bacterial targets. Our laboratory group is dedicated to identify novel compounds that interfere with bacterial pathogenic processes with the final aim of developing anti-virulence drugs that can be used as antimicrobials for therapeutic use.
From the early beginning, our team was dedicated to the analysis of the two-component family of regulatory systems, and focused on the Salmonella typhimuriumPhoP/PhoQ-dependent signal transduction regulatory cascade. This signal transduction system controls the expression of more than 60 genes involved in magnesium homeostasis, resistance to antimicrobial peptides, non-phagocytic cells invasion and survival along the infective cycle within the host. By using biochemical, genetic and molecular biology approaches, we have defined essential aspects of the PhoP/PhoQ-dependent transduction mechanism: signals detection, PhoP/PhoQ specific recognition, phosphotransfer reactions and dynamic cytolocalization of the components of the system. In parallel, we have determined the identity and expression mechanisms of PhoP/PhoQ-dependent genes. With this knowledge, and taking into account that two-component systems are ubiquitous in bacteria but absent in mammalian cells, our group focuses in the identification of new compounds that are able to modulate the PhoP/PhoQ system , with potential pharmacological use.
Gene regulation and bacterial pathogenesis in Serratiamarcescens
S. marcescens is an etiological agent of urinary tract and respiratory infections, endocarditis, osteomyelitis, eye and wound infections, meningitis and sepsis. Serratia shows a high incidence in immunocompromised patients and also in neonatal intensive care units. The analysis of pathogenic mechanisms of opportunistic bacteria such as Serratia, with high impact in under-developed and developing countries, has given priority to the understanding of antibiotic multiple resistances. This information is valuable to adopt strategies to fight infectionsin the short term. However, for most opportunistic bacteria, the analysis of virulence mechanisms and factors involved in the long term pathogen-host interaction (colonization, invasion and dissemination) has been relegated. This knowledge constitutes the basis for the development of alternative therapeutic targets.
Our laboratory has demonstrated that Serratiamarcescens has the capacity not only to promote its internalization in non-phagocytic cells but also to survive and proliferate intracellularly. We showed that the Serratia-containing vacuole deploys a traffic pathway that diverges from the canonical route, avoiding the physiological mechanisms that promote bacterial elimination. After intracellular proliferation, the expression of a bacterial effector allows Serratia to egress from the invaded cell by a non-lytic process, granting its dissemination. In parallel, we have demonstrated that phenotypes such as swimming and swarming motility, adherence to host cells, the expression levels of the cytolysinsShlA and PhlA, and the ability to produce outer membrane vesicles depend on the activation status of the Rcs signal transduction system. These results revealed that the Rcs system is key for the proper temporal and spatial regulation of the Serratia pathogenic traits expression.
In sum, our group has elucidated a repertoire of adaptive regulatory mechanisms and specific effectors deployed by Serratia to survive, proliferate and disseminate outside and inside the host. Because all of these features, Serratiamarcescens constitutes an optimal model to further contribute to the understanding of bacteria-host interaction.
- Lazzaro M, Feldman MF, García Véscovi E. (2017) A Transcriptional Regulatory Mechanism Finely Tunes the Firing of Type VI Secretion System in Response to Bacterial Enemies. MBio. Aug 22;8(4). doi: 10.1128/mBio.00559-17.
- Di Venanzio G, Lazzaro M, Morales ES, Krapf D, García Véscovi E. (2016) A pore-forming toxin enables Serratia a non-lytic egress from host cells. Cell Microbiol. Feb;19(2). doi: 10.1111/cmi.12656.
- Hover T, Maya T, Ron S, Sandovsky H, Shadkchan Y, Kijner N, Mitiagin Y, Fichtman B, Harel A, Shanks RMQ, Bruna RE, García-Véscovi E and Osherov N. (2016) Mechanisms of Bacterial (Serratia marcescens) Attachment to, Migration along, and Killing of Fungal Hyphae. Appl. Environ. Microbiol. 18;82(9):2585-94, doi:10.1128/AEM.04070-15
- Bruna RE, Revale S, García Véscovi E and Mariscotti JF. (2015) Draft Whole-Genome Sequence of Serratia marcescens Strain RM66262, Isolated from a Patient with a Urinary Tract Infection. Genome Announc. Dec 3;3(6). pii: e01423-15. doi: 10.1128/genomeA.01423-15.
- Di Venanzio G, Stepanenko T and García Véscovi, E. (2014) Serratia marcescens ShlA pore-forming toxin is responsible for early induction of autophagy in host cells and is transcriptionally regulated by RcsB. Infection and Immunity, 82 (9):3542-54. doi: 10.1128/IAI.01682-14.
- Salazar; MO; Viarengo, G; Sciara, MI ; Kieffer, PM; García Véscovi, E and Furlán, RL. (2014) A thin-layer chromatography autographic method for the detection of inhibitors of the Salmonella PhoP-PhoQ regulatory system. Phytochem. Anal., 25(2):155-60. doi: 10.1002/pca.2482.
- Viarengo G, Sciara MI, Salazar MO, Kieffer PM, Furlán RL, García Véscovi, E. (2013) Unsaturated Long Chain Free Fatty Acids Are Input Signals of the Salmonella enterica PhoP/PhoQ Regulatory System. Journal of Biological Chemistry, Aug 2;288(31):22346-58. doi: 10.1074/jbc.M113.472829. EDITOR´s CHOICE: A. M. VanHook, Fatty Acids as Environmental Cues? Sci. Signal. 6, ec189 (2013).
- Barchiesi J, Castelli ME, Di Venanzio G, Colombo MI, García Véscovi E. (2012) The PhoP/PhoQ System and Its Role in Serratia marcescens Pathogenesis. Journal of Bacteriology, 194, 2949-61. doi: 10.1128/JB.06820-11.
- Fedrigo, G.V., Campoy, E., Di Venanzio, G., Colombo, M.I. García Véscovi, E. (2011) Serratia marcescens is able to survive and proliferate in autophagic-like vacuoles inside non-phagocytic cells. PLoS ONE, 6(8):e24054. doi: 10.1371/journal.pone.0024054.
- Castelli, M.E. García Véscovi, E. (2011) The Rcs signal transduction pathway is triggered by ECA structure alterations in Serratia marcescens. Journal of Bacteriology, 193, 63-74. doi: 10.1128/JB.00839-10.
- Castelli, M.E.; Sciara, M.I, García Véscovi, E. (2010) Two Component Systems in the Spatial Program of Bacteria. Current Opinion in Microbiology, 13, 210–218. doi: 10.1016/j.mib.2009.12.012.
- Sciara, M.I., Spagnuolo, C., Jares-Erijman, E. García Véscovi, E. (2008) Cytolocalization of the PhoP response regulator in Salmonella enterica: modulation by extracellular Mg2+ and by the SCV environment. Molecular Microbiology, 70, 479–493. doi: 10.1111/j.1365-2958.2008.06427.x.
- Castelli, M.E., Cauerhff, A., Amongero, M., Soncini, F.C., García Véscovi, E. (2003) The H box-harboring domain is key to the function of the Salmonella enterica PhoQ Mg2+-sensor in the recognition of its partner PhoP. Journal of Biological Chemistry, 278, 23579-85. doi: 10.1074/jbc.M303042200.
- Castelli, M. E., García Véscovi, E., and Soncini, F. C. (2000) The phosphatase activity is the target for Mg2+ regulation of the sensor protein PhoQ in Salmonella. Journal of Biological Chemistry, 275, 22948-22954. doi: 10.1074/jbc.M909335199.
- Ricardo L. Furlán, IIDEFAR-CONICET, Área Farmacognosia, Facultad Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario.
- María Isabel Colombo, IHEM-CONICET, Laboratorio de Biología Celular y Molecular; Facultad Ciencias Médicas, Universidad Nacional de Cuyo.
- Darío Krapf, Laboratorio de Cascadas de Señalización Celular, IBR; Facultad Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario.
- Mario F. Feldman, Washington University School of Medicine, St. Louis, MO, United States of America.
- Francisco García-del Portillo, Centro Nacional de Biotecnología, CSIC, Madrid, España.
- Dominique Ferrandon, Institut de BiologieMoléculaire et Cellulaire, CNRS, Strasbourg, France.
- Nir Osherov, School of Medicine, Tel Aviv University, Ramat Aviv, Israel.
- Agencia Nacional de Promoción Científica y Tecnológica (ANPCyT).
- Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET).
- Secretaria de Estado de Ciencia Tecnología e Innovación (SECTEI), Provincia de Santa Fe
- Universidad Nacional de Rosario
García Véscovi, Eleonora
Phone: +54 341 4237070
Office Extension: 657
Laboratory Extension: 622
- Roberto Bruna
- Martina Lazzaro
- Ma. Ayelen Carabajal
- Ma. Victoria Molino
- Carlos Lobertti
Screening and identification of new antibacterial compounds. Natural products are source of structural diversity and complexity, and this potential is amplified by means of chemical transformations. Natural and chemically modified extracts are subjected to a bioautographic assay to determine their effect over the Salmonella enterica PhoP/PhoQ two-component system. From those extracts that exhibit inhibitory action, we proceed to the identification of the compound/s responsible for the effect and the mechanism of action over the system. The compounds are also further used as a platform for the design of new molecules with potential antimicrobial action.
Extra- and intra-cellular life cycle of Serratia marcescens. Microscopical images show (1) Serratia marcescens growing in planktonic culture, (2 and 3) in two sequential post-infection stages, 120 and 350 min p.i., which denote the bacterial capacity to survive and proliferate intracellularly, and (4) a time-lapse where the exocytic process that allows Serratia to egress from the intravacuolar compartment to the external milieu is observed. The confocal laser microscopy images were obtained by Martina Lazzaro and Gisela Di Venanzio