KatE from the bacterial plant pathogen ralstonia solanacearumis a monofunctional catalase controlled by HrpG that plays a major role in bacterial survival to hydrogen peroxide

Ralstonia solanacearum is the causative agent of bacterial wilt disease on a wide range of plant species. Besides the numerous bacterial activities required for host invasion, those involved in the adaptation to the plant environment are key for the success of infection. R. solanacearum ability to cope with the oxidative burst produced by the plant is likely one of the activities required to grow parasitically. Among the multiple reactive oxygen species (ROS)-scavenging enzymes predicted in the R. solanacearum GMI1000 genome, a single monofunctional catalase (KatE) and two KatG bifunctional catalases were identified. In this work, we show that these catalase activities are active in bacterial protein extracts and demonstrate by gene disruption and mutant complementation that the monofunctional catalase activity is encoded by katE. Different strategies were used to evaluate the role of KatE in bacterial physiology and during the infection process that causes bacterial wilt. We show that the activity of the enzyme is maximal during exponential growth in vitro and this growth-phase regulation occurs at the transcriptional level. Our studies also demonstrate that katE expression is transcriptionally activated by HrpG, a central regulator of R. solanacearum induced upon contact with the plant cells. In addition, we reveal that even though both KatE and KatG catalase activities are induced upon hydrogen peroxide treatment, KatE has a major effect on bacterial survival under oxidative stress conditions and especially in the adaptive response of R. solanacearum to this oxidant. The katE mutant strain also exhibited differences in the structural characteristics of the biofilms developed on an abiotic surface in comparison to wild-type cells, but not in the overall amount of biofilm production. The role of catalase KatE during the interaction with its host plant tomato is also studied, revealing that disruption of this gene has no effect on R. solanacearum virulence or bacterial growth in leave tissues, which suggests a minor role for this catalase in bacterial fitness in planta. Our work provides the first characterization of the R. solanacearum catalases and identifies KatE as a bona fide monofunctional catalase with an important role in bacterial protection against oxidative stress

Light modulates important physiological features of Ralstonia pseudosolanacearum during the colonization of tomato plants

Ralstonia pseudosolanacearum GMI1000 (Rpso GMI1000) is a soil-borne vascular phytopathogen that infects host plants through the root system causing wilting disease in a wide range of agro-economic interest crops, producing economical losses. Several features contribute to the full bacterial virulence. In this work we study the participation of light, an important environmental factor, in the regulation of the physiological attributes and infectivity of Rpso GMI1000. In silico analysis of the Rpso genome revealed the presence of a Rsp0254 gene, which encodes a putative blue light LOV-type photoreceptor. We constructed a mutant strain of Rpso lacking the LOV protein and found that the loss of this protein and light, influenced characteristics involved in the pathogenicity process such as motility, adhesion and the biofilms development, which allows the successful host plant colonization, rendering bacterial wilt. This protein could be involved in the adaptive responses to environmental changes. We demonstrated that light sensing and the LOV protein, would be used as a location signal in the host plant, to regulate the expression of several virulence factors, in a time and tissue dependent way. Consequently, bacteria could use an external signal and Rpsolov gene to know their location within plant tissue during the colonization process.Fil: Tano, María Josefina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Ripa, Maria Belén. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Tondo, Maria Laura. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; ArgentinaFil: Carrau, Analía. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Petrocelli, Silvana. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; ArgentinaFil: Rodriguez, María Victoria. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; ArgentinaFil: Ferreira, Virginia. Universidad de la República. Facultad de Química; UruguayFil: Siri, María Inés. Universidad de la República. Facultad de Química; UruguayFil: Piskulic, Laura. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas; ArgentinaFil: Orellano, Elena Graciela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; Argentin

Identification of pesticide-degrading Pseudomonas strains as poly-β-hydroxybutyrate producers

Polyhydroxybutyrate (PHB) is an ecological promising substitute for polypropylene because it is biocompatible, biodegradable and can be produced by renewable sources. This study investigated PHB accumulation on pesticide-degrading Pseudomonas. 14 strains of pesticide-degrading Pseudomonas isolated from subtropical lowland soil in southern Brazil were analyzed using optical microscopy. Lipid inclusions were identified in four strains, and three of them, which degraded the pesticide carbofuran, had extensive granules accumulation which was detected by transmission electron microscopy. These strains were cultivated in a shaker at 28°C and the polymer was characterized by Fourier transform infrared (FTIR) spectroscopy. Strain CMM43 had the best accumulation after 48 h. The biopolymer was identified as poly-β-hydroxybutyrate.Key words: Polyhydroxybutyrate (PHB), pesticide-degrading Pseudomonas sp., microscopy, subtropical lowland soil

The LOV Protein of Xanthomonas citri subsp. citri Plays a Significant Role in the Counteraction of Plant Immune Responses during Citrus Canker

Pathogens interaction with a host plant starts a set of immune responses that result in complex changes in gene expression and plant physiology. Light is an important modulator of plant defense response and recent studies have evidenced the novel influence of this environmental stimulus in the virulence of several bacterial pathogens. Xanthomonas citri subsp. citri is the bacterium responsible for citrus canker disease, which affects most citrus cultivars. The ability of this bacterium to colonize host plants is influenced by bacterial blue-light sensing through a LOV-domain protein and disease symptoms are considerably altered upon deletion of this protein. In this work we aimed to unravel the role of this photoreceptor during the bacterial counteraction of plant immune responses leading to citrus canker development. We performed a transcriptomic analysis in Citrus sinensis leaves inoculated with the wild type X. citri subsp. citri and with a mutant strain lacking the LOV protein by a cDNA microarray and evaluated the differentially regulated genes corresponding to specific biological processes. A down-regulation of photosynthesis-related genes (together with a corresponding decrease in photosynthesis rates) was observed upon bacterial infection, this effect being more pronounced in plants infected with the lov-mutant bacterial strain. Infection with this strain was also accompanied with the up-regulation of several secondary metabolism- and defense response-related genes. Moreover, we found that relevant plant physiological alterations triggered by pathogen attack such as cell wall fortification and tissue disruption were amplified during the lov-mutant strain infection. These results suggest the participation of the LOV-domain protein from X. citri subsp. citri in the bacterial counteraction of host plant defense response, contributing in this way to disease development.Fil: Kraiselburd, Ivana. Consejo Nacional de Invest.cientif.y Tecnicas. Centro Cientifico Tecnol.conicet - Rosario. Instituto de Biologia Molecular y Celular de Rosario;Fil: Daurelio, Lucas Damian. Consejo Nacional de Invest.cientif.y Tecnicas. Centro Cientifico Tecnol.conicet - Rosario. Instituto de Biologia Molecular y Celular de Rosario;Fil: Tondo, Maria Laura. Consejo Nacional de Invest.cientif.y Tecnicas. Centro Cientifico Tecnol.conicet - Rosario. Instituto de Biologia Molecular y Celular de Rosario;Fil: Merelo, Paz. INSTITUT VALENCIÀ D'INVESTIGACIONS AGRÀRIES (IVIA);Fil: Cortadi, Adriana Amalia. Consejo Nacional de Invest.cientif.y Tecnicas. Centro Cientifico Tecnol.conicet - Rosario. Instituto de Biologia Molecular y Celular de Rosario;Fil: Talón, Manuel. INSTITUT VALENCIÀ D'INVESTIGACIONS AGRÀRIES (IVIA);Fil: Tadeo, Francisco R.. INSTITUT VALENCIÀ D'INVESTIGACIONS AGRÀRIES (IVIA);Fil: Orellano, Elena Graciela. Consejo Nacional de Invest.cientif.y Tecnicas. Centro Cientifico Tecnol.conicet - Rosario. Instituto de Biologia Molecular y Celular de Rosario

Micro-Maravillas: Introducción a los microorganismos

Realizando talleres experimentales en escuelas de nuestra ciudad vimos que en las aulas existía la oportunidad de acercar el conocimiento científico de manera lúdica y didáctica. Fue por eso que nos propusimos diseñar cuadernillos que faciliten el proceso de aprendizaje de las Ciencias Naturales en los niveles inicial y primario. Como parte de este trayecto decidimos realizar un abordaje interdisciplinario incluyendo cuentos de autoría propia que puedan ser utilizados como disparador de debates y reflexiones sobre la temática en estudio. Así surge nuestra primera serie llamada Micro-Maravillas que invita a descubrir algunos aspectos del fascinante mundo de la Microbiología.Fil: Di Capua, Cecilia Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas; ArgentinaFil: Bortolotti, Ana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas; ArgentinaFil: Campero, María Belén. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Nacional de Rosario. Facultad de Humanidades y Artes; ArgentinaFil: Tondo, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas; ArgentinaFil: Ceccoli, Romina Denis. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas; ArgentinaFil: Balaban, Cecilia Lucía. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas; Argentin

Analysis of sulfur compounds present in Natural Gas, Liquefied Petroleum Gas and biogas using Gas Chromatography with Pulsed Flame Photometric Detector (PFPD)

Los compuestos volátiles azufrados, presentes en el gas natural y en el biogas, entre otros, son altamente corrosivos y ocasionan serios problemas en distintos equipos empleados en su procesamiento y producción. La adecuada determinación de dichos compuestos presentes en matrices que contienen grandes cantidades de hidrocarburos es un desafío, especialmente cuando se deben determinar concentraciones a niveles de ppm. La cromatografía gaseosa con detectores específicos para azufre es la técnica mayormente elegida para su cuantificación. En el presente trabajo se optimizó un método que permite determinar compuestos volátiles de azufre utilizando un Cromatógrafo Gaseoso con Detector Fotométrico de Llama Pulsada, PFPD de sus siglas en inglés. Las temperaturas, flujos, condiciones y parámetros del detector fueron fijados con el objetivo de determinar diferentes compuestos azufrados presentes en tres matrices: Gas Natural, Gas Licuado de Petróleo (GLP) y biogas.Volatile sulfur compounds, present in natural gas and biogas among others, are highly corrosive corrosive and can cause severe damage in the equipment employed in their processing and production. It is a challenge to analyze these compounds adequately in the presence of large amounts of hydrocarbons, specially when concentration of sulfur in ppm level must be determined. Gas Chromatography is an analytical technique widely chosen for this purpose, employing specific sulfur detectors. In this work a method for the determination of volatile sulfur compounds was optimized using gas chromatography with a Pulsed Flame Photometric Detector (PFPD). Temperature, flows, conditions, and detector parameters were set, in order to determine different sulfur compounds present in three hydrocarbon samples: Natural Gas, Liquified Petroleum Gas and Biogas.Fil: Herrero, Maria Sol. Pontificia Universidad Católica Argentina "Santa María de los Buenos Aires". Facultad de Química e Ingeniería-Rosario; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; ArgentinaFil: Hamann, Diana Elizabeth. Pontificia Universidad Católica Argentina "Santa María de los Buenos Aires". Facultad de Química e Ingeniería-Rosario; ArgentinaFil: Olivera, Camila. Pontificia Universidad Católica Argentina "Santa María de los Buenos Aires". Facultad de Química e Ingeniería-Rosario; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; ArgentinaFil: Girardi, Valentina. Pontificia Universidad Católica Argentina "Santa María de los Buenos Aires". Facultad de Química e Ingeniería-Rosario; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; ArgentinaFil: Tondo, Maria Laura. Pontificia Universidad Católica Argentina "Santa María de los Buenos Aires". Facultad de Química e Ingeniería-Rosario; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; ArgentinaFil: Perez, Leonardo Martin. Pontificia Universidad Católica Argentina "Santa María de los Buenos Aires". Facultad de Química e Ingeniería-Rosario; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; ArgentinaFil: Salvatierra, Lucas Matías. Pontificia Universidad Católica Argentina "Santa María de los Buenos Aires". Facultad de Química e Ingeniería-Rosario; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentin

Protein O-linked glycosylation in the plant pathogen Ralstonia solanacearum

Ralstonia solanacearum is one of the most lethal phytopathogens in the world. Due to its broad host range, it can cause wilting disease in many plant species of economic interest. In this work, we identified the O-oligosaccharyltransferase (O-OTase) responsible for protein O-glycosylation in R. solanacearum. An analysis of the glycoproteome revealed that 20 proteins, including type IV pilins are substrates of this general glycosylation system. Although multiple glycan forms were identified, the majority of the glycopeptides were modified with a pentasaccharide composed of HexNAc-(Pen)-dHex3, similar to the O antigen subunit present in the lipopolysaccharide of multiple R. solanacearum strains. Disruption of the O-OTase led to the total loss of protein glycosylation, together with a defect in biofilm formation and reduced pathogenicity towards tomato plants. Comparative proteomic analysis revealed that the loss of glycosylation is not associated with widespread proteome changes. Only the levels of a single glycoprotein, the type IV pilin, were diminished in the absence of glycosylation. In parallel, disruption of glycosylation triggered an increase in the levels of a surface lectin homologous to Pseudomonas PA-IIL. These results reveal the important role of glycosylation in the pathogenesis of R. solanacearum.Fil: Elhenawy, Wael. University of Alberta; CanadáFil: Scott, Nichollas E.. University of British Columbia; CanadáFil: Tondo, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Orellano, Elena Graciela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Foster, Leonard J.. University of British Columbia; CanadáFil: Feldman, Mario F.. University of Alberta; Canadá. University of Washington; Estados Unido

Microbial characterization of a facultative residual sludge obtained from a biogas plant with ability to degrade commercial B10 diesel oil

Biodegradation of commercial B10 diesel oil (DB10) by the microbial consortium present in a biogas residual sludge was first investigated. Maximum removal efficiencies of the petrochemical and fatty acids methyl ester (FAME) fractions of DB10 were 55 ± 3% and 94 ± 6%, respectively. Taxonomic profiling of the microbial consortium present in the residual sludge was carried out in order to identify potential hydrocarbon-degrading bacteria. After mild selection pressure using DB10, three Pseudomonas species (P. aeruginosa, P. stutzeri and P. mendocina) were mostly found in the treated sample. However, only P. aeruginosa and P. stutzeri showed a rapid growth and high dehydrogenase activity at the expense of DB10 as the sole carbon source. In addition, both strains removed 42 ± 5% and 53 ± 2% of the petrochemical fraction, and >75% of the FAME fraction, respectively. Similar results were found with a bacterial consortium of the three isolated Pseudomonas, indicating no synergistic hydrocarbon degradation by these strains. As expected, biosurfactant production by the pseudomonads was directly associated with the bacterial DB10 biodegradation performance. These results are the first describing petroleum-based hydrocarbon biodegradation ability of a crude facultative residual sludge obtained from a biogas facility; and represent a rational first step in order to understand which bacteria in the sludge may act in petroleum-based fuels degradation.Fil: Loureiro, Dana Belen. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Católica de Rosario; Argentina. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas; ArgentinaFil: Olivera, Camila. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Católica de Rosario; ArgentinaFil: Tondo, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas; Argentina. Universidad Católica de Rosario; ArgentinaFil: Herrero, María Sol. Universidad Católica de Rosario; ArgentinaFil: Salvatierra, Lucas Matías. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Católica de Rosario; ArgentinaFil: Perez, Leonardo Martin. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas; Argentina. Universidad Católica de Rosario; Argentin

Dynamics of the active site architecture in plant-type ferredoxin-NADP + reductases catalytic complexes

Kinetic isotope effects in reactions involving hydride transfer and their temperature dependence are powerful tools to explore dynamics of enzyme catalytic sites. In plant-type ferredoxin-NADP+ reductases the FAD cofactor exchanges a hydride with the NADP(H) coenzyme. Rates for these processes are considerably faster for the plastidic members (FNR) of the family than for those belonging to the bacterial class (FPR). Hydride transfer (HT) and deuteride transfer (DT) rates for the NADP+ coenzyme reduction of four plant-type FNRs (two representatives of the plastidic type FNRs and the other two from the bacterial class), and their temperature dependences are here examined applying a full tunnelling model with coupled environmental fluctuations. Parameters for the two plastidic FNRs confirm a tunnelling reaction with active dynamics contributions, but isotope effects on Arrhenius factors indicate a larger contribution for donor–acceptor distance (DAD) dynamics in the Pisum sativum FNR reaction than in the Anabaena FNR reaction. On the other hand, parameters for bacterial FPRs are consistent with passive environmental reorganisation movements dominating the HT coordinate and no contribution of DAD sampling or gating fluctuations. This indicates that active sites of FPRs are more organised and rigid than those of FNRs. These differences must be due to adaptation of the active sites and catalytic mechanisms to fulfil their particular metabolic roles, establishing a compromise between protein flexibility and functional optimisation. Analysis of site-directed mutants in plastidic enzymes additionally indicates the requirement of a minimal optimal architecture in the catalytic complex to provide a favourable gating contribution.Fil: Sanchez Azqueta, Ana. Universidad de Zaragoza; España. Consejo Superior de Investigaciones Científicas; EspañaFil: Catalano Dupuy, Daniela Luján. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Lopez Rivero, Arleth Susana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Tondo, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Orellano, Elena Graciela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Ceccarelli, Eduardo Augusto. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Medina, Milagros. Universidad de Zaragoza; España. Consejo Superior de Investigaciones Científicas; Españ