Caracterización del sistema de secreción de tipo VI en Rhizobium etli Mim1

La simbiosis rizobio-leguminosa es altamente específica. La translocación de proteínas denominadas efectores desde el citoplasma bacteriano a la célula vegetal es un elemento relacionado con dicha especificidad. Los efectores pueden ser translocados a través de diferentes sistemas de secreción. El análisis de genomas de rizobios ha permitido identificar en algunos la presencia de sistemas de secreción de tipo VI (T6SS). El T6SS tiene como componente principal una nanoestructura similar a las que utilizan los bacteriófagos1 para inyectar su ADN y que las bacterias usan para secretar proteínas. Los genes implicados en la formación de T6SS están agrupados y los que codifican para componentes estructurales del sistema presentan mayor grado de conservación entre rizobios y frente a otras bacterias en comparación a los genes que codifican para efectores y reguladores del sistema. En nuestro grupo se está estudiando el T6SS de Rhizobium etli bv mimosae Mim12 aislada de nódulos de Mimosa affinis y capaz de nodular además Phaseolus vulgaris y Leucaena leucocephala. La cepa Mim1 contiene una agrupación de 28 genes en el plásmido f no simbiótico, relacionados con la formación de un T6SS, presentando una organización similar a la descrita en Agrobacterium tumefaciens C583 que consiste en dos operones divergentes. Se ha descrito para varios microorganismos que cuando el T6SS está activo, las proteínas Hcp y VgrG que forman parte del aparato de secreción pueden detectarse en el medio extracelular3. Los genes que codifican proteínas estructurales en las dos bacterias presentan una gran similitud, así Hcp muestra un 94% de identidad entre ambas permitiendo que los anticuerpos que detectan Hcp de Agrobacterium3 también reaccionen con Hcp de Mim1. Utilizando anticuerpos contra Hcp de Agrobacterium se ha identificado esta proteína en el medio extracelular de cultivos de Mim1 en fase estacionaria y débilmente en fase exponencial. También se ha demostrado su presencia en nódulos de judía y en cultivos crecidos en presencia de exudados de L. leucocephala, P. vulgaris y Pisum sativum. Además, con el fin de conocer en qué condiciones se activa el T6SS de Mim1, se analizó una región de ADN presumiblemente promotora comprendida entre las dos agrupaciones de genes orientados de forma divergente de Mim1. Esta región se fusionó transcripcionalmente a un gen b-gal delator sin promotor del vector pMP220 en las dos posibles orientaciones, una de las orientaciones (P1) controlaría la expresión de genes como hcp y posibles efectores y la otra (P2) de otros genes estructurales. Los resultados mostraron que ambas orientaciones se expresaban a altas DO600 (0,8-1) aunque los valores de P1 fueron entre dos y tres veces superiores a los de P2. Sin embargo a bajas DO600 (0,1-0,2) la actividad de P1 ser redujo a la mitad y la de P2 a niveles del control sin promotor. Con el objetivo de conocer el papel del T6SS en simbiosis se han realizado 3 mutantes que afectan a genes estructurales del T6SS de Mim1, uno en el gen hcp, otro en tssM y el tercero es una deleción de todos los genes presumiblemente dependientes de P2. Se examinó el fenotipo producido en P. vulgaris y L. leucocephala y se observó que los tres mutantes produjeron nódulos blancos y plantas con un porte similar a plantas no inoculadas, con menor tamaño que las inoculadas con la cepa parental y con un color más amarillento. En este trabajo se ha mostrado por primera vez que la presencia de un T6SS en rizobios tiene un efecto beneficioso en la simbiosis con varios hospedadores. En estos momentos se esta trabajando en la caracterización de posibles efectores. Referencias. 1. Records AR. 2011 The type VI secretion system: a multipurpose delivery sustem with a phage-like machinery. Mol Plant Microbe Interact 24: 751-757. 2. Rogel MA et al. 2014. Genomic basis of symbiovar mimosae in Rhizobium etli. BMC Genomics 15: 575 3. Wu, HY et al. 2012. Acid-induced type VI secretion system is regulated by ExoR-ChvG/Chv

Characterization of type VI secretion systems (T6SS) of endosymbionts from mimosa or lupine

The T6SS is a nanosyringe that injects proteins into prokaryotic or eukaryotic cells, and it is encoded in the genomes of more than 25% of Gram-negative bacteria (1). We are studying the T6SS of Rhizobium etli Mim1 and Bradyrhizobium sp. LmicA16, symbionts of Phaseolus vulgaris/Leucaena leucocephala and Lupinus micranthus/Lupinus angustifolius/Spartium junceum, respectively. R. etli Mim1 contains a T6SS gene cluster organized in two divergent operons. When the T6SS is active, Hcp, a constituent of the secretory apparatus, can be detected in the extracellular medium (2). Hcp has been immunologically detected in the supernatant of Mim1 cultures. This protein was also detected in bean nodule extracts and in cultures grown in the presence of different legumes exudates. The putative divergent promoters located between the two T6SS gene clusters were analysed by ?- gal fusions. The results showed high levels of expression of the two promoters at high OD and low values at lower ODs. Mutants affected in structural genes induced white nodules with P. vulgaris and L. leucocephala. On the other hand, mutagenesis of T6SS structural genes from LmicA16 strain produced different symbiotic phenotypes. An LmicA16 tssC mutant showed reduced levels of nitrogen fixation on L. micranthus, whereas the same mutant induced the formation of few white, non-fixing nodules on L. angustifolius and S. junceum. (1) Ho et al. (2013) Cell Host Microbe 15:9-21. (2) Wu et al. (2012) PLoS Pathog. 8:1-18 Funded by grants BIO2013-43040-P (MINECO), CGL2011-26932 (MICINN) and AL16-PID-06 (UPM)

Cannabinoid signaling modulation through JZL184 restores key phenotypes of a mouse model for Williams-Beuren syndrome

Williams-Beuren syndrome (WBS) is a rare genetic multisystemic disorder characterized by mild-to-moderate intellectual disability and hypersocial phenotype, while the most life-threatening features are cardiovascular abnormalities. Nowadays, there are no pharmacological treatments to directly ameliorate the main traits of WBS. The endocannabinoid system (ECS), given its relevance for both cognitive and cardiovascular function, could be a potential druggable target in this syndrome. We analyzed the components of the ECS in the complete deletion (CD) mouse model of WBS and assessed the impact of its pharmacological modulation in key phenotypes relevant for WBS. CD mice showed the characteristic hypersociable phenotype with no preference for social novelty and poor short-term object-recognition performance. Brain cannabinoid type-1 receptor (CB1R) in CD male mice showed alterations in density and coupling with no detectable change in main endocannabinoids. Endocannabinoid signaling modulation with subchronic (10 days) JZL184, a selective inhibitor of monoacylglycerol lipase, specifically normalized the social and cognitive phenotype of CD mice. Notably, JZL184 treatment improved cardiovascular function and restored gene expression patterns in cardiac tissue. These results reveal the modulation of the ECS as a promising novel therapeutic approach to improve key phenotypic alterations in WBS

Clinical and Epidemiological Characteristics of Streptococcus suis Infections in Catalonia, Spain

Introduction: Streptococcus suis (S. suis) is a human zoonotic pathogen of occupational origin, with infection acquired through contact with live pigs or pig meat. Pig farming is one of Catalonia's biggest industries and as a result this region of Spain has one of the highest density pig populations per km2. The aim of our study was to describe the infections caused by S. suis occurring in that area over a 9-year period. Materials and Methods: A retrospective, multi-center study was carried out by searching records from 15 hospitals in Catalonia for the period between 2010 and 2019. Results: Over the study period altogether nine cases of S. suis infection were identified in five hospitals, with five of these cases occurring in the 2018-2019 period. The mean age of patients was 48 ± 8.9 years and all of them were males. Five patients (55.6%) worked in pig farms. The most frequent manifestation of infection was meningitis (5 cases; 55.6%) followed by septic arthritis (3 cases; 33.3%). None of the patients died at 30 days; nonetheless, 4 developed hearing loss as a long-term complication. Conclusion: The most commonly identified S. suis infection was meningitis. Over 50% of the episodes occurred in the last 2 years and have affected pig farm workers. Further surveillance is needed in order to know its prevalence

Clinical and Epidemiological Characteristics of Streptococcus suis Infections in Catalonia, Spain

Streptococcus suis (S. suis) is a human zoonotic pathogen of occupational origin, with infection acquired through contact with live pigs or pig meat. Pig farming is one of Catalonia's biggest industries and as a result this region of Spain has one of the highest density pig populations per km 2. The aim of our study was to describe the infections caused by S. suis occurring in that area over a 9-year period. A retrospective, multi-center study was carried out by searching records from 15 hospitals in Catalonia for the period between 2010 and 2019. Over the study period altogether nine cases of S. suis infection were identified in five hospitals, with five of these cases occurring in the 2018-2019 period. The mean age of patients was 48 ± 8.9 years and all of them were males. Five patients (55.6%) worked in pig farms. The most frequent manifestation of infection was meningitis (5 cases; 55.6%) followed by septic arthritis (3 cases; 33.3%). None of the patients died at 30 days; nonetheless, 4 developed hearing loss as a long-term complication. The most commonly identified S. suis infection was meningitis. Over 50% of the episodes occurred in the last 2 years and have affected pig farm workers. Further surveillance is needed in order to know its prevalence

Endoscopic treatment (endoscopic balloon dilation/self-expandable metal stent) vs surgical resection for the treatment of de novo stenosis in Crohn's disease (ENDOCIR study): an open-label, multicentre, randomized trial.

Background: Stenosis is one of the most common complications in patients with Crohn's disease (CD). Endoscopic balloon dilation (EBD) is the treatment of choice for a short stenosis adjacent to the anastomosis from previous surgery. Self-expandable metal stents (SEMS) may be a suitable treatment option for longer stenoses. To date, however, there is no scientific evidence as to whether endoscopic (EBD/SEMS) or surgical treatment is the best approach for de novo or primary stenoses that are less than 10 cm in length. Methods/design: Exploratory study as "proof-of-concept", multicentre, open-label, randomized trial of the treatment of de novo stenosis in the CD; endoscopic treatment (EBD/SEMS) vs surgical resection (SR). The type of endoscopic treatment will initially be with EDB; if a therapeutic failure occurs, then a SEMS will be placed. We estimate 2 years of recruitment and 1 year of follow-up for the assessment of quality of life, costs, complications, and clinical recurrence. After the end of the study, patients will be followed up for 3 years to re-evaluate the variables over the long term. Forty patients with de novo stenosis in CD will be recruited from 15 hospitals in Spain and will be randomly assigned to the endoscopic or surgical treatment groups. The primary aim will be the evaluation of the patient quality of life at 1 year follow-up (% of patients with an increase of 30 points in the 32-item Inflammatory Bowel Disease Questionnaire (IBDQ-32). The secondary aim will be evaluation of the clinical recurrence rate, complications, and costs of both treatments at 1-year follow-up. Discussion: The ENDOCIR trial has been designed to determine whether an endoscopic or surgical approach is therapeutically superior in the treatment of de novo stenosis in CD

Blood RNA sequencing reveals immunological processes associated with the response to abatacept in rheumatoid arthritis.

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