Novel Approaches To Fight Streptococcus pneumoniae

Streptococcus pneumoniae affects millions of people worldwide. It is responsible for a wide spectrum of serious illnesses such as pneumonia, meningitis and bacteraemia. The highest rate of pneumococcal disease (and the highest mortality) occurs in young children, as well as in the elderly and the immunocompromised patients. Identification of S. pneumoniae in diagnostic procedures may significantly improve thanks to the descripion of new PCR-derived techniques. Vaccination based on the polysaccharidic capsule, together with benzylpenicillin-derived drugs, constitute the current choices to tackle pneumococcal diseases. However, the wide serotype diversity of S. pneumoniae and the emergence of antibiotic-resistant strains is fostering the development of new methods to fight this microorganism. In this sense, patents documenting the use of novel antibiotics of the fluoroquinolone or tetracycline families have recently been described. Moreover, surface-associated proteins are receiving an increasingly special attention, as they are synthesized by most pneumococcal strains and play an important role in virulence. New patented protein-based vaccines take into consideration these polypeptides. In this article we present the main relevant characteristics of this pathogen and review the most recent methods that have been patented for the prevention, diagnostic and treatment of the pneumococcal diseases

Extensive unfolding of the C-LytA choline-binding module by submicellar concentrations of sodium dodecyl sulphate

We have investigated the stability of the choline-binding module C-LytA against sodium dodecyl sulphate (SDS)-induced unfolding at pH 7.0 and 20 C. A major intermediate with an unfolded N-terminal region accumulates at around 0.75 mM SDS, whereas 2.0 mM SDS was sufficient for a complete unfolding. This might be the first report of a protein being extensively unfolded by submicellar concentrations of SDS, occurring through formation of detergent clusters on the protein surface. All transitions were reversible upon SDS complexation with b-cyclodextrin, allowing the calculation of thermodynamic parameters. A model for the unfolding of C-LytA by SDS is presented and compared to a previous denaturation scheme by guanidine hydrochloride

Inhibition of pneumococcal choline-binding proteins and cell growth by esters of bicyclic amines

Streptococcus pneumoniae is one of the major pathogens worldwide. The use of currently available antibiotics to treat pneumococcal diseases is hampered by increasing resistance levels; also, capsular polysaccharide-based vaccination is of limited efficacy. Therefore, it is desirable to find targets for the development of new antimicrobial drugs specifically designed to fight pneumococcal infections. Choline-binding proteins are a family of polypeptides, found in all S. pneumoniae strains, that take part in important physiologic processes of this bacterium. Among them are several murein hydrolases whose enzymatic activity is usually inhibited by an excess of choline. Using a simple chromatographic procedure, we have identified several choline analogs able to strongly interact with the choline-binding module (C-LytA) of the major autolysin of S. pneumoniae. Two of these compounds (atropine and ipratropium) display a higher binding affinity to C-LytA than choline, and also increase the stability of the protein. CD and fluorescence spectroscopy analyses revealed that the conformational changes of C-LytA upon binding of these alkaloids are different to those induced by choline, suggesting a different mode of binding. In vitro inhibition assays of three pneumococcal, choline-dependent cell wall lytic enzymes also demonstrated a greater inhibitory efficiency of those molecules. Moreover, atropine and ipratropium strongly inhibited in vitro pneumococcal growth, altering cell morphology and reducing cell viability, a very different response than that observed upon addition of an excess of choline. These results may open up the possibility of the development of bicyclic amines as new antimicrobials for use against pneumococcal pathologies

Fraude de ley en las decisiones empresariales de reestructuración empresarial con pérdida de empleo.

En este trabajo se van a analizar los despidos en fraude de ley en las reestructuraciones empresariales. El inestable contexto económico y laboral vivido en los últimos años ha permitido que los empresarios abusen de los despidos colectivos, muchos de ellos sin causa legal, o en fraude de ley, que los justifique. En algunos casos los despidos son para beneficio del empresario y en otros para los trabajadores que, a pesar de la pérdida del empleo, acceden a las prestaciones de desempleo o jubilación de forma indebida o fraudulenta. Para ello se hará una revisión de la normativa actual y un análisis empírico del despido colectivo y por causas objetivasDepartamento de Derecho Mercantil, Derecho del Trabajo e Internacional PrivadoGrado en Derech

Drug Repositioning as a Therapeutic Strategy against Streptococcus pneumoniae: Cell Membrane as Potential Target

A collection of repurposing drugs (Prestwick Chemical Library) containing 1200 compounds was screened to investigate the drugs’ antimicrobial effects against planktonic cultures of the respiratory pathogen Streptococcus pneumoniae. After four discrimination rounds, a set of seven compounds was finally selected, namely (i) clofilium tosylate; (ii) vanoxerine; (iii) mitoxantrone dihydrochloride; (iv) amiodarone hydrochloride; (v) tamoxifen citrate; (vi) terfenadine; and (vii) clomiphene citrate (Z, E). These molecules arrested pneumococcal growth in a liquid medium and induced a decrease in bacterial viability between 90.0% and 99.9% at 25 µM concentration, with minimal inhibitory concentrations (MICs) also in the micromolar range. Moreover, all compounds but mitoxantrone caused a remarkable increase in the permeability of the bacterial membrane and share a common, minimal chemical structure consisting of an aliphatic amine linked to a phenyl moiety via a short carbon/oxygen linker. These results open new possibilities to tackle pneumococcal disease through drug repositioning and provide clues for the design of novel membrane-targeted antimicrobials with a related chemical structure.Depto. de Sanidad AnimalFac. de VeterinariaTRUEMCIN/AEI/ 10.13039/501100011033pu

Rational stabilization of the C-LytA affinity tag by protein engineering

The C-LytA protein constitutes the choline-binding module of the LytA amidase from Streptococcus pneumoniae. Owing to its affinity for choline and analogs, it is regularly used as an affinity tag for the purification of proteins in a single chromatographic step. In an attempt to build a robust variant against thermal denaturation, we have engineered several salt bridges on the protein surface. All the stabilizing mutations were pooled in a single variant, C-LytAm7, which contained seven changes: Y25K, F27K, M33E, N51K, S52K, T85K and T108K. The mutant displays a 7 degrees C thermal stabilization compared with the wild-type form, together with a complete reversibility upon heating and a higher kinetic stability. Moreover, the accumulation of intermediates in the unfolding of C-LytA is virtually abolished for C-LytAm7. The differences in stability become more evident when the proteins are bound to a DEAE-cellulose affinity column, as most of wild-type C-LytA is denatured at approximately 65 degrees C, whereas C-LytAm7 may stand temperatures up to 90 degrees C. Finally, the change in the isoelectric point of C-LytAm7 enhances its solubility at acidic pHs. Therefore, C-LytAm7 behaves as an improved affinity tag and supports the engineering of surface salt bridges as an effective approach for protein stabilization

Evaluación de la calidad de vida en personas con enfermedad inflamatoria intestinal en Teruel

Introducción. El término enfermedad inflamatoria intestinal (EII) incluye una amplia variedad de presentaciones y manifestaciones clínicas cuya característica principal es la inflamación crónica del tubo digestivo en diferentes localizaciones. Actualmente engloba tres entidades: Enfermedad de Crohn (EC), Colitis Ulcerosa (CU) y Colitis Indeterminada (CI). La EII afecta desfavorablemente la percepción de la salud de las personas que las padece y limitan su calidad de vida. Objetivo: evaluar el impacto que tiene la EII en la calidad de vida relacionada con la salud en pacientes diagnosticados de EII mediante el cuestionario CCVEII-9. Material y métodos: se realizó un estudio descriptivo transversal donde se evaluó la calidad de vida en los pacientes diagnosticados de enfermedad inflamatoria intestinal en la Provincia de Teruel. Se registraron variables sociodemográficas y clínicas de cada paciente. La calidad de vida relacionada con la salud se investigó utilizando el cuestionario reducido de la calidad de vida de enfermedad inflamatoria intestinal (CCVEII-9). Resultados: un total de 42 pacientes, 16 (38,1%) con enfermedad de Crohn y 26 con colitis ulcerosa (61,9%) fueron encuestados. La media de edad fue de 47 años. No se apreciaron diferencias significativas en la calidad de vida en función de la enfermedad, sin embargo, tras el análisis bivariado, el sexo, el número de brotes, el tiempo de evolución de la enfermedad y presentar ansiedad se asociaron negativamente con la CVRS. Conclusiones: nuestra población analizada refleja una relativa buena CVRS similar para ambas enfermedades