Analysis of the immunological and functional features of the Neisserial Heparin Binding Antigen (NHBA)

Neisserial Heparin Binding Antigen (NHBA) is a surface-exposed lipoprotein ubiquitously expressed by genetically diverse Neisseria meningitidis strains and is an antigen of the multicomponent protein-based 4CMenB vaccine, able to induce bactericidal antibodies in humans and to bind heparin-like molecules. The aim of this study is to characterize the immunological and functional properties of NHBA. To evaluate immunogenicity and the contribution of aminoacid sequence variability to vaccine coverage, we constructed recombinant isogenic strains that are susceptible to bactericidal killing only by anti-NHBA antibodies and engineered them to express equal levels of selected NHBA peptides. In these recombinant strains, we observed different titres associated with the different peptide variants. These recombinant strains were then further engineered to express NHBA chimeric proteins to investigate the regions important for immunogenicity. In natural strains, anti-NHBA antibodies were found to be cross-protective against strains expressing different peptides. To investigate the functional properties of this antigen, the recombinant purified NHBA protein was tested in in vitro binding studies and was found to be able to bind epithelial cells. The binding was abolished when cells were treated specifically with heparinase III, suggesting that the interaction with the cells is mediated by heparan sulfate proteoglycans (HSPG). Mutation of the Arg-rich tract of NHBA abrogated the binding, confirming the importance of this region in mediating the binding to heparin-like molecules. In a panel of N. meningitidis strains, the deletion of nhba resulted in a reduction of adhesion with respect to each isogenic wild type strain. Furthermore, the adhesion of the wild-type strain was prevented by using anti-NHBA polyclonal sera, demonstrating the specificity of the interaction. These results suggest that NHBA could be a novel meningococcal adhesin contributing to host-cell interaction. Moreover, we analysed NHBA NalP-mediated cleavage in different NHBA peptides and showed that not all NHBA peptides are cleaved.

Adsorption of Components of the Plasma Kinin-forming System on the Surface of Porphyromonas gingivalis Involves Gingipains as the Major Docking Platforms

Enhanced production of proinflammatory bradykinin-related peptides, the kinins, has been suggested to contribute to the pathogenesis of periodontitis, a common inflammatory disease of human gingival tissues. In this report, we describe a plausible mechanism of activation of the kinin-generating system, also known as the contact system or kininogen-kallikrein-kinin system, by the adsorption of its plasma-derived components such as high-molecular-mass kininogen (HK), prekallikrein (PK), and Hageman factor (FXII) to the cell surface of periodontal pathogen Porphyromonas gingivalis. The adsorption characteristics of mutant strains deficient in selected proteins of the cell envelope suggested that the surface-associated cysteine proteinases, gingipains, bearing hemagglutinin/adhesin domains (RgpA and Kgp) serve as the major platforms for HK and FXII adhesion. These interactions were confirmed by direct binding tests using microplate-immobilized gingipains and biotinylated contact factors. Other bacterial cell surface components such as fimbriae and lipopolysaccharide were also found to contribute to the binding of contact factors, particularly PK. Analysis of kinin release in plasma upon contact with P. gingivalis showed that the bacterial surface-dependent mechanism is complementary to the previously described kinin generation system dependent on HK and PK proteolytic activation by the gingipains. We also found that several P. gingivalis clinical isolates differed in the relative significance of these two mechanisms of kinin production. Taken together, these data show the importance of this specific type of bacterial surface-host homeostatic system interaction in periodontal infections.

Physiologic cold shock of Moraxella catarrhalis affects the expression of genes involved in the iron acquisition, serum resistance and immune evasion

Background Moraxella catarrhalis, a major nasopharyngeal pathogen of the human respiratory tract, is exposed to rapid downshifts of environmental temperature when humans breathe cold air. It was previously shown that the prevalence of pharyngeal colonization and respiratory tract infections caused by M. catarrhalis are greatest in winter. The aim of this study was to investigate how M. catarrhalis uses the physiologic exposure to cold air to upregulate pivotal survival systems in the pharynx that may contribute to M. catarrhalis virulence. Results A 26°C cold shock induces the expression of genes involved in transferrin and lactoferrin acquisition, and enhances binding of these proteins on the surface of M. catarrhalis. Exposure of M. catarrhalis to 26°C upregulates the expression of UspA2, a major outer membrane protein involved in serum resistance, leading to improved binding of vitronectin which neutralizes the lethal effect of human complement. In contrast, cold shock decreases the expression of Hemagglutinin, a major adhesin, which mediates B cell response, and reduces immunoglobulin D-binding on the surface of M. catarrhalis. Conclusion Cold shock of M. catarrhalis induces the expression of genes involved in iron acquisition, serum resistance and immune evasion. Thus, cold shock at a physiologically relevant temperature of 26°C induces in M. catarrhalis a complex of adaptive mechanisms that enables the bacterium to target their host cellular receptors or soluble effectors and may contribute to enhanced growth, colonization and virulence.

Outer membrane protein UspA1 and lipooligosaccharide are involved in invasion of human epithelial cells by Moraxella catarrhalis

Invasion of non-professional phagocytes is a strategy employed by several mucosal pathogens, but has not been investigated in detail for Moraxella catarrhalis, a major cause of human respiratory tract infections. We investigated the role of outer membrane protein (OMP) UspA1 and lipooligosaccharide (LOS) in M. catarrhalis invasion into epithelial cells. An isogenic mutant of strain O35E, which lacked expression of the UspA1 adhesin, demonstrated not only severely impaired adherence (86%) to but also reduced invasion (77%) into Chang conjunctival cells in comparison with the wild-type strain. The isogenic, LOS-deficient mutant strain O35E.lpxA was attenuated in adherence (93%) and its capacity to invade was severely reduced (95%), but not abolished. Inhibition assays using sucrose and cytochalasin D, respectively, demonstrated that clathrin and actin polymerization contribute to internalization of M. catarrhalis by Chang cells. Furthermore, inhibition of UspA1-mediated binding to cell-associated fibronectin and alpha5beta1 integrin decreased invasion of M. catarrhalis strain O35E (72% and 41%, respectively). These data indicate that OMP UspA1 and LOS profoundly affect the capacity of M. catarrhalis to invade epithelial cells.

Physiologic cold shock increases adherence of Moraxella catarrhalis to and secretion of interleukin 8 in human upper respiratory tract epithelial cells

Moraxella catarrhalis, a major nasopharyngeal pathogen of the human respiratory tract, is exposed to rapid and prolonged downshifts of environmental temperature when humans breathe cold air. In the present study, we show that a 26 degrees C cold shock up-regulates the expression of UspA1, a major adhesin and putative virulence factor of M. catarrhalis, by prolonging messenger RNA half-life. Cold shock promotes M. catarrhalis adherence to upper respiratory tract cells via enhanced binding to fibronectin, an extracellular matrix component that mediates bacterial attachment. Exposure of M. catarrhalis to 26 degrees C increases the outer membrane protein-mediated release of the proinflammatory cytokine interleukin 8 in pharyngeal epithelial cells. Furthermore, cold shock at 26 degrees C enhances the binding of salivary immunoglobulin A on the surface of M. catarrhalis. These data indicate that cold shock at a physiologically relevant temperature of 26 degrees C affects the nasopharyngeal host-pathogen interaction and may contribute to M. catarrhalis virulence.

Host-pathogen interactions of secreted and surface Staphylococcus aureus factors

Staphylococcus aureus is an opportunistic bacterial pathogen that can infect humans and other species. It utilizes an arsenal of virulence factors to cause disease, including secreted and cell wall anchored factors. Secreted toxins attack host cells, and pore-forming toxins destroy target cells by causing cell lysis. S. aureus uses cell-surface adhesins to attach to host molecules thereby facilitating host colonization. The Microbial Surface Components Recognizing Adhesive Matrix Molecules (MSCRAMMs) are a family of cell-wall anchored proteins that target molecules like fibronectin and fibrinogen. The Serine-aspartate repeat (Sdr) proteins are a subset of staphylococcal MSCRAMMs that share similar domain organization. Interestingly, the amino-terminus, is composed of three immunoglobulin-folded subdomains (N1, N2, and N3) that contain ligand-binding activity. Clumping factors A and B (ClfA and ClfB) and SdrG are Sdr proteins that bind to fibrinogen (Fg), a large, plasma glycoprotein that is activated during the clotting cascade to form fibrin. In addition to recognizing fibrinogen, ClfA and ClfB can bind to other host ligands. Analysis of S. aureus strains that cause osteomyelitis led to the discovery of the bone-sialoprotein-binding protein (Bbp), an Sdr protein. Because several MSCRAMMs target more than one molecule, I hypothesized that Bbp may recognize other host proteins. A ligand screen revealed that the recombinant construct BbpN2N3 specifically recognizes human Fg. Surface plasmon resonance was used to determine the affinity of BbpN2N3 for Fg, and a dissociation constant of 540 nM was determined. Binding experiments performed with recombinant Fg chains were used to map the binding of BbpN2N3 to the Fg Aalpha chain. Additionally, Bbp expressed on the surface of Lactococcus lactis and S. aureus Newman bald mediated attachment of these bacteria to Fg Aalpha. To further characterize the interaction between the two proteins, isothermal titration calorimetry and inhibition assays were conducted with synthetic Fg Aalpha peptides. To determine the physiological implications of Bbp binding to Fg, the effect of Bbp on fibrinogen clotting was studied. Results show that Bbp binding to Fg inhibits the formation of fibrin. The consequences of this interaction are currently under investigation. Together, these data demonstrate that human Fg is a novel ligand for Bbp. This study indicates that the MSCRAMM Bbp may aid in staphylococcal attachment by targeting both an extracellular matrix and a blood plasma protein. The implications of these novel findings are discussed.

Role played by serum, a biological cue, in the adherence of Enterococcus faecalis to extracellular matrix proteins, collagen, fibrinogen, and fibronectin.

BACKGROUND: Most previous studies have found that Enterococcus faecalis isolates do not show significant adherence to fibronectin and fibrinogen. METHODS: The influence of various conditions on E. faecalis adherence to extracellular matrix (ECM) proteins was evaluated using a radiolabeled-cell adherence assay. RESULTS: Among the conditions studied, growth in 40% horse serum (a biological cue with potential clinical relevance) elicited adherence of all 46 E. faecalis strains tested to fibronectin and fibrinogen but not to elastin; adherence levels were independent of strain source, and adherence was eliminated by treating cells with trypsin. As previously reported, serum also elicited adherence to collagen. Although prolonged exposure to serum during growth was needed for enhancement of adherence to fibrinogen, brief exposure (<5 >min) to serum had an immediate, although partial, enhancing effect on adherence to fibronectin and, to a lesser extent, collagen; pretreatment of bacteria with chloramphenicol did not decrease this enhanced adherence to fibronectin and collagen, indicating that protein synthesis is not required for the latter effect. CONCLUSION: Taken together, these data suggest that serum components may serve (1) as host environmental stimuli to induce the production of ECM protein-binding adhesin(s), as previously seen with collagen adherence, and also (2) as activators of adherence, perhaps by forming bridges between ECM proteins and adhesins.

Ligand-signaled upregulation of Enterococcus faecalis ace transcription, a mechanism for modulating host-E. faecalis interaction.

Enterococcus faecalis, the third most frequent cause of bacterial endocarditis, appears to be equipped with diverse surface-associated proteins showing structural-fold similarity to the immunoglobulin-fold family of staphylococcal adhesins. Among the putative E. faecalis surface proteins, the previously characterized adhesin Ace, which shows specific binding to collagen and laminin, was detectable in surface protein preparations only after growth at 46 degrees C, mirroring the finding that adherence was observed in 46 degrees C, but not 37 degrees C, grown E. faecalis cultures. To elucidate the influence of different growth and host parameters on ace expression, we investigated ace expression using E. faecalis OG1RF grown in routine laboratory media (brain heart infusion) and found that ace mRNA levels were low in all growth phases. However, quantitative reverse transcription-PCR showed 18-fold-higher ace mRNA amounts in cells grown in the presence of collagen type IV compared to the controls. Similarly, a marked increase was observed when cells were either grown in the presence of collagen type I or serum but not in the presence of fibrinogen or bovine serum albumin. The production of Ace after growth in the presence of collagen type IV was demonstrated by immunofluorescence microscopy, mirroring the increased ace mRNA levels. Furthermore, increased Ace expression correlated with increased collagen and laminin adhesion. Collagen-induced Ace expression was also seen in three of three other E. faecalis strains of diverse origins tested, and thus it appears to be a common phenomenon. The observation of host matrix signal-induced adherence of E. faecalis may have important implications on our understanding of this opportunistic pathogen.

A trilocus sequence typing scheme for hospital epidemiology and subspecies differentiation of an important nosocomial pathogen, Enterococcus faecalis.

In this study, we present a trilocus sequence typing (TLST) scheme based on intragenic regions of two antigenic genes, ace and salA (encoding a collagen/laminin adhesin and a cell wall-associated antigen, respectively), and a gene associated with antibiotic resistance, lsa (encoding a putative ABC transporter), for subspecies differentiation of Enterococcus faecalis. Each of the alleles was analyzed using 50 E. faecalis isolates representing 42 diverse multilocus sequence types (ST(M); based on seven housekeeping genes) and four groups of clonally linked (by pulsed-field gel electrophoresis [PFGE]) isolates. The allelic profiles and/or concatenated sequences of the three genes agreed with multilocus sequence typing (MLST) results for typing of 49 of the 50 isolates; in addition to the one exception, two isolates were found to have identical TLST types but were single-locus variants (differing by a single nucleotide) by MLST and were therefore also classified as clonally related by MLST. TLST was also comparable to PFGE for establishing short-term epidemiological relationships, typing all isolates classified as clonally related by PFGE with the same type. TLST was then applied to representative isolates (of each PFGE subtype and isolation year) of a collection of 48 hospital isolates and demonstrated the same relationships between isolates of an outbreak strain as those found by MLST and PFGE. In conclusion, the TLST scheme described here was shown to be successful for investigating short-term epidemiology in a hospital setting and may provide an alternative to MLST for discriminating isolates.

Comportamiento y expectativas de los jóvenes frente a la crisis de la política

Es indiscutible la relevancia que tienen los jóvenes como sujetos políticos, por ser uno de los sectores más dinámicos de la sociedad, contribuyendo a través de sus orientaciones y prácticas a superar la crisis de la política y recuperar su centralidad. Se detectan en las nuevas generaciones manifestaciones de rechazo y distanciamiento hacia la política: desinterés, escasa adhesión a los partidos políticos y personalización de la opción electoral, crítica a la dirigencia y emergencia de nuevos actores sociales que amplían los canales de participación. Estos factores emergentes muestran el agotamiento de cierta concepción de la política y una redefinición de los sentidos de la misma.

El concepto de persona en el comentario de las sentencias de Guillermo de Ockham

En un primer examen de las auctoritates Ockham formula un claro acercamiento al esquema aristotélico-boeciano y a la definición de persona como sustancia en cuanto suppositum intellectualis, definición que encuentra conveniente aplicar tanto a lo creado como a Dios. Comienza luego una discusión más próxima y contemporánea con los moderni, que está centrada, por un lado en Escoto para quien la persona se ha de definir a partir de la relación; y por otro, con santo Tomás de Aquino. “Persona", para el Aquinate, no significa una naturaleza común quidditas, ousía o sustancia segunda, por el contrario, indica al individuo: “esta carne y estos huesos" pero lo significa de un modo vago e indeterminado. Precisamente, éste es el punto que Ockham discute: qué denota esta significación indeterminada; le dedica a la cuestión un amplio análisis que lo conduce a equiparar los conceptos de naturaleza y de persona. En un paso subsiguiente Ockham propone examinar las personas in divinis: no es posible establecer in divinis ninguna diferencia o distinción; si se afirma en Dios la presencia de tres personas y de una sola naturaleza la adhesión se presta por la fe sin que medie un acercamiento racional al tema. El aparato conceptual y metafísico para abordar el problema de la persona en sede divina, ha pasado por la criba de un examen que concluye, para Ockham, en la verdadera imposibilidad de elaborar una teología trinitaria.

El dilema tiempo-creación en el pensamiento de Alberto Magno

La cuestión sobre la creación y su relación con el tiempo constituye una de las problemáticas fundamentales del siglo XIII y un punto de encuentro entre el pensamiento filosófico y teológico, entre una representación científica del mundo que se descubre en la transmisión del pensamiento griego y lo que la palabra revelada ofrece como adhesión de fe. El presente trabajo intenta mostrar el aporte de Alberto Magno a partir del análisis de de las siguientes obras: In II Sententiarum, Physica, Metaphysica y Summa Theologiae. Alberto expone su pensamiento a partir de la interpretación del Génesis 1, 1: «en un principio Dios creó el cielo y la tierra». En su opinión, la noción de principio señala un comienzo temporal, de modo que un universo creado, necesariamente debe haber comenzado a ser en el tiempo, asumiendo como contradictoria la posibilidad de un universo creado y eterno. Admite, sin embargo, que no es posible demostrar racionalmente que el universo creado tuvo un comienzo. Por esta razón, la problemática del inicio temporal del universo creado pertenece propiamente al ámbito teológico y de la fe, ya que no puede llegar a ser resuelta desde la filosofía y el conocimiento natural.

Restauración de incisivo inferior fracturado mediante procedimientos de invasión mínima : reporte de un caso clínico

La odontología está inmersa en un nuevo paradigma: no se puede pensar en ninguna técnica restauradora sin que participen fenómenos de adhesión. El desarrollo de pernos de fibra de vidrio, sumado a los procedimientos de restauraciones adhesivas puede utilizarse como uno de los tantos recursos de la odontología de invasión mínima. Los pernos de fibra de vidrio ofrecen varias ventajas: comportamiento anisótropo, módulo de elasticidad bajo, buena resistencia mecánica, el lecho que aloja al perno de fibra requiere de una mínima preparación y se cementan con cementos adhesivos con carga, permitiendo de esta manera obtener una superficie homogénea que se interpone entre el perno de fibra y los tejidos dentales, conectándolo a los tejidos del conducto y sustituyendo mecánicamente la dentina. El caso clínico que se reporta se presentó para su resolución en la Clínica Integrada III F. O. UNCuyo durante el año lectivo 2009. El paciente presentaba una fractura amelodentinaria desde hacía cuatro años, con compromiso de la vitalidad y un proceso periapical. Durante los procedimientos endodónticos se realizó una perforación de la pared del conducto que se selló mediante la colocación de hidróxido de calcio y la obturación del mismo con conos de gutapercha. Se efectuó el seguimiento clínico y radiográfico del caso en donde se constató la reparación del proceso apical y luego se procedió a la restauración del elemento dentario con resinas compuestas con la ayuda de un poste de fibra de vidrio cementado con cemento resinoso. Dadas las características del tratamiento endodóntico realizado, se decidió dejar más porción del cono de gutapercha a pesar de lo aconsejado por numerosos autores, ya que de esta manera se aseguró el sellado de la perforación radicular para evitar de esta manera la nanofiltración hacia el interior del elemento dentario.

La educación del diabético

En este trabajo se plantea una problemática de salud observada en el hospital Antonio J. Scaravelli, en el servicio de clínica médica y traumatología. Dicho problema es la falta de adhesión al tratamiento en los pacientes diabéticos, lo que genera reingresos en el servicio con grandes complicaciones, de las cuales muchas prevenibles. El contenido del informe responde a la investigación realizada durante tres meses del año 2013 y tomando como muestra los pacientes internados en los servicios de clínica médica y traumatología durante ese lapso. Para la recolección de los datos se utilizó como herramienta la entrevista, se ordenó la información en tablas y gráficos para la interpretación de resultados.

¿Cuáles son las causas de abandono del tratamiento de pacientes diabéticos que se atienden en Clínica Médica del Hospital Malargüe período 2008-2009?

Los objetivos de este trabajo son: determinar el grado de cumplimiento del tratamiento y su relación con la educación recibida por enfermería, de pacientes diabéticos internados en el Hospital Malargüe durante los meses de enero a diciembre de 2010, caracterizar a los pacientes diabéticos incluidos en la muestra, determinar el grado de adhesión al tratamiento en cuanto a ejercicios, dieta, medicación y controles, conocer la forma en que realizan la consulta médica y planificar acciones de educación desde el área de enfermería para contribuir a un tratamiento y cuidados integrales e intedisciplinarios.