José Antonio Caride Gómez

Cooperation between the components of the meningococcal transferrin receptor, TbpA and TbpB, in the uptake of transferrin iron by the 37-kDa ferric-binding protein (FbpA).

Meningococcal TbpAB complexes TbpA, TbpB and FbpA were purified and used to study their role in the uptake of iron from transferrin to FbpA. Purification was achieved by affinity chromatography techniques, yielding homogeneous, non-denatured and functional material. TbpA could not be separated from TbpB and had to be purified from a TbpB-defective mutant strain. FbpA was able to bind iron from transferrin only when TbpAB complexes, TbpA and/or TbpB, were also present during the interaction. The highest uptake efficiences were obtained with TbpAB complexes or TbpA/TbpB mixtures. We conclude that the TbpA and TbpB molecules form true functional transferrin receptors, that FbpA is able to take iron directly from transferrin when in the presence of the components of the receptor, and that both Tbps are necessary for an optimal operation of the uptake system.

The neisserial 37 kDa ferric binding protein (FbpA)

The ferric binding protein (FbpA) is one of the major proteins regulated by the level of environmental iron in the genus Neisseria. Its conservation in all species of pathogenic Neisseria has been demonstrated, and the possible role that it plays in the iron uptake mechanisms in these bacteria has been postulated. Similar proteins in Haemophilus influenzae (HitA) and in Serratia marcescens (SfuA) have been described, but relationships with the meningococcal FbpA could not be proven. Although supposedly periplasmic, the exact location of FbpA remains controversial because some molecules, or parts of them, have been found exposed to the bacterial outer surface. The DNA sequence downstream of the fbpA gene has been recently analysed, finding an operon composed of three open reading frames: fbpA, encoding for FbpA; fbpB, that codifies a cytoplasmic permease, and fbpC, that contains the information for a nucleotide binding protein. These proteins would form an iron transport system through the periplasmic space. FbpA is highly antigenic in mice when injected in purified form, shows intraspecies and interspecies antigenic homogenicity, and specific anti-FbpA antibodies are fully cross-reactive; nevertheless, the in vivo induction of anti-FbpA antibodies in man is still polemical. Recent studies reveal that the purified FbpA induces a fair response of bactericidal antibodies in mice.

Analysis of TbpA and TbpB functionality in defective mutants of Neisseria meningitidis

Iron uptake analysis suggested that the Neisseria meningitidis transferrin (Tf) binding proteins, TbpA and TbpB, form only one type of receptor complex. Mutants defective in the synthesis of either TbpA or TbpB, but not defective in both proteins, can bind Tf, suggesting that both proteins are surface exposed and function in Tf binding. Also, iron uptake from Tf into the meningococci did not require the presence of both Tbps. The TbpB-defective mutant incorporated c. 37% of the iron taken up by the wild-type strain, but this was insufficient for bacterial growth. The TbpA-defective mutant incorporated c. 50% of the iron taken up by the wild-type strain and was able to grow with Tf as the only iron source. Mouse antibodies specific for TbpA were able to block c. 70% of the iron uptake from Tf in the wild-type strain, whereas they blocked only 22% of iron uptake in the TbpB-defective mutant and did not block uptake in the TbpA-defective strain. These results emphasise that TbpA should be considered in future vaccine trials in which iron-restricted proteins are to be included in the vaccine formulation.

Effect of adjuvants in the isotypes and bactericidal activity of antibodies against the transferrin-binding proteins of Neisseria meningitidis

Twenty-eight Neisseria meningitidis strains of different serogroups, serotypes, and TbpB isotypes were used to test the effect of five adjuvant formulations on the immune response to the meningococcal transferrin-binding proteins (Tbps) in mice. Levels of anti-Tbps antibodies were relatively low when purified TbpA-TbpB complexes were used for immunization, those obtained with the RAS adjuvant being the highest, and the isotype distribution reveals a prevalence of the non-bactericidal IgG1. Specific anti-Tbps antibody levels were five to 125 times higher immunizing with whole outer membrane vesicles, with bactericidal isotypes prevailing, which suggests that presentation of these antigens in their natural conformation is crucial to elicit a good response. Nevertheless, bactericidal activity did not correlate with these characteristics, confirming that it must be also influenced by other factors, and direct evaluation of the killing ability is necessary to draw conclusions about the efficacy of antigens or adjuvants in vaccine design.

Antigenicity, cross-reactivity and surface exposure of the Neisseria meningitidis 37 kDa protein (Fbp)

The 37 kDa iron-repressible protein, Fbp, was purified from two Neisseria meningitidis strains by metal-affinity chromatography and used to obtain mouse monospecific polyclonal immune sera. Dot-blot, immunoblotting and whole cell ELISA results demonstrate that the Fbp is present in all 16 N. meningitidis and four commensal Neisseria species tested, is highly antigenic in mouse when injected in pure form, and shows intra- and inter-species antigenic homogeneity, anti-Fbp antibodies being fully cross-reactive using the techniques mentioned. We also found that Fbp molecules (or parts of them) are surface exposed, in disagreement with the proposed exclusively periplasmic localization, although anti-Fbp antibodies seem unable to block iron uptake or to induce complement-mediated killing of the meningococci. Taken along with the high immunogenicity of the purified protein and the complete cross-reactivity of the antibodies elicited, this suggests that the protective effect of the purified Fbp must be further studied to evaluate its inclusion in future vaccine trials.

Blocking of iron uptake by monoclonal antibodies specific for the Neisseria meningitidis transferrin- binding protein 2

The existence of epitopes common to different strains in the Neisseria meningitidis transferrin (Tf)-binding protein 2 (TBP2), combined with the ability of polyclonal anti-TBP2 antibodies to inhibit Tf binding and block iron uptake in this species, led to this study on the effect of anti-TBP1+2 monoclonal antibodies (MAbs) to determine the presence of epitopes inside the Tf-binding region. All MAbs used reacted exclusively with the homologous strain when tested by dot-blots of outer membrane vesicles, with the reaction being specific for TBP2 after SDS-PAGE and electroblotting. In contrast, ELISA and iron-uptake blocking assays were also positive with heterologous strains belonging to Rokbis group II (high mol.wt TBP2). The results confirmed the two group classification proposed by Rokbi and, in contrast to other studies, indicated the existence of epitopes in the Tf-binding region that are common only to strains of Rokbis group II. These epitopes may become denatured after drying for dot-blot assays or after SDS-PAGE and electroblotting.

In the Name of Environmental Education: Words and Things in the Complex Territory of Education-Environment-Development Relations.

The proclamation of the Decade of Education for Sustainable Development by the United Nations has placed education in general, and environmental education in particular, at the front of a future full of important and uncertain meanings. On the one hand, those inviting a conceptual, theoretical and praxiological revision of the education–environment–development relationship appeal to the role of education in the construction of ‘sustainability’ and lifestyles that will make it possible. On the other hand, there are those that anticipate new and different readings of the environmental educative task. They range from the questioning of its historic entity and identity (more than 30 years of initiatives, plans and programmes across the world) to the firm demand for its proposals to provide an ‘education’ that is essential for the renewal of human action and thought. The article subscribes to the latter position, arguing in favour of the necessity of an environmental education that does not contradict itself, neither in its critical-reflexive discourses nor in its emancipative practices, as a fundamental pillar of any development that aims at being ‘human’ and ‘sustainable’ from a pedagogical, ecological and social point of view.

One-step metal-affinity method for the purification of the iron-binding protein Fbp of Neisseria meningitidis

A one-step method for the purification of the iron-binding protein, Fbp, from Neisseria meningitidis has been developed using affinity chromatography on iron-Sepharose. This method allowed the purification of the protein in non-denaturing conditions and in a similar way to those used previously. Our results demonstrate the homogeneity of the purified protein as tested by SDS-PAGE and IEF.

La Pedagogía Social ante el proceso de convergencia europea de la Educación Superior.

En los inicios del siglo XXI las universidades europeas afrontan uno de sus procesos de reforma estructural y curricular mas decisivos, con objetivos y lineas de actuacion que tienen como principal soporte la Declaracion de Bolonia (junio de 1999). Un documento con el que se propone crear un sistema de grados academicos facilmente comprensibles y comparables, fomentar la movilidad de los estudiantes, docentes e investigadores, garantizar la calidad de la ensenanza y crear un Espacio Europeo de la Educacion Superior (EEES). En este contexto, el articulo argumenta acerca de las connotaciones pedagogicas y sociales inherentes a estas reformas, valorando los principios y actuaciones en los que se sustentan como una nueva oportunidad para el reencuentro de las universidades consigo mismas y con la sociedad. Tambien se analiza el estado de cuestion actual de la Pedagogia Social y de la Educacion Social en Espana ante el proceso de convergencia, de sus realidades y desafios en la construccion de una sociedad educadora, garante de los derechos civicos y la condicion ciudadana. Para concluir con una serie de recomendaciones y propuestas destinadas a mejorar las decisiones que se adoptan en el futuro, ante la previsible reconversion de la Diplomatura en Educacion Social en un Grado con la misma denominacion. -------------------------------------------------------------------------- In the beginnings of the 21st century, European Universities confront one of the processes of structural and curricular reform more decisive, with aims and lines of action that take the Bologna Declaration as a principal support (June, 1999). A document that is designed to introduce a system of academic degrees that are easy to read and compare, to foment the mobility of students, teachers and researchers, to guarantee the quality in education and to take into account the European dimension of higher Education. In this context, the article argues about the pedagogic and social connotations inherent in these reforms, valuing the principles and actions that sustain them, as a new opportunity for the meeting of the Universities with himself and with the society. Also it is analyzed the condition of current question about the Social Pedagogy and about the Social Education in Spain before the process of convergence, about his realities and challenges in the construction of an educational and responsible society that guarantees the civic rights and the civil condition. To conclude with a recommendations and offers destined to improve the decisions that are adopted in the future, before the predictable restructuring of the Social Education Diplomature in a Degree with the same denomination.

Prevention of meningococcal disease: present and future

Meningococcal infections are an important health care problem in underdeveloped areas, and their incidence is growing in developed countries. Despite the great advances in the treatment of bacterial infections, mortality due to meningitis and meningococcaemia is high, about 10%, morbidity also being important. Meningococcal diseases can be prevented in close contacts of cases by using chemoprophylaxis, to eliminate the carrier state, or vaccination. Those vaccines which are actually approved are made from purified capsular polysaccharides and are effective only for meningococcal serogroups A and C (also Y and W135 in some cases), but they present some important problems - particularly their ineffectivity in children under 2 years and their inability to induce immunological memory. The most promising alternatives are the vaccines based on outer membrane proteins; these vaccines can induce memory, can be effective at all ages and can act against all serogroups, particularly serogroup B, the serogroup which causes most of the cases in developed countries.