Hijacking bacterial glycosylation for the production of glycoconjugates, from vaccines to humanised glycoproteins.

OBJECTIVES: Glycosylation or the modification of a cellular component with a carbohydrate moiety has been demonstrated in all three domains of life as a basic post-translational process important in a range of biological processes. This review will focus on the latest studies attempting to exploit bacterial N-linked protein glycosylation for glycobiotechnological applications including glycoconjugate vaccine and humanised glycoprotein production. The challenges that remain for these approaches to reach full biotechnological maturity will be discussed. KEY FINDINGS: Oligosaccharyltransferase-dependent N-linked glycosylation can be exploited to make glycoconjugate vaccines against bacterial pathogens. Few technical limitations remain, but it is likely that the technologies developed will soon be considered a cost-effective and flexible alternative to current chemical-based methods of vaccine production. Some highlights from current glycoconjugate vaccines developed using this in-vivo production system include a vaccine against Shigella dysenteriae O1 that has passed phase 1 clinical trials, a vaccine against the tier 1 pathogen Francisella tularensis that has shown efficacy in mice and a vaccine against Staphylococcus aureus serotypes 5 and 8. Generation of humanised glycoproteins within bacteria was considered impossible due to the distinct nature of glycan modification in eukaryotes and prokaryotes. We describe the method used to overcome this conundrum to allow engineering of a eukaryotic pentasaccharide core sugar modification within Escherichia coli. This core was assembled by combining the function of the initiating transferase WecA, several Alg genes from Saccharomyces cerevisiae and the oligosaccharyltransferase function of the Campylobacter jejuni PglB. Further exploitation of a cytoplasmic N-linked glycosylation system found in Actinobacillus pleuropneumoniae where the central enzyme is known as N-linking glycosyltransferase has overcome some of the limitations demonstrated by the oligosaccharyltransferase-dependent system. SUMMARY: Characterisation of the first bacterial N-linked glycosylation system in the human enteropathogen Campylobacter jejuni has led to substantial biotechnological applications. Alternative methods for glycoconjugate vaccine production have been developed using this N-linked system. Vaccines against both Gram-negative and Gram-positive organisms have been developed, and efficacy testing has thus far demonstrated that the vaccines are safe and that robust immune responses are being detected. These are likely to complement and reduce the cost of current technologies thus opening new avenues for glycoconjugate vaccines. These new markets could potentially include glycoconjugate vaccines tailored specifically for animal vaccination, which has until today thought to be non-viable due to the cost of current in-vitro chemical conjugation methods. Utilisation of N-linked glycosylation to generate humanised glycoproteins is also close to becoming reality. This 'bottom up' assembly mechanism removes the heterogeneity seen in current humanised products. The majority of developments reported in this review exploit a single N-linked glycosylation system from Campylobacter jejuni; however, alternative N-linked glycosylation systems have been discovered which should help to overcome current technical limitations and perhaps more systems remain to be discovered. The likelihood is that further glycosylation systems exist and are waiting to be exploited

Antibody Responses to <i>Schistosoma mansoni</i> Schistosomula Antigens

While antigens from Schistosoma schistosomula have been suggested as potential vaccine candidates, the association between antibody responses with schistosomula antigens and infection intensity at reinfection is not well known. Schistosoma mansoni-infected individuals were recruited from a schistosomiasis endemic area in Uganda (n = 372), treated with 40 mg/kg praziquantel (PZQ) and followed up at five weeks and at one year post-treatment. Pre-treatment and five weeks post-treatment immunoglobulin (Ig) E, IgG1 and IgG4 levels against recombinant schistosomula antigens rSmKK7, rSmLy6A, rSmLy6B and rSmTSP7 were measured using ELISA. Factors associated with detectable pre-treatment or post-treatment antibody response against the schistosomula antigens and the association between five-week antibody responses and one year post-treatment reinfection intensity among antibody responders were examined. Being male was associated with higher pre-treatment IgG1 to rSmKK7, rSmLy6a and AWA. Five weeks post-treatment antibody responses against schistosomula antigens were not associated with one year post-treatment reinfection intensity among antibody responders' antibody levels against rSmKK7, rSmLy6B and rSmTSP7 dropped, but increased against rSmLy6A, AWA and SEA at five weeks post-treatment among antibody responders. S. mansoni-infected individuals exhibit detectable antibody responses to schistosomula antigens that are affected by treatment. These findings indicate that schistosomula antigens induce highly varied antibody responses and could have implications for vaccine development

Engineering Eukaryotic-Like Protein Glycosylation In Escherichia Coli

N-linked glycosylation is a common protein post-translational modification where glycans are attached to asparagine residues located on a consensus sequence. Structure of these glycans varies widely among species, which is of particular importance given the role that glycans play on protein folding, functionality and recognition. Notably, glycans are often necessary to maintain stability and efficacy of therapeutic proteins in the human body. Production of therapeutic glycoproteins in Escherichia coli has the potential to become a flexible and relatively cheap alternative to current production systems, even though E. coli has no native protein glycosylation machinery. Recently, the glycosylation machinery from Campylobacter jejuni, a gram negative bacterium, was transferred to E. coli, making protein glycosylation possible for the first time in this host. Unfortunately glycans synthesized by C. jejuni, and bacteria in general, are not appropriate for use in human therapeutics, hence engineering the synthesis of human-like glycans in E. coli becomes necessary. This study is focused on recombinant expression and engineering of eukaryotic glycosyltransferases to enable E. coli to synthesize eukaryotic-like glycans suitable for in vivo production of therapeutic glycoproteins. Initially, we expressed glycosyltransferases from Saccharomyces cerevisiae (Alg13, Alg14, Alg1 and Alg2) for the synthesis of a core glycan structure (trimannose core) common to many human glycans. Later, the work was ori- ented towards the extension of this trimannose core for production of authentic human-like glycans and improvement of the current system using directed evolution of glycosyltransferases. In the future, we expect to be able to produce glycosylated proteins in vivo, displaying fully sialylated human-like glycans

Estilos de enseñanza de los docentes de arquitectura en las asignaturas de taller de diseño en la Universidad Francisco de Paula Santander – seccional Cúcuta

Esta investigación se centró en la necesidad de establecer un marco de referencia investigativo sobre los estilos de enseñanza, ya que dentro de la Universidad Francisco de Paula Santander muy poco se ha indagado esta temática y por tanto la inexistencia de referentes que permita la creación de estrategias que refuercen el mejoramiento de las prácticas educativas en el Programa de Arquitectura de la Universidad Francisco de Paula Santander. El objeto central se basó en analizar y describir los estilos de enseñanza de los docentes de Taller de Diseño en el programa de Arquitectura de la Universidad, teniendo en cuenta variables como la edad, sexo, experiencia docente y programa académico, esto mediante el Cuestionario estilos de enseñanza (CEE)EspecializaciónEspecialista en Práctica Pedagógic

Design of a low budget lab-scale UASB reactor for research and teaching in low income laboratories

Upflow anaerobic sludge blanket (UASB) reactors are most widely implemented setup for anaerobic treatment of wastewaters with a high organic load. Even though these are very efficient systems and methane production from organic waste makes them energetically attractive, its implementation in developing countries is still hampered not just because of financial reasons, but because of its relatively difficult operation. For this reason, it is crucial to maintain a number of installed lab-scale UASB reactors for training of technicians and professionals. Unfortunately, the installation costs might be prohibitive for low income laboratories. As scientific literature usually focuses on results obtained using UASB reactors, but rarely focuses on construction details, here we present a detailed description of the building of a low budget UASB reactor for research and instruction in the Antonio Nariño University environmental engineering labs

Importancia del regente de farmacia en la implementación del programa de farmacovigilancia en establecimientos independientes

Autoria propia.El presente proyecto busca establecer la importancia del regente de farmacia en la implementación del programa de farmacovigilancia del establecimiento naturista y homeopático independiente “Distri Naturals Diaz” en Pitalito Huila; el cual es regulado por el Instituto Nacional de Vigilancia de Medicamentos Invima. Su justificación se basa en la importancia de implementar actividades relacionadas con la detección, evaluación, entendimiento y prevención de los eventos adversos o cualquier otro problema relacionado con medicamentos en los establecimientos independientes, ligadas a las orientaciones de Organización Mundial de la Salud (OMS), al igual que la Organización Panamericana de la Salud (OPS) así como la normatividad legal y vigente colombiana Se realiza el soporte teórico, contextual y legal desde la función objetiva de la farmacovigilancia abordando aspectos relacionados con las actividades de análisis y gestión de riesgos para la salud pública, permitiendo identificar, cuantificar y evaluar los riesgos asociados con el uso de medicamentos permite a los pacientes tomar las medidas necesarias para prevenir o reducir el daño y, cuando sea necesario, implementar medidas regulatorias o de mitigación. Por ende, se planteó de manera metodológica la aplicación de una encuesta direccionada de manera cualitativa descriptiva a usuarios y trabajadores, obteniendo como discusión y recomendación la responsabilidad ética y social en la identificación de riesgos, promoción de medidas preventivas y contribución a la mejora de la salud pública por parte del establecimiento independiente sujeta del presente estudio.This project seeks to establish the importance of the pharmacy manager in the implementation of the pharmacovigilance program of the independent naturopathic and homeopathic establishment “Distri Naturals Diaz” in Pitalito Huila; which is regulated by the National Institute for Drug Surveillance Invima. Its justification is based on the importance of implementing activities related to the detection, evaluation, understanding and prevention of adverse events or any other problem related to medications in independent establishments, linked to the guidelines of the World Health Organization (WHO), as well as the Pan American Health Organization (PAHO) as well as the Colombian legal and current regulations. Theoretical, contextual and legal support is provided from the objective function of pharmacovigilance, addressing aspects related to the analysis and risk management activities for public health, allowing the identification, quantification and evaluation of the risks associated with the use of medicines. Patients take necessary measures to prevent or reduce harm and, where necessary, implement regulatory or mitigation measures. Therefore, the application of a qualitatively descriptive survey to users and workers was proposed in a methodological manner, obtaining as a discussion and recommendation the ethical and social responsibility in the identification of risks, promotion of preventive measures and contribution to the improvement of the public health by the independent establishment subject of this study