Use of recombinant virus replicon particles for vaccination against Mycobacterium ulcerans disease

Buruli ulcer, caused by infection with Mycobacterium ulcerans, is a necrotizing disease of the skin and subcutaneous tissue, which is most prevalent in rural regions of West African countries. The majority of clinical presentations seen in patients are ulcers on limbs that can be treated by eight weeks of antibiotic therapy. Nevertheless, scarring and permanent disabilities occur frequently and Buruli ulcer still causes high morbidity. A vaccine against the disease is so far not available but would be of great benefit if used for prophylaxis as well as therapy. In the present study, vesicular stomatitis virus-based RNA replicon particles encoding the M. ulcerans proteins MUL2232 and MUL3720 were generated and the expression of the recombinant antigens characterized in vitro. Immunisation of mice with the recombinant replicon particles elicited antibodies that reacted with the endogenous antigens of M. ulcerans cells. A prime-boost immunization regimen with MUL2232-recombinant replicon particles and recombinant MUL2232 protein induced a strong immune response but only slightly reduced bacterial multiplication in a mouse model of M. ulcerans infection. We conclude that a monovalent vaccine based on the MUL2232 antigen will probably not sufficiently control M. ulcerans infection in humans

Dynamic effluents : a political economy analysis of the water sector in Lebanon

After years of neglect, Lebanon is facing the consequences of a struggling water sector. Existing water networks are deficient; water supply continuity is low; most wastewater is discharged into rivers or the Mediterranean Sea without adequate treatment; and inter-agency coordination remains weak and fragmented. In 2000, the government together with international donor agencies initiated a reform process to re-structure the water sector, merging local water authorities into four regional Water Establishments and giving more weight to the Ministry of Water and Energy. The water sector reforms pushed for good governance and the marketization of water services. The implementation process of the reforms, however, has been contested, modified and transformed by different political actors. This thesis takes a closer look at the water sector reforms in Lebanon, mapping key actors, their roles and responsibilities, their interests and interrelationships, and their influence in the sector. It seeks to understand the dynamics that underpin the institutional and policy set-up in the sector. For this purpose, the thesis makes use of an analytical framework for political economy analysis, contextualizing the reforms in the broader political and economic context. What drives and what constrains changes in water governance in Lebanon? A particular focus is put on the World Bank and other donor agencies, the Council for Development and Reconstruction (CDR) and the municipalities. They are key actors in the water sector and yet they have been largely disregarded by the reforms. The thesis explores how these actors interact and influence decision-making processes in the Lebanese sector. The Foucauldian notion of governmentality thereby serves as a method of inquiry, explaining how forms of political rationality inscribe themselves in practices, and by extension, in the political economy

Europe in local news

Local newspapers are rarely in the focus of research on the European public sphere. However, due to their various democratic functions, they should not be neglected and may help to build a European public sphere. This article presents the results of an in-depth study of local newspapers by asking: a) How and to what extent can local newspapers contribute to the construction of a European public sphere by Europeanisation of the local public through reporting on the EU? b) Which factors have an effect on how local newspapers report on EU topics? This paper synthesises theories and models of the European public to develop hypotheses, which are then tested in interviews with six local German journalists. It finds that local newspapers can promote the emergence of a European public sphere and that news factors, resources, individual attitudes and editorial frameworks do have an impact on their EU reporting

SNP-Typisierung des Buruli-Ulkus-Erregers

Zusammenfassung: Mithilfe vergleichender Genomanalysen konnten wir eine Feintypisierungsmethode für Mycobacterium ulcerans, dem genetisch monomorphen Erreger des Buruli-Ulkus, entwickel

Repurposing of Tuberculosis Drug Candidates for the Treatment of Mycobacterium ulcerans Disease

Buruli ulcer (BU) is a chronic necrotizing skin disease caused by Mycobacterium ulcerans. Historically, the disease was treated by surgical excision of the skin lesions, until an 8-week combination therapy of rifampicin and streptomycin was introduced in 2004. This treatment modality was effective and reduced recurrence rates. Rifampicin is the most efficacious antibiotic for the treatment of BU and, should rifampicin-resistant M. ulcerans strains emerge, there is currently no replacement for it. As for mycobacterial diseases in general, there is a pressing need for the development of novel, fast-acting drugs. Under market economy conditions, repurposing of new tuberculosis drug candidates is the most promising avenue for alternative BU treatments. Our drug repurposing activities have led to the identification of several actives against M. ulcerans. In particular, the cytochrome bc1 complex inhibitor telacebec (Q203) is a promising drug candidate for the treatment of BU in Africa and Australia. While an active cytochrome-bd oxidase bypass limits the potency of the cytochrome-bc1-specific inhibitor telacebec against M. tuberculosis, classical lineage M. ulcerans strains rely exclusively on cytochrome-bc1 to respire. Hence, telacebec is effective at nanomolar concentration against M. ulcerans, and a high treatment efficacy in an experimental mouse infection model indicates that treatment of BU could be substantially shortened and simplified by telacebec

Efficacy of an acid-oxidising solution (AOS) against Mycobacterium ulcerans

For the treatment of chronic wounds, acid-oxidising solutions (AOSs) with broad-spectrum microbicidal activity without disturbing granulation tissue formation, have been developed. We found AOSs to efficiently kill Mycobacterium ulcerans, the causative agent of Buruli ulcer, which is able to survive harsh decontamination treatments. Topical AOS treatment of Buruli ulcer lesions may support the recommended antibiotic therapy (oral rifampicin and clarithromycin), prevent contamination of the environment by the mycobacteria, and control secondary infections, which are a prevalent wound management problem in resource-poor Buruli ulcer endemic settings

Experimental infection of the pig with Mycobacterium ulcerans : a novel model for studying the pathogenesis of Buruli ulcer disease

Buruli ulcer (BU) is a slowly progressing, necrotising disease of the skin caused by infection with Mycobacterium ulcerans. Non-ulcerative manifestations are nodules, plaques and oedema, which may progress to ulceration of large parts of the skin. Histopathologically, BU is characterized by coagulative necrosis, fat cell ghosts, epidermal hyperplasia, clusters of extracellular acid fast bacilli (AFB) in the subcutaneous tissue and lack of major inflammatory infiltration. The mode of transmission of BU is not clear and there is only limited information on the early pathogenesis of the disease available.; For evaluating the potential of the pig as experimental infection model for BU, we infected pigs subcutaneously with different doses of M. ulcerans. The infected skin sites were excised 2.5 or 6.5 weeks after infection and processed for histopathological analysis. With doses of 2×107 and 2×106 colony forming units (CFU) we observed the development of nodular lesions that subsequently progressed to ulcerative or plaque-like lesions. At lower inoculation doses signs of infection found after 2.5 weeks had spontaneously resolved at 6.5 weeks. The observed macroscopic and histopathological changes closely resembled those found in M. ulcerans disease in humans.; Our results demonstrate that the pig can be infected with M. ulcerans. Productive infection leads to the development of lesions that closely resemble human BU lesions. The pig infection model therefore has great potential for studying the early pathogenesis of BU and for the development of new therapeutic and prophylactic interventions

Mycobacterium ulcerans mouse model refinement for pre-clinical profiling of vaccine candidates

Buruli Ulcer is a neglected tropical disease leading to extensive disabilities and morbidity in West Africa. In this paper we sought to characterize various strains of Mycobacterium ulcerans (M.ulcerans) with different origins and laboratory passage records while refining a mouse model for Buruli ulcer. We described, compared and followed the kinetics of the histo-pathological outcome of infection of a collection of strains at various anatomical sites of infection in order to find a suitable model for further immunization studies. Moreover we compared the outcome of infection in C57Bl/6 and Balbc/J mice. Specifically we described thoroughly one M. ulcerans strain characterized by slow growth rate and limited tissue necrosis, which presents close ressemblance with the infection kinetics in humans. This strain caused macrophages as well as T and B cells infiltration, correlating with mycobacterial proliferation at the site of infection as well as in the draining lymph nodes, making it a suitable strain to screen vaccine candidates efficacy

An efficient system to generate monoclonal antibodies against membrane-associated proteins by immunisation with antigen-expressing mammalian cells

ABSTRACT: BACKGROUND: The generation of monoclonal antibodies specific for protein antigens usually depends on purified recombinant protein for both immunisation and hybridoma screening. Purification of recombinant protein in sufficient yield and purity is a tedious undertaking and can be demanding especially in the case of membrane proteins. Furthermore, antibodies generated against a purified recombinant protein are frequently incapable of binding to the endogenous protein in its native context. RESULTS: We describe a strategy to generate monoclonal antibodies against membrane or membrane-associated proteins that completely bypasses any need for purified recombinant antigen. This approach utilises stably transfected mammalian cells expressing recombinant antigens on their cell surface for immunisation of mice. The transfected cells are also used for measuring seroconversion, hybridoma selection and antibody characterisation. By presenting the antigen in its native conformation for immunisation and hybridoma selection, this procedure promotes the generation of antibodies capable of binding to the endogenous protein. In the present study, we applied this approach successfully for three predicted GPI-anchored proteins of the malaria parasite Plasmodium falciparum. CONCLUSIONS: The described entirely cell-based technology is a fast and efficient approach for obtaining antibodies reactive with endogenous cell-surface proteins in their native conformatio

Improved Protective Efficacy of a Species-Specific DNA Vaccine Encoding Mycolyl-Transferase Ag85A from Mycobacterium ulcerans by Homologous Protein Boosting

Vaccination with plasmid DNA encoding Ag85A from M. bovis BCG can partially protect C57BL/6 mice against a subsequent footpad challenge with M. ulcerans. Unfortunately, this cross-reactive protection is insufficient to completely control the infection. Although genes encoding Ag85A from M. bovis BCG (identical to genes from M. tuberculosis) and from M. ulcerans are highly conserved, minor sequence differences exist, and use of the specific gene of M. ulcerans could possibly result in a more potent vaccine. Here we report on a comparison of immunogenicity and protective efficacy in C57BL/6 mice of Ag85A from M. tuberculosis and M. ulcerans, administered as a plasmid DNA vaccine, as a recombinant protein vaccine in adjuvant or as a combined DNA prime-protein boost vaccine. All three vaccination formulations induced cross-reactive humoral and cell-mediated immune responses, although species-specific Th1 type T cell epitopes could be identified in both the NH2-terminal region and the COOH-terminal region of the antigens. This partial species-specificity was reflected in a higher—albeit not sustained—protective efficacy of the M. ulcerans than of the M. tuberculosis vaccine, particularly when administered using the DNA prime-protein boost protocol