R Phillips, KNUST Ghana. Pathogenesis and management of M. ulcerans disease, Buruli ulcer

Buruli ulcer is a neglected tropical disease caused by infection with Mycobacterium ulcerans (Mu) which is common in rural parts of West African countries including Ghana. It causes large, disfiguring skin ulcers mainly in children aged 5 to 15 years although any age can be affected. Access to treatment in rural areas is limited and many cases present late because of fear, suspicion about conventional medicine and the economic consequences for poor families. The disease is highly focal and in Ghana, where these investigations will be carried out, it is not seen at all in the North, most cases occurring in particular parts of the Ashanti Region. The mode of transmission remains unknown but there have been major advances in understanding the mechanism of disease since the establishment of the WHO Buruli ulcer initiative in 1998 together with improved diagnosis and management. The aim of the proposed studies is to identify markers predictive of a rapid response to antibiotic treatment and to investigate the pathogenesis of paradoxical reactions and oedematous lesions in Mu disease.
When someone is infected with Mu, the first sign is a nodule under the skin which gradually enlarges and breaks down in the centre to form an ulcer. This is because Mu produces a toxin that spreads outwards and damages subcutaneous tissue. In about 15% of cases the skin around the ulcer is swollen with oedema, that is fluid in the tissue, and in this situation the ulcer enlarges more rapidly so that it may involve a whole limb for example.
In recent years it has been found that instead of removing ulcers by extensive surgery it is possible to heal them by antibiotic treatment for 8 weeks with daily tablets and intramuscular injections. This is unpleasant, particularly for children, and it would be better if the treatment could be limited to a shorter time. Our previous studies suggest that it may be possible to do investigations at the beginning of treatment that distinguish people who need only 4 weeks antibiotic treatment. A wide range of tests will be investigated in detail to identify markers for people in whom the infection is at an early stage with low numbers of the Mu bacteria and low levels of the toxin in the skin. During antibiotic treatment the rate of healing will be measured carefully to find out which markers are most reliable and practical for use in countries where Buruli ulcer is endemic.
After antibiotic treatment some patients develop new areas of inflammation or a sudden increase in the size of a healing ulcer, giving the impression that the infection is still active. It is not known what causes these paradoxical reactions but it is thought that there is an increasing immune response to residual dead Mu organisms in the tissue. In the proposed studies the immune response to Mu will be investigated serially during antibiotic treatment and when patients develop a paradoxical reaction.
Oedematous disease is the most severe form of Buruli ulcer. The cause of it is not known but a likely explanation is that larger than normal amounts of the Mu toxin mycolactone are produced and diffuse away from the initial site of infection. This will be investigated by measuring mycolactone in the skin of patients and studying mycolactone production by the strain of Mu cultured from patients with this form of the disease.

Buruli ulcer is a neglected tropical disease caused by infection with Mycobacterium ulcerans (Mu) which is common in remote rural parts of West African countries including Ghana. The disease results initially in a subcutaneous painless nodule tethered to the skin or an intradermal plaque both of which ulcerate centrally. Oedematous disease is the most severe form of disease and it occurs in about 15% of cases. It may extend to a whole limb.
Antibiotic treatment with rifampicin and streptomycin for 8 weeks is now standard. The main aim of the proposed studies is to investigate markers for patients with early infection in whom the bacterial load is low and production of the M. ulcerans toxin mycolactone is limited. It is proposed that such patients will respond rapidly to antibiotic treatment and that a shortened course of antibiotics will be adequate.
A prospective observational study will be undertaken in patients diagnosed accurately to have Mu infection. Biomarkers will be investigated in serum using multianalyte profiling by Luminex. The immune response to Mu antigens will be studied and lymphocyte populations will be analysed by flow cytometry. Tissue mycolactone concentration will be measured by a cytotoxicity assay and by mass spectrometry.
Paradoxical reactions are seen in 10% of cases during or after antibiotic treatment. The pathogenesis of these reactions and of oedematous disease will be investigated using the same techniques to measure the immune response to Mu antigens, serum biomarkers and tissue mycolactone concentration.

The results of this research will be made available to the Buruli ulcer community including scientists studying the disease and health workers of all levels. The potential benefits are that patients will receive better informed management in the future and in particular it may be possible to shorten the course of antibiotic treatment with consequent savings in resources as well as reduced discomfort for patients. The management of paradoxical reactions and oedematous disease may improve as a result of knowledge acquired during the investigation.
The research will be largely carried out in Ghana where Buruli ulcer is endemic and there will be benefits in the development of infrastructure and capacity building.
The African Research Leader will acquire new knowledge and skills that will enable him to compete in obtaining funding for future research. Support during the tenure of the award will allow him to continue building a team of researchers in Kumasi. The link between KNUST and SGUL will be strengthened and new research ideas will be generated at both institutions.