Mycolactone is the exotoxin produced by Mycobacterium ulcerans and is the virulence factor behind the neglected tropical disease Buruli ulcer. The toxin has a broad spectrum of biological effects within the host organism, stemming from its interaction with at least two molecular targets and the inhibition of protein uptake into the endoplasmic reticulum. Although it has been shown that the toxin can passively permeate into host cells, it is clearly lipophilic. Association with lipid carriers would have substantial implications for the toxin's distribution within a host organism, delivery to cellular targets, diagnostic susceptibility, and mechanisms of patho-genicity. Yet the toxin's interactions with, and distribution in, lipids are unknown. Herein we have used coarse-grained molecular dynamics simulations, guided by all-atom simulations, to study the interaction of mycolactone with pure and mixed lipid membranes. Using established techniques, we calculated the toxin's preferential localization, membrane translocation, and impact on membrane physical and dynamical properties. The computed water-octanol partition coefficient indicates that mycolactone prefers to be in an organic phase rather than in an aqueous environment. Our results show that in a solvated membrane environment the exo-toxin mainly localizes in the water-membrane interface, with a preference for the glycerol moi-ety of lipids, consistent with the reported studies that found it in lipid extracts of the cell. The calculated association constant to the model membrane is similar to the reported association constant for Wiskott-Aldrich syndrome protein. Mycolactone is shown to modify the physical properties of membranes, lowering the transition temperature, compressibility modulus, and critical line tension at which pores can be stabilized. It also shows a tendency to behave as a linactant, a molecule that localizes at the boundary between different fluid lipid domains in membranes and promotes inter-mixing of domains. This property has implications for the tox-in's cellular access, T-cell immunosuppression, and therapeutic potential.

Mosquitoes positive for M. ulcerans were linked to outbreaks of Buruli ulcer in humans.

Buruli ulcer is a disease of skin and soft tissue caused by Mycobacterium ulcerans. It can leave affected people scarred and disabled. What are the prospects for disease control?

Mycobacterium ulcerans infection, or Buruli ulcer, is the third most frequent mycobacterial disease in humans, often causing serious deformities and disability. The disease is most closely associated with tropical wetlands, especially in west and central Africa. Most investigators believe that the aetiological agent proliferates in mud beneath stagnant waters. Modes of transmission may involve direct contact with the contaminated environment, aerosols from water surfaces, and water-dwelling fauna (e.g. insects). Person-to-person transmission is rare. Trauma at the site of skin contamination by M. ulcerans appears to play an important role in initiating disease. Once introduced into the skin or subcutaneous tissue, M. ulcerans multiplies and produces a toxin that causes necrosis. However, the type of disease induced varies from a localised nodule or ulcer, to widespread ulcerative or non-ulcerative disease and osteomyelitis.Although culture of M. ulceransfrom a patient was first reported in 1948, attempts to culture the mycobacterium from many specimens of flora and fauna have been unsuccessful. Failure to cultivate this organism from nature may be attributable to inadequate sampling, conditions of transport, decontamination and culture of this fastidious heat-sensitive organism, and to along generation time relative to that of other environmental mycobacteria. Nevertheless, recent molecular studies using specific primers have revealed M. ulcerans in water, mud, fish and insects. Although no natural reservoir has been found, the possibility that M. ulcerans may colonise microfauna such as free-living amoebae has not been investigated. The host range of experimental infection by M. ulcerans includes lizards, amphibians, chick embryos, possums, armadillos, rats, mice and cattle. Natural infections have been observed only in Australia, in koalas, ringtail possums and a captive alpaca. The lesions were clinically identical to those observed in humans. Mycobacterium ulcerans infection is a rapidly re-emerging disease in some developing tropical countries. The re-emergence may be related to environmental and socioeconomic factors, for example, deforestation leading to increased flooding, and population expansion without improved agricultural techniques, thus putting more people at risk. Eradication of diseases related to these factors is difficult. Whether wild animals have a role in transmission is an important question that, to date, has been virtually unexplored. To address this question, surveys of wild animals are urgently required in those areas in which Buruli ulcer is endemic. KeywordsAlpacas -Aquatic insects -Buruli ulcer -Environment -Fish -Koalas -Mycobacteria -Mycobacterium ulcerans -Possums-Water. Bev. sci. teck Off.int.Epa.. 20 (1)

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SummaryMycobacterium ulcerans produces an extracellular cutaneous infection (Buruli ulcer) characterized by immunosuppression. This is in stark contrast to all other pathogenic Mycobacteria species that cause intracellular, granulomatous infections. The unique mycobacterial pathology of M. ulcerans infection is attributed to a plasmid-encoded immunomodulatory macrolide toxin, mycolactone. In this article we explore the role of mycolactone in the virulence of M. ulcerans using mycolactone and genetically defined mycolactone negative mutants. In a guinea pig infection model wild-type (WT) M. ulcerans produces an extracellular infection whereas mycolactone negative mutants produce an intracellular inflammatory infection similar to that of Mycobacterium marinum . Although mycolactone negative mutants are avirulent, they persist for at least 6 weeks. Chemical complementation of M. ulcerans mutants with mycolactone restores WT M. ulcerans pathology. Mycolactone negative mutants are capable of growth within macrophages in vitro whereas macrophages are killed by WT M. ulcerans . The ability of mycolactone to caused delayed cell death via apoptosis has been reported. However, mycolactone also causes cell death via necrosis. In vitro mycolactone has antiphagocytic properties. Neither WT M. ulcerans nor mycolactone negative strains are strong neutrophil attractants. These results suggest that mycolactone is largely responsible for the unique pathology produced by M. ulcerans .

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Mycobacterium ulcerans is a slow-growing, acid-fast bacillus that causes chronic necrotizing skin ulcers known as Buruli ulcers. Previously reported information on immunity to this mycobacterium is limited. We examined immune responses to M. ulcerans and M. bovis BCG in patients with M. ulcerans disease and in 20 healthy control subjects (10 tuberculin test positive and 10 tuberculin test negative). Cell-mediated immunity was assessed by stimulating peripheral blood mononuclear cells (PBMC) with whole mycobacteria and then measuring PBMC proliferation and the production of gamma interferon (IFN-␥). Humoral immunity was assessed by immunoblotting. PBMC from all subjects showed significantly greater proliferation and IFN-␥ production in response to stimulation with living mycobacteria compared with killed cells. However, PBMC from subjects with past or current M. ulcerans disease showed significantly reduced proliferation and production of IFN-␥ in response to stimulation with live M. ulcerans or M. bovis than PBMC from healthy, tuberculin test-positive subjects (P < 0.001) and showed results in these assays comparable to those of tuberculin testnegative subjects (P > 0.2). Serum from 9 of 11 patients with M. ulcerans disease, but no control subject, contained antibodies to M. ulcerans. The results indicate that patients with M. ulcerans infection mount an immune response to M. ulcerans as evidenced by antibody production, but they demonstrate profound systemic T-cell anergy to mycobacterial antigens. These findings may explain some of the distinct clinical and pathological features of M. ulcerans-induced disease.Mycobacterium ulcerans is the third most frequent cause of mycobacterial infections in immunocompetent individuals, after M. tuberculosis and M. leprae (18). M. ulcerans-induced disease generally manifests itself as indolent cutaneous ulcers, known as Buruli ulcers, accompanied by minimal systemic symptoms. Unlike other mycobacterial diseases, infections with M. ulcerans are characterized by extracellular bacteria, a lack of inflammatory cells, and extensive tissue necrosis at the site of infection (18). The indolent course, prominent extracellular bacteria, paucity of mononuclear cell infiltrate, and lack of systemic symptoms suggest a depressed or absent T-cell immune response.There are no published studies on the in vitro immune response of subjects with M. ulcerans infection. Although some individuals demonstrate a delayed-type hypersensitivity response to an extract of M. ulcerans (burulin) on skin testing, indicating a degree of T-cell sensitization, the close correlation between reactors to burulin and purified protein derivative suggests that sensitization is due to cross-reactivity with other mycobacterial species (17).A soluble lipid product, called mycolactone, produced by M. ulcerans produces cutaneous histological lesions in guinea pigs similar to those observed in patients with M. ulcerans disease (5). Mycolactone also exhibits immunosuppressive properties in vitro, manifested by suppression...