Mycobacterium ulcerans is an emerging environmental pathogen which causes chronic skin ulcers (i.e., Buruli ulcer) in otherwise healthy humans living in tropical countries, particularly those in Africa. In spite of epidemiological and PCR data linking M. ulcerans to water, the mode of transmission of this organism remains elusive. To determine the role of aquatic insects in the transmission of M. ulcerans, we have set up an experimental model with aquariums that mimic aquatic microenvironments. We report that M. ulcerans may be transmitted to laboratory mice by the bite of aquatic bugs (Naucoridae) that are infected with this organism. In addition, M. ulcerans appears to be localized exclusively within salivary glands of these insects, where it can both survive and multiply without causing any observable damage in the insect tissues. Subsequently, we isolated M. ulcerans from wild aquatic insects collected from a zone in the Daloa region of Ivory Coast where Buruli ulcer is endemic. Taken together, these results point to aquatic insects as a possible vector of M. ulcerans.
Dermatophytes are keratinolytic fungi responsible for a large variety of diseases that can affect glabrous skin, nails and hair. In many cases, the diagnosis is not clinically obvious, and mycological analysis is required. This includes both direct microscopic examination and cultures. First of all, clinical specimens have to be sampled according to localization and characteristics of the lesions. Direct microscopic examination is usually performed using clearing reagents (KOH or Amman's chloral-lactophenol), but its sensitivity may be greatly enhanced by the use of stains or fluorochromes such as Congo red or Calcofluor white. Histological analysis is an efficient method, but it is constraining for the patients and, as direct examination, it does not allow precise identification of the pathogen. Cultures are therefore needed, and specific culture media may be used to overcome the growth of rapidly growing contaminating moulds which may hamper the recovery of dermatophytes. Identification at the species level which may be useful to initiate an appropriate treatment or for setting prophylactic measures, relies on macroscopic and microscopic morphology. Subcultures on culture media which stimulate conidiation and, for some species, the production of pigments, are often necessary. Additionally, in case of atypical isolates, some biochemical or physiological tests may be performed such as the search for urease activity or the in vitro hair perforation test. However, their contribution to species identification is rather limited, and progress is still needed for the development of biochemical or immunological tests allowing an accurate identification at the species level, pending for the availability of molecular biology-based kits.
Mycobacterium ulcerans is the causative agent of Buruli ulcer, one of the most common mycobacterial diseases of humans. Recent studies have implicated aquatic insects in the transmission of this pathogen, but the contributions of other elements of the environment remain largely unknown. We report here that crude extracts from two green algae added to the BACTEC 7H12B culture medium halved the doubling time of M. ulcerans and promoted biofilm formation. Using the 7H12B medium, modified by the addition of the algal extract, and immunomagnetic separation, we also demonstrate that M. ulcerans is associated with aquatic plants in an area of the Ivory Coast where Buruli ulcer is endemic. Genotype analysis showed that plant-associated M. ulcerans had the same profile as isolates recovered in the same region from both aquatic insects and clinical specimens. These observations implicate aquatic plants as a reservoir of M. ulcerans and add a new potential link in the chain of transmission of M. ulcerans to humans.Mycobacterium ulcerans is an emerging environmental pathogen (2). It is the etiologic agent of Buruli ulcer, a necrotic skin disease highly prevalent in many countries throughout west Africa and one of the most common mycobacterial diseases in humans after tuberculosis and leprosy (2). M. ulcerans is the only mycobacterium known to produce a toxin, a polyketide-derived macrolide called mycolactone (10, 11). The infection begins with a painless nodule or papule that spreads over the surrounding tissue. Ischemic and necrotized tissue disappears and is replaced by centralized ulceration of the limb. At present, the only effective treatment consists of excision of the lesions, and this is often followed by extensive skin grafting (6).Until very recently M. ulcerans had never been isolated in culture from the environment. Indirect evidence from epidemiological studies suggests that M. ulcerans is an environmental mycobacterium present in swampy areas. Humans are thought to be infected through minor wounds or skin abrasions via contact with mycobacterium-containing water (33). In a previous study we demonstrated that aquatic insects are a possible mode of transmission of M. ulcerans to humans. We showed that, in an experimental mouse model, aquatic insects (Naucoridae) were able to transmit an infection by biting, thereby inoculating bacilli that had accumulated in the salivary glands of the insects (16). We were then able to isolate in pure culture M. ulcerans from the salivary glands of Naucoridae captured in a region of endemicity in the Ivory Coast. However it seems unlikely that these insects are the only environmental source of M. ulcerans. In another recent study, data gathered using an M. ulcerans-specific PCR to survey environments of endemicity in southeastern Australia identified aquatic plants as a possible reservoir of this pathogen (30).In this report, we test the hypothesis that M. ulcerans is associated with aquatic plants by studying the effects of crude aquatic plant extracts on the growth of M. ulcer...
Recent evidence for the role of laminin in cell adhesion and in the pathogenesis of several bacterial infections has led us to investigate the existence of receptors for this extracellular matrix component in Candida albicans. At first, immunofluorescence demonstrated the presence of laminin-binding sites at the surface of germ tubes. Electron microscopy confirmed this result and permitted precise localization of the binding sites on the outermost fibrillar layer of the germ tube cell wall. By using 1251-radiolabeled laminin, the binding was shown to be saturable and specific, hence demonstrating characteristics of true receptors. Analysis of the data by the Scatchard equation indicated that there were about 8,000 binding sites per cell, with a dissociation constant 48
Germ tubes of Candida albicans produced an additional fibrillar surface layer responsible for enhanced adherence to plastic. The correlation between germination of C. albicans and adherence of germ tubes to a plastic matrix led us to consider the existence of germ tube-specific adhesive components involved in the attachment process. Using concanavalin A-sensitized latex microspheres, we first detected extracellular molecules on the plastic surface after removal of the adherent germ tubes. Electron microscopy confirmed that fibrils of the germ tube involved in cell-substratum interconnections were retained on the plastic surface. Cytochemistry with concanavalin A-gold labeling demonstrated that these fibrillar structures contained mannoproteins. Dithiothreitol and iodoacetamide treatment of washed plastic allowed us to further characterize these fibrillar adhesins. Through analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, two components with molecular weights (MWs) of 68,000 and 60,000 were detected on the plastic surface. The 68,000-MW component appeared to be one of the major constituents of the germ tube surface layers. Biosynthetic labeling experiments performed with L-[35S]methionine revealed two additional proteins: a high-MW component (greater than 200,000), and a 200,000-MW component. These four proteins, strongly labeled on the plastic surface and on the germ tube cell wall layers, were in contrast slightly labeled or even nonidentified in the culture supernatant, suggesting their involvement in germ tube adherence.
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