Ketogenic Diet Impairment of Mycobacterium ulcerans Growth and Toxin Production and Enhancement of Host Response to Infection in an Experimental Mouse Model.

Ketogenic diets have been used to treat diverse conditions, and there is growing evidence of their benefits for tissue repair and in inflammatory disease treatment. However, their role in infectious diseases has been little studied. Buruli ulcer (Mycobacterium ulcerans infection) is a chronic infectious disease characterized by large skin ulcerations caused by mycolactone, the major virulence factor of the bacillus. In the current study, we investigated the impact of ketogenic diet on this cutaneous disease in an experimental mouse model. This diet prevented ulceration, by modulating bacterial growth and host inflammatory response. ß-hydroxybutyrate, the major ketone body produced during ketogenic diet and diffusing in tissues, impeded M. ulcerans growth and mycolactone production in vitro underlying its potential key role in infection. These results pave the way for the development of new patient management strategies involving shorter courses of treatment and improving wound healing, in line with the major objectives of the World Health Organization.

Purification and Characterization of Extracellular Vesicles from Mycobacterium ulcerans Culture.

Extracellular vesicles (EVs) from both eukaryotic and prokaryotic cells have been characterized over decades and present many biological properties. Since it has been shown that mycobacterial extracellular vesicles (MEVs) of M. ulcerans contain the macrolide toxin mycolactone, MEVs are known to be associated with the pathogenesis of mycobacteria. This chapter describes a method for purifying and characterizing vesicles from in vitro cultures of M. ulcerans. We also describe how purified vesicles can be used in cellular tests, to determine their role in the pathophysiology of M. ulcerans infection.

Transcriptional adaptation of Mycobacterium ulcerans in an original mouse model: New insights into the regulation of mycolactone.

Mycobacterium ulcerans is the causal agent of Buruli ulcer, a chronic infectious disease and the third most common mycobacterial disease worldwide. Without early treatment, M. ulcerans provokes massive skin ulcers, caused by the mycolactone toxin, its main virulence factor. However, spontaneous healing may occur in Buruli ulcer patients several months or years after the disease onset. We have shown, in an original mouse model, that bacterial load remains high and viable in spontaneously healed tissues, with a switch of M. ulcerans to low levels of mycolactone production, adapting its strategy to survive in such a hostile environment. This original model offers the possibility to investigate the regulation of mycolactone production, by using an RNA-seq strategy to study bacterial adaptation during mouse infection. Pathway analysis and characterization of the tissue environment showed that the bacillus adapted to its new environment by modifying its metabolic activity and switching nutrient sources. Thus, M. ulcerans ensures its survival in healing tissues by reducing its secondary metabolism, leading to an inhibition of mycolactone synthesis. These findings shed new light on mycolactone regulation and pave the way for new therapeutic strategies.

Skin-specific antibodies neutralizing mycolactone toxin during the spontaneous healing of Mycobacterium ulcerans infection.

Buruli ulcer, a neglected tropical infectious disease, is caused by Mycobacterium ulcerans. Without treatment, its lesions can progress to chronic skin ulcers, but spontaneous healing is observed in 5% of cases, suggesting the possible establishment of a host strategy counteracting the effects of M. ulcerans. We reveal here a skin-specific local humoral signature of the spontaneous healing process, associated with a rise in antibody-producing cells and specific recognition of mycolactone by the mouse IgG2a immunoglobulin subclass. We demonstrate the production of skin-specific antibodies neutralizing the immunomodulatory activity of the mycolactone toxin, and confirm the role of human host machinery in triggering effective local immune responses by the detection of anti-mycolactone antibodies in patients with Buruli ulcer. Our findings pave the way for substantial advances in both the diagnosis and treatment of Buruli ulcer in accordance with the most recent challenges issued by the World Health Organization.

Lactobacillus paracasei CNCM I-3689 reduces vancomycin-resistant Enterococcus persistence and promotes Bacteroidetes resilience in the gut following antibiotic challenge.

Enterococci, in particular vancomycin-resistant enterococci (VRE), are a leading cause of hospital-acquired infections. Promoting intestinal resistance against enterococci could reduce the risk of VRE infections. We investigated the effects of two Lactobacillus strains to prevent intestinal VRE. We used an intestinal colonisation mouse model based on an antibiotic-induced microbiota dysbiosis to mimic enterococci overgrowth and VRE persistence. Each Lactobacillus spp. was administered daily to mice starting one week before antibiotic treatment until two weeks after antibiotic and VRE inoculation. Of the two strains, Lactobacillus paracasei CNCM I-3689 decreased significantly VRE numbers in the feces demonstrating an improvement of the reduction of VRE. Longitudinal microbiota analysis showed that supplementation with L. paracasei CNCM I-3689 was associated with a better recovery of members of the phylum Bacteroidetes. Bile salt analysis and expression analysis of selected host genes revealed increased level of lithocholate and of ileal expression of camp (human LL-37) upon L. paracasei CNCM I-3689 supplementation. Although a direct effect of L. paracasei CNCM I-3689 on the VRE reduction was not ruled out, our data provide clues to possible anti-VRE mechanisms supporting an indirect anti-VRE effect through the gut microbiota. This work sustains non-antibiotic strategies against opportunistic enterococci after antibiotic-induced dysbiosis.