Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2021 Dec;12(1):1438-1451.
doi: 10.1080/21505594.2021.1929749.

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

Marie Robbe-Saule  1 , Mélanie Foulon  1 , Isabelle Poncin  2 , Lucille Esnault  1 , Hugo Varet  3   4 , Rachel Legendre  3   4 , Alban Besnard  1 , Anna E Grzegorzewicz  5 , Mary Jackson  5 , Stéphane Canaan  2 , Laurent Marsollier  1 , Estelle Marion  1
Affiliations

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

Marie Robbe-Saule et al. Virulence. 2021 Dec.

Abstract

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.

Keywords: Mycobacterium ulcerans; host-bacterium interaction; metabolism; mycolactone; rna-sequencing.

PubMed Disclaimer

Figures

Figure 1.
Figure 1.
RNA-seq analysis of M. ulcerans in the FVB/N mouse spontaneous healing model. (a) MA-plot of the data for the comparison of the healing (n = 3 mouse) and edema (n = 5 mouse) stages. Differentially expressed genes, identified with a false discovery rate of < 0.05, are highlighted in red. (b) Heatmap of the 707 differentially expressed genes (DEGs) of M. ulcerans isolated at the healing and edema stages in a mouse model. The heatmap is based on the variance-stabilized transformed count matrix, the rows and columns of which were re-ordered by hierarchical clustering (using the correlation and Euclidean distances, respectively, and the Ward aggregation criterion). The color scale ranges from −2 to +2 as the rows of the matrix have been centered. (c) The pie chart shows the percentage of differentially expressed genes in each functional category as per the BuruList database. (d) The numbers of differentially expressed genes are shown by functional category, according to BuruList database. Circle sizes are proportional to the number of genes with significant differential expression (adjusted p-value < 0.05) between the edema and healing stages. The total number of genes per functional category listed in BuruList is indicated in parentheses. (e) Pathway enrichment analysis for genes that are induced (left panel) or repressed (right panel) during the healing stage. Negative log10(p-values) were calculated from the difference between the observed and expected numbers of induced or repressed genes annotated in the Biocyc database. Only significant pathways (Fisher’s exact test) with p-value < 0.05 are represented
Figure 2.
Figure 2.
Changes in the lipid-associated phenotype of M. ulcerans during spontaneous healing. (a) Three bacillus populations were observed by transmission electron microscopy (TEM) in glutaraldehyde-fixed mouse tissues at the edema and healing stages (n = 2 mouse/stage): M. ulcerans without ILI, M. ulcerans containing ILI and damaged M. ulcerans. (b) The proportion of M. ulcerans bacteria containing ILI increased during spontaneous healing, reaching 13% of total bacillus counts (n = 67), versus 0.8% at the edema stage (n = 118)
Figure 3.
Figure 3.
Regulation of mycolactone synthesis under nutrient variation is associated with an induction of lipid metabolism and stress response pathway genes. (a) The amount of mycolactone produced by M. ulcerans in vitro was 3.7 times higher in the presence of glucose. The results are expressed as mean of two independent experiments. (b) Validation of RNA-seq data with in vitro cultures, based on an analysis of gene expression. The expression of genes involved in the methylcitrate cycle, methylmalonyl/malonyl-CoA pathway, nutrient stress and mycolactone synthesis was compared between conditions of high levels of mycolactone production (edema in vivo and 0.2% glucose in vitro) and low levels of mycolactone production (healing stage in vivo and 0.2%acetate in vitro). The in vivo/in vitro results are expressed as fold induction/repression. For in vitro results, fold-changes in gene expression were calculated by comparison with M. ulcerans grown in the presence of glucose, for two independent experiments with two to four biological replicates. Statistical analyses comparing cultures in the presence of acetate and glucose culture were performed with nonparametric Mann–Whitney U tests. For in vivo and in vitro fold changes, significant differences are illustrated as *p < 0.05, **p < 0.01, ***p < 0.001
Figure 4.
Figure 4.
Lactate availability promotes changes in lipid metabolism of M. ulcerans. (a) An increase in lactate availability, and a simultaneous decrease in glucose availability are observed in tissues during spontaneous healing. The amounts of lactate and glucose are expressed per 1 g of tissue (n = 8 mice for each stage, *p-value<0.05, Mann–Whitney U test). (b) The amount of mycolactone produced by M. ulcerans in vitro was 3.5 times higher in the presence of glucose. The results are expressed as mean of two independent experiments. (c) An accumulation of ILIs were observed by transmission electron microscopy (TEM) in the presence of lactate. Black arrows indicate the ILIs in M. ulcerans cells
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

References

    1. Portaels F, Silva MT, Meyers WM.. Buruli ulcer. Clin Dermatol. 2009;27(3):291–305. - PubMed
    1. Marion E, Chauty A, Kempf M, et al. Clinical features of spontaneous partial healing during mycobacterium ulcerans Infection. Open Forum Infect Dis. 2016;3(1):ofw013. - PMC - PubMed
    1. Barogui Y, Johnson RC, Van Der Werf TS, et al. Functional limitations after surgical or antibiotic treatment for Buruli ulcer in Benin. Am J Trop Med Hyg. 2009;81(1):82–87. - PubMed
    1. Vincent QB, Ardant MF, Adeye A, et al. Clinical epidemiology of laboratory-confirmed Buruli ulcer in Benin: a cohort study. Lancet Glob Health. 2014;2(7):e422–30. - PubMed
    1. Wadagni AC, Barogui YT, Johnson RC, et al. Delayed versus standard assessment for excision surgery in patients with Buruli ulcer in Benin: a randomised controlled trial. Lancet Infect Dis. 2018;18(6):650–656. - PubMed

Publication types

Grants and funding

This work was supported by the Agence Nationale de la Recherche [Grant ANR-15-IFEC-0006]; Agence Nationale de la Recherche [Grant ANR- 16-CE12-0023]; Fondation pour la Recherche Médicale [Equipe FRM]; Region Pays de la Loire [Grant Starter]; Fondation Raoul Follereau; INSERM.