Cellular Immunity Confers Transient Protection in Experimental Buruli Ulcer following BCG or Mycolactone-Negative Mycobacterium ulcerans Vaccination

Fraga, Alexandra G.; Martins, Teresa G.; Torrado, Egídio; Huygen, Kris; Portaels, Françoise; Silva, Manuel T.; Castro, António G.; Pedrosa, Jorge

doi:10.1371/journal.pone.0033406

Cited by 39 publications

(52 citation statements)

References 45 publications

(79 reference statements)

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“…As previously described [38]–[40], we found that M. ulcerans disseminates to the DLN after footpad infection (Figure 1C ), probably due to continuous lymphatic dissemination of bacteria either freely or shuttled within phagocytes. Here we show a significant reduction in CFU counts (P<0.05) in the DLN of mycobacteriophage D29 treated mice, as compared with non-treated counterparts, at day 68 post-infection (day 35 post-treatment) (Figure 1C), correlating with the reduction of M. ulcerans numbers in the footpads.…”

Section: Resultssupporting

confidence: 86%

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Phage Therapy Is Effective against Infection by Mycobacterium ulcerans in a Murine Footpad Model

et al. 2013

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BackgroundBuruli Ulcer (BU) is a neglected, necrotizing skin disease caused by Mycobacterium ulcerans. Currently, there is no vaccine against M. ulcerans infection. Although the World Health Organization recommends a combination of rifampicin and streptomycin for the treatment of BU, clinical management of advanced stages is still based on the surgical resection of infected skin. The use of bacteriophages for the control of bacterial infections has been considered as an alternative or to be used in association with antibiotherapy. Additionally, the mycobacteriophage D29 has previously been shown to display lytic activity against M. ulcerans isolates.Methodology/Principal findingsWe used the mouse footpad model of M. ulcerans infection to evaluate the therapeutic efficacy of treatment with mycobacteriophage D29. Analyses of macroscopic lesions, bacterial burdens, histology and cytokine production were performed in both M. ulcerans-infected footpads and draining lymph nodes (DLN). We have demonstrated that a single subcutaneous injection of the mycobacteriophage D29, administered 33 days after bacterial challenge, was sufficient to decrease pathology and to prevent ulceration. This protection resulted in a significant reduction of M. ulcerans numbers accompanied by an increase of cytokine levels (including IFN-γ), both in footpads and DLN. Additionally, mycobacteriophage D29 treatment induced a cellular infiltrate of a lymphocytic/macrophagic profile.Conclusions/SignificanceOur observations demonstrate the potential of phage therapy against M. ulcerans infection, paving the way for future studies aiming at the development of novel phage-related therapeutic approaches against BU.

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Section: Resultssupporting

confidence: 86%

“…As previously described [38]–[40] and confirmed in this study, the tissue destruction of the DLN was associated with bacterial colonization, which is consistent with the spreading of M. ulcerans from the site of infection via afferent lymphatic drainage [55], [56] …”

Section: Discussionsupporting

confidence: 90%

Phage Therapy Is Effective against Infection by Mycobacterium ulcerans in a Murine Footpad Model

et al. 2013

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“…Anti-mycobacterial immunity is largely governed by responses from CD4 + T helper cells [40, 41]. In order to determine if antigen-specific adaptive immune responses could be generated by vaccination with BCG-MU-Ag85B-EsxH, C57BL/6 mice were either subcutaneously or intravenously or primed with 10 7 bacilli from thawed quality controlled vaccine lots prepared as previously described [39].…”

Section: Resultsmentioning

confidence: 99%

“…There is experimental evidence to support this strategy, whereby subcutaneous vaccination with the mycolactone negative strain, MU5114, yielded a delay to footpad swelling similar to that induced by BCG vaccination [41]. Furthermore, species of mycobacteria possessing greater genetic homology to MU than BCG may represent a richer source of protective antigens and lack the potential pathological features of an MU-based vaccine.…”

Section: Discussionmentioning

confidence: 99%

Overexpression of a Mycobacterium ulcerans Ag85B-EsxH Fusion Protein in Recombinant BCG Improves Experimental Buruli Ulcer Vaccine Efficacy

Hart

Lee

2016

PLoS Negl Trop Dis

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Buruli ulcer (BU) vaccine design faces similar challenges to those observed during development of prophylactic tuberculosis treatments. Multiple BU vaccine candidates, based upon Mycobacterium bovis BCG, altered Mycobacterium ulcerans (MU) cells, recombinant MU DNA, or MU protein prime-boosts, have shown promise by conferring transient protection to mice against the pathology of MU challenge. Recently, we have shown that a recombinant BCG vaccine expressing MU-Ag85A (BCG MU-Ag85A) displayed the highest level of protection to date, by significantly extending the survival time of MU challenged mice compared to BCG vaccination alone. Here we describe the generation, immunogenicity testing, and evaluation of protection conferred by a recombinant BCG strain which overexpresses a fusion of two alternative MU antigens, Ag85B and the MU ortholog of tuberculosis TB10.4, EsxH. Vaccination with BCG MU-Ag85B-EsxH induces proliferation of Ag85 specific CD4+ T cells in greater numbers than BCG or BCG MU-Ag85A and produces IFNγ+ splenocytes responsive to whole MU and recombinant antigens. In addition, anti-Ag85A and Ag85B IgG humoral responses are significantly enhanced after administration of the fusion vaccine compared to BCG or BCG MU-Ag85A. Finally, mice challenged with MU following a single subcutaneous vaccination with BCG MU-Ag85B-EsxH display significantly less bacterial burden at 6 and 12 weeks post-infection, reduced histopathological tissue damage, and significantly longer survival times compared to vaccination with either BCG or BCG MU-Ag85A. These results further support the potential of BCG as a foundation for BU vaccine design, whereby discovery and recombinant expression of novel immunogenic antigens could lead to greater anti-MU efficacy using this highly safe and ubiquitous vaccine.

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“…M. ulcerans then kills the host macrophage by producing mycolactone, a lipid toxin, initiating an extracellular stage, in which local mycolactone concentrations increase considerably, leading to massive host tissue destruction. During these two stages, mycolactone is not only cytotoxic, it also modulates the immune system, modifying cytokine production and acting on the peripheral nervous system to induce the formation of a painless lesion (George et al, 1999; Coutanceau et al, 2005; Oliveira et al, 2005; Torrado et al, 2007, 2010; Silva et al, 2009; Fraga et al, 2010, 2012; Marion et al, 2014b). These pleiotropic effects of mycolactone facilitate host colonization by this bacillus.…”

Section: Introductionmentioning

confidence: 99%

An Optimized Method for Extracting Bacterial RNA from Mouse Skin Tissue Colonized by Mycobacterium ulcerans

et al. 2017

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Bacterial transcriptome analyses during host colonization are essential to decipher the complexity of the relationship between the bacterium and its host. RNA sequencing (RNA-seq) is a promising approach providing valuable information about bacterial adaptation, the host response and, in some cases, mutual tolerance underlying crosstalk, as recently observed in the context of Mycobacterium ulcerans infection. Buruli ulcer is caused by M. ulcerans. This neglected disease is the third most common mycobacterial disease worldwide. Without treatment, M. ulcerans provokes massive skin ulcers. A healing process may be observed in 5% of Buruli ulcer patients several months after the initiation of disease. This spontaneous healing process suggests that some hosts can counteract the development of the lesions caused by M. ulcerans. Deciphering the mechanisms involved in this process should open up new treatment possibilities. To this end, we recently developed the first mouse model for studies of the spontaneous healing process. We have shown that the healing process is based on mutual tolerance between the bacterium and its host. In this context, RNA-seq seems to be the most appropriate method for deciphering bacterial adaptation. However, due to the low bacterial load in host tissues, the isolation of mycobacterial RNA from skin tissue for RNA-seq analysis remains challenging. We developed a method for extracting and purifying mycobacterial RNA whilst minimizing the amount of host RNA in the sample. This approach was based on the extraction of bacterial RNA by a differential lysis method. The challenge in the development of this method was the choice of a lysis system favoring the removal of host RNA without damage to the bacterial cells. We made use of the thick, resistant cell wall of M. ulcerans to achieve this end.

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Cellular Immunity Confers Transient Protection in Experimental Buruli Ulcer following BCG or Mycolactone-Negative Mycobacterium ulcerans Vaccination

Cited by 39 publications

References 45 publications

Phage Therapy Is Effective against Infection by Mycobacterium ulcerans in a Murine Footpad Model

Phage Therapy Is Effective against Infection by Mycobacterium ulcerans in a Murine Footpad Model

Overexpression of a Mycobacterium ulcerans Ag85B-EsxH Fusion Protein in Recombinant BCG Improves Experimental Buruli Ulcer Vaccine Efficacy

An Optimized Method for Extracting Bacterial RNA from Mouse Skin Tissue Colonized by Mycobacterium ulcerans

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