Spontaneous Healing of Mycobacterium ulcerans Lesions in the Guinea Pig Model

Silva-Gomes, Rita; Marcq, Elly; Trigo, Gabriela; Gonçalves, Carine M.; Longatto‐Filho, Adhemar; Castro, António G.; Pedrosa, Jorge; Fraga, Alexandra G.

doi:10.1371/journal.pntd.0004265

Cited by 19 publications

(23 citation statements)

References 36 publications

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“…This lower dose is likely to be more physiologically relevant, in terms of reflecting the bacterial inoculum that occurs during M. ulcerans transmission to humans (10, 11, 59). Sporadic healing of BU disease was also seen in this model, an observation that has been noted in humans and other animals (29, 66-68).…”

Section: Discussionsupporting

confidence: 80%

Vaccine-specific immune responses againstMycobacterium ulceransinfection in a low-dose murine challenge model

Mangas

Buultjens

Porter

et al. 2019

Preprint

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25The neglected tropical disease Buruli ulcer (BU) is an infection of subcutaneous tissue with 26 Mycobacterium ulcerans. There is no effective BU vaccine. Here, we assessed an experimental 27 prime-boost vaccine in a low-dose murine tail infection model. We used the enoyl-reductase (ER) 28 domain of the M. ulcerans mycolactone polyketide synthases electrostatically coupled with a 29 previously described TLR-2 agonist-based lipopeptide adjuvant, R4Pam2Cys. Mice were vaccinated 30 and then challenged via tail inoculation with 14-20 colony forming units (CFU) of an engineered 31 bioluminescent strain of M. ulcerans. Mice receiving either the experimental ER vaccine or 32Mycobacterium bovis Bacille Calmette-Guérin (BCG) were equally well protected, with both groups 33 faring significantly better than non-vaccinated animals (p<0.05). A suite of 29 immune parameters 34 were assessed in the mice at the end of the experimental period. Multivariate statistical approaches 35were then used to interrogate the immune response data to develop disease-prognostic models. 36High levels of IL-2 and low IFN-! produced in the spleen best predicted control of infection across 37 all vaccine groups. Univariate logistic regression then revealed vaccine-specific profiles of 38 protection. High titres of ER-specific IgG serum antibodies together with IL-2 and IL-4 in the draining 39 lymph node (DLN) were associated with protection induced by the experimental ER vaccine. In 40 contrast, high titres of IL-6, TNF-", IFN-! and IL-10 in the DLN and low IFN! titres in the spleen were 41 associated with protection following BCG vaccination. This study suggests an effective BU vaccine 42 must induce localized, tissue-specific immune profiles with controlled inflammatory responses at 43 the site of infection. 44 45 46 47 Results: 121 Formulation of the ER-R4Pam2Cys subunit vaccine candidate 122Mycolactone is the key virulence factor produced by M. ulcerans and an attractive vaccine target, 123 but the molecule is poorly immunogenic (55). However, the PKS enzymes used by the bacterium to 124 synthesize mycolactones -are highly conserved and immunogenic (49,(56)(57)(58). Therefore, we 125 hypothesized that targeting the conserved enzymatic domains of the mycolactone PKS could be an 126 effective vaccine strategy. One domain in particular, the enoyl reductase (ER) protein domain, elicits 127 serum antibodies in BU patients and healthy controls in BU endemic regions (57). The ER protein 128 expressed as an antigen in a DNA-protein prime-boost vaccine has also been shown to reduce 129 bacterial burden in a mouse footpad M. ulcerans challenge model (49). Here, we utilised the ER 130 protein to create a novel BU vaccine candidate by electrostatically associating it with the TLR-2 131 agonist-based lipopeptide R4Pam2Cys. To formulate this vaccine, recombinant 6xHis-tagged ER 132 protein (37 kDa) was first produced and confirmed by SDS-PAGE ( Fig. 1A) and western blotting (Fig. 133 1B). This protein antigen was formulated with R4Pam2Cys at various rat...

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Section: Discussionsupporting

confidence: 80%

Vaccine-specific immune responses againstMycobacterium ulceransinfection in a low-dose murine challenge model

Mangas

Buultjens

Porter

et al. 2019

Preprint

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“…In humans, serological screening for Hsp18 antibodies indicates that large parts of the population in endemic areas are exposed to M. ulcerans but only some develop the disease [59]. Guinea pigs infected with M. ulcerans appear to self-heal as do some mice [60]. It is conceivable that humans infected with certain doses of M. ulcerans develop either no disease, limited disease, or even unnoticed disease that self-resolves.…”

Section: Resultsmentioning

confidence: 99%

Bioluminscent Mycobacterium ulcerans, a tool to study host-pathogen interactions in a murine tail model of Buruli ulcer

Omansen

Marcsisin

Chua

et al. 2018

Preprint

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25 Buruli ulcer is a neglected tropical disease caused by infection with Mycobacterium 26 ulcerans. In this study we used a previously reported strain of M. ulcerans, genetically 27 engineered to constitutively produce bioluminescence, to follow the progression of Buruli 28 ulcer in mice using an in-vivo imaging (IVIS®) system. We aimed to characterize a mouse 29 tail infection model for pathogenesis, as well as for pre-clinical vaccine and drug 30 development research for Buruli ulcer. Immune parameters, such as antibody titers and 31 cytokine levels, were determined throughout the course of the infection and histology 32 specimens were examined for comparison with human pathology. Nine out of ten (90%) 33 BALB/c mice infected subcutaneously with 10 5 M. ulcerans JKD8049 (containing 34 pMV306 hsp16+luxG13) exhibited light emission from the site of infection over the course 35 of the experiment indicating M. ulcerans growth in-vivo. Five out of ten (50%) animals 36 developed clinical signs of disease. Antibody titers were overall low and their onset was 37 late, as measured by responses to both heterogenous (bacterial whole cell lysate) and single 38 antigen (Hsp18) targets. IFN-γ, and IL-10 are reported to play a vital role in host control 39 of Buruli ulcer and these cytokines were elevated in animals with pathology. For mice with 40 advanced pathology, histology revealed clusters of acid-fast bacilli within subcutaneous 41 tissue 300-400 μm beneath the epidermis of the tail, with macrophage infiltration and 42 granuloma-formation resembling human Buruli ulcer. This study has shown the utility of 43 using bioluminescent M. ulcerans and IVIS® in a mouse tail infection model to study 44 Buruli ulcer infection.45 3 46 Author summary 47 Buruli ulcer is one of the so called neglected tropical diseases. It is an infectious disease, 48 mainly occurring in West Africa but also in Australia. It manifests as skin lesion and ulcer.49 Up to date, the way of transmission is inadequately understood. Also, there is no vaccine 50 to protect against the disease. Buruli ulcer is treatable with a course of antibiotics that need 51 to be given for the duration of two months. More laboratory research is needed to elucidate 52 the mechanism of transmission, develop a vaccine and improve and shorten antibiotic 53 therapy. For this, animal (mouse) models of disease are used. The aim of this study was to 54 refine and improve the mouse tail infection model of Buruli ulcer. For this, we used a 55 genetically modified Mycobacterium ulcerans strain that emits light. After infection of 56 animals, light emitted from the bacteria was read out with an in-vivo imaging (IVIS) 57 camera. This allowed us to monitor the location of bacteria in the living animal over time 58 without the need to kill the animal. We also measured parameters of the immune system 59 such as antibodies and cytokines as a baseline for future studies into immunology, vaccine 60 development and pathology of Buruli ulcer. We successfully improved and characterized 61 the m...

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“…What causes lie between these two extremes? Two recent studiesaimed to shed some light on this question, by exploring the ability of the Hartley guinea pigs and the FVB/N mice to spontaneously heal BU [21,184]. Strikingly, whereas the guinea pigs were able to achieve sterilizing immunity, FVB/N mice appear to halt the production of mycolactone by M. ulcerans without being able to completely eliminate the pathogen [21,184].…”

Section: Novel Models For Investigation and Future Perspectivesmentioning

confidence: 99%

“…Two recent studiesaimed to shed some light on this question, by exploring the ability of the Hartley guinea pigs and the FVB/N mice to spontaneously heal BU [21,184]. Strikingly, whereas the guinea pigs were able to achieve sterilizing immunity, FVB/N mice appear to halt the production of mycolactone by M. ulcerans without being able to completely eliminate the pathogen [21,184]. Considering for instance, that changes in the sugar content of the growth medium of M. ulcerans can affect its production of mycolactone and antigens, this raises the hypothesis whether FVB/N mice deal with the infection by manipulating the local environment [185].…”

Section: Novel Models For Investigation and Future Perspectivesmentioning

confidence: 99%

“…Considering for instance, that changes in the sugar content of the growth medium of M. ulcerans can affect its production of mycolactone and antigens, this raises the hypothesis whether FVB/N mice deal with the infection by manipulating the local environment [185]. It is intriguing to observe that both models control the infection regardless of the initial inoculum, which most likely suggests that there are key host factors that can effectively hamper the progression of the infection and that have never been so far considered as preventive or therapeutic approaches [21,184]. Hence, it is highly likely that answers to this conundrum lie within the genetic makeup of BU patients that spontaneously recover from the disease, but with the current widespread use of antibiotics, the task of identifying such cases is extremely difficult.…”

Section: Novel Models For Investigation and Future Perspectivesmentioning

confidence: 99%

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The Immunology of Buruli Ulcer

2019

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Buruli ulcer (BU) represents a unique human mycobacteriosis. Caused by Mycobacterium ulcerans, the disease spectrum is dominated by the activity of mycolactone, a dermanecrotic toxin that has shown the ability to interfere with the immune response. This poses an additional difficulty to the understanding of the immunological determinants for the outcome of infection, a fundamental step to develop better preventive or curative strategies. In this chapter, the immune response against M. ulcerans is reviewed, both with a series of clinical observations and experimental infection models and going through several other lines of evidence, including epidemiological and genetic studies. This holistic approach is expected to shed further light on the intriguing pathophysiology of this disease and help guide future research efforts.

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Spontaneous Healing of Mycobacterium ulcerans Lesions in the Guinea Pig Model

Cited by 19 publications

References 36 publications

Vaccine-specific immune responses againstMycobacterium ulceransinfection in a low-dose murine challenge model

Vaccine-specific immune responses againstMycobacterium ulceransinfection in a low-dose murine challenge model

Bioluminscent Mycobacterium ulcerans, a tool to study host-pathogen interactions in a murine tail model of Buruli ulcer

The Immunology of Buruli Ulcer

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