Deciphering and Imaging Pathogenesis and Cording of &lt;em&gt;Mycobacterium abscessus&lt;/em&gt; in Zebrafish Embryos

Bernut, Audrey; Dupont, Christian; Sahuquet, Alain; Herrmann, Jean-Louis; Lutfalla, Georges; Kremer, Laurent

doi:10.3791/53130

Cited by 54 publications

(81 citation statements)

References 41 publications

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“…Making targeted genetic deletions are particularly challenging in the R variant due to its extremely high aggregative properties. Herein, we combined the larval zebrafish as a tractable host to study the innate immune response to Mabs (16,17) and the successful achievement of the first deletion mutant in the R background. We demonstrate that the recombineering method (31) can be successfully applied to inactivate MAB_4780 in the R variant, leading to multiple and remarkable in vitro and in vivo phenotypes.…”

Section: Discussionmentioning

confidence: 99%

“…bolletii (15), a subspecies of the Mabs complex. We recently used the zebrafish embryo as a tractable infection model to study, at a spatiotemporal level, the physiopathology of Mabs infection and demonstrated the high propensity of virulent R variant Mabs to produce cords in vivo (16,17). We showed that extracellular cording is an example of morphological plasticity allowing bacteria to escape the host immune system.…”

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confidence: 99%

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Deletion of a dehydratase important for intracellular growth and cording renders rough Mycobacterium abscessus avirulent

Halloum

Carrère-Kremer

Blaise

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Mycobacterium abscessus (Mabs) is a rapidly growing Mycobacterium and an emerging pathogen in humans. Transitioning from a smooth (S) high-glycopeptidolipid (GPL) producer to a rough (R) low-GPL producer is associated with increased virulence in zebrafish, which involves the formation of massive serpentine cords, abscesses, and rapid larval death. Generating a cord-deficient Mabs mutant would allow us to address the contribution of cording in the physiopathological signs of the R variant. Herein, a deletion mutant of MAB_4780, encoding a dehydratase, distinct from the β-hydroxyacyl-ACP dehydratase HadABC complex, was constructed in the R morphotype. This mutant exhibited an alteration of the mycolic acid composition and a pronounced defect in cording. This correlated with an extremely attenuated phenotype not only in wild-type but also in immunocompromised zebrafish embryos lacking either macrophages or neutrophils. The abolition of granuloma formation in embryos infected with the dehydratase mutant was associated with a failure to replicate in macrophages, presumably due to limited inhibition of the phagolysosomal fusion. Overall, these results indicate that MAB_4780 is required for Mabs to successfully establish acute and lethal infections. Therefore, targeting MAB_4780 may represent an attractive antivirulence strategy to control Mabs infections, refractory to most standard chemotherapeutic interventions. The combination of a dehydratase assay with a highresolution crystal structure of MAB_4780 opens the way to identify such specific inhibitors.M. abscessus | zebrafish | cording | dehydratase | virulence

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Deletion of a dehydratase important for intracellular growth and cording renders rough Mycobacterium abscessus avirulent

Halloum

Carrère-Kremer

Blaise

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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“…abscessus sensu stricto strain CIP104536 T (ATCC19977T) carrying pTEC15 (Addgene, plasmid 30174), pTEC27 (Addgene, plasmid 30182) or pTEC19 (Addgene, plasmid 30178) that express green fluorescent protein (Wasabi), red fluorescent protein (tdTomato) or bright far-red fluorescent protein (E2-Crimson), respectively, were prepared and microinjected in zebrafish embryos, according to procedures described earlier [17,55]. Briefly, systemic infections were carried by the injection of 100–200 CFU into the caudal vein of 30 hours post-fertilization (hpf) embryos.…”

Section: Methodsmentioning

confidence: 99%

“…To generate neutrophil depleted-embryos, csf3r translation morpholino (5’-GAAGCACAAGCGAGACGGATGCCA T-3’ ) targeting the csf3r gene was used [31]. For the selective depletion of macrophages into embryos, lipo-clodronate (Lipo-C) [56] was injected into the caudal vein of 24 hpf embryos as previously reported [17,55]. …”

Section: Methodsmentioning

confidence: 99%

Mycobacterium abscessus-Induced Granuloma Formation Is Strictly Dependent on TNF Signaling and Neutrophil Trafficking

et al. 2016

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Mycobacterium abscessus is considered the most common respiratory pathogen among the rapidly growing non-tuberculous mycobacteria. Infections with M. abscessus are increasingly found in patients with chronic lung diseases, especially cystic fibrosis, and are often refractory to antibiotic therapy. M. abscessus has two morphotypes with distinct effects on host cells and biological responses. The smooth (S) variant is recognized as the initial airway colonizer while the rough (R) is known to be a potent inflammatory inducer associated with invasive disease, but the underlying immunopathological mechanisms of the infection remain unsolved. We conducted a comparative stepwise dissection of the inflammatory response in S and R pathogenesis by monitoring infected transparent zebrafish embryos. Loss of TNFR1 function resulted in increased mortality with both variants, and was associated with unrestricted intramacrophage bacterial growth and decreased bactericidal activity. The use of transgenic zebrafish lines harboring fluorescent macrophages and neutrophils revealed that neutrophils, like macrophages, interact with M. abscessus at the initial infection sites. Impaired TNF signaling disrupted the IL8-dependent neutrophil mobilization, and the defect in neutrophil trafficking led to the formation of aberrant granulomas, extensive mycobacterial cording, unrestricted bacterial growth and subsequent larval death. Our findings emphasize the central role of neutrophils for the establishment and maintenance of the protective M. abscessus granulomas. These results also suggest that the TNF/IL8 inflammatory axis is necessary for protective immunity against M. abscessus and may be of clinical relevance to explain why immunosuppressive TNF therapy leads to the exacerbation of M. abscessus infections.

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“…The increasing success of this vertebrate model relies on unique advantages that motivated its use to increase our knowledge of many viral and bacterial infections (Davis et al, 2002; Prajsnar et al, 2008; Phennicie et al, 2010; Alibaud et al, 2011; Mostowy et al, 2013; Palha et al, 2013). Zebrafish embryos have also recently been exploited to visualize, by non-invasive imaging, the progression and fate of Mabs infection, allowing to observe host-pathogen interactions in a live animal at a high resolution level (Bernut et al, 2015; Figure 1). Methods were specifically adapted for assessing Mabs virulence by measuring distinct parameters, such as embryo survival and bacterial burden and by monitoring the chronology of the infection using video microscopy.…”

Section: Introductionmentioning

confidence: 99%

The Diverse Cellular and Animal Models to Decipher the Physiopathological Traits of Mycobacterium abscessus Infection

Bernut

Herrmann

Ordway

et al. 2017

Front. Cell. Infect. Microbiol.

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Mycobacterium abscessus represents an important respiratory pathogen among the rapidly-growing non-tuberculous mycobacteria. Infections caused by M. abscessus are increasingly found in cystic fibrosis (CF) patients and are often refractory to antibiotic therapy. The underlying immunopathological mechanisms of pathogenesis remain largely unknown. A major reason for the poor advances in M. abscessus research has been a lack of adequate models to study the acute and chronic stages of the disease leading to delayed progress of evaluation of therapeutic efficacy of potentially active antibiotics. However, the recent development of cellular models led to new insights in the interplay between M. abscessus with host macrophages as well as with amoebae, proposed to represent the environmental host and reservoir for non-tuberculous mycobacteria. The zebrafish embryo has also appeared as a useful alternative to more traditional models as it recapitulates the vertebrate immune system and, due to its optical transparency, allows a spatio-temporal visualization of the infection process in a living animal. More sophisticated immunocompromised mice have also been exploited recently to dissect the immune and inflammatory responses to M. abscessus. Herein, we will discuss the limitations, advantages and potential offered by these various models to study the pathophysiology of M. abscessus infection and to assess the preclinical efficacy of compounds active against this emerging human pathogen.

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Deciphering and Imaging Pathogenesis and Cording of <em>Mycobacterium abscessus</em> in Zebrafish Embryos

Cited by 54 publications

References 41 publications

Deletion of a dehydratase important for intracellular growth and cording renders rough Mycobacterium abscessus avirulent

Deletion of a dehydratase important for intracellular growth and cording renders rough Mycobacterium abscessus avirulent

Mycobacterium abscessus-Induced Granuloma Formation Is Strictly Dependent on TNF Signaling and Neutrophil Trafficking

The Diverse Cellular and Animal Models to Decipher the Physiopathological Traits of Mycobacterium abscessus Infection

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