In Vivo Assessment of Drug Efficacy against Mycobacterium abscessus Using the Embryonic Zebrafish Test System

Abstract

c Mycobacterium abscessus is responsible for a wide spectrum of clinical syndromes and is one of the most intrinsically drug-resistant mycobacterial species. Recent evaluation of the in vivo therapeutic efficacy of the few potentially active antibiotics against M. abscessus was essentially performed using immunocompromised mice. Herein, we assessed the feasibility and sensitivity of fluorescence imaging for monitoring the in vivo activity of drugs against acute M. abscessus infection using zebrafish embryos. A protocol was developed where clarithromycin and imipenem were directly added to water containing fluorescent M. abscessusinfected embryos in a 96-well plate format. The status of the infection with increasing drug concentrations was visualized on a spatiotemporal level. Drug efficacy was assessed quantitatively by measuring the index of protection, the bacterial burden (CFU), and the number of abscesses through fluorescence measurements. Both drugs were active in infected embryos and were capable of significantly increasing embryo survival in a dose-dependent manner. Protection from bacterial killing correlated with restricted mycobacterial growth in the drug-treated larvae and with reduced pathophysiological symptoms, such as the number of abscesses within the brain. In conclusion, we present here a new and efficient method for testing and compare the in vivo activity of two clinically relevant drugs based on a fluorescent reporter strain in zebrafish embryos. This approach could be used for rapid determination of the in vivo drug susceptibility profile of clinical isolates and to assess the preclinical efficacy of new compounds against M. abscessus. The emerging pathogen Mycobacterium abscessus is the etiological agent of a wide spectrum of infections in humans, including severe chronic pulmonary and disseminated infections, mostly in immunosuppressed and cystic fibrosis (CF) patients (1), and cutaneous diseases, often posttraumatic and postsurgical. This neglected pathogen causes a higher fatality rate than other rapidly growing mycobacteria (RGM), and CF patient infections is becoming a major threat in most CF centers worldwide (2). M. abscessus infections occur in early childhood (3), are severe and sometimes fatal, especially following transplantation (4-6), and may lead to outbreaks of infection (6). It is also the main RGM responsible for nosocomial and iatrogenic infections in humans (postinjection abscesses, cardiac surgery, and plastic surgery) (7-9). It has been reported to cross the blood-brain barrier and cause important central nervous system (CNS) lesions. Although a rapid grower, M. abscessus possesses several important pathogenic traits, such as the ability to (i) persist silently for years and even decades in the human host (10) and to (ii) induce lung disease with caseous lesions and granuloma formation in the parenchyma (11, 12).The major issue with M. abscessus relies on its intrinsic resistance to the most available antibiotics. The American Thoracic Society has recommended different...

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

“…However, due to the inherent limitations of the currently available cellular/animal models used to study M. abscessus infection, our knowledge regarding the pathophysiological events of the R or S variants remains obscure. Infection of immuno-competent mice via intravenous or aerosol routes leads to transient colonization, impeding the use of mice to study persistent infections and for in vivo drug susceptibility testing 17,20 .…”

Regard à travers le danio pour mieux comprendre les interactions hôte/pathogène

“…Ces infections sont associées au développe-ment de lésions granulomateuses pseudotuberculeuses. Cependant, les déterminants de virulence requis pour le développement de la maladie demeurent inconnus, en raison du nombre restreint de modèles expérimen-taux disponibles, modèles par ailleurs peu pertinents -notamment la souris -, les souches immunocompé-tentes n'étant pas permissives à l'infection [19]. La description de certaines caractéristiques physio-et immunopathologiques de ces infections a néanmoins été possible grâce au développement du danio comme nouveau modèle d'étude de la virulence de M. abscessus [12].…”

“…Grâce à la transparence des danios, il est possible de visualiser et quantifier les éventuels effets toxiques ou tératogènes survenant à la suite du traitement, ainsi que l'efficacité microbicide ou microbiostatique des composés sur des embryons infectés ; ceci grâce à un suivi de la mortalité et de la progression de l'infection via l'utilisation de bactéries fluorescentes. Ces dernières années ont vu le succès grandissant du danio pour tester l'efficacité de différents composés antimicrobiens [19,26,27] avec notamment le développement de plates-formes automatisées propices au criblage à haut débit [28]. En testant rapidement de larges chimiothèques 2 , il est dorénavant possible d'identifier de nouvelles molécules antimicrobiennes, première étape vers de futurs traitements.…”

Activity of LCB01-0371, a Novel Oxazolidinone, against Mycobacterium abscessus

“…Originally, M. abscessus was considered to belong to the M. chelonae group, but in 1992, M. abscessus was reclassified as an individual species (1). M. abscessus causes a variety of infections with a high fatality rate (2,3). Over the past 10 years, this species has been highlighted as an important human pathogen responsible for a wide spectrum of soft tissue infections, disseminated infections, and severe chronic pulmonary infections, mostly in immunosuppressed and cystic fibrosis (CF) patients (4).…”