Protection and pathology in TB: learning from the zebrafish model

Semin Immunopathol DOI 10.1007/s00281-015-0522-4 REVIEW Protection and pathology in TB: learning from the zebrafish model Annemarie H. Meijer 1 Received: 24 June 2015 / Accepted: 11 August 2015 # The Author(s) 2015. This article is published with open access at Springerlink.com Abstract Zebrafish has earned its place among animal models of tuberculosis. Its natural pathogen, Mycobacterium marinum, shares major virulence factors with the human pathogen Mycobacterium tuberculosis. In adult zebrafish, which possess recombination-activated adaptive immunity, it can cause acute infection or a chronic progressive disease with containment of mycobacteria in well-structured, caseating granulomas. In addition, a low-dose model that closely mimics human latent infection has recently been developed. These models are used alongside infection of optically transparent zebrafish embryos and larvae that rely on innate immunity and permit non-invasive visualization of the early stages of developing granulomas that are inaccessible in other animal models. By microinjecting mycobacteria intravenously or into different tissues, systemic and localized infections can be induced, each useful for studying particular aspects of early pathogenesis, such as phagocyte recruitment, granuloma expansion and maintenance, vascularization of granulomas, and the phagocyte-mediated dissemination of mycobacteria. This has contributed to new insights into the mycobacteria-driven mechanisms that promote granuloma formation, the doubleedged role of inflammation, the mechanisms of macrophage cell death that favor disease progression, and the hostprotective role of autophagy. As a result, zebrafish models are now increasingly used to explore strategies for adjunctive therapy of tuberculosis with host-directed drugs. This article is a contribution to the Special Issue on Immunopathology of Mycobacterial Diseases - Guest Editor: Stefan Kaufmann * Annemarie H. Meijer 1 Institute of Biology, Leiden University, Einsteinweg 55, 2333 CC Leiden, The Netherlands Keywords Mycobacterium marinum . Tuberculosis . Granuloma . Innate immunity . Inflammation . Autophagy Introduction Mycobacterium tuberculosis (Mtb) is one of the most successful human pathogens that is estimated to have infected one third of the human population and to be responsible for nine million new cases of tuberculosis (TB) in 2013 (WHO Global Tuberculosis report 2014). Mtb parasitizes macrophages and can persist for decades as a latent infection inside its human host [1]. The formation of granulomas is central to the pathology of TB and the development of latency [2, 3]. TB granulomas are highly organized host cellular structures that contain an inner core of infected macrophages and necrotic cell debris (the caseum) where bacteria persist extracellular. In the surrounding cell layers, other immune cells, including dendritic cells, neutrophils, and T and B cells, wall off the bacteria inside the granuloma [2, 4]. A latent infection in granulomas has the ability to reactivate after many years, and the disease can be transmitted when granuloma integrity is lost. An alarming rise in antibiotic resistances and the lack of an effective vaccine against latent or reactivated TB emphasize the need for novel therapeutic strategies to control TB [5]. Animal models are indispensable for studying the host and bacterial factors involved in TB pathology and for evaluating new drug and vaccine candidates. Important insights into human TB pathology have been inferred from experimental Mtb infections in mice, guinea pigs, rabbits, and non-human primates, particularly macaques [6, 7]. In addition, now for over 10 years, the zebrafish has become widely used as an alternative animal model for TB [8–10]. Zebrafish can be infected with Mycobacterium marinum (Mm), a natural pathogen of cold-blooded vertebrates. The genomes of Mtb and Mm share Semin Immunopathol 3000 orthologs with an average amino acid identity of 85 % [11]. Mm occasionally causes a granulomatous skin infection in humans known as Bfish tank granuloma^ [12]. In zebrafish, Mm causes a systemic disease with containment of bacteria in granulomas that show strong structural similarity with the human TB granuloma [13–15]. Although differences in the adaptation of Mtb and Mm to different hosts must not be ignored, the important virulence factors of Mtb are functionally able to complement mutations in Mm genes and vice versa [16, 17]. Studies using the zebrafish-Mm model have contributed importantly to the changed view of the role of the granuloma in TB pathogenesis that has emerged over the recent years [2, 10]. Historically, the granuloma has been regarded as a static host defense structure. However, granuloma formation is driven by bacterial virulence, and it is now widely accepted that granulomas are highly dynamic structures that, especially during early stages of pathogenesis, can promote the dissemination of mycobacteria [2, 18]. Work in zebrafish has shown that the presence of macrophages is sufficient to initiate granuloma formation [19]. This means that the early stages of granuloma formation can be observed in optically transparent zebrafish embryos and larvae that have a functional innate immune system but have not yet developed adaptive immunity. The use of these zebrafish early life stages has shown that secondary granulomas can be seeded by the egression of infected macrophages from a primary granuloma [20]. That granulomas are not impenetrable is evidenced by experiments with superinfecting mycobacteria that are found to be transported by infected macrophages into established granulomas. This was initially observed during Mm infection of zebrafish and frogs and has subsequently been confirmed during Mtb infection in mice [21, 22]. Intravital imaging in both zebrafish and mice has demonstrated the migration of immune cells throughout the process of granuloma development [20, 23]. The heterogeneity and dynamic nature of granulomas observed in zebrafish and mice is in perfect agreement with serial PET-CT imaging data from Mtb-infected cynomolgus macaques showing that individual granulomas within the same host can regress and even be sterilized, while other granulomas progress during the same time [24]. This review will discuss how studies either in adult zebrafish or in embryos and larvae have advanced our understanding of mycobacterial virulence factors and of host genes implicated in immune protection or TB pathogenesis. TB in adult zebrafish While entry via de gastrointestinal tract is most likely the primary route of Mm infection in the natural environment, experimental infection of adult zebrafish is commonly achieved by intraperitoneal injection [13–15, 25]. Dependent on the particular dose and strain, the infection manifests with acute symptoms or develops as a chronic progressive disease [13–15]. Acute disease is characterized by rapid lethal inflammation and is more frequently obs (...truncated)