Zebrafish model for mycobacterial infection

Abstract

M. marinum infection in the zebrafish (Danio rerio) is a well-established model of human tuberculosis. In this project, methodology for experimental infections and assessment of the disease progression was set up and optimized in Mika Rämet?s laboratory. The methods include bacterial cultures, experimental infection by injection, assessment of survival, quantification of mycobacterial load from infected fish by plating and an in-house-developed qPCR method, and quantification of granulomas in different organs by histology. In addition, methods were established to induce immune suppression in the context of mycobacterial infection. The methods were utilized in a longitudinal study (32 weeks) on the progression of the disease in adult fish. I.p. infection with 1575 cfu of the ATCC 927 type strain of M. marinum resulted in a progressive disease with high mortality (64 %), increasing bacterial load and number of granulomas in various target organs. On the contrary, with 35 cfu, a chronic/latent disease developed in most individuals. The low-dose infection was characterized by limited mortality (25 %), static bacterial loads after 4 weeks of infection and constant numbers of highly organized granulomas in few target organs. Similar latency is also the usual outcome of the human disease. In approximately 5-10 % of the latent cases in humans, the disease gets reactivated as a consequence of immune suppression. Also this feature can be conveniently replicated in the adult zebrafish model. The disease can now be re-induced in chronically infected zebrafish by gamma irradiation that depletes granulo/monocyte and lymphocyte pools, as shown by flow cytometry (FCM). This weakening causes a rapid outgrowth of mycobacteria (100-fold increase compared to the levels seen in chronic infection) leading to death of 88 % of the fish in less than a month. The adult zebrafish presents itself as a unique low vertebrate model for studying the development of latency as well as re-induction of latent tuberculosis.