With six interactive research work packages FishForPharma aims to deliver the proof-of-principle for drug discovery using zebrafish infectious disease models and to increase understanding of host-pathogen interaction mechanisms to identify new drug targets for infectious disease treatment.
Infectious diseases caused by pathogenic micro-organisms are major causes of death, disability, and social and economic disruption for millions of people. During evolution these pathogens have developed intricate strategies to manipulate host defense mechanisms and outwit the immune system. To reduce the burden of infectious diseases it is important to increase understanding of these host-pathogen interaction mechanisms and to develop more effective strategies for drug discovery.
The zebrafish holds much promise as a high throughput drug screening model. In the last few years, zebrafish models for studying human pathogens or closely related animal pathogens have emerged at a rapid pace. The fact that zebrafish produce large amounts of embryos, which develop externally and are optically transparent, gives unprecedented possibilities for live imaging of disease processes and is the basis of novel high-throughput drug screening approaches.
WP1: Development and optimisation of infection models.
Many human infectious diseases, for example tuberculosis and hospital-acquired infections can be modeled in the zebrafish. FishForPharma participants have established zebrafish models for infections with different bacteria (Mycobacteria, Salmonella, Staphylococci, Aeromonas, Yersinia, Burkholderia) and viral pathogens (rhabdo- and toga-viruses). These models will be further optimized and new models for parasitic infections will be developed.
WP2: Development of novel imaging and analysis tools.
Transparent zebrafish embryos are ideal for live imaging of host-pathogen interactions. To this end, we use transgenic zebrafish lines with fluorescently labelled immune cells in combination with pathogens in other fluorescent colors. In WP2 we will generate new marker lines for different subsets of immune cells and for subcellular compartments where pathogens reside. We will also develop new antibodies to detect components of the immune system.
WP3: Development of high-throughput methods.
Zebrafish produce large amounts of embryos that develop externally and are very suitable for high-throughput screening. Participants ofFishForPharma (Leiden University and ZF-screens) have developed a robotic infection system for zebrafish embryos, which has been successfully applied for pilot anti-tuberculosis drug screens. In WP3, this high-throughput screening platform will be fully standardised, extended to other infection models, and integrated with automated fluorescence image acquisition and recognition tools.
WP4: Host immune response mechanisms.
The zebrafish model is a valuable addition to mammalian models for vertebrate immunology. Efficient gene knockdown and knockout methods can be used to discover novel factors that participate in host-pathogen interactions. In WP4 we will use deep sequencing technology to better characterize immune responses to different infections and will perform functional studies of genes involved in central pathways of the immune system, such as Toll-like, Nod-like, and chemokine receptor signaling.
WP5: Pathogen virulence mechanisms.
A better understanding of the intricate interactions between virulence factors of pathogens and the immune system of the host is important to develop novel treatment strategies and reduce the burden of infectious diseases. This WP uses the advantages of live imaging of infection in zebrafish to determine the precise stage(s) of action of key virulence factors. Virulence of clinical isolates in zebrafish will be compared with epidemiological findings, to further demonstrate that zebrafish can be faithfully used as a model for human disease and drug screening.
WP6: Anti-microbial drug screening.
In this WP we will demonstrate the use of high-throughput technology and infection models in chemical library screens. The main focus will be on drug screens with the mycobacterium infection model for tuberculosis, which is already at an advanced stage of development. Several recent studies underscore the potential for immunomodulatory drugs or drugs that target host pathways manipulated by pathogens. Our screens will therefore focus not only on candidate antimicrobial compounds, but also on candidate compounds that modulate the host immune response.