Background

Host-pathogen interaction studies are important to understand disease pathogenesis and also for the development of effective treatments. Multiple aspects of both the host and pathogen need to be considered to determine potential risk factors in the host and key virulence factors in the pathogen. Innate immunity represents the first line of defense against infections, triggering a cascade of responses, including inflammation, neutralization and recruitment of components of the adaptive immune system (Akira et al., 2006). Neutrophils are rapidly recruited to the site of bacterial infection in response to chemoattractant gradients of host chemokines released from damaged cells as well as bacterial products themselves, where they initiate the immune response by phagocytosis and netolysis (Renshaw and Trede, 2012; Deng et al., 2013). Macrophage recruitment follows, with these cells focused on removing dead cells, remodeling injured tissue and coordinating adaptive immune cells (Weiss and Schaible, 2015).

The development and function of innate immune cells are controlled by a range of specific genes, the dysregulation of which can lead to a number of pathological states, including enhanced susceptibility to infection and chronic inflammation. Being short-lived, neutrophils are depleted rapidly in response to exposure to a pathogen, with so-called ‘emergency granulopoiesis’ being initiated to generate additional neutrophils (Hall et al., 2012; Manz and Boettcher, 2014). Granulocyte colony-stimulating factor (G-CSF) acting via its receptor, G-CSFR, is a key mediator of emergency granulopoiesis through its action promoting the proliferation and differentiation of relevant hematopoietic progenitor cells (Panopoulos and Watowich, 2008; Liongue et al., 2009). Understanding the role of factors that regulate the innate immune response during bacterial infection in an appropriate in vivo model can provide unique insights into infection and immunity.

Zebrafish represents a very attractive in vivo model to perform host-pathogen studies, with its transparent embryos allowing in vivo imaging at high-resolution to track infection in real-time, by tagging the pathogens with fluorescent markers (Basheer et al., 2019). In addition, using transgenic approaches in zebrafish, neutrophils and macrophages can also be monitored via fluorescent tags (Ellett et al., 2011; Gray et al., 2011). Importantly, these cells along with other components of the immune system share remarkable similarity with those of humans (Meeker and Trede, 2008). The zebrafish innate immune system forms early during their development with the generation of macrophages at 24 h post fertilization (hpf) and neutrophils by 32-48 hpf (Herbomel et al., 1999; Willett et al., 1999; Bennett et al., 2001). The zebrafish adaptive immune system develops later, allowing the innate immune system to be studied independently. Zebrafish infection models of different pathogen–bacterial, viral and fungal–have been established (Gratacap and Wheeler 2014; Masud et al., 2017; Varela et al., 2017).

Our research has employed a zebrafish bacterial infection model to understand the role of key genes involved in innate immunity in defense against infection. Escherichia coli (E. coli) bacteria expressing green fluorescent protein (GFP) were microinjected into 72 hpf larvae derived from either wild-type zebrafish or those mutant for the gene encoding G-CSFR (Figure 2). These were monitored by fluorescence microscopy over a time-course to determine the relative infection kinetics (Basheer et al., 2019). Overall, this procedure helps in understanding different host-pathogen interactions and unravel the functions of key immune genes involved in the process using zebrafish as an animal model.