Search

Severe, often fatal, opportunistic fungal infections frequently arise following mucosal damage caused by trauma or cytotoxic chemotherapy. Interaction of fungal pathogens with epithelial cells that comprise mucosa is a key early event associated with invasion, and therefore, enhancing epithelial defense mechanisms may mitigate infection. Here we establish a model of mold and yeast infection mediated by inducible epithelial cell loss in larval zebrafish. Epithelial cell loss by extrusion promoted exposure of laminin that was associated with increased fungal attachment, invasion and larval lethality, while fungi defective in adherence or filamentation did not impact survival. Transcriptional profiling identified significant upregulation of the epidermal growth factor receptor ligand epigen (EPGN) upon mucosal damage. Treatment with recombinant human EPGN suppressed epithelial cell extrusion, leading to reduced fungal invasion and significantly enhanced survival. Together, these data support the concept of augmenting epithelial restorative capacity to attenuate pathogenic invasion of fungi associated with human disease.

Due to their transparency, genetic tractability, and ease of maintenance, zebrafish (Danio rerio) have become a widely-used vertebrate model for infectious diseases. Larval zebrafish naturally prey on the unicellular protozoan Paramecium caudatum. This protocol describes the use of P. caudatum as a vehicle for food-borne infection in larval zebrafish. P. caudatum internalize a wide range of bacteria and bacterial cells remain viable for several hours. Zebrafish then prey on P. caudatum, the bacterial load is released in the foregut upon digestion of the paramecium vehicle, and the bacteria colonize the intestinal tract. The protocol includes a detailed description of paramecia maintenance, loading with bacteria, determination of bacterial degradation and dose, as well as infection of zebrafish by feeding with paramecia. The advantage of using this method of food-borne infection is that it closely mimics the mode of infection observed in human disease, leads to more robust colonization compared to immersion protocols, and allows the study of a wide range of pathogens. Food-borne infection in the zebrafish model can be used to investigate bacterial gene expression within the host, host-pathogen interactions, and hallmarks of pathogenicity including bacterial burden, localization, dissemination and morbidity.