Abstract
The larval zebrafish is an increasingly popular host model for the study of Aspergillosis. The visual accessibility, genetic resources, small size, and ease of handling make zebrafish larvae compatible with higher-throughput investigation of fungal virulence and host resistance mechanisms. This article provides the protocols needed to prepare Aspergillus fumigatus spore inocula and use microinjection to infect the hindbrain ventricle of zebrafish larvae. Furthermore, we include protocols for analyzing host survival, immobilizing larvae for live imaging, and suggestions for image analysis. © 2021 Wiley Periodicals LLC. Support Protocol 1: Preparing Aspergillus spores Support Protocol 2: Dechorionating zebrafish embryos Support Protocol 3: Generating transparent larvae with 1-phenyl 2-thiourea (PTU) Basic Protocol 1: Hindbrain microinjection of zebrafish larvae with Aspergillus spores Basic Protocol 2: Survival analysis Basic Protocol 3: Multi-day imaging of infected larvae Alternate Protocol: Embedding larvae in low-melting-point agarose.