Infection of zebrafish larvae with human norovirus and evaluation of the in vivo efficacy of small-molecule inhibitors

This Protocol Extension details the use of zebrafish larvae as a simple and robust in vivo system for studying human norovirus infection, enabling evaluation of the antiviral effects and toxicities of small molecules. We have recently established that human norovirus (HuNoV) replicates efficiently in zebrafish larvae after inoculation of a clinical sample into the yolk, providing a simple and robust in vivo system in which to study HuNoV. In this Protocol Extension, we present a detailed description of virus inoculation by microinjection, subsequent daily monitoring and harvesting of larvae, followed by viral RNA quantification. This protocol can be used to study viral replication of genogroup (G)I and GII HuNoVs in vivo within 3-4 d. Additionally, we describe how to evaluate the in vivo antiviral effect and toxicity of small molecules using HuNoV-infected zebrafish larvae, in multi-well plates and without the need for specific formulations. This constitutes a great advantage for drug discovery efforts, as no specific antivirals or vaccines currently exist to treat or prevent norovirus gastroenteritis.

Automated summary

This study utilizes zebrafish larvae as an in vivo model to study human norovirus infection and evaluate antiviral effects and toxicities. Through steps such as virus injection and RNA quantification, viral replication of HuNoV can be studied within 3-4 days, and the antiviral effects and toxicities of small molecules can be evaluated in HuNoV-infected zebrafish larvae.