Abstract Title
Comparison of human norovirus replication in two zebrafish systems: embryos and larvae
Presenter
Charlène Plénière, Université Laval
Co-Author(s)
Charlène Plénière(1), Valérie Goulet-Beaulieu(1), Éric Jubinville(1), Paul De Koninck(2), Julie Jean(1)
(1) Laboratoire Julie Jean, Department of Food Sciences, Université Laval, Québec, Canada
(2) Department of Biochemistry, Microbiology and Bioinformatics, Université Laval, Québec, Canada
Abstract Category
Entry and Replication
Abstract
Among human noroviruses (HuNoVs), genotype GII.4 is associated with most gastroenteritis outbreaks worldwide. Despite its significant impact, HuNoV research has long been hindered by the absence of an efficient and reproducible replication system. Recently, two protocols using zebrafish embryos and larvae infected with contaminated feces have enabled HuNoV replication.
These two protocols differ slightly in experimental parameters, including incubation temperature, duration, and the composition of the aqueous medium. To date, only a single comparison using one GII.4 strain has been conducted, showing enhanced viral RNA detection in the embryo-based system. We aim to evaluate whether both protocols yield consistent results across multiple genotypes under the same laboratory conditions and using the same samples, while also identifying their respective technical advantages and limitations.
Different HuNoV genotypes (GII.3, GII.7, and several GII.4) circulating in Canada were microinjected (3 nL) into zebrafish embryos (3 hours post-fertilization) and larvae (3 days post-fertilization). Embryos were incubated at 29,5 °C for 3 days; larvae at 32 °C for 2 days. Samples were snap-frozen in liquid nitrogen and stored at -80 °C. Viral RNA was quantified by RT-qPCR 1-hour post-injection and after incubation to assess replication levels.
Both systems supported replication across all genotypes, reaching similar viral loads (~6 log RNA copies per zebrafish). These findings differ from previous literature, potentially due to strain-specific factors. Although both systems are effective, the embryo-based protocol offers a simpler and faster workflow. This system represents a valuable tool for advancing HuNoV research and supporting future applications in food safety.
These two protocols differ slightly in experimental parameters, including incubation temperature, duration, and the composition of the aqueous medium. To date, only a single comparison using one GII.4 strain has been conducted, showing enhanced viral RNA detection in the embryo-based system. We aim to evaluate whether both protocols yield consistent results across multiple genotypes under the same laboratory conditions and using the same samples, while also identifying their respective technical advantages and limitations.
Different HuNoV genotypes (GII.3, GII.7, and several GII.4) circulating in Canada were microinjected (3 nL) into zebrafish embryos (3 hours post-fertilization) and larvae (3 days post-fertilization). Embryos were incubated at 29,5 °C for 3 days; larvae at 32 °C for 2 days. Samples were snap-frozen in liquid nitrogen and stored at -80 °C. Viral RNA was quantified by RT-qPCR 1-hour post-injection and after incubation to assess replication levels.
Both systems supported replication across all genotypes, reaching similar viral loads (~6 log RNA copies per zebrafish). These findings differ from previous literature, potentially due to strain-specific factors. Although both systems are effective, the embryo-based protocol offers a simpler and faster workflow. This system represents a valuable tool for advancing HuNoV research and supporting future applications in food safety.