Testing Alternative Surface Disinfection Agents for Zebrafish (Danio rerio) Embryos

Pathogen transmission into zebrafish colonies is controlled through vigilant biosecurity practices. One such practice is embryo surface disinfection, which often uses sodium hypochlorite. However, if sodium hypochlorite is used at an inappropriate pH, concentration, or exposure time, zebrafish embryos can experience significant mortality and morbidity. Reagent-grade sodium hypochlorite is often used for embryo surface disinfection because commercial-grade sodium hypochlorite has additional ingredients that may have deleterious effects on the embryo. In addition, chlorine dioxide and the combination of sodium chloride and potassium peroxymonosulfate (SCPP) are effective equipment disinfectants; however, the effects of these chemical agents on zebrafish embryos during surface disinfection are unknown. In this study, we exposed strain 5D zebrafish embryos (ages, 6 and 24 h postfertilization) to 4 chlorine-containing agents (reagent-grade sodium hypochlorite [bleach], commercial-grade sodium hypochlorite [bleach], SCPP, and chlorine dioxide) at either 50- or 100- ppm for 5 or 10 min. All groups were evaluated at 5 d postfertilization for survival, hatching rate, and morphologic defect rate. The experimental group with the highest survival and hatching rates and the lowest morphologic defect rate was the 24-h postfertilization embryos exposed to 50 ppm SCPP for 5 min. The survival, hatching rate, and defect rate did not differ significantly among age-matched controls; however, the hatching rate after exposure to 50 ppm SCPP was significantly higher than that of embryos exposed to 50 ppm reagent-grade sodium hypochlorite for 5 min (100% compared with 23% respectively). SCPP solution may provide an alternative surface disinfectant for zebrafish embryos because it maximizes survival and hatching rates and minimizes morphologic defect rates. However, in vivo efficacy against common zebrafish pathogens requires further testing. Use of chlorine dioxide at 50 ppm or greater is not recommended for zebrafish embryo surface disinfection due to significant mortality among 6 and 24 h postfertilization embryos.