Neutrophils play important roles in innate immunity and are mainly dependent on various enzyme-containing granules to kill engulfed microorganisms. Zebrafish nephrosin (npsn) is specifically expressed in neutrophils; however, its function is largely unknown. Here, we generated an npsn mutant (npsnsmu5 ) via CRISPR/Cas9 to investigate the in vivo function of Npsn. The overall development and number of neutrophils remained unchanged in npsn-deficient mutants, whereas neutrophil antibacterial function was defective. Upon infection with Escherichia coli, the npsnsmu5 mutants exhibited a lower survival rate and more severe bacterial burden, as well as augmented inflammatory response to challenge with infection when compared with wild-type embryos, whereas npsn-overexpressing zebrafish exhibited enhanced host defence against E. coli infection. These findings demonstrated that zebrafish Npsn promotes host defence against bacterial infection. Furthermore, our findings suggested that npsn-deficient and -overexpressing zebrafish might serve as effective models of in vivo innate immunity.