Most fish embryos that develop externally are exposed to an environment full of microbes. How they survive microbial attacks are not understood to date. Here we demonstrated that the egg cytosol prepared from the newly fertilized eggs of zebrafish Danio rerio is capable of killing the Gram-negative bacterium Escherichia coli, via in vitro assay system of the complement activity established. All findings indicate that it is the complement system operating via the alternative pathway that is attributable to the bacteriolytic activity. This is the first report providing the evidence for the functional role of the maternal complement components in fish eggs, paving the way for study of maternal immunity in other organisms whose eggs are fertilized in vitro.