Awareness of the consistently degraded state of urban streams, with lower biodiversity has heightened worldwide, increasing restoration initiatives to ameliorate the adverse effects. The present study examined anthropogenically-derived contaminants in water and sediments and bioaccumulation in tissues and bile of shortfin eels (Anguilla australis) in Hamilton City urban gully streams. Bioassays were also conducted with two native Crustacea; the amphipod Paracorophium lucasi and freshwater crayfish (kōura) (Paranephrops planifrons) to test endpoints of survival, reburial behaviour and growth. Results of dissolved Cu, Pb and Zn showed the industrial Waitawhiriwhiri catchment, with higher impervious surfaces, to have the greatest potential for generating contaminants in stream waters. Relationships between sediment contaminant concentrations and upstream % impervious area suggest an association with stormwater runoff and metal accumulation. Shortfin eels from Gibbons Creek and Lake Rotoroa bioaccumulated high concentrations of Pb and As respectively in livers, and PAH metabolite pyrene-1-glucuronide in bile was found in many eels, highlighting the bioavailability of these contaminants. Muscle tissues cocnentrations of Pb and Hg in some sites triggered food safety guidelines presenting a low risk for human consumption. Amphipods exposed to sediments from Lake Rotoroa had significantly reduced survival compared with those exposed to other site sediments in a 10-day sediment toxicity test. Photo-induced toxicity is not of concern for biota exposed to these sediments. Sediments did not induce any sub-lethal effect on reburial behaviour. Growth rates of kōura fed Salix fragilis leaf material incubated in Hamilton streams were not significantly different, and the short duration of the study meant conclusions could not be made on the significance of observed accumulated metals and metalloids on differences in growth rates. Results of this study highlight a number of locations in the Hamilton stream network, where contaminants are of concern, especially in streams with fully urbanised catchments with high effective imperviousness and legacies of past land use or pollution. Bioavailability of some metals, metalloids and PAHs is constraining the diversity of some species present in these streams, although not apparent for shortfin eels, as they are very common.