In a 4-mo study (June 2004–September 2004), round gobies (Neogobius melanostomas) were dosed orally every 72 hr for up to 21 days with Clostridium botulinum neurotoxin type E (BoNT/E) at one of four doses: 0, 50, 250, and 500 mouse lethal doses (MLD). Fish were observed for changes in pigmentation and behavior for the duration of the experiment. Mortality was observed with all treatments, with the exception of the 0 MLD control. Clinical signs observed were consistent with prior research and appeared to occur in a threshold manner. The mean times to death and percent mortalities were dose dependant. Hazard ratios were determined to have a significant positive (parameter estimate=0.03) linear relationship with dose. The hazard ratio showed that per one unit dose increase, the instantaneous probability of a fish dying increased 1.02%. Postmortem analysis of experimental fish demonstrated that 11% (3/27) of fish contained detectable BoNT/E in their visceral fraction. The other 89% tested negative for BoNT/E, despite the fact that all fish died as a result of BoNT/E exposure. Therefore, botulism should not necessarily be ruled out as the cause of a fish kill, even if the fish test negative for BoNT/E.