Chlorothalonil (2,4,5,6-tetrachloroisophthalonitrile) is the second most widely used fungicide in the United States. Due to the widespread use of chlorothalonil, it is important to investigate the effects chlorothalonil may have on estuarine species such as the grass shrimp, Palaemonetes pugio. This study examined the toxicity of chlorothalonil to three life-history stages (embryo, larvae, adult) of the grass shrimp. Also, molting frequency, growth response and metamorphosis from a larval life cycle pulsed exposure assay were examined as sub-lethal indicators of chlorothalonil exposure. Results showed embryos were the least sensitive with a 96-h Median Lethal Concentration (LC50) of 396.0 µg/L (95% Confidence Interval [CI] 331.3-472.4 µg/L). The adult 96-h LC50 was 152.9 µg/L (95% CI 120.3-194.5 µg/L). Larvae were the most sensitive to chlorothalonil exposure with a 96-h LC50 of 49.5 µg/L (95% CI 44.4-55.27 µg/L). In the life cycle pulsed exposure assay, all surviving larvae in the treatments required significantly more molts to reach postlarvae than the control. Other measured parameters showed differences between treatments and control but there was no statistical significance. This research demonstrated that chlorothalonil is highly toxic to grass shrimp and that larval grass shrimp would be the most appropriate life stage to use for chlorothalonil risk assessments since that stage is the most sensitive.