Ecdysteroid responses of estuarine crustaceans exposed through complete larval development to juvenile hormone agonist insecticides.

Fenoxycarb and pyriproxyfen are insecticides that gain their toxicity by specifically acting as insect juvenile hormone agonists (JHA), and so are endocrine disruptors by design and effectively prevent larvae from maturing into adults. Efforts to assess the environmental effects of JHAs on nontarget populations of invertebrates have resulted in the utilization of several established estuarine crustacean models. This work was conducted to test the hypothesis that the mortality, inhibition of development and decreased fecundity reported previously in these animals from JHA exposure coincides with abnormal circulating titers of ecdysteroids. Gravid female grass shrimp (Palaemonetes pugio) and mud crabs (Rhithropanopeus harrisii), species with different developmental plasticity and JHA tolerances, were collected and held at wet lab conditions (20 ppt salinity, 25°C) until larval release. Larvae were collected <12 hr after hatch and exposed to JHAs during a static renewal test through end of development with seawater or nominal concentrations of JHA previously shown to induce significant developmental delays and/or decreased body weights. Larvae were subsampled (10 larvae/sample, n = 2 to 8) at each developmental stage, lyophilized, and ecdysteroids extracted by homogenization in 80% methanol and elution from C18 Sep-Pak cartridges with 25%, 60% and 100% methanol to capture the polar, free, and apolar conjugates, respectively, and then quantified by ELISA. As was expected significant differences in successful completion of development (larval survival), developmental duration, and growth (dry weight) were observed. These physiological perturbations were linked with significantly altered ecdysteroid titers, supporting a newly emerging theory that juvenoids possibly act as anti-ecdysteroids through a novel molecular mechanism involving inhibition of ecdysteroid signaling.