Biomarkers of Heavy Metal Effects in Two Species of Caddisfly Larvae from Clark Fork River, Montana: Stress Proteins (HSP70) and Lysosomal Membrane Integrity.

 Werner I.*, Broeg K., Cain D., Wallace W., Hornberger M., Hinton D.E. and Luoma S., UC Davis, Davis, CA; Biologische Anstalt Helgoland, Hamburg, Germany;
US Geological Survey, Menlo Park, CA. 

Potential sublethal effects of heavy metals in stream macroinvertebrates were examined with two cellular and biochemical biomarkers in larvae of two caddisflies indigenous to the Clark Fork River, Montana, - Hydropsyche spp. and Arctopsyche grandis. Stress proteins, in particular members of the HSP70 family, are involved in cellular protein homeostasis and repair, and are induced by a variety of stressors, which either damage cellular proteins directly or cause cells to synthesize aberrant proteins. Lysosomes are intracellular organelles that play key roles in the detoxification of both organic and inorganic xenobiotic compounds. Larvae of Hydropsyche spp. were collected from four sites on the Clark Fork (Galen Gage--4.7 km, Goldcreek--85.6 km, Turah--189.7 km, above Flathead--381 km) and a reference site (the Blackfoot River). Larvae of A. grandis were collected from the same sites minus the Galen site. Samples were immediately frozen in liquid nitrogen for HSP70 analysis, or preserved with Tissue Tek, then frozen in liquid nitrogen for the lysosomal stability assay. HSP70 was analyzed by western blotting using monoclonal antibodies. Lysosomal integrity was measured in cryosections by acid labilization with acid phosphatase as a marker enzyme. Results to date show elevated tissue concentrations of Cd, Cu, Pb and Zn and significantly increased levels of HSP70 in Arctopsyche from Goldcreek compared to reference samples. Lysosomal integrity also was compromised in samples from Goldcreek.  In Hydropsyche, tissue concentrations of Cd, Cu and Pb from Galen Gage were elevated (4-7 times) relative to the Blackfoot River, but levels of HSP70 did not differ between the two sites. These preliminary results indicate that sublethal effects of metal exposure may differ between species.

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