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

Sublethal biological effects of heavy metals accumulated in stream macro-invertebrates were examined 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 role in the detoxification of both organic and inorganic xenobiotic compounds. Integrity of the lysosomal membrane is essential for proper function of these structures. Following a heavy metal pollution gradient, larvae of Hydropsyche spp. and A. grandis were collected from 4 or 3 stations, respectively, in the Clark Fork River, and a reference site (Blackfoot River). 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 show good correlation between elevated tissue concentrations of Cd, Cu, Pb and Zn, significantly increased levels of hsp70, and reduced lysosomal membrane stability in both species collected at the Goldcreek site.