Mining, whether current or inactive, generally increases salt concentrations in catchment watersheds due to precipitation on and through exposed rock surfaces. Practices like mountaintop removal mining have exacerbated this issue, with measurements of salt concentrations in nearby catchment systems well above normal levels. Nevertheless, the impact of the ionic composition of mining effluent on aquatic animal health is not well understood. This is a particularly important issue in Appalachia because it is home to an enormous diversity of organisms, including a huge array of amphibians that live in streams that receive mining effluent from operating and abandoned mines. To investigate this issue, we examined the effects of reconstituted mining effluent on the development of wild-caught wood frog (Lithobates sylvaticus) tadpoles. We collected day-old fertilized eggs from a creek near Blacksburg, VA in early March, 2018 and raised them to hatch. Tadpoles were then assigned to either sulfate or chloride-based reconstituted mining effluent diluted to six different conductivities (100 µS/cm -2,400 µS/cm). After 7 or 14 days of treatment, tadpoles were euthanized and fixed in paraformaldehyde. We imaged the heads and bodies of tadpoles for morphometric analysis before dissecting out brains and immunostaining them for phospho-histone H3, which labels dividing progenitor cells in the brain. We found that sulfate-based reconstituted mining effluent significantly lowered progenitor cell division at 1200 µS/cm at Day 7 and at 600 µS/cm at Day 14 relative to control. Chloride-based reconstituted mining effluent was less impactful, with no significant differences observed at Day 7 and significantly lowered progenitor cell division at 2400 µS/cm at Day 14. In addition, both treatments slowed growth of some head morphological features, including head size and interocular distance. Chloride treatment slowed growth of body length at Day 14 at 600 µS/cm, whereas sulfate-based reconstituted mining effluent had no effect on body length. These data show that sulfate-based mining effluent has a substantial
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