Document Type

Dissertation

Publication Date

Summer 7-9-2004

Abstract

Acid mine drainage (AMD) contaminates thousands of miles of mountain streams worldwide. At the same time, nitrate loading to many watersheds is increasing. Little is known about nitrogen cycling in acidic, heavy metal-laden streams, however it has been reported that denitrification, the reduction of nitrate to nitrogen gas, is inhibited under low pH. Snowmelt dominated mountain streams, bringing a pulse of nitrate, dissolved organic carbon, and in AMD sites iron, sulfate, heavy metals, and acidity. The objectives of this research were to determine if denitrifying microorganisms are present and active in AMD streams, investigate the effects of pH, electron donors, and iron on denitrification in acidic and circumneutral streams, examine seasonal affects on denitrification, and compare microbial diversity in AMD streams with that in a naturally acidic stream and a pristine mountain stream. Nitrogen gas was the major end-product of denitrification in sediments from several AMD impacted streams in Colorado with pHs ranging from 2.60 to 4.54, with no correlation between rate and initial pH. Denitrification increased pH in all microcosms. Carbon stimulated denitrification, as well as the presence of high DOC in stream waters, showing the importance of heterotrophic denitrifiers. Additional soluble iron (Fe2+ and Fe3+) decreased denitrification. Denitrifiers were present and active throughout the year in sediments from a direct mine effluent stream, a higherorder AMD-impacted stream, a naturally acidic stream, and a pristine, near-neutral pH mountain stream. Stream nitrous oxide concentrations correlated with highest denitrification potential rates during snow cover indicating microcosm studies complemented in situ denitrification activity. rRNA genes amplified from environmental DNA included members from all three domains and 21 different bacterial divisions and 715 previously unidentified strains. No common denitrifiers were found, however rate constants significantly correlated with the number of clones found in a-Proteobacteria, Actinobacteria, and Chloroflexi, indicating that denitrification may be widespread within these groups. Denitrifiers are active in acid-impacted streams and are influenced by environmental and hydrologic conditions. The results presented here have increased our understanding of nitrogen cycling in extreme environments as well as environments were multiple stressors are present and may be important for nitrogen and carbon budgets in mountain catchments.

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