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Environmental oxidation and microbial metabolism drive production of acid mine drainage (AMD). Understanding changes in the microbial community, due to geochemical and seasonal characteristics, is fundamental to AMD monitoring and remediation. Using direct sequencing of the 16S and 18S rRNA genes to identify bacterial, archaeal, and eukaryotic members of the microbial community at an AMD site in Northern Ontario, Canada, we found a dynamic community varying significantly across winter and summer sampling times. Community composition was correlated with physical and chemical properties, including water temperature, pH, conductivity, winter ice thickness, and metal concentrations. Within Bacteria, Acidithiobacillus was the dominant genus during winter (11%-57% of sequences) but Acidiphilium was dominant during summer (47%-87%). Within Eukarya, Chrysophyceae (1.5%-94%) and Microbotrymycetes (8%-92%) dominated the winter community, and LKM11 (4%-62%) and Chrysophyceae (25%-87%) the summer. There was less diversity and variability within the Archaea, with similar summer and winter communities mainly comprising Thermoplasmata (33%-64%) and Thermoprotei (5%-20%) classes but also including a large portion of unclassified reads (∼40%). Overall, the active AMD community varied significantly between winter and summer, with changing community profiles closely correlated to specific differences in AMD geochemical and physical properties, including pH, water temperature, ice thickness, and sulfate and metal concentrations. [ABSTRACT FROM AUTHOR]