The broad objective of this research was to determine the environmental drivers of macroinvertebrate and microbial assemblages in acidic pit lakes. This is important because pit lake ecosystem development is influenced by prevailing environmental characteristics. Three lakes (Stockton, Kepwari, WO5H) within a larger pit-lake district in Collie, Western Australia were surveyed for spatial variability of benthic macroinvertebrate and microbe (Archaea, Bacteria) assemblage composition as well as potential environmental drivers (riparian condition, aquatic habitat, sediments, and aquatic chemistry) of assemblages. With the exception of sediment chemistry, biophysical variables were significantly different across lakes and reflected riparian condition and groundwater chemistry. Microbial assemblages in pit lakes were significantly different across lakes and correlated with water chemistry, particularly metals in Lake WO5H. However, the most abundant microbes were not readily identified beyond class, making it difficult to speculate on their ecological function. Macroinvertebrate assemblage composition and species richness were also significantly different across all lakes, and in Lake WO5H (a lake with low pH and high metal concentrations), taxa were correlated with benthic organic matter as well as water chemistry. Results indicated that despite poor water quality, input of nutrients from terrestrial leaf litter can support or augment pit lake ecosystems. This is a demonstration of the concept that connection of pit lakes to catchments can positively affect aquatic ecosystems, which can inform management actions for remediation.