Native plasmids constitute a major category of extrachromosomal DNA elements responsible for harboring and transferring genes important in survival and fitness. A focused evaluation of plasmidomes can reveal unique adaptations required by microbial communities. We examined the plasmid DNA from two pristine wells at the Oak Ridge Field Research Center. Using a cultivation-free method that targets plasmid DNA, a total of 42,440 and 32,232 (including 67 and 548 complete circular units) scaffolds > 2 kb were obtained from the two wells. The taxonomic distribution of bacteria in the two wells showed greater similarity based on their plasmidome sequence, relative to 16S rRNA sequence comparison. This similarity is also evident in the plasmid encoded functional genes. Among functionally annotated genes, candidates providing resistance to copper, zinc, cadmium, arsenic, and mercury were particularly abundant and common to the plasmidome of both wells. The primary function encoded by the most abundant circularized plasmid, common to both wells, was mercury resistance, even though the current ground water does not contain detectable levels of mercury. This study reveals that the plasmidome can have a unique ecological role in maintaining the latent capacity of a microbiome and being important for rapid adaptation to environmental stresses.