Background: Environmental monitoring for pharmaceuticals and endocrine disruptors in the aquatic environment traditionally employs a variety of methods including analytical chemistry, as well as a variety of histological and biochemical endpoints that correlate with the fish fitness. It is now clear that analytical chemistry alone is insufficient to identify aquatic environments that are compromised because these measurements do not identify the biologically available dose. The biological endpoints that are measured are important because they relate to known impairments; however, they are not specific to the contaminants and often focus on only a few known endpoints. These studies can be enhanced by looking more broadly at changes in gene expression, especially if the analysis focuses on biochemical pathways. The present study was designed to obtain additional information for well-characterized sites adjacent to sewage treatment plants in MN that are thought to be impacted by endocrine disruptors. Results: Here we examine five sites that have been previously characterized and examine changes in gene expression in fathead minnows (Pimephales promelas) that have been caged for 48 h in each of the aquatic environments. We find that the gene expression changes are characteristic and unique at each of the five sites. Also, fish exposed to two of the sites, 7 and 12, present a more aggressive behavior compared to control fish. Conclusion: Our results show that a short-term exposure to sewage treatment plant effluents was able to induce a site-specific gene expression pattern in the fathead minnow gonad and liver. The short-term exposure was also enough to affect fish sexual behavior. Our results also show that microarray analysis can be very useful at determining potential exposure to chemicals, and could be used routinely as a tool for environmental monitoring.