This study was conducted to investigate whether exposure to wastewater treatment works (WWTW) effluent affects the adaptive stress axis of fish resident within the receiving water.Three-spined sticklebacks (Gasterosteus aculeatus) were sampled from sites downstream of ten WWTWs in north-west England, selected to represent a range of human population equivalents between 1,000 and 125,000. Following capture, indices of stress (whole-body cortisol and glucose concentrations) were measured both prior to, and following, the imposition of a standardised stressor to establish both baseline and stress-induced concentrations of cortisol and glucose. There was considerable between-site variation in size, and to a lesser extent condition, of the fish. Pre-and post-stress cortisol and glucose concentrations also varied significantly between-sites. A large proportion of the variation in both the somatic data and the stress response was explained by variation in the proportion of effluent contributing to total river flow at the study sites. Mass (r 2 = 0.35, P < 0.001) and length (r 2 = 0.37, P < 0.001) of the fish, and cortisol (r 2 = 0.26, P < 0.001) and glucose (r 2 = 0.12, P < 0.01) concentrations in unstressed sticklebacks, were positively related to the concentration of effluent across the sample sites. However, in stressed fish, cortisol (r 2 = 0.32, P < 0.001) and glucose (r 2 = 0.14, P < 0.001) concentrations exhibited a negative trend in relation to the effluent concentrations across sites. Individual variation in fish size did not account for the variation in either cortisol or glucose levels. These data provide the first indication that modulation of the stress axis in fish by anthropogenic factors might be widespread and of greater significance than hitherto assumed.