In the Central United States, the Arkansas darter (Etheostoma cragini) is listed as a threatened fish species by the State of Kansas. Survival of the darter is threatened by loss of habitat caused by changing streamflow conditions, in particular flow depletion. Future management of darter populations and habitats requires an understanding of streamflow conditions and how those conditions may have changed over time in response to natural and anthropogenic factors. In Kansas, streamflow alteration was assessed at 9 U.S. Geological Survey streamgages in 6 priority basins with no pronounced long-term trends in precipitation. The assessment was based on a comparison of observed (O) and predicted expected (E) reference conditions for 29 flow metrics. The O/E results indicated a likely or possible diminished flow condition in 2 basins; the primary cause of which is groundwater-level declines resulting from groundwater pumping for irrigated agriculture. In these 2 basins, habitat characteristics adversely affected by flow depletion may include stream connectivity, pools, and water temperature. The other 4 basins were minimally affected, or unaffected, by flow depletion and therefore may provide the best opportunity for preservation of darter habitat. Through the O/E analysis, anthropogenic streamflow alteration was quantified and the results will enable better-informed decisions pertaining to the future management of darters in Kansas.