Many headwater streams in the midwestern United States were channelized for agricultural drainage. Conservation practices are implemented to reduce nutrient, pesticide, and sediment loadings within these altered streams. The impact of these practices is not well understood because their ecological impacts have not been evaluated and the relationships between water chemistry and fishes are not well understood. We evaluated relationships between water chemistry and fish communities within channelized headwater streams of Cedar Creek, Indiana, and Upper Big Walnut Creek, Ohio. Measurements of water chemistry, hydrology, and fishes have been collected from 20 sites beginning in 2005. Multiple regression analyses indicated that the relationships between water chemistry and fish communities were weak, but significant (P < 0·05). Fish communities exhibited negative relationships with ammonium and nitrate plus nitrite and positive relationships with dissolved oxygen, pH, and metolachlor. The strongest observed relationships occurred within those regression models that included a combination of nutrients, herbicides, and physicochemical variables. Multiple regression analyses also indicated that five water chemistry variables exhibited significant relationships (P < 0·05) with hydrology. Our results suggest that if water chemistry is the focus of a conservation plan, then the most effective conservation practices may be those that have a combined influence on nutrients, herbicides, and physicochemical variables. Additionally, the use of a combination of conservation practices to address physical habitat and water chemistry degradation is most likely to provide the greatest benefits for fish communities within channelized headwater streams. Copyright © 2009 John Wiley & Sons, Ltd.