2012 Annual Report 1a.Objectives (from AD-416): The objective of this research project will be to evaluate the influence of nutrient and pesticide mixtures on the biota within agricultural headwater streams in the midwestern United States. 1b.Approach (from AD-416): To fulfill the objective of this project, a three tiered approach will be taken. a) We will initiate and conduct a multi-scaled evaluation of the prevalence of endocrine disruption in fishes in the midwestern United States. Specifically, this research component will consist of two parts – a three year study within St. Joseph River watershed study sites and a large scale one year study encompassing ten watersheds. This will involve the use of physiological and morphological biomarkers to measure the degree of endocrine disruption in selected fish species within the St. Joseph River watershed. This component will be conducted in FY 2011 and FY 2012 in conjunction with the ecological field sampling and the streamside bioassays. b) We will initiate and conduct streamside bioassays at selected locations in the St. Joseph River CEAP study sites in FY 2011 and FY 2012. The streamside bioassays will enable us to expose laboratory reared fishes and invertebrates to actual nutrient and pesticide mixtures that occur in the study watersheds. The test organisms have not experienced prior exposure to nutrient and pesticide mixtures. Thus the streamside bioassays enable us to evaluate how survival, growth, and reproductive characteristics of “naïve” stream organisms respond to the different nutrient and pesticide mixtures that occur among our study watersheds. c) We will conduct ecological sampling of the St. Joseph River watershed study sites in FY 2011 through FY 2013 as described in Smiley et al. (2008) and Smiley et al. (2009). The additional three years of data collection will result in an eight year database on nutrients, pesticides, fishes, and invertebrates. 3.Progress Report: As of May, FY12, we have completed six years of annual ecological assessments in the Cedar Creek Watershed Conservation Effects Assessment Project (CEAP) study area. Data for fish communities and habitat for sampling years 2006-2011 have been entered into an Excel database. Data for macroinvertebrate communities for sampling years 2006 and 2007 have been entered into an Excel database. During FY12 we also conducted year two of biomarker surveys at four sites in the Cedar Creek Study Area and four sites in the Upper Big Walnut Creek Study Area to assess the effects of exposure to water from agricultural ditches on the biology of stream fishes. Preliminary analyses of relationships among water chemistry, habitat variables and aquatic communities have been made.