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Streams draining karst areas with rapid groundwater transit times may respond relatively quickly to nitrogen reduction strategies, but the complex hydrologic network of interconnected sinkholes and springs is challenging for determining the placement and effectiveness of management practices. This study aims to inform nitrogen reduction strategies in a representative agricultural karst setting of the Chesapeake Bay watershed (Fishing Creek watershed, Pennsylvania) with known elevated nitrate contamination and a previous documented groundwater residence time of less than a decade. During baseflow conditions, streamflow did not increase with drainage area. Headwaters and the main stem lost substantial flow to sinkholes until eventually discharging along large springs downstream. Seasonal hydrologic conditions shift the flow and nitrogen load spatially among losing and gaining stream sections. A compilation of nitrogen source inputs with the geochemistry and the pattern of enrichment of δ 15 N and δ 18 O suggest that the nitrogen in streams and springs during baseflow represents a mixture of manure, fertilizer, and wastewater sources with low potential for denitrification. The pH and calcite saturation index increased along generalized flow paths from headwaters to springs and indicate shorter groundwater residence times in baseflow during the spring versus summer. Given the substantial investment in management practices, fixed monitoring sites could incorporate synoptic water sampling to properly monitor long-term progress and help inform management actions in karst watersheds. Although karst watersheds have the potential to respond to nitrogen reduction strategies due to shorter groundwater residence times, high nitrogen inputs, effectiveness of conservation practices, and release of legacy nutrients within the karst cavities could confound progress of water quality goals., (© ([0‐9]+) The Authors. Journal of Environmental Quality © ([0‐9]+) American Society of Agronomy, Crop Science Society of America, and Soil Science Society of America.)