The U.S. Geological Survey, in cooperation with the U.S. National Park Service, conducted a study to determine the presence of algal toxins and Mycobacterium avium ssp. paratuberculosis (MAP) within Pipestone National Monument located in Pipestone County in southwestern Minnesota. Three algal toxins (microcystins, saxitoxin, and anatoxin-a) were analyzed in 12 water-quality samples collected from 8 surface and 2 quarry-water sample sites by enzyme-linked immunosorbent assay (ELISA) methods. Two of the three analyzed algal toxins (microcystins and anatoxin-a) were also analyzed using immunosorbent strip test methods. All analysis kits were provided by Abraxis Inc. (Warminister, Pa) and analyses were completed at the U.S. Geological Survey - Upper Midwest Water Science Center in Mounds View, Minnesota. MAP was analyzed in 16 water-quality samples and 8 sediment samples collected from 8 surface-water sample sites by multiple real-time polymerase chain reaction (PCR) methods at the Wisconsin Veterinary Diagnostic Laboratory at the University of Wisconsin-Madison in Madison, Wisconsin. These data are a part of a larger study and an associated report documented in the U.S. Geological Survey Investigations Report 2023-XXXX (http://doiXXXXXplaceholder).