The vegetation of the PPR consists of emergent plants and tall grasses, while the prairie surrounding the region has dense grassland vegetation. The composition of a local plant community is heavily affected by the amount of water available. In wetter wetlands that retain water through the summer, the common plant is hard-stem bulrush, along with soft-stem bulrush and common threesquare in slightly drier regions of the wetlands. The vegetation in permanently flooded wetlands is more aquatic; duckweeds, pondweeds, aquatic buttercups, and aquatic smartweeds are some of the most common. In drier wetlands of the PPR, the vegetation varies from spikerush, which is found in the wetter areas of the wetland, to foxtail barley and wheatgrass on the outer edges of the wetland. The variable availability of water in the Prairie Pothole Region is buffered by an abundant seed bank under the soil, comprising species that thrive under different water regimes. During times of drought, shortgrass species increase and expand their range, while during wetter periods, tallgrass and mixed prairie communities become more common.

The Prairie Pothole Region provides important habitats for migratory waterfowl and other wildlife, supporting more than 50% of North America's migratory waterfowl. In particular, the region is one of North America's most important breeding areas for ducks. Although the region contains only about one-tenth of the continent's habitat area for breeding of waterfowl, roughly half the primary species of game ducks on the continent breed there. The region accounts for more than 60% of the breeding populations of mallard, gadwall, blue-winged teal, northern shoveler, northern pintail, redhead, and canvasback ducks.

Conversion of land for agriculture is a significant factor in the loss of wetlands in the PPR. More than half of the wetlands have been drained for farming. In particular, 90% of the prairie wetlands of the Minnesota River basin have been lost as habitat. The wetlands that do persist, surrounded as they are by agricultural lands, are also affected. Chemical runoff, sedimentation, and nutrient flow into the wetlands have adverse impacts.

Climate change is an adverse factor in the long-term viability of PPR wetlands for breeding ducks and other birds. Without mitigation, severe droughts and rising temperatures will cause many pothole wetlands to dry up sooner in the spring. In turn, due to the timing of waterfowl migrations, these dried wetlands will not present suitable breeding habitat. Warming-related drought may affect as much as 90 percent of the PPR's remaining wetlands. Simulations suggest that climate change will shift the most productive wetlands from the center of the region (southeastern Saskatchewan and the Dakotas) to edges of the PPR in the east and north. However, research has suggested that the effect of global warming is overshadowed by that of intensified land use and drainage of wetlands.