Nitrate concentration and flux were estimated from 1980 through 2010 at eight sites in the Mississippi River Basin as part of the National Water-Quality Assessment (NAWQA) Program of the U.S. Geological Survey (USGS). These estimates extend the results from a previous investigation that provided nitrate estimates from 1980 through 2008 at the same sites. From 1980 through 2010, annual flow-normalized (FN) nitrate concentration and flux in the Iowa and Illinois Rivers decreased by 11 to 15 percent. These two rivers had the highest FN nitrate concentration in 1980 (5.3 milligrams per liter [mg/L] and 3.9 mg/L, respectively) of any of the study sites. Nitrate increased in the Missouri River (79 and 45 percent increase in FN concentration and flux, respectively), and at the four sites on the Mississippi River (17 to 70 percent increase in FN concentration and 8 to 55 percent increase in FN flux) from 1980 through 2010. Nitrate in the Ohio River was generally stable during this time. Historically, nitrate was high and changed little in the Iowa and Illinois Rivers; however, nitrate concentrations began to decrease around 2000, and this decrease continued through 2010. Also during this time, near-flat nitrate trends in lower sections of the Mississippi River began increasing, likely reflecting the acceleration of already increasing nitrate trends in the upper Mississippi and Missouri Rivers, in addition to increases in inputs from other tributaries in the lower part of the Mississippi River Basin. Spring trends (April through June) generally parallel annual trends at all sites from 1980 through 2010, except in the Iowa River where decreasing nitrate during the spring was not observed. In general, most sites had increases in nitrate concentration at low streamflows, which suggests increases in legacy nitrate from groundwater or point source contributions. In aggregate, the decreases in nitrate concentrations from the Iowa and Illinois Rivers, which largely occurred during high flows, appear to be overshadowed by increasing nitrate concentrations across much of the Mississippi River Basin.