The Middle Mississippi River (MMR) is unimpounded and free-flowing and extends about 310 rkm from the Missouri River confluence to the confluence with the Ohio River, at Cairo, Illinois. The Lower Mississippi River (LMR) is also free-flowing and extends 1,600 rkm from the Ohio River confluence to its outlet into the Gulf of Mexico.

Sampling the Mississippi River network {#sec004}
--------------------------------------

Water samples were collected in sequence from a series of thirteen sample sites on the Mississippi River extending from just above St. Louis, Missouri to Natchez, Mississippi, a distance of 1,300 rkm. Additionally, samples were collected from the mouths of five tributaries to the Mississippi, each of which drains an important sub-basin. Only one site per day was sampled on the Mississippi starting on July 22, 2013 at the north-mostand Ohio Rivers, the three sections of the Mississippi River (the Upper Mississippi, Middle Mississippi, and Lower Mississippi River) are distinguishable from each other in physical and chemical properties. After one day of flow below confluences with these major tributaries there were usually major shifts in diversity of the Mississippi River microbiome compared to pre-tributary diversity. This is in contrast with stretches of the river more distant in distance and time from major confluences, where changes between adjacent sites in diversity tended to be more gradual. We conclude that while flowing downriver, diversity of the Mississippi River microbiome is modified by point source inputs of external microbiomes from major tributaries draining distinct sub-watersheds, complemented by a gradual process of community succession along intervening stretches of more stable environmental