Like other native groups decimated by European diseases and warfare, the Five Nations of the Iroquois suffered as a result of colonization, but they were able to capitalize on their strategic location in central New York to dominate the region between the French and Dutch colonies. Obtaining guns and goods from Dutch merchants at Fort Orange, Iroquois warriors inflicted terror on their neighbors. Partly in response to a virulent smallpox epidemic in 1633, which cut their number by one-third, the Iroquois waged a series of devastating wars against the Hurons (1649), Neutrals (1651), Eries (1657), and Susquehannocks (1660) — all Iroquoian-speaking peoples. They razed villages, killing many residents and taking many more captive. The conquered Hurons ceased to exist as a distinct people; survivors trekked westward with displaced Algonquian peoples and formed a new nation, the Wyandots. Iroquois warriors pressed still farther — eastward into New England, south to the Carolinas, north to Quebec, and west via the Great Lakes to the Mississippi — dominating Indian groups along the way. Collectively known as the Beaver Wars, these Iroquois campaigns dramatically altered the map of northeastern North America.

Many Iroquois raids came at the expense of French-allied Algonquian Indians, and in the 1660s New France committed to all-out war against the Iroquois. In 1667, the Mohawks were the last of the Five Nations to admit defeat. As part of the peace settlement, the Five Nations accepted Jesuit missionaries into their communities. A minority of Iroquois — perhaps 20 percent of the population — converted to Catholicism and moved to the St. Lawrence Valley, where they settled in mission communities near Montreal (where their descendants still live today).

The Iroquois who remained in New York did not collapse, however. Forging a new alliance with the Englishmen who had taken over New Netherland, they would continue to be a dominant force in the politics of the Northeast for generations to come.