The Order of Knights of the Hospital of Saint John of Jerusalem (Latin: Ordo Fratrum Hospitalis Sancti Ioannis Hierosolymitani), commonly known as the Knights Hospitaller (), was a medieval and early modern Catholic military order. It was founded in the Kingdom of Jerusalem in the 12th century and was headquartered there until 1291, thereafter being based in Kolossi Castle in Cyprus (1302–1310), the island of Rhodes (1310–1522), Malta (1530–1798), and Saint Petersburg (1799–1801).

The Hospitallers arose in the early 12th century during the height of the Cluniac movement, a reformist movement within the Benedictine monastic order that sought to strengthen religious devotion and charity for the poor. Earlier in the 11th century, merchants from Amalfi founded a hospital in Jerusalem dedicated to John the Baptist to care for sick, poor, or injured pilgrims to the Holy Land. Blessed Gerard, a lay brother of the Benedictine order, became its head in 1080. After the conquest of Jerusalem in 1099 during the First Crusade, a group of crusaders formed a religious order to support the hospital. Some scholars consider the Amalfitan order and hospital to have been distinct from Gerard's order and its hospital.

The organization became a military religious order under its own papal charter, charged with the care and defense of the Holy Land. Following the conquest of the Holy Land by Islamic forces, the knights operated from Rhodes, over which they were sovereign, and later from Malta, where they administered a vassal state under the Spanish viceroy of Sicily. The Hospitallers were one of the smallest groups to have colonized parts of the Americas, briefly acquiring four Caribbean islands in the mid-17th century, which they turned over to France in the 1660s.

The knights became divided during the Protestant Reformation, when rich commanderies of the order in northern Germany and the Netherlands became Protestant and largely separated from the Catholic main stem, remaining separate to this day; modern ecumenical relations between the descendant chivalric orders are amicable. The order was suppressed in England, Denmark, and other parts of northern Europe, and was further damaged by Napoleon's capture of Malta in 1798, after which it became dispersed throughout Europe.

Today several organizations continue the Hospitaller tradition, specifically the mutually recognized orders of St. John, which are the Sovereign Military Order of Malta, the Most Venerable Order of the Hospital of Saint John, the Bailiwick of Brandenburg of the Chivalric Order of Saint John, the Order of Saint John in the Netherlands, and the Order of Saint John in Sweden.