Anna Isabella Gonzaga was the daughter of Ferrante III Gonzaga, Duke of Guastalla, and Margherita d'Este (1619–1692), daughter of Duke Alfonso III d'Este. Her marriage was arranged by the assistance of empress dowager Eleanor Gonzaga, and took place in 1671. Anna Isabella Gonzaga was the heir of the Duchy of Guastalla and Luzzara Reggiolo, and her rights transferred these areas, which had been a long concern of conflicts between the two Gonzaga lines, to the Mantua line of the Gonzaga dynasty. The marriage was childless. In 1701, during the War of the Spanish Succession, her consort fled Mantua and appointed her regent of the state in his absence.