The pilot of the plane was also killed in the crash Saturday in Guntersville.
The death toll rose to three Sunday morning when 4-year-old Hailee Day succumbed to her injuries at Huntsville Hospital, WHNT-TV, Huntsville, Ala., said.
Her brother, Gavin Whitacre, was listed in critical condition.
WHNT said the victims, all from Xenia, Ohio, were flying from Nashville to Florida. The plane went down after takeoff following a refueling stop in Guntersville.
Senate Democrats to pull all-nighter on climate change
Susan Sarandon 'very excited' about daughter's pregnancy