Di Matteo was brought in over the close season with the target of leading Villa back into the Premier League following relegation in May.
But the former Schalke, West Brom and Chelsea boss – he led the London club to the Champions League title in 2012 – departs with Villa in 19th place, having won just one of their opening 11 games.
A club statement read: “Aston Villa Football Club has parted company with manager Roberto Di Matteo.
“The club decided to act following a run of disappointing results which has the left the team occupying 19th position in the Championship.
“The club would like to place on record its appreciation to Roberto for all his efforts in helping rebuild the squad and wish him well for the future.
“Steve Clarke will take over as caretaker manager during the search for a new manager.”
Should Villa fail to appoint a full-time successor to Di Matteo in the meantime, Clarke’s first game in charge will be the derby clash with Wolves on October 15.