Fiorentina coach Vincenzo Montella spent 10 years at Roma during his playing days and also had a spell as caretaker coach at Stadio Olimpico, but there will be no room for sentiment when his side attempt to dump the Eternal City club out of Europe.
Montella's side, who knocked out Tottenham in the round of 32, will be at home for the first leg on March 12 and travel to Rome a week later.
Fiorentina knocked Roma out of the Coppa Italia earlier this month, so Rudi Garcia's side have an opportunity to exact revenge.
Sevilla were crowned champions when they beat Benfica last season and Unai Emery's side knocked out Borussia Monchengladbach to keep their hopes of lifting the trophy again alive.
Everton v Dynamo Kiev
Dnipro v Ajax
Zenit v Torino
Villarreal v Sevilla
Napoli v Dinamo Moscow
Fiorentina v Roma
