The backbone of the Italy team is the same as in their opening group-stage victories over Belgium and Sweden, with Leonardo Bonucci, Andrea Barzagli and Giorgio Chiellini starting in defence.
Central midfielders Daniele De Rossi, Marco Parolo and Emanuele Giaccherini will line up in front of the back three while the striking duties fall to Eder and Graziano Pelle as Lorenzo Insigne starts on the bench.
Alessandro Florenzi is on the right flank with Mattia De Sciglio retaining his spot on the left after being included in the starting team for their last group game against Ireland following Antonio Candreva's injury.
Ten Italy players, including six in the starting line-up, are one booking away from missing the quarter-finals.
Spain coach Vicente Del Bosque kept faith in the side that started the three previous games, with Alvaro Morata up front supported by David Silva and Nolito.
Sergio Busquets, Cesc Fabregas and Andres Iniesta start in midfield while Sergio Ramos and Gerard Pique marshal the defence.
Spain, who thrashed Italy 4-0 in the Euro 2012 final, have not lost against the Azzurri in a competitive game since 1994, when Conte was in the team.
Original source: #Euro2016: Italy field five-man mid against Spain