But Enrico Castellacci refused to rule out the Juventus centre-back recovering from a left-thigh injury if the Azzurri progress further.
Chiellini, the most experienced defender in Italy's squad with 53 caps, was forced off during the second half of Monday's 2-0 win over Republic of Ireland, which secured a last-eight place.
"The scan he underwent confirmed it's a strain," Castellacci said.
"He will certainly miss the game on Sunday.
"In a competition like this you have to try everything to have him back as quickly as possible but we will not take any unnecessary risks with his muscles."
The 27-year-old had been a doubt for the tournament after suffering a similar injury in Juve's Serie A win against Atalanta last month.
Chiellini was replaced by Juventus team-mate Leonardo Bonucci against Ireland.
Meanwhile, Louis Saha is hoping Laurent Blanc stays as France coach beyond Euro 2012 despite being strongly linked with the vacant position at Tottenham.
The former Spurs striker said: "He's a top manager.
"He'll always have interest. I can imagine him being in the Premier League because of his experience.
"I want him to stay in the France job because he has been doing a terrific job so I wouldn't like him to go."