Mattia Zaccagni became the latest player to leave beleaguered Italy's Euro 2024 qualifying camp with an ankle injury ruling the Lazio midfielder out of next week's game against England.
Zaccagni had already been sidelined for Saturday's Group C fixture with Malta but had been hoping to be available for the European champions' trip to Wembley to take on runaway leaders England.
"Following tests carried out yesterday (Friday) and today, Mattia Zaccagni cannot be available for Tuesday's match in London against England," Italy's football federation (FIGC) said in a short statement.
The FIGC did not say if coach Luciano Spalletti would be calling up another player to replace Zaccagni, 28, who has three international caps.
Spalletti has already lost striker Federico Chiesa to a thigh problem and suffered the ignomy of Sandro Tonali and Nicolo Zaniolo being questioned for two hours by criminal prosecutors at Coverciano over alleged gambling offences.
Stephan El Shaarawy was brought in on Friday to plug the gap left by Newcastle United midfielder Tonali and Aston Villa forward Zaniolo, who were both released from Italy's training camp.
Spalletti's team began the day second in the group on seven points, six behind England and level with Ukraine and North Macedonia who face off on Saturday afternoon in Prague.
Italy also have a game in hand on every other team in the five-team group.