Rossi had been out for six months and hoped to find form in time to take part in this summer's tournament, but damaged the same right knee ligament today and will be sidelined for another six months.
"I was just on the phone to Beppe. He's distraught and really expected to make a comeback with a chance for the Euros, but now I think he'll go back to the United States," Italy chief medic Professor Enrico Castellacci told Radio Sportiva.
"This is a terrible blow for the athlete and the man, who I really care about. He's a splendid lad and this setback gives us all great pain. I had spoken to him only two days ago and he said he felt good. We were happy to hear his enthusiasm and I probably would've gone to see him personally.
"I perform hundreds of operations on cruciate ligaments every year. He said it was a banal trauma that caused it, but there could be many reasons for the fresh injury and it doesn't seem right to discuss them until we know more details. Meanwhile, the drama of this poor lad is what stands out.
"He will have to undergo another operation and wait those six months to get back to playing football."