Willingham was slotted to be the designated hitter in the series opener Monday night. But the Twins say he experienced increasing discomfort in his knee as he went through his pregame routine. He is scheduled to have an MRI on Tuesday.
"It hasn't got any better, might have even got a touch worse, I don't know," Willingham told the Minneapolis Star Tribune. "I thought [a cortisone shot] helped there for a while, but then I tweaked it again, and it hasn't gotten better.
"Today I didn't run during batting practice, and I thought I felt OK," Willingham added to the Star Tribune. "I went to run before the game, and it wasn't happening."
Information from The Associated Press was used in this report.