Villanueva was scheduled to undergo an MRI on Thursday but the results were not immediately available. Even without the final diagnosis, the club knew a roster move needed to be made.
"This is probably as much fatigue as it is a strain," Blue Jays manager John Farrell said following the move. "I don't have the current results ... but the move was clear that we had to make a spot with Carlos going to the disabled list."
Villanueva surrendered a season-high eight runs on nine hits in just 2 2/3 innings against Tampa Bay on Wednesday. He has now thrown 97 2/3 innings on the year, which is his highest total since 2008.
The 27-year-old began the season in the bullpen, and it's possible the high number of innings has finally caught up to him.
There weren't any obvious signs of a major injury, but the club wanted Villanueva to be examined for an MRI just to make sure there isn't any structural damage in his right arm.
"He wasn't feeling discomfort," Farrell said earlier in the day. "I think just physically he is starting to fatigue. You see it with a little bit less velocity, particularly the action to the secondary stuff.
"That's where you see the reduced arm speed not allowing that good bottoming out action to his changeup or any sharpness to his slider."
Toronto recalled left-hander Luis Perez from Triple-A Las Vegas to take Villanueva's spot on the 25-man active roster. Perez will move into the bullpen, where he posted a 3.54 ERA in 35 1/3 innings with the club earlier this season.
The Blue Jays have an off-day scheduled for Monday and the club has opted to move forward with a four-man rotation on a temporary basis.
A fifth starter would not be needed until Aug. 13 against the Angels. Prior to Thursday's 7-6 loss to the Rays, Farrell mentioned Perez and top pitching prospect Henderson Alvarez as potential candidates to fill the vacancy left behind by Villanueva in the rotation.
Alvarez began the season with Class A Dunedin before making the jump to Double-A New Hampshire. The 21-year-old is 7-4 with a 2.82 ERA and 61 strikeouts in 14 games with the Fisher Cats.