Willis was placed on the season-ending injured reserve list Tuesday for the injury suffered in the third quarter at St. Louis on Oct. 13.
A Pro Bowler all seven seasons in the NFL, Willis had missed six games in his career before this latest injury sidelined him for the final 10 games in 2014 for San Francisco (5-4). He had 34 tackles and an interception this season for the Niners.
Willis is a captain and leader of a 49ers team that has reached the NFC championship game in each of the past three seasons.
"We haven't lost him in the locker room," linebacker Michael Wilhoite said. "We're still all together here. It hurts but at same time we need to keep going out and win games."
Willis had hoped the injury wasn't so serious to sideline him long term. He missed practice last week after returning in a limited role the previous week.
After resting through the Oct. 26 bye, Willis was back as part of the 49ers' full active squad on the field for the start of practice Oct. 29, but he had been limited primarily to individual work.
This is a big loss to one of the NFL's leading defences, though the 49ers are expected to have Aldon Smith back Sunday on the road against the New York Giants.
Smith is set to begin practicing this week after serving his nine-game suspension by the NFL for violations of the league's substance-abuse and personal-conduct policies.
In addition, 2013 All-Pro NaVorro Bowman could be cleared to begin practice this week, opening a 21-day window for the team to activate him or place him on IR.
Bowman is recovering from surgery on his left knee after he tore the anterior cruciate and medial collateral ligaments in a devastating injury in the NFC championship game loss at Seattle last January.
Rookie Chris Borland has filled in for Willis and been spectacular, recording 18 tackles in a loss to the Rams two weeks ago then 17 more tackles in an overtime win at New Orleans on Sunday.
The NFL Network first reported Willis would need toe surgery.
The Niners signed running back Alfonso Smith to replace Willis on the roster and added linebacker Shayne Skov to the practice squad.
AP freelancer Rick Eymer in Santa Clara, California, contributed to this report.
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