The Germany international had returned to training after initially tearing the cruciate in his right knee against Borussia Dortmund in December last year.
But while his team-mates were concluding their title-winning Bundesliga season with a 4-3 away win over Borussia Monchengladbach on Saturday afternoon, Badstuber was reeling from the news of a fresh rupture which will require additional surgery.
Bayern director of sport Matthias Sammer told the club's website: "Obviously, we'll give him all the time in the world to get fit. We're optimistic. I'm confident he'll be back, stronger than ever."