The forward landed awkwardly during Saturday's 4-1 defeat at Manchester City and Southampton confirmed he has ruptured his anterior cruciate ligament.
The 24-year-old was in the running for a spot in England's 23-man squad, having enjoyed a fine season with Saints in which he netted 17 goals.
Such form saw Rodriguez handed his Three Lions debut in November's friendly against Chile, but his hopes of a spot on the plane to Brazil have been cruelly taken away.
Team-mate and fellow England hopeful Adam Lallana said: "Forget the decision for the offside, forget the result, forget everything. We're just disappointed for our team-mate getting injured. "We're all absolutely devastated for him – he's an unbelievable lad. He's proven this season what a player he is and he's got his rewards internationally."
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