In front of the watching Roy Hodgson the 24-year-old, who has scored 15 Premier League goals this season, landed awkwardly after jumping for the ball after 26 minutes at the Etihad Stadium.
Team-mates and fans watched on with concern as a distraught Rodriguez's right leg was put in a knee brace before he was taken from the field to a sympathetic ovation from the Etihad Stadium crowd.
England Under-21 international James Ward-Prowse came on to replace the injured Rodriguez and speaking after the game Southampton manager Mauricio Pochettino admitted the injury looked serious.
"To be completely honest, it's not looking good," he told reporters.
"We need to assess him today and tomorrow but it's not looking good for him it's not going to be good for the team.
"All the players are really affected, not just about the result not just about the decisions but more about how severe his injury is."Rodriguez has made one appearance for the senior England side, in a friendly defeat to Chile at Wembley in November.