Leighton Baines has pulled out of the England squad to face Slovenia and Scotland because of a hamstring injury.
The Everton defender's withdrawal leaves Arsenal's Kieran Gibbs as the only left-back available for Saturday's Euro 2016 qualifier against Slovenia at Wembley.
Manchester United's Luke Shaw was part of the original 26-man squad but was not retained when it was cut to 23.
England face Scotland in a friendly at Celtic Park next Tuesday.
Uefa regulations do not allow Shaw to be added to the squad against Slovenia, but he is eligible to play against Scotland.
England manager Roy Hodgson suggested that Baines's injury might be "something which has been niggling away because Everton have played a lot of games recently".
Hodgson also expressed frustration that he was not allowed to restore Shaw to the squad, saying: "My hope was that Uefa would look at it and show some flexibility but they decided that the rule was the rule."
Baines, 29, was England's first-choice left-back at this summer's World Cup in Brazil and has started three of their four matches since the tournament. He has won 29 caps.
Gibbs played in the 5-0 Euro 2016 qualifying win over San Marino at Wembley in October, his fourth England appearance.
Midfielders Andros Townsend and Michael Carrick withdrew from the squad this week with ankle and groin injuries respectively.
Baines's injury will concern Everton manager Roberto Martinez, who is missing defenders John Stones and Antolin Alcaraz with ankle and shoulder injuries respectively, while reserve left-back Bryan Oviedo has recently returned from a broken leg.