Gareth Southgate confirmed Eric Dier will miss England's European Championship qualifier against Montenegro on Monday through injury.
The 25-year-old managed to carry on briefly but was eventually forced off after 26 minutes and replaced by Ross Barkley.
The former Sporting Lisbon youngster will now return to Tottenham for further treatment with the club facing a nervous wait to see if he will recover in time for their trip to leaders Liverpool in the Premier League next weekend.
Speaking to reporters after the game Southgate confirmed Dier would definitely miss Monday’s trio to Podgorica but said he would wait to assess the rest of the squad before deciding whether to call-up any replacements.
"I think he will go back to his club. Hopefully not too serious, bit of a muscular injury but he won't be right for our second game,” said the Three Lions boss.
"If Eric is the only injury I don't think we'll call anyone else in. If we have any other concerns we'll have to review that on Saturday.”
Chelsea midfielder Barkley is in line to start Monday’s game after replacing Dier at Wembley. Another option though could be Declan Rice, who came on as a second-half substitute for his England debut after switching allegiance from the Republic of Ireland.
Southgate has already had to deal with a number of withdrawals to his squad, with Luke Shaw, Ruben Loftus-Cheek, John Stones and Fabian Delph all returning to their clubs due to injury.
James Ward-Prowse and Callum-Hudson Odoi were added to the squad, with the latter coming on as a substitute for his Three Lions debut.
Despite those setbacks, England comfortably overcame the Czechs at Wembley, with Raheem Sterling leading the way by scoring a hat-trick.
The Manchester City winger converted Jadon Sancho’s cross for the opener with Harry Kane’s penalty doubling their lead shortly before half-time.
Two goals in six second-half minutes completed Sterling’s hat-trick before a late own goal from defender Tomas Kalas’ rounded off a convincing win.