England are currently sitting top of Group A on 12 points, having won all four of their fixtures to date.
The striker is joined in the 25-man squad by Chelsea team-mate Tomori, who has also broken into Frank Lampard's first team at Stamford Bridge in recent weeks.
There is, however, no place for City right-back Walker, who misses out for the second time in the space of two months, despite his impressive displays for Pep Guardiola's side this term.
Southgate has placed his faith in the likes of Ben Chilwell, Declan Rice, Mason Mount and Callum Wilson once again, while James Maddison and Tyrone Mings will hope to earn their first caps after being called up for a second consecutive round of fixtures.
United striker Marcus Rashford makes the final cut despite his recent injury problems, with Borussia Dortmund's Jadon Sancho and City's Raheem Sterling poised to link up once again in the attack either side of Tottenham talisman Harry Kane.
If England pick up another win in Prague next Friday, their spot in next summer's European championships will be guaranteed, which might give Southgate the opportunity to blood the younger members of his squad in Bulgaria the following Monday.
Goalkeepers: Tom Heaton, Jordan Pickford, Nick Pope
Defenders: Trent Alexander-Arnold, Ben Chilwell, Joe Gomez, Michael Keane, Harry Maguire, Tyrone Mings, Danny Rose, Fikayo Tomori, Kieran Trippier
Midfielders: Ross Barkley, Fabian Delph, Jordan Henderson, James Maddison, Mason Mount, Declan Rice, Harry Winks
Forwards: Tammy Abraham, Harry Kane, Marcus Rashford, Jadon Sancho, Raheem Sterling, Callum Wilson