Everton's Barkley, 19, and Tottenham Hotspur's Townsend, 22, received their first senior call-ups and Southampton forward Rickie Lambert was given another chance after scoring on his debut against Scotland two weeks ago.
Liverpool striker Daniel Sturridge, who missed that match due to injury, was recalled to the squad and Celtic goalkeeper Fraser Forster replaced the injured Ben Foster.
England face Moldova at Wembley on September 6 before travelling to Kiev to play Ukraine on September 10.
England squad
Goalkeepers: Fraser Forster (Celtic), Joe Hart (Manchester City), John Ruddy (Norwich City).
Defenders: Leighton Baines (Everton), Gary Cahill (Chelsea), Ashley Cole (Chelsea), Phil Jagielka (Everton), Glen Johnson (Liverpool), Phil Jones (Manchester United), Chris Smalling (Manchester United), Kyle Walker (Tottenham Hotspur).
Midfielders: Ross Barkley (Everton), Michael Carrick (Manchester United), Tom Cleverley (Manchester United), Steven Gerrard (Liverpool), Frank Lampard (Chelsea), James Milner (Manchester City), Raheem Sterling (Liverpool), Andros Townsend (Tottenham Hotspur), Theo Walcott (Arsenal), Jack Wilshere (Arsenal).
Forwards: Jermain Defoe (Tottenham Hotspur), Rickie Lambert (Southampton), Wayne Rooney (Manchester United), Daniel Sturridge (Liverpool), Danny Welbeck (Manchester United), Ashley Young (Manchester United).
(Reporting by Tom bartlett, editing by Ed Osmond)
