Match ends, Czech Republic 2, England 1.
England must wait to confirm their place in Euro 2020 after the Czech Republic deservedly ended their 43-game unbeaten run in qualifiers stretching back 10 years.
Gareth Southgate's side would have secured their place in next summer's tournament with victory in Prague but they can have no complaints after a wretched display against a Czech side who were a different proposition from that swept aside 5-0 at Wembley in March.
England can still qualify on Monday if they beat Bulgaria and Kosovo do not beat Montenegro.
England were handed the perfect start when captain Harry Kane put them ahead from the penalty spot in the fifth minute after Lukas Masopust fouled Raheem Sterling but they were well below their best and slumped to a bitterly disappointing defeat.
The Czechs were swiftly level when Jakub Brabec scored following a corner. England could not muster further inspiration and substitute Zdenek Ondrasek pounced to score the winner with four minutes left.
England were desperately poor throughout and were grateful to goalkeeper Jordan Pickford, who produced fine saves from Vladimir Coufal, Masopust and Alex Kral, while Czech counterpart Tomas Vaclik did well to deny Sterling and Kane.
They can put things right against Bulgaria in Sofia on Monday but this was a serious reality check for a side hoping to do great things next summer, losing a qualifier for the first time since they went down 1-0 to Ukraine on 10 October 2009.
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England deserve no better
This must rank as one of the worst performances of Southgate's reign and the manager himself has to take his own share of the responsibility.
No-one can complain about his decision to give Chelsea's in-form Mason Mount his debut ahead of team-mate Ross Barkley but the youngster never flourished in an advanced midfield position in the first half.
Mount was barely able to influence the game and with Jordan Henderson and Declan Rice exposed and pedestrian, England found themselves often overrun by the sprightly Czechs.
England's potent attacking trio of Kane, Sterling and Jadon Sancho were starved of service, leaving them under-performing in every area of the pitch.
The Czech Republic, backed by a noisy crowd in Prague, gathered momentum and confidence and it was no surprise when Ondrasek finally broke Pickford's resistance late on.
This was simply not good enough from England and the concerns that surfaced about their true quality when they won 5-3 in chaotic style against Kosovo in Southampton will only increase after this.
Defensive woes continue
Any hope the defensive uncertainty that characterised England's victory against Kosovo had been successfully addressed was banished inside the first 10 minutes in Prague.
Pickford saved brilliantly from Coufal as the Czech Republic responded to England's early salvo but there was the trademark confusion at the resulting corner which ended with Brabec stabbing home from close range.
England lived dangerously throughout and it was not a shock when they finally conceded late on.
Everton's Michael Keane struggles desperately at this level while full-backs Danny Rose and Kieran Trippier hardly covered themselves in glory either.
Harry Maguire looks the one staple in defence but he does not exude confidence either.
In other words, quality sides will relish facing this England defence and Southgate is running out of time to apply the fix.
Southgate = Hodgson - the stats
- Czech Republic had 17 shots in this match, the most England have faced in a qualifying match since March 2013, when Montenegro had 19 in a World Cup qualifier.
- This was England's first defeat in a European Championship qualifier since losing 3-2 at Wembley against Croatia in November 2007.
- Harry Kane has scored 20 goals in 21 matches for England when starting as captain - the only player with more is Vivian Woodward (23 goals between 1908 and 1911).
- Excluding shootouts, no player has scored more penalties for England than Harry Kane (9, level with Frank Lampard).
- Zdenek Ondrasek scored on his international debut for Czech Republic, scoring with his first shot in international football.
- Gareth Southgate (W21 D9 L8) has lost as many games as England manager in 38 matches as Roy Hodgson lost in 56 matches as manager (W33 D15 L8).
England travel to Bulgaria on Monday (kick-off 19:45 BST) looking to secure a spot at Euro 2020.
- 20MasopustSubstituted forZmrhalat 90'minutes
- 14JanktoBooked at 8minsSubstituted forKopicat 83'minutes
- 19SchickSubstituted forOndrasekat 65'minutes
- 3RoseBooked at 10mins
- 8HendersonBooked at 90mins
- 4RiceSubstituted forAbrahamat 88'minutes
- 10SanchoSubstituted forRashfordat 73'minutes
- 11MountSubstituted forBarkleyat 72'minutes
- 7SterlingBooked at 68mins
- Damir Skomina
- Shots on Target
Second Half ends, Czech Republic 2, England 1.
Attempt saved. Jan Kopic (Czech Republic) left footed shot from the centre of the box is saved in the top left corner. Assisted by Jaromir Zmrhal.
Jordan Henderson (England) is shown the yellow card for a bad foul.
Ross Barkley (England) wins a free kick in the defensive half.
Foul by Vladimír Darida (Czech Republic).
Harry Maguire (England) wins a free kick in the defensive half.
Foul by Zdenek Ondrasek (Czech Republic).
Substitution, Czech Republic. Jaromir Zmrhal replaces Lukas Masopust.
Substitution, England. Tammy Abraham replaces Declan Rice.
Goal! Czech Republic 2, England 1. Zdenek Ondrasek (Czech Republic) right footed shot from the centre of the box to the bottom right corner. Assisted by Lukas Masopust.
Danny Rose (England) wins a free kick on the left wing.
Foul by Alex Kral (Czech Republic).
Substitution, Czech Republic. Jan Kopic replaces Jakub Jankto.
Corner, England. Conceded by Jan Boril.
Attempt saved. Harry Kane (England) right footed shot from a difficult angle on the right is saved in the centre of the goal. Assisted by Ross Barkley with a through ball.
Raheem Sterling (England) wins a free kick on the right wing.
Foul by Jan Boril (Czech Republic).
Corner, Czech Republic. Conceded by Jordan Pickford.
Attempt saved. Alex Kral (Czech Republic) right footed shot from the right side of the box is saved in the top right corner. Assisted by Vladimír Darida.