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Lyon surprised virtually everyone in the ground when he hobbled out to bat at No. 11 as his team looked to add more runs to their lead over England.
Lyon, 35, who became the first bowler of any type to play 100 consecutive Tests when he took the field at Lord's, pulled up in pain while jogging in from the boundary at square leg to field a top-edged pull from England opener Ben Duckett on Thursday.
Lyon, who is playing in his 100th consecutive Test, was running in to take a catch of Ben Duckett off the bowling of Cameron Green in the leg-side after tea, when he pulled up sharply while running towards the ball.
The Indians had their back to the wall once they lost five wickets inside the first hour and were bundled out for 109 in just 33.2 overs after opting to bat in Indore.
Needing only five runs to win, opener David Warner hit Ramesh Mendis for a four and followed it with a six to seal Australia's victory in four deliveries.
The 34-year-old took the prized wickets of Imam-ul-Haq, Azhar Ali and Babar Azam to end Pakistan's resistance and finished with figures of 5/83 in the second innings.
Lyon became only the third Australian bowler to join the 400-club after legspinner Shane Warne (708) and paceman Glenn McGrath (563).
Resuming on 220/2 with the match still within their grasp, England lost their last eight wickets for 77 runs to be bowled out for 297 by lunch, leaving Australia needing only 20 runs to win.
Lyon, who has played 100 Test matches, is just one short of 400 wickets while Ashwin has taken 409 wickets in 78 Test matches.
Several former England players questioned the suitability of the pitch after 30 wickets fell in five sessions in what was the shortest completed Test match since 1935.