The opening day at Edgbaston came just 18 days after an amazing Lord's final against New Zealand, as England became world champions following a dramatic Super Over.
Six members of that ODI side featured in Birmingham, although their stand-out performers against Australia, Stuart Broad – who took 5-86 in the first innings – and centurion Rory Burns, were not among them.
Root, however, instead blamed an inability to capitalise on an early position of strength, having reduced Australia to 122-8 in their first innings, for England's 251-run defeat.
"No, I think that [World Cup fatigue] is a bit of an excuse if I'm being honest," he said at the post-match presentation.
"As I said we played some good stuff throughout the game, we just didn't do it for long enough and we didn't take our chance right at the start of the game when we had them in that position."
The returning Steve Smith proved the difference by becoming just the third Australian to make centuries in each innings of an away Ashes Test.
England were also hindered by a calf injury sustained by James Anderson, which Root felt had an influence on the outcome of the Test.
"Yeah it was two brilliant innings [from Steve Smith], we have to keep working hard on getting him out," Root added.
"It was hard losing Jimmy early on, it becomes a balancing act, these things happen in cricket."
Anderson's selection was a dilemma for England before the Test with the seamer having sustained a similar injury last month, but Root defended the decision to select their all-time leading wicket taker.
"It was a group decision in terms of selection, he passed a fitness test [and] it's just one of those freak things that can happen in cricket," Root said.
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