The on-loan Norwich midfielder hurt his ankle in training and could now miss the crunch qualifier at the start of next month as Scotland bid to book a place in Qatar.
Gilmour is expected to miss Norwich's final game of the Premier League season against Tottenham on Sunday after being sent back to parent club Chelsea for a scan.
The 20-year-old has endured a difficult campaign with the relegated Canaries, but the former Rangers youngster has been an influential figure for Scotland since making his international breakthrough at Euro 2020 last year.
His injury will be of concern to national team boss Steve Clarke, who is already set to be without Arsenal left-back Kieran Tierney after his season was ended by a knee injury.
Norwich boss Dean Smith said: "Gilmour rolled his ankle and has gone back to Chelsea for an assessment today.
"We’ll wait and see if he gets back or not."
Scotland face Ukraine at Hampden on June 1 with the play-off final against Wales taking place four days later.