Star Ospreys trio Shane Williams, Lee Byrne and Jerry Collins will all miss the must-win Magners League finale with Aironi Rugby.
The Magners League Champions need a bonus-point win against Aironi on Friday and other results to go their way if they are to sneak into the play-offs on the final weekend.
Lee Byrne is the biggest injury worry and is also a major doubt for Wales ahead of their clash with the Barbarians on June 4.
The 30-year-old suffered a minor tear to cartilage in his right knee during the Ospreys' defeat to Munster 11 days ago.
It now looks almost certain that the Clermont Auvergne-bound fullback has played his last game for the Swansea-based region and he is unlikely to feature against the Baa-Baas in five week's time.
"We are currently seeking specialist advise for a right knee problem and a decision will be made in the next couple of days regarding management and prognosis," said Ospreys physio Chris Towers.
"It looks unlikely that he will be available to take any further part for the Ospreys this season, even if we make the play-offs, but we remain positive that this will not impact on his availability for the Rugby World Cup."
They will also be without Welsh record try-scorer Williams. The wing wizard has failed in his race to recover from the knee injury he picked up in the Six Nations, before the regular season. But he could come into contention if the Ospreys finish in the top four.
"We've tried but Shane just hasn't made it for this week," added Ospreys head coach Sean Holley.
"If we are fortunate to make the play-offs then he maybe available for that. But this game has come too early for him and us and we'll have to reassess of we qualify but I don’t think there are any issues for later in the summer."
Meanwhile former All Black Collins also looks to have played his last game for the Liberty Stadium outfit.
The fearsome backrower picked up his third yellow card of the season in the defeat to Munster and has subsequently been given a one-match suspension with a disciplinary hearing still to come.
"It's a sad way for Jerry to go out. He's been a good servant for us and he's worked hard here and been a big influence in big games.
"It wasn’t a great one in that game but it's a sad way to go and it’s been a great experience to have Jerry here."