Sunderland have confirmed striker Steven Fletcher and midfielder Lee Cattermole will miss the rest of the season due to injury.
Leading scorer Fletcher suffered ankle ligament damage on international duty with Scotland last week, and midfielder Cattermole underwent surgery on a persistent knee problem earlier today.
A club statement said: "Sunderland have confirmed today that striker Steven Fletcher and captain Lee Cattermole will miss the rest of the current campaign due to injury."
The statement added: "Cattermole underwent an operation on his knee earlier today and Fletcher sustained ankle ligament damage whilst on international duty with Scotland.
"It is not known at this stage if Fletcher will require an operation, with the injury requiring further assessment in the coming weeks.
"Both players will play no further part in the Black Cats' 2012-13 Barclays Premier League season."
The news could hardly have come at a worse time for O'Neill, whose side finds themselves firmly entrenched in a battle for top-flight survival with eight games of the campaign to play.
They face leaders Manchester United at the Stadium of Light on Saturday desperate to end a run of seven league games without a victory which has yielded just three points and left them only four clear of the drop zone.
Fletcher in particular will be a huge loss after paying off a large proportion of the £12million the club invested in him during the summer with 11 goals.
However, having scored five in his first four games for the Black Cats, he has managed only one in his last seven, with the supply lines having dried up alarmingly.
The 26-year-old former Wolves frontman was injured just two minutes into Scotland's 2-1 home defeat by Wales in Friday night's World Cup qualifier at Hampden Park and was carried from the field on a stretcher.
If he does need surgery, player and manager alike will hope he can return in time for the start of the new campaign.
O'Neill at least has January signing Danny Graham, who has played alongside Fletcher in recent weeks, at his disposal.
Cattermole has been plagued by injuries for much of his time on Wearside and has managed only two senior appearances since the end of November.
He had an injection in his injured knee several weeks ago in a bid to solve the problem once and for all, although O'Neill admitted at the time that an operation was a possibility.
Having visited his specialist again, the decision was taken to perform the surgery and write off the rest of the campaign.
Meanwhile, it has been confirmed that David Miliband is to remain a member of the club's board despite stepping down as MP for South Shields.
The former Foreign Secretary has announced he is leaving UK politics to take up a post with the US-based charity the International Rescue Committee.
However, Mr Miliband, who joined the board as vice-chairman and non-executive director in 2011, will maintain his links with the club.
Chief executive Margaret Byrne told Sunderland's official website, www.safc.com: "David's knowledge and experience have been a tremendous benefit to Sunderland AFC.
"Naturally we are delighted that he is to continue in his role at the club, and we also wish him every success with this new chapter of his life."