The Waratahs have suffered another blow with the news that Berrick Barnes will not be available for his showdown with Melbourne Rebels star James O'Connor at inside centre this weekend.

Barnes missed the Waratahs' opening clash against the Reds, which ended in late heartbreak for the Sydney team as the Reds snatched a last-second victory, and there were high hopes that he would return to bolster them against the Rebels this weekend.

However, the injury-prone Wallaby playmaker pulled up with a sore groin at training on Tuesday and as a result will not be available to take on O'Connor and his new teammates in Melbourne on Friday.

He told AAP: "It's frustrating really ... I didn't think it was going to be an issue this week but after running yesterday (Monday) and being put through a bit of a fitness test it just hasn't pulled up as well as myself and the medical staff more importantly wanted."

The injury means that Tom Carter will probably line up at inside centre against O'Connor and Barnes said the key to shutting down the young backline ace would be to get in his face as early as possible.

"Defensively ... if you don't come one out against him it eliminates his ability to use footwork on people and get offloads away which he craves," Barnes said.

"He's a hard man to stop and I suppose we've got to nullify them first up front first to limit the amount of opportunities he gets to use his feet and that sort of thing but it's a hard task," he added.

Waratahs coach Michael Foley said that at this early stage of the season he would rather take a conservative approach to Barnes' injury, rather than risk losing him for a more lengthy period.

"He came close last week and trained really hard yesterday and we thought he'd ticked all of the boxes but when he came in this morning he was a little bit sore," Foley said.

"The brief to the medical staff was that if there was any doubt what so ever then we were to rule him out. I'm loathe to say he'll be right for Highlanders (next week) but I believe he will be," he added.

With Wallabies Rocky Elsom, Drew Mitchell and Lachlan Turner already on the sidelines the Waratahs are having their depth seriously tested early in the season, and the Rebels will be hopeful that their new stars O'Connor and Kutely Beale are able to manufacture an upset this weekend.