Fonte handed in a transfer request on Thursday amid speculation over a January move to Manchester United, serving as a major blow to the Saints’ hopes this season.
The Portuguese has played the full 90 minutes in each of his 19 appearances so far in 2016/17, but that won’t be the case on Wednesday night.
“Jose won’t participate for this game. We will see for the next game,” he told reporters on Monday.
“We have an exciting game on Wednesday, and concentrating on this is the most important thing. We are not favourites, but we have nothing to lose.
“It’s important to try to have a good result in the first game to prepare us for the second one. My players are ready.
“This competition is an opportunity for us to win something, and this is important.”
Southampton are also without Charlie Austin (shoulder), Sofiane Boufal (knee), Matty Targett (hamstring) and Jeremy Pied (ACL), though Alex McCarthy (hamstring) and Cedric Soares (head) could return.
Boufal was forced to pull out of this month’s Africa Cup of Nations tournament, and has left the Morocco squad to return to Hampshire for treatment.
Liverpool’s treatment room is significantly less busy heading into Wednesday night’s matchup, with Jurgen Klopp confirming Joel Matip, James Milner, Jordan Henderson and Philippe Coutinho were all recovering well.
Serbian midfielder Marko Grujic has also returned to training alongside Milner and Coutinho, while Matip and Henderson are likely to join their team-mates in the coming days.