The 44-year-old stepped up from assistant to replace club hero Pep Guardiola at the end of last season, after having had surgery on a tumour in his saliva glands in November last year, and had made a record-breaking start to his tenure.
"While monitoring his parotid gland a change has been detected in his condition that requires surgery which will be carried out on Thursday," Barca said in a statement.
"His time in hospital will be about three to four days. During the period after his operation, depending on how he recovers, an attempt will be made to balance his treatment with his working life."
At a later news conference, Barca president Sandro Rosell and sports director Andoni Zubizarreta put an end to speculation that a replacement would be brought in from outside on an interim basis.
"Tito is the coach, and continues to be so," Zubizarreta said, dismissing rumours they may have tried to contact the out-of-work Guardiola.
"(Assistant) Jordi Roura will take charge against Valladolid. Tito I am sure will be watching us on television and wanting us to win."
Barca visit Valladolid in La Liga on Saturday before a two-week mid-season break.
Rosell added: "Tito has an incredible mental strength. He was worried for his family. This was his main concern. We have to leave them alone.
"Tito is the leader of the group, he won't be here for a few weeks, but we will continue working just the same."
The news of Vilanova's illness had been filtering out during the day after the club suddenly cancelled some scheduled events.
Messages of support came in from around the soccer and sporting world.
"Real Madrid wish to express their love and support to Barcelona coach Tito Vilanova, to whom we wish a prompt recovery," Barca's arch-rivals said in a statement.
"Real extend this support to the club and his family."
Former tennis world number one Rafa Nadal said: "All my strength and support for Tito Vilanova! We are all with you to overcome this next hurdle."
Vilanova was out of action for around three weeks after surgery last year but returned to the dugout to help Guardiola for the last six months of the 2011-12 campaign.
The quietly-spoken Catalan, at his presentation in June, said the experience had left him with a different perspective on life.
"I had an important operation a few months ago," he said. "I spoke with the medics and my family about the job, but they all said I was perfectly okay and there was no problem.
"Having overcome this operation it gives me a different view on life. Perhaps, after what has happened, training Barca will be like child's play."
Vilanova has guided the team to the top of La Liga with 15 wins in 16 games, nine points clear of second-placed Atletico Madrid, and into the last 16 of the Champions League as group winners.
He is the second member of the Barcelona squad to be afflicted with such an illness, after France defender Eric Abidal had to undergo surgery to remove a tumour from his liver in March 2011.
The 33-year-old returned to the field of play later that year but also suffered a relapse and underwent a liver transplant in April, from which he has yet to return to action.
Abidal was given the all-clear to return to full training on Wednesday and joined up with the rest of the squad for the first time after nine months out - in a session which Vilanova did not take.
"Abidal was emotional," Zubizarreta said when asked about the return of the Frenchman. "There was an exchange of glances between him and Tito in the dressing room that was very special."
- Sports & Recreation
- Tito Vilanova
- Pep Guardiola