Ignazio Marino on Monday resigned in writing as Rome's mayor, the speaker of the city parliament said. He announced last week he would step down amid an expenses scandal.
"A short while ago I received a letter from Marino stating that he has resigned," said Valeria Baglio.
"His resignation will be effective in 20 days' time," she added.
A commissioner will be appointed to run the city from 2 November until fresh mayoral elections are held next Spring.
Marino, announced last week he would stand down after he lost the support of his centre-left Democratic Party over an expenses scandal in which he allegedly used his mayor's credit card to pay for some private dinners.
He said his resignation should not be taken as an admission of guilt, but said "political conditions" had made it impossible for him to carry on.
Rome prosecutors have begun an investigation into Marino's expenses, after petitions filed by two opposition parties, the rightwing Brothers of Italy party and the grassroots Five-Star Movement.
Marino offered to pay back 20,000 euros of expenses opposition parties allege he racked up using a town hall credit card, but late on Thursday said he would stand down.
In a video posted on Facebook and in a letter on Rome City Council's website, Marino said he feared the return of graft to the capital under future administrations.
Rome City Council will form the plaintiff in a corruption trial of five people including the ex-head of the municipal garbage collection service, which opens on 20 October and Marino has said he plans to attend the first hearings of the trial.
Supporters of Marino - considered by many to be honest but bumbling and unpopular - suspect a concerted smear campaign linked to his role in exposing a criminal network with links to the far right which had thrived under his conservative predecessor Gianni Alemanno.
Alemanno was among dozens of politicians and businessmen implicated in a major probe into the alleged mafia network, which is suspected of rigging tenders and siphoning off millions of euros.
Marino claims that his bid to clean up city corruption by ending all public contracts without tender was the nail in his coffin.
The 60 year-old doctor was already under pressure to resign amid mounting public anger over the litter-strewn and pot-holed state of the city's crumbling streets, and a failing public transport system.
Marino also allegedly left a series of parking tickets unpaid and drove his car in a restricted area with an expired permit.
He assumed office on 12 June, 2013 with 64 percent of votes cast.
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