There is no reason for concern, and there is no need to feed alarmism," Vatican spokesman Father Federico Lombardi said.
He added that the Vatican - and, by extension, the Pope - was "extremely calm" regarding the alleged threat.
The warning was voiced by Nicola Gratteri, a respected state prosecutor in the southern Calabria region, who said the vicious local mafia, the 'Ndrangheta, is "nervous" the Pope is threatening its interests.
"Those who up to now have fed off the power and wealth coming directly from the church are nervous, upset," he said in an interview published by the newspaper Il Fatto Quotidiano this week.
The Pope, Gratteri said, "is dismantling the Vatican's economic centres. If the mafia bosses can trip him up, they won't hesitate".
Gratteri didn't substantiate his warning. But his words reverberated through the Italian and foreign media, sparking fears for the pontiff's safety.
Initially the Vatican tried to dismiss the allegation. But from Thursday it started saying it was simply taking the warning in its stride.
Implied in Gratteri's comments is that Italy's mafia has its tentacles in the Vatican's obscure financial dealings and agencies, some of which have been marred by scandal.
Since taking the papacy in March, Pope Francis has set about cleaning up the Holy See's vast holdings and making them more transparent.
One of his first steps was to install a special commission tasked with investigating the Vatican's bank and another to probe Vatican finances in general.
The Pope has also called in a US consultancy, Promontory Financial Group, to conduct an external review of the Vatican bank's money-laundering rules and, more recently, to look into the internal agency handling its many real estate holdings.
The Vatican's bank, known as the Institute for Religious Works, was notably the main shareholder of the Banco Ambrosiano, which collapsed in 1982 amid accusations of laundering money for the mafia.
Banco Ambrosiano's chairman Roberto Calvi - dubbed "God's Banker" - was found hanging from a London bridge that year in a suspected murder by mobsters.
The Vatican's agency handling its real estate assets, the Administration of the Patrimony of the Apostolic See, came under scrutiny when a prelate, Nunzio Scarano, was arrested in June on suspicion of acting as a front for dubious international payments made through the Vatican bank.
Scarano wrote to the Pope to defend himself, accusing cardinals of covering up irregular financial activities carried out by his superiors.
Italy's various crime syndicates have been held responsible for several high-profile assassinations and abductions.
Although the Sicilian mafia Cosa Nostra is perhaps the best-known, the Calabrian 'Ndrangheta, whose name comes from the Greek for courage or loyalty, is considered by many as more dangerous and difficult to predict.
It has a tight clan structure which has made it famously difficult to penetrate, and specializes in drug and arms trafficking, prostitution, extortion and illegal construction.
The 'Ndrangheta runs an international crime network from its base in Calabria and has been linked to operations across western and northern Europe and as far afield as the Americas and Australia.