Ilary Blasi claims she received a phone call from their daughter whilst Totti was away.
The couple are currently going through a divorce having separated two years ago.
Totti and Blasi tied the knot in 2005 live on Italian TV and have three children together.
Their oldest child, 19-year-old Cristian, is also a footballer in the Italian fourth tier with Olbia.
However, his World Cup-winning father is now the subject of a serious allegation by Blasi.
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Speaking about the story, Italian journalist Daniele Fisichella told Hawksbee and Jacobs: "The latest chapter is the accusation from the wife to Totti of having left their eight-year-old daughter, Isabel, alone at home while he went away.
"So now there is an official investigation from the authorities in Rome.
"It allegedly happened almost a year ago. And apparently the daughter, this young girl, called (her) mom, who is also a famous TV presenter, by the way, in Italy, goes by the name of Ilary Blasi.
"The police came knocking on Totti's door and a woman, allegedly a babysitter, opened the door.
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"It's not clear whether it was actually a babysitter. Totti was away, or it was the concierge working at Totti's residence. But yes, it's just the last chapter.
"And if anyone wants to write a TV series, 'The Totti's', that obviously could be part of it. It's been going on for the last two years, to be fair.
"They've been seeing judges, lawyers, and yeah, it's been very difficult for him and for his wife as well. 20 years together and three children.
"One of them is 19 and actually plays football professionally, but yeah, it seems that the end, it's very far away."
A source close to Totti said, as quoted by The Times: "The allegation was untrue. His daughter was at home with a professional babysitter and after Totti returned home he spoke to the police to explain this."
The Times' report adds that Totti could choose to sue Blasi for defamation over her allegations that he left their youngest daughter alone at home.
Totti and his wife were dubbed 'the Italian Beckhams' during their 17-year marriage.
Blasi is also known for her career as a model and TV presenter in Italy.
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Meanwhile, the Roma legend retired in 2017 aged 40 after scoring 307 goals in 785 appearances for the club.