French Foreign Ministry spokeswoman Agnes von der Muhll said the ambassador was being brought back for "consultations" and urged Italy in a statement to work to restore friendly relations worthy of "our common destiny."
Italian Deputy Prime Minister Luigi Di Maio met with supporters of France's yellow vest protest movement running as candidates for the European Parliament. Di Maio has said the populist 5-Star Movement he leads was ready to help the French protesters and has accused France of fueling Europe's immigration difficulties.
That came after Italian Interior Minister Matteo Salvini called Macron "a terrible president" in January. He said he hoped French voters would send Macron a message during the European elections by showing their support for far-right leader Marine Le Pen, with whom Salvini is allied in European politics.
Von der Muhll called the incidents an "unacceptable" interference in French democracy, and said they were unprecedented since the two neighbors joined together after World War II to help create the European Union.
"The campaign for the European elections cannot justify the lack of respect for each people or for their democracy," she said. "For several months, France has been the subject of repeated accusations, unfounded attacks and outrageous declarations," she added. "To have disagreements is one thing, to exploit the relationship for electoral purposes is another."
Italy's foreign minister, Enzo Moavero Milanesi, sought to tamp down the dispute, stressing the "profound friendship" between the two allies. But he acknowledged that differences were coming to the fore ahead of May's European Parliament elections.
"The defense of each one's interests and points of view, as well as the political debate ahead of the upcoming European Parliament elections, cannot influence on the solid relations that have united us for decades," Milanesi said in a statement.
In response to France's move, Salvini said he was open to meeting with Macron and the French government, but insisted that France must stop sending back migrants at the border and stop penalizing Italian workers in France.
"We don't want to fight with anyone. We are not interested in polemics. We are concrete people and we defend the interests of Italians," he said. Di Maio had already sparked annoyance in January when he accused France of leading colonial-style policies in Africa, prompting the French Foreign Ministry to summon the Italian ambassador. And the Italian government last fall accused France of dumping underage migrants over the border without authorization.
After meeting with members of the Citizens' Initiative Rally group of yellow vests on Tuesday, Di Maio boasted on Twitter that "the wind of change has crossed the Alps."
Colleen Barry in Milan contributed to this report.