As has been the norm, Marquez followed the Ducati knowing full well his Honda was no match for the pace of the Italian manufacturer and championship leader Bagnaia made his feelings known when they came close together on track.
The Italian rider caught Marquez's attention and gestured angrily with his hand but the Honda rider was unperturbed and merely shook a finger, staying on the Ducati's tail to seal a spot on the front row.
The rush of adrenaline from the encounter clearly had an effect as the two men became the first riders to breach the one minute 45 second barrier at Mugello, with Bagnaia setting a lap record with a time of 1:44.855.
"To be on the front row in Mugello is not easy. I wasn't looking for Pecco (Bagnaia)... He decided to stop in my sight and speak with me," Marquez said in a pit lane interview.
"Somebody brought the opportunity to follow Pecco and I did it. We achieved our target - it wasn't the front row, it was top nine."
Gresini Ducati's Alex Marquez (27) - the younger brother of Marc - and Red Bull KTM's Jack Miller (28), who both moved to Q2 after topping the charts in Q1, finished third and fourth respectively.
Jorge Martin (25) and Marco Bezzecchi (24) complete the second row while Aprilia's Aleix Espargaro (33), racing with pain in his right heel after a bicycle crash in training, remarkably finished seventh fastest.
Ducati rider Enea Bastianini (25), who was returning from injury to race in his home grand prix, crashed early in Q2. He returned to the pits and came back out for another go but did not set a fast time and will start 12th on the grid.
Espargaro's teammate Maverick Vinales (28) did not move into Q2 and will start 13th on the grid while the Yamahas of Franco Morbidelli (28) and Fabio Quartararo (24) struggled for pace and will start 14th and 15th.
