Bezzecchi clocked one minute 50.028 seconds at the Lusail International Circuit to take pole, 0.266 seconds ahead of Marquez, while 2023 race winner Fabio di Giannantonio of VR46 Racing joins them on the front row after going third fastest.
"I made a mistake in the last corner; I tried a little bit too much. But I can't complain at all," Bezzecchi said.
"Perfect qualifying. I wanted to start in the front row, and the pole position is even better. So, very happy."
Aprilia's Jorge Martin, who is second in the championship and 44 points behind Marquez, was left frustrated as he could go only 11th fastest - more than a second slower than his teammate.
Marquez can be crowned champion in Qatar if he finishes 30 points ahead of Martin and eight points ahead of Bezzecchi after Sunday's race.
"It was the main target, as I said yesterday, the front row. It was not easy," Marquez said.
"We know that Marco has another pace this weekend. He's super fast. We must understand this and try to do our best."
Marquez's teammate Francesco Bagnaia was fifth fastest, and he will be sandwiched between the two Trackhouse riders Raul Fernandez and Ai Ogura.
The Yamaha factory team and its satellite team, Prima Pramac, did not fly to Qatar after withdrawing from the Grand Prix, citing "company-wide travel restrictions" in the region due to the war against Iran, leaving only 18 riders on the grid.