The photo, taken last October by Javier Manzano in Syria's largest city Aleppo, shows two men gripping assault rifles, one of them stuck out of a hole in the nest, as light beams at different angles through more than a dozen bullet holes in a corrugated tin wall behind them.
Otherwise the rebels' lair is largely dark. Both men have their heads covered with scarves. The Pulitzer committee called the photo "extraordinary."
Manzano was born in Mexico and is based on Istanbul.
The photo was submitted to AFP by Manzano himself.
Manzano, who left for the United States at the age of 18, has focused much of his work on issues involving the US-Mexican border, the Pulitzer web site said.
He began his career in the newspaper industry as a photo and videojournalist, and later got into television and electronic media.
He has been a freelancer covering the drug war in Mexico and the wars in Afghanistan and Syria since his last employer, The Rocky Mountain News, shut down in 2009, the site said.