Hard-liners accuse Rouhani of selling out on Iran's ideological values.
One of the organizers of the anti-Israeli conference, dubbed New Horizon, said the cancellation was a "disaster" and a "big mistake."
"Cancellation of the anti-Zionist New Horizon festival is a disaster. It was the most powerful anti-Zionist conference in Iran," website mashreghnews.ir quoted the organizer, Nader Talebzadeh, as saying.
Talebzadeh said 63 foreign scholars and 50 Iranian speakers were expected to address the gathering, which was supposed to have taken place either in late September or early October. He added that he began inviting foreign speakers back in May, when Ahmadinejad was in office.
Rouhani's short-term goal is seeking to ease Western sanctions as part of negotiations over Iran's controversial nuclear program. Talks with the U.S. and other world powers are scheduled to resume in Geneva next week, but Iran has not yet given details on what new proposals it would bring.
The West and its allies fear Iran's ability to make nuclear fuel will eventually lead to weapons-grade material. Iran repeatedly denies it seeks nuclear weapons and says its atomic program is only for energy and medical applications.
Israel views a nuclear-armed Iran as a threat to its very existence, citing Iran's repeated calls for Israel's destruction, its long-range missile program and its support for violent anti-Israel groups like the Hezbollah in Lebanon.