Iran charged on Monday three detained American citizens with espionage, the official IRNA news agency quoted a judiciary official as saying, while U.S. Secretary of State Hilary Clinton said there was "no evidence" for Tehran to charge its citizens.
The three were held after they strayed into Iran from northern Iraq at the end of July.
"We believe strongly that there is no evidence to support any charge whatsoever," Clinton said on a visit to Berlin on Monday.
The three, Shane Bauer, 27, Sarah Shourd, 31, and Josh Fattal, 27, crossed into Iran from Iraq and their families say they strayed across the border accidentally.
"The three are charged with espionage. Investigations continue into the three detained Americans in Iran," Tehran general prosecutor Abbas Jafari Dolatabadi told IRNA.
Iranian President Mahmoud Ahmadinejad suggested in an interview with the American television network NBC in September that the Americans' release might be linked to the release of Iranian diplomats he said were being held by U.S. troops in Iraq.
According to Iranian law espionage is punishable by death.
The United States has sent strong messages to Iran urging the release of the three hikers, calling Iranian authorities to exercises "compassion" towards the three Americans.
Some Iranian authorities have linked the illegal entry of the Americans, to unrest that erupted after Iran's June presidential election.
Ahmadinejad's re-election on June 12, sparked Iran's worst unrest since the 1979 Islamic revolution. Authorities deny vote-rigging and portrayed the unrest as a foreign-backed bid to undermine the Islamic state.
Dolatabadi said the case of a Danish student, detained during a rally on Nov. 4 to mark the anniversary of the seizure of the U.S. embassy, was under investigation.
"This accused Danish citizen has introduced himself as a reporter but he holds no official press accreditation. Investigations about him continue," he said."Today the Danish embassy lawyer was allowed to meet the prisoner."
Police clashed with supporters of Iran's opposition leader Mirhossein Mousavi in Tehran on Wednesday when they used the anti-U.S. rallies to revive protests against the clerical establishment after June's vote.