Arabic television channel Al-Jazeera played a message on Thursday purportedly from Al-Qaeda deputy leader Ayman al-Zawahiri, calling on Pakistanis to overthrow President Pervez Musharraf.
There was no immediate independent verification of the audio recording.
"I call on the Pakistani people to unite under the banner of Islam," said the voice.
"All Muslims in Pakistan should attempt to get rid of this traitor government, which has surrendered to the Americans, leading to the destruction of Pakistan so the Indians can be in control," he added.
"I call on the Pashtun and Balushi tribes, (on) the two sides of the border (with Afghanistan)... Oh tribes, your Taliban brothers and their followers are your brothers and your hosts.
"How can you allow the agents of the Crusaders and the Jews to strike you? The Pakistani people in general and the Pashtun tribes in particular should take revenge," added the voice.
"I call on the Pakistani army, the poor army that was put in a miserable state by Musharraf, the Indians are in front of you and behind you in Afghanistan... Musharraf has disposed of your nuclear arms. Are you going to remain silent until the partition of Pakistan once again?" it said.
"Fight the supporters of the devil," the voice says, also calling on Islamic ulema, or religious leaders, "to do their duty and reveal the real face of Musharraf".
There was speculation that Zawahiri was a "high-value target" being protected by tribesmen and hundreds of fighters who have been locked in battle with Pakistani forces since March 16.
Some 7,500 troops have been battling an estimated 500 Al-Qaeda suspects and their tribal supporters, entrenched in mud-walled fortresses near Wana, close to the Afghan border, in Pakistan's largest ever assault in its two-year Al-Qaeda hunt.
But Zawahiri, who has a 25-million-dollar bounty on his head, was never seen and his whereabouts are unknown.
It is unclear when the tape was recorded and whether it was meant to signal that Zawahiri had escaped the Pakistani dragnet or had never been endangered by it.
There was no immediate reaction from Islamabad, but the message was likely aimed at the Pakistani military because Musharraf requires its support to maintain his dictatorship.
The voice also accused Musharraf of "killing thousands of Muslims in Afghanistan" and being put in charge by the United States "of new missions, most importantly the destruction of Pakistan."
"Among these plans is the freezing of the Pakistani nuclear programme," it added.
On March 19, a Taliban spokesman, Abdul Samad, said Zawahiri and Al-Qaeda chief Osama bin Laden are safe and in Afghanistan, not Pakistan where the intense manhunt was underway.
Zawahiri, a founder of the militant Egyptian Islamic Jihad organisation, is rated number two in the US government's list of the 22 most wanted terrorists.
Last seen in the Afghan town of Khost in October 2001, he went into hiding after US forces ousted the Taliban government in Kabul.
A veteran of the resistance against the Soviet forces who invaded Afghanistan in 1979, he was subsequently identified as the head of the Vanguards of Conquest group that killed foreign tourists in the Egyptian city of Luxor in 1997.
Two years later an Egyptian court sentenced him to death in absentia.
With Inputs from Shyam Bhatia in London