Anwar al-Awlaki's 23-minute video was released by Islamic Web sites Monday, ABC News reported.
He says jihadists who want to kill Americans don't need a fatwa (special religious permission).
"Don't consult with anybody in killing the Americans," says Awlaki, sitting at a desk with a dagger in his belt. "Fighting the devil doesn't require consultation or prayers seeking divine guidance. They are the party of the devils."
He calls Americans and Muslims "two opposites who will never come together" and that it is "either us or them."
Awlaki also attacks Arab leaders, including Yemen's, as corrupt and traitors to Islam.
Though Awlaki often records in English, this video is in Arabic. It seems to be an excerpt from a longer video, from which other clips were released in October.
Awlaki is being tried in absentia in Yemen for his alleged role in the kidnapping and murder of a French national. On Saturday, Judge Moshen Allwan ordered him "arrested by force, dead or alive" when he failed to appear.