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While there always was some evidence of the existence of alien life forms, it was very scarce. Until the events of the Fallout 3 add-on Mothership Zeta, there was no record of direct contact between humanity and the creatures. Despite this, it is apparent that, with evidence found on Zeta, the alien's have quite possibly been watching Earth and humanity evolve for nearly a thousand years.
The Brotherhood of Steel and Shi have encountered the bodies of alien life forms or their spaceships, as did the pre-War United States government, which kept its alien specimens mostly in Area 51. By extension, the Enclave and Brotherhood Outcasts also have access to alien technology. Finally being encountered in Mothership Zeta, they are small, greenish humanoids that are presented as malevolent creatures that seem to view humanity with contempt and will gladly vivisect their test subjects or keep them in cryogenic storage after torturing them for their scientific pursuits. The aliens' language cannot be understood by the Lone Wanderer.