The United States has ordered the expulsion of a senior Venezuelan envoy in a tit-for-tat move that has drawn criticism from Caracas.
Hugo Chavez has said any US spies will be arrested
On Thursday, Venezuelan President Hugo Chavez announced the expulsion of a US naval attaché for spying - a charge the US described as "baseless".
Venezuela attacked the US decision to expel its diplomat on Friday as "disproportionate and incongruous".
Relations between Washington and Mr Chavez have long been under strain.
Mr Chavez frequently accuses the US of trying to overthrow him, which the US denies.
On Thursday, Venezuela said it was expelling US naval attaché John Correa for allegedly passing secret information from Venezuelan military officers to the Pentagon.
Friday saw US State Department spokesman Sean McCormack announce that Jeny Figueredo Frias, chief of staff to Venezuela's ambassador, must leave the country within 72 hours.
Mr McCormack said the move was a direct response to the action taken against Capt Correa, and Ms Frias was "the most appropriate" choice.
"We don't like to get into tit-for-tat games like this with the Venezuelan government, but they initiated this and the US chose to respond," Mr McCormack said.
Caracas' vice foreign minister for North America accused the US of acting without proof in pursuit of "political retaliation".
"The decisions that Venezuela takes are taken based on facts and proof, not simply for retaliation," Mari Pili Hernandez said.
Mr Chavez has also said his government will not rule out further expulsions if more evidence of spying is uncovered.
The expulsions take the animosity between Venezuela and the US to a new level, says the BBC's State Department correspondent Jonathan Beale.
Washington is deeply opposed to the government of left-wing Mr Chavez, who is a vocal critic of the US.
The US has expressed concerns about Venezuelan democracy under Mr Chavez and about the effect of his government's military purchases on regional stability.
Washington has tried to block his attempts to buy military equipment that contain American technology.
Earlier this week, US defence secretary Donald Rumsfeld likened Mr Chavez to Adolf Hitler, describing both as leaders who were "elected legally and then consolidated power".
Venezuelan Vice-President Jose Vicente Rangel responded on Friday by describing US President George W Bush as the "North American Hitler" and his defence secretary as a "delinquent" and "arms dealer".