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Thousands of employees at the US Agency for International Development (USAID) will be placed on leave from Friday night, the agency says.
A USAID statement said the order would affect all "direct-hire personnel" except those on "mission-critical functions, core leadership and specially designated programs".
It's unclear exactly which jobs are affected. The statement, posted on the agency's website, said employees would be notified by Thursday afternoon.
The Trump administration has said USAID is wasting money and needs to align with the president's policy priorities. Agency staff, backed by Democratic lawmakers, have protested over the cuts, saying they will put lives in danger and hamper national security.
President Donald Trump's earlier decision to freeze foreign assistance has upended the global aid system, with effects already felt in multiple countries.
The agency, which provides humanitarian aid to more than 100 countries, employs 10,000 people worldwide. Two-thirds of those people work overseas, according to the Congressional Research Service.
USAID, founded in 1961, has a budget of around $40bn (£32.25bn) per year, amounting to roughly 0.6% of federal spending, according to official figures.
In a statement on its website on Tuesday, USAID said it would work with the US Department of State to arrange and pay for return travel for personnel posted outside the US within 30 days.
Those employees who are part of the exceptions will be told by 15:00 EDT (20:00 GMT) on Thursday, it said.
The statement concluded with the message: "Thank you for your service."
Given that some USAID staff work in active conflict zones, it was not immediately clear how they would be withdrawn.
The measures are part of a string of cuts that Trump has enacted since returning to office last month through co-ordination with Elon Musk's unofficial Department of Government Efficiency.
Among the countries affected by the aid freeze is Haiti. The United Nations said Tuesday that it had been notified by the US that it was freezing $13.3m in aid destined for the country.
But on Wednesday the US State Department issued a waiver exempting $40.7m of foreign assistance for Haiti's national police and the UN-backed international security support mission.
Earlier this week, the White House put out a statement listing a number of projects that it called evidence of "waste and abuse", including a grant of $1.5m to an LGBTQ group in Serbia and $2.5m for electric vehicles in Vietnam.
However, critics have called the moves to close the agency an illegitimate power grab spearheaded by Musk, an unelected billionaire.
Earlier this week, Trump's top diplomat, Marco Rubio, became the acting head of USAID as part of its reported merger into the State Department.
Closing USAID altogether would require an act of Congress.
On Tuesday, many USAID staffers received an email notifying them they had been placed on paid administrative leave.
The email, obtained by BBC News, told employees they must remain "available" by telephone and email during business hours, but were not allowed to enter USAID buildings.
Musk, the CEO of Tesla and Space X, has suggested USAID should be shut down entirely, as it is "beyond repair".
Many have cautioned that closing the agency's doors would have devastating effects on vulnerable populations across the world.
USAID's activities range from providing prosthetic limbs to soldiers injured in Ukraine, to clearing landmines and containing the spread of Ebola in Africa.
An HIV patient in Nigeria told the BBC that she and others are worried that free anti-retroviral drugs, funded by USAID, would be cut off.
"I am really scared about what the future holds given all that is going on in America," she said. "I got my last set of HIV drugs in November and I am supposed to go back in April... but I don't know what would happen by then.
"These free drugs are really important for me and most of us (HIV patients) would be in trouble if it is affected," she said.
Democrats in Washington DC have been particularly critical of changes to the agency, calling them "illegal" and "unconstitutional".
"[USAID is] a foreign policy tool with bipartisan origins that is critical in this dangerous global environment," New Jersey Democratic Senator Andy Kim, who previously worked at USAID, wrote on social media.
"Gutting it means gutting our ability to compete and keep America safe."
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