The 5th U.S. Circuit Court of Appeals granted an emergency stay of the requirement by the federal Occupational Safety and Health Administration that those workers be vaccinated by Jan. 4 or face mask requirements and weekly tests.
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Louisiana Attorney General Jeff Landry said the action stops Democrat President Joe Biden "from moving forward with his unlawful overreach."
"The president will not impose medical procedures on the American people without the checks and balances afforded by the constitution," said a statement from Landry, a Republican.
Such circuit decisions normally apply to states within a district - Mississippi, Louisiana and Texas, in this case - but Landry said the language employed by the judges gave the decision a national scope.
"This is a great victory for the American people out there. Never before has the federal government tried in a such a forceful way to get between the choices of an American citizen and their doctor. To me that's the heart of the entire issue," he said.
The Biden administration had no immediate comment.
At least 27 states filed lawsuits challenging the rule in several circuits.
The Biden administration has been encouraging widespread vaccinations as the quickest way to end the pandemic that has claimed more than 750,000 lives in the United States.
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The administration says it is confident that the requirement, which includes penalties of nearly $14,000 per violation, will withstand legal challenges in part because its safety rules pre-empt state laws.
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The 5th Circuit, based in New Orleans, said it was delaying the federal vaccine requirement because of potential "grave statutory and constitutional issues" raised by the plaintiffs. The government must provide an expedited reply to the motion for a permanent injunction Monday, followed by the petitioners' reply on Tuesday.