People must work from home unless there is “a reasonable excuse” with fines for bosses and workers.
Fines will be handed out to employees in Wales if they are not working from home with good reason.
From Monday, workers will receive a £60 fixed penalty notice and companies hit with fines of £1,000 every time they break the rule.
Until now there has just been guidance encouraging home working.
The union GMB said it would affect “the poorest, most vulnerable workers” while the TUC said it was “at best naïve” to think responsibility is shared.
“Additional measures have been introduced to limit the spread of the virus and protect public health,” said a Welsh government spokesman.
“Further to our long-standing advice for people to work from home wherever possible, from Monday this will now be a legal requirement to work from home unless there is a reasonable excuse not to.
“We expect employers to take all reasonable steps to facilitate home working and provide employees with the support they need.”
The enforcement approach will include improvement or closure notices for premises, with a business owner liable to be fined £1,000 for not complying.
This could rise to a maximum of £10,000, with individual employees liable to be hit with a £60 fine.
Wales TUC general secretary Shavanah Taj expressed “shock and concern” at the decision and hoped it would be
“A worker is not responsible for their place of work, their employer is,” she added.
“This sets a really worrying precedent that the responsibility is somehow shared, and is at best naive.”
The GMB said it was concerned the poorest and most vulnerable workers would be hardest hit, with “bad employers” able to “protect themselves from fines by placing liability on workers”.
Senior organiser Kelly Andrews said: “We think this strikes the wrong chord.
“We have major worries that this could lead to bad employers pressuring their workers to work from home without a paper trail and place any financial risk on them.
“Those workers are also the most vulnerable and can least afford to take the financial hit.
“But the truth is for a lot of families a £60 fine over Christmas will have a severe financial impact.”
The Welsh Conservatives economy spokesman, Paul Davies MS, said the new rules had been brought in under the radar.
“This Labour administration seems to be getting rather comfortable governing by late night press releases and tinkering with the rules without letting the public know until they’ve already come into force,” he said.
Welsh Liberal Democrats called on the Welsh government to scrap the £60 fines for employees.
Party leader Jane Dodds MS said: “Employees should not be punished for potentially following the direction of a bad boss or employer who is not complying with instructions.”