All of the South Island has been declared a "controlled area" for the invasive algae didymo, which is also known as rock snot.
Travellers and freight companies crossing Cook Strait from the south must clean any items that have been in lakes or rivers.
People in the South Island will have to clean all items - clothing, rods, waders, boats, 4WD vehicles - that have been in a stream, river or lake before they enter another waterway.
"Nothing we can do cancels the need to clean between waterways anywhere in New Zealand," said Biosecurity New Zealand spokesman Peter Thomson.
"It applies to everyone, everywhere, every time."
The new controlled-area regulation takes effect today.
Spot checks will be made of people entering and leaving waterways, such as at boat-launching sites, but in effect the biosecurity agency has given up on the idea of "isolating" infested rivers.
Didymo has been found in the Mararoa, upper and lower Waiau, Oreti, Buller, Hawea, upper Clutha, and Von rivers. Cells of the algae have also been found in Manapouri and they are expected to grow into plants.