The Environment Agency (EA) has said it is investigating a fuel spill in upper Wensleydale.
Residents of Hawes and Gayle reported that oil was entering Gayle Beck.
They said there was a strong smell of fuel in the air along the length of beck from early Friday morning.
Local residents say the fuel appears to be entering Gayle Beck via a stream off Gayle Lane, which flows under the Beulah Bank footpath, part of the Pennine Way.
Oil could still be seen on the water surface on Monday lunchtime.
A slick has gathered a few hundred metres downstream at Haylands Bridge, where Gayle Beck meets the River Ure.
Stones lifted from the beck smell strongly of fuel.
Five years ago, Hawes and High Abbotside Angling Association recorded a similar significant pollution incident, with the fuel entering the river system at the same place.
In a statement the Environment Agency said: “Our officers are currently investigating several reports of an oil spillage on Gayle Beck in Hawes, Upper Wensleydale, North Yorkshire.
“We know seeing pollution in our local watercourses can be distressing. We are tracing the source of the spill and will assess its impact to minimise any damage caused to wildlife and the environment.
“Anyone who spots water pollution is encouraged to report it to us via our 24-hour freephone incident hotline on 0800 807060.”
Wensleydale Creamery owner Saputo said it had also notified the EA.
“On the afternoon of Friday, February 11, we identified some fuel in Gayle Beck. We immediately notified the Environment Agency who visited site yesterday.
“Preliminary findings suggest the creamery is not the source of the contamination but we will continue to work with the EA to support their investigation.”