Investigators suspect pesticides are the reason for the deaths of about 1,000 fish, said Wayne MacKinnon, a spokesman for the Environment Department.
"We probably think it's pesticide runoff," he added..
Brook trout, rainbow trout and Atlantic salmon were found along a 3.5-kilometre stretch of the North River, MacKinnon said.
Existing regulations require a buffer zone between streams and planted fields, he said, and there are also rules that prevent planting on slopes that run towards waterways.
He said conservation officers and Environment Canada officials will collect samples of both fish and water, and they will also investigate conditions on surrounding farms.
Last year, fish also died in Barclay Brook in western P.E.I., where thousands of fish died in 2011 and 2012 after pesticides from farmers' fields seeped into the water.
NDP Leader Mike Redmond says the damage to tourism and the fishery is significant, but the province hasn't been firm enough in its enforcement of existing regulations.
MacKinnon said there is a review of existing regulations on the use of pesticides.
He said a committee that includes the P.E.I. Potato Board, the farming industry and the provincial government is looking at measures to reduce pesticide runoffs.
"We've gone so far as to purchase one of the fields that was involved to take it out of production," he said, referring to last summer's incident in the Barclay Brook area.
— By Michael Tutton in Halifax.