Police officers, paramedics and firefighters responded to the scene at about 2:30 p.m. ET on Tuesday to the highway near the County Road 34 exit, which is about 120 kilometres southeast of Ottawa.
Police reported about 15 people hurt in the crash, including a teenage boy who was taken to hospital with serious injuries.
The bus began its route in Montreal and was destined for Kingston, Ont., when it collided with the tractor-trailer, Megabus confirmed.
"Safety is our number 1 priority and Megabus is fully co-operating with the authorities with their investigation into the incident," the company's statement read.
The highway's westbound lanes remain closed between County Road 2 and County Road 27. Detours are in place.