Three people were killed and at least 20 vehicles were destroyed when terrorists ambushed the convoy near Islamabad in Pakistan.
In the latest incident, the trucks were on their way to a border crossing at Torkham along the Khyber Pass.
The route is used to bring fuel, military vehicles, spare parts and other supplies for troops in Afghanistan.
However, the checkpoint has been closed temporarily by Islamabad in protest over a Nato helicopter strike that killed three Pakistani troops last week.
|Pakistani driver looks for items inside an oil tanker that was attacked by suspected militants at a terminal in Rawalpindi, Pakistan, on Monday, Oct. 4, 2010.|
|Pakistani driver stands beside an oil tanker that was attacked by suspected militants at a terminal in Rawalpindi, Pakistan, on Monday, Oct. 4, 2010|
Sky News foreign affairs editor Tim Marshall said that has resulted in a very big traffic jam.
"It appears the Taliban is taking advantage of the back-up of tankers caused by Pakistan closing the route to Nato convoys," he said.
"It's thought that having made their point, the Pakistanis will re-open the pass later this week, but that still leaves a backlog of tankers and that means extra targets for the Taliban."
On Friday, a day after the closure of the Khyber Pass route, there were two attacks on oil tankers heading to Afghanistan, one of which was claimed by the Pakistani Taliban.
The terror group has warned it would launch further attacks on the supply route.
This latest incident is likely to further strain relations between Pakistan and the US, which has consistently called on Islamabad to do more to root out terrorists within its borders.