Five Sudanese teenagers riding in the back of a pick-up belonging to the German Agency for Technical Co-operation (GTZ) and one attacker were shot dead in the attack, they said on Wednesday.
The ambush occurred on Monday, about 20km west of Juba, the provisional capital of southern Sudan.
In addition to the casualties, a Kenyan surveyor working for GTZ was reported missing.
About 30 assailants, thought to be members of Uganda's Lord's Resistance Army (LRA) stormed the truck carrying non-German GTZ employees and security guards and about 20 passengers, the witnesses said.
"They were shooting bullets all around us," said Paul Agos, a Sudanese security guard for GTZ who was in the vehicle. "I fell off the truck and shot one of them. Then they all fled into the bush."
Witnesses said the attackers, some of whom were uniformed, were rebel fighters with the LRA, which has waged a nearly 20-year war in northern Uganda and southern Sudan but is preparing for peace talks with Kampala.
Witnesses said that the assailants spoke the Acholi dialect of northern Uganda and that the attack took place in an area known to be frequented by the rebels, but the identities of the attackers could not be independently confirmed.
A spokesman for the LRA delegation in Juba awaiting the start of Wednesday's peace talks with the Ugandan government denied the rebels were involved in any such attack.
"I can categorically deny that the LRA is responsible for the attack," spokesman Obonyo Olweny said.
Herbert Kremeier, GTZ's Kenya-based programme director for southern Sudan, said the attack underscored the agency's concern for its employees' safety in the region where it is building a road from Juba to the town of Bor.
"GTZ is concerned about security," he said. "It has been an issue."
Kremeier said a search, backed by UN helicopters, was under way for the missing Kenyan surveyor, Daniel Wekesa.