The explosion, apparently from a bomb planted in the bus, wounded five others as the bus passed through Nadh Ali district in Helmand state on its daily route to the provincial capital Lashkargah.
The bus bombing along with two explosions and gunbattles in two provinces left the death toll for Wednesday at 58.
The deaths were part of a trend of stepped-up attacks and killings that are increasing the pressure on the fragile Afghan government and creating an atmosphere of constant fear in the country.
"It was a powerful explosion. The bus was destroyed," a district administrator Ghulam Mahauddin said in a satellite telephone interview. "Six of the dead were children, eight were men and one was a woman."
No one claimed responsibility, and no arrests were made. But Helmand provincial deputy governor Haji Pir Mohammed and Mahauddin blamed al Qaeda insurgents and remnants of the Taliban militia, ousted by a U.S.-led coalition in late 2001.
"They are killing innocent people," Mohammed said.
Anti-government insurgents have been stepping up attacks in recent months, particularly in the south and east of the country. Authorities say insurgents have also infiltrated the Afghan capital, Kabul, to carry out terrorist attacks.
Twenty people were on the bus in Helmand when the blast went off, Mahauddin said. The driver of the bus was among the wounded.
"Right now we don't think it was a suicide bomber or that it was a remote controlled devise," he said. "It seems that someone placed the bomb in the bus. Whoever did this is an enemy of Afghanistan."
In Kabul on Wednesday, two university students were killed and one was seriously wounded when a bomb they were making — apparently in preparation for a terrorist attack — went off by accident, police said.
The blast occurred at the home of one of the students in western Kabul, said deputy police chief Khalil Aminzada.
On Monday, NATO took command of the 5,000-strong international peacekeeping force that patrols Kabul's crumbling streets.
The alliance took over from Germany and the Netherlands, which have jointly led the International Security Assistance Force, known as ISAF, since Feb. 10.
"ISAF's name and mission will not change," said NATO Deputy Secretary General Alessandro Minuto Rizzo. "But what will change as of today is the level of commitment and capability NATO provides."
German Defense Minister Peter Struck said in a speech that the handover showed the world's commitment to rebuilding war-shattered Afghanistan.
"Afghanistan must not lapse back into anarchy and chaos and must not again become the home of global terror, as was the case under the rule of the Taliban," Struck said.
But Wednesday's bus attack was a stark reminder that remnants of the Taliban and al Qaeda remained intent on disrupting any efforts to achieve stability in the country.
Past sporadic bombings in the capital have been blamed on a mix of Taliban rebels, al Qaeda fighters and supporters of renegade warlord Gulbuddin Hekmatyar.
A suicide bombing in eastern Kabul in June killed four German peacekeepers and wounded 29. In July, a suspected terrorist was killed when a homemade bomb he was planting in a dirt road exploded prematurely 9 miles east of Kabul.
Elsewhere, attackers late Tuesday fired two 107 mm rockets at a U.S. base in the eastern city of Asadabad, in Kunar province, the U.S. military said in a statement from Bagram Air Base, north of the capital.
The rockets fell harmlessly like scores of others have in the past, causing no damage or casualties.
In a separate incident on Wednesday morning, two laborers rebuilding a ruined house in southern Kabul were wounded when they set off a mine left over from years of war, said Ghulam Ghawis, a police official.
The mine, hidden at the base of a wall made of hardened mud, exploded when the workers tried to collapse the wall, Ghawis said.
Much of Afghanistan is littered with the debris of war. Unexploded ordnance and mines kill or maim over a 120 people in the country every month.