US Defence Secretary Robert Gates has said "substantial" quantities of Iranian weapons are finding their way into Afghanistan.
Gates said Iranian arms were headed for the Taleban
Speaking on a visit to Germany, Mr Gates said it was difficult to believe the Iranian government was unaware, given the scale of arms involved.
Washington has recently expressed growing concern about Iran's involvement with its eastern neighbour.
Iran has denied accusations it is helping arm Taleban insurgents.
Mr Gates' comments follow those of Undersecretary of State Nicholas Burns, who told reporters that the weapons were coming from the Iranian Revolutionary Guard.
Tehran 'must know'
Mr Gates said he had seen recent intelligence which made it "pretty clear there's a fairly substantial flow of weapons" into Afghanistan.
"Given the quantities that we're seeing, it is difficult to believe that it's associated with smuggling or the drug business or that it's taking place without the knowledge of the Iranian government," he said.
Mr Gates said this was ironic, because Iran had good relations with the Afghan leadership, adding that it was anybody's guess what Iran's motives might be beyond causing trouble for the United States.
He said he believed the weapons were intended for the Taleban.
Earlier, Mr Burns was more direct, telling CNN there was "irrefutable evidence" that Iran was supplying the Taleban with arms.
"It's certainly coming from the government of Iran," he said.
"It's coming from the Iranian Revolutionary Guard corps command, which is a basic unit of the Iranian government."
In April, US Joint Chiefs Chairman Gen Peter Pace said US troops had intercepted Iranian-made arms intended for use by the Taleban.
It was not clear if the Iranian government was directly to blame, Gen Pace said.
Iran, a Shia Islamic state, supported the Northern Alliance against the Sunni Taleban in a decade of conflict which ended with the Taleban's overthrow in December 2001.
Tehran initially co-operated with US forces fighting insurgents in Afghanistan, and as late as February this year the US military said there was no evidence Iran had sent any weapons to the Taleban.