Iran's main cultural centre in the Sudanese capital was padlocked on Sunday, an AFP correspondent reported, after Khartoum ordered it closed in a shock to generally close relations.
The Iranian flag still flew above the building on one of Khartoum's main roads across from the airport.
But an AFP reporter found the gate padlocked and no sign of anybody inside.
Tehran last week denied the centre and two others had been shut on Khartoum's orders.
A Sudanese official, speaking on condition of anonymity, said on Thursday that the centres would close their doors by Sunday.
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Sudan's foreign ministry told the centres to close and ordered their Iranian staff to leave the country.
Foreign ministry officials could not immediately be reached on Sunday to confirm the closure had taken effect.
Sudan said it was acting in response to the centre's spreading of Shiite Islam, Iran's majority faith.
Sudan is overwhelmingly Sunni.
A Sudanese analyst told AFP the move by Khartoum might be in response to pressure from Riyadh, which further isolated the indebted and sanctions-hit Sudanese economy earlier this year by denying access to major Saudi Arabian banks.
Iranian warships have periodically made stops in Port Sudan, across the Red Sea from Iran's regional rival Saudi Arabia.
Evidence suggests Iran has played a key role in supporting war-torn Sudan's weapons production, said a May report from the Small Arms Survey, a Swiss-based independent research project.