Saudi Arabia's King Salman opened the annual one-day summit of the Gulf Cooperation Council, urging fellow member states Kuwait, Oman, Bahrain, the UAE and Qatar to maintain a united front against Iran and terrorism.
"This requires all of us to maintain our countries' gains and to work with our partners to preserve security and stability in the region and the world," he said in a speech. King Salman also accused Iran of "continuing to interfere in the affairs of the countries in the region".
Iranian Foreign Minister Mohammad Javad Zarif responded swiftly, tweeting that the "region has had far too many strongmen who have only caused war & misery."
But the Qatari Emir did not respond to an invitation from the Saudi king to attend the summit and instead sent his foreign minister.
The summit is overshadowed by the Khashoggi murder and the continuing disputes with Qatar, which is under multiple boycotts by the other members, although Kuwait has tried to mediate the dispute.
Saudi Arabia, UAE and Bahrain severed relations with Doha in 2017, accusing the tiny state of supporting "terrorism" and maintaining close relations with Iran.
A U.S. official on Sunday called on Gulf states to mend fences to confront Iran and pave the way for a proposed new Middle East security alliance that would include the Gulf bloc, Egypt and Jordan.
"We'd like to see that unity restored, not on our terms, but on terms of the countries that are involved," Timothy Lenderking, Deputy Assistant Secretary for Arabian Gulf Affairs, told reporters at a security forum in the UAE capital Abu Dhabi.
While the boycotting states insist the row is not a priority for them and that the GCC remains valid, Qatar maintains the dispute harms regional security by weakening the bloc.