"The main (actor) responsible for the problems in Libya since the crisis started in 2011 is France," Turkish foreign ministry spokesman Hami Aksoy said in a statement.
"It's no secret that this country has given unconditional support to (military strongman Khalifa) Haftar in order to have a say regarding natural resources in Libya," he added.
Macron claimed earlier on Wednesday at a meeting with Greek Prime Minister Kyriakos Mitsotakis that Turkish ships accompanying Syrian mercenaries arrived on Libyan territory in recent days.
The French leader said the action was a "clear violation" of what Turkish President Recep Tayyip Erdogan promised at the Berlin conference on January 19 where world leaders vowed to stay out of the Libyan conflict.
"It is a failure to keep his word," Macron said.
But Aksoy said Macron "was once again trying to set the agenda with fanciful claims."
Turkey supports the UN-recognised Government of National Accord in Tripoli against Haftar, who controls much of the south and east of Libya.
The strongman launched an assault in April 2019 to seize Tripoli, and has the support of the United Arab Emirates and Egypt, countries with whom Turkey's relations are tense.
Aksoy said France's support alongside other countries giving military assistance to Haftar who is attacking "the legitimate government" was "the most serious threat to Libya's territorial integrity and sovereignty."
The Turkish ministry spokesman added: "If France wants to contribute to decisions of the (Berlin) conference being applied, it should first end its support for Haftar."
Ties between Paris and Ankara are increasingly strained over multiple issues including Syria and the eastern Mediterranean.
When Macron declared NATO "brain dead" last year, Erdogan said the French leader was "in a state of brain death".
And Aksoy in his statement also accused France of welcoming "terrorists who threaten Syria's territorial integrity" to the Elysee in a reference to Syrian Kurdish officials meeting Macron last year.
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