The bus was attacked by unidentified gunmen as it traveled its twice-weekly route from the village of Songho to a market in Bandiagara, 10 kilometres (6 miles) away, said Moulaye Guindo, mayor of the nearby town of Bankass.
"Armed men ... shot at the vehicle, slashed the tires, and shot at the people," Guindo said.
He and another local official, who spoke on condition of anonymity, said at least 31 were confirmed dead with many more wounded or missing.
The villages sit in the heart of the Mopti region, an epicentre of violence in Mali fuelled by insurgents linked to al Qaeda and Islamic State.
An internal security memo, shared by a source, said first responders to the scene of the latest attack discovered 25 burned bodies in the truck.
Images circulated on social media showed the smouldering frame of a passenger bus filled with bodies, still slumped over in their seats. Reuters could not independently verify the authenticity of the images.
Jihadist attacks have surged across Africa's Sahel region, killing thousands and displacing millions across Mali, Burkina Faso and Niger.