The bus was travelling from Montecatini toward Siena when it crashed through a guard rail at around 9:30 am.
The victim, identified as a Russian woman in her 40s, was removed from the wreckage following the accident on the Siena-Florence highway, ANSA said.
The woman died at the scene.
Another 31 people are reported injured, with 15 believed to be in a serious, though not life-threatening, condition.
The cause of the accident was not immediately known.
The bus driver tested negative for alcohol, but “we’re not ruling distraction as one of the causes”, said regional highway police chief Paolo Maria Pomponio.
“It could have been worse, the vegetation cushioned the vehicle’s fall,” he added.
An investigation was underway on Wednesday to determine the cause of the crash.