The image of 81-year-old Stefano Bozzini playing the accordion from an Italian street below his wife's hospital window stole hearts around the world.
Carla Sacchi was allowed out of the hospital near Piacenza a few days ago but has now died at her home.
Although she had not contracted coronavirus, hospital rules meant her husband was unable to visit her.
Piacenza Mayor Patrizia Barbieri said the disease had "broken their embrace".
However, in a Facebook message, she paid tribute to Mr Bozzini's gesture of "tenderness".
"In that serenade we all recognised love, in the simplicity and immediacy of its universal language."