The Duke of Ferrara describes the duchess's habit of riding her white mule "round the terrace." He describes her as smiling and speaking to the mule with great sweetness. The duchess's excessive kindness towards her mule enrages the duke, since he feels she holds him in no higher regard than that animal. He believes her love should be for him alone because of his position as her husband and because of his high standing in society. He is so insecure about this that even her smiling at an animal sets him off, leading to her mysterious death.

The white mule also serves as a symbol of the duchess's innocence and humility. The white color is linked with the former idea, but the very species of the mule presents the duchess as someone who is not prideful or arrogant. Mules were often seen as beasts of burden and not at all beautiful like horses. That the duchess is okay with riding a mule shows she is not vain. The mule image also evokes the biblical story of Jesus Christ entering Jerusalem on the back of a donkey, once again emphasizing the duchess's goodness and her eventual unjust death at the hands of a cruel husband.