The play opens in Parma, as Giovanni and the Friar are having a philosophical discussion concerning the romantic feelings that Giovanni has developed toward his sister, Annabella. Giovanni argues that the prohibition that bars love between a brother and sister is merely an old custom. The Friar counsels him to pray for three days straight on his knees, repenting and weeping, to cure him of his love. Giovanni resolves to try.
In front of Giovanni and Annabella’s house, the servant Vasquez challenges Grimaldi, a roman soldier, to a fight. Just as they begin to fight, Giovanni and Annabella’s father, Florio, emerges from the house with his friend Donado and Soranzo, and asks for an explanation. Annabella and her servant Puttana watch unseen from the balcony of the house. Soranzo explains that he and Grimaldi are rivals competing for Annabella’s love, and Grimaldi had been spreading lies about Soranzo, so he sent his servant Vasquez to challenge Grimaldi. Florio breaks up the fight and insists that no blood be shed over Annabella.
Puttana asks if Annabella is excited by these men fighting over her, but Annabella seems disinterested. Puttana says that she prefers Soranzo because he is wealthy, handsome, and noble. Another of Annabella’s suitors, Bergetto, and his servant Poggio enter below. They speak foolishly about how Bergetto plans to win Annabella, and Annabella makes it clear to Puttana that she thinks he is an idiot. When Annabella spots Giovanni, without realizing who it is, she remarks how attractive and sad the man looks. When she recognizes the man as her brother, she goes down to meet him. Giovanni confesses his love for Annabella, and even though she initially expresses worry about the idea of a relationship between them, she also admits that she loves him. The two resolve to love each other, or else to kill each other, and go off to consummate their relationship in the bedchamber.
Donado convinces Florio to let his nephew Bergetto continue to try to win Annabella’s heart. Florio says that because he has enough wealth, he does not need to force Annabella to marry against her will, and will defer to her choice. Bergetto and Poggio seem more concerned with ponies and puppet shows than with love, so Donado decides that he must write a letter to Annabella on Bergetto’s behalf.
Annabella and Giovanni return from the bedchamber, and she promises that she will not marry another man. Giovanni leaves, and Puttana expresses her support for the relationship even though they are siblings. Florio introduces Annabella to the doctor he has called to look at her because she has not been feeling well. This “doctor” is in fact Richardetto in disguise, who is accompanied by his niece Philotis.
Soranzo is lost in a soliloquy about how deeply he is in love with Annabella when he is interrupted by a former lover, Hippolita. She is enraged because she and Soranzo had carried on an affair while her husband was alive and Soranzo had promised that if her husband ever died, he would marry her. She then encouraged her husband to go on a dangerous journey to find his niece and, as she had hoped, he died on the journey. However, Soranzo did not keep his promise to her, and now Hippolita curses him. Soranzo grows angry at her accusations and scorns her as he leaves.
Vasquez tells Hippolita that she should try to talk with Soranzo at another time, and with less malice. Hippolita tries to seduce Vasquez into helping her get revenge against Soranzo. He agrees, but in asides he reveals that his true intention is to remain loyal to his master.
Richardetto reveals that he is Hippolita’s husband, who is thought to be dead. He begins to plot against Soranzo and Hippolita because the two carried on an affair under his nose. Grimaldi enters, asking for a love potion to make Annabella fall in love with him, but Richardetto comes up with a different plan: he convinces Grimaldi to kill Soranzo, and supplies him with a poisoned dagger.
Giovanni visits the Friar and confesses what has happened between him and Annabella. The Friar counsels him to marry her, but Giovanni says that that would ruin her reputation. Meanwhile, Donado writes a letter to Annabella on Bergetto’s behalf, but Bergetto and Poggio still seem more interested in puppet shows than in marriage. Donado delivers the letter and Annabella takes it graciously, but says that she cannot love Bergetto. Bergetto enters and says that he has fallen in love with Philotis. He reveals to Poggio that he intends to marry her in secret because his uncle would object to the match.
Soranzo then tries his hand at wooing Annabella, but she is indifferent to him as he pleads with her. She says that she has no intention of marrying, but assures him that if she were to marry, she would marry him. He is temporarily appeased. Suddenly Annabella falls ill, and Puttana reveals to Giovanni that Annabella is pregnant with his child.
Richardetto looks at Annabella and tells Florio that she has the maid’s sickness (a type of anemia that often affected young women in puberty). Florio resolves that she should marry Soranzo immediately, and the Friar convinces Annabella that this is the best course of action to atone for her sins.
That same night, Grimaldi hides with the poisoned dagger Richardetto gave him, and waits for Soranzo to visit Annabella. Bergetto and Philotis pass, on their way to the Friar’s cell to be married, and Grimaldi mistakes Bergetto for Soranzo. He stabs Bergetto, who then dies. Donado, Florio, Richardetto, and several officers go to find Bergetto’s murderer, but the Cardinal protects Grimaldi, so Bergetto receives no justice.
Two days later, at Soranzo and Annabella’s wedding, feast, Hippolita interrupts the festivities. She claims to want to lay to rest her anger towards Soranzo, but when she makes a toast to the couple, Vasquez gives her the poisoned wine that she had meant for Soranzo. Before she dies, she curses Soranzo and Annabella. Richardetto tells Philotis that she should avoid men and sex entirely and become a nun. She agrees and leaves to join a nunnery.
Some days later, Soranzo has found out that Annabella had sex before their marriage and is pregnant. He drags her across the room and threatens with a sword to tell him who the father is. She refuses to reveal her lover’s name, and Vasquez advises Soranzo to be gentler with Annabella. Meanwhile, he finds out from Puttana that it was Giovanni, and he then has the Banditti (a gang of robbers) gag and blind Puttana for her complicity in the incest. Vasquez and Soranzo plot to get revenge on Giovanni at Soranzo’s birthday feast.
Annabella writes a letter to Giovanni about what has happened and how she wants to repent for her sins, which the Friar delivers to him. Giovanni realizes that Soranzo has found out about her pregnancy and says that he will get his own revenge on Soranzo at his birthday feast. Before he attends the feast, he visits his sister in her bedchamber where, in order to save her reputation, he stabs her to death, hoping that he will see her in heaven.
The Cardinal, Florio, Donado, Soranzo, Richardetto, and Vasquez are all in attendance at the feast. Suddenly, Giovanni bursts in with Annabella’s heart on his dagger. Giovanni confesses to his incestuous relationship, and Florio dies from a broken heart. Giovanni then stabs Soranzo. Vasquez jumps in and stabs Giovanni, then cries out “Vengeance!” – the watchword for the Banditti, who have been hiding. The Banditti attack and severely wound Giovanni. Soranzo is pleased that he was able to get revenge on Giovanni, and he dies. Giovanni is also happy to have gotten his own revenge on Soranzo and dies quickly after. Vasquez is satisfied that he has carried out a servant’s duty for Soranzo.
The Cardinal asks if anyone else knew about the siblings’ relationship. Vasquez tells him about the now-blind Puttana, and the Cardinal demands that she be taken outside the city and burned to death. Vasquez, for his own crimes, shall simply be banished. Richardetto finally reveals himself, satisfied that he has seen Soranzo killed and thereby been avenged. The Cardinal surveys the bodies and concludes the strange tale, lamenting that it is a pity that Annabella was a whore.
