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The play was first published in , in a quarto printed by Nicholas Okes for the bookseller Richard Collins. Giovanni, recently returned to Parma from university in Bologna, has developed an incestuous passion for his sister Annabella and the play opens with his discussing this ethical problem with Friar Bonaventura. Bonaventura tries to convince Giovanni that his desires are evil despite Giovanni's passionate reasoning and eventually persuades him to try to rid himself of his feelings through repentance.
Annabella, meanwhile, is being approached by a number of suitors including Bergetto, Grimaldi, and Soranzo. She is not interested in any of them, however. Giovanni finally tells her how he feels obviously having failed in his attempts to repent and finally wins her over. Annabella's tutoress Putana literally, "Whore" encourages the relationship.
The siblings consummate their relationship. Hippolita, a past lover of Soranzo, verbally attacks him, furious with him for letting her send her husband Richardetto on a dangerous journey she believed would result in his death so that they could be together, then declining his vows and abandoning her. Soranzo leaves and his servant Vasques promises to help Hippolita get revenge on Soranzo, and the pair agree to marry after they murder him. However, Richardetto is not dead but also in Parma with his niece Philotis.
Richardetto is also desperate for revenge against Soranzo, and convinces Grimaldi that to win Annabella, he should stab Soranzo with a poisoned sword. Unfortunately, Bergetto and Philotis, now betrothed, are planning to marry secretly in the place Richardetto orders Grimaldi to wait.
Grimaldi mistakenly stabs and kills Bergetto instead, leaving Philotis, Poggio, and Donado distraught. Annabella resigns herself to marrying Soranzo, knowing she has to marry someone other than her brother. She subsequently falls ill and it is revealed that she is pregnant. Friar Bonaventura then convinces her to marry Soranzo before her pregnancy becomes apparent. Meanwhile, Donado and Florio go to the cardinal's house, where Grimaldi has been in hiding, to beg for justice.