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Rizzio enters, and Lady Argyle attempts to seduce and coerce Rizzio to give up the treaty banishing Lennox, but he, loyal to Mary, refuses, and at Lady Argyle's signal Murry and his armed men attack Rizzio, but are foiled by the return of Darnley. Mary enters from Mass with a group of Catholic priests and nuns. Darnley, who has been stung by Murray's accusations, asks Mary when his coronation will take place; the queen, disgusted by his drunkenness, sends him away; she also sends away Murray, who attempts to seduce her. Alone, she bemoans her position, alone in an alien country. ("Sola, abbandonata") David Rizzio enters, and Darnley, hidden there by Murray and Argyle, witnesses a tender scene between them: Mary declares her love for Rizzio, who begs her to remain her servant. Mary, angrily, sends them away, but Darnley, enraged by what he has seen, signs the compact.
An intermezzo follows, after which Act II begins. Mary and her court are at dinner, with a storm beginning outside. Darnley enters and insults Mary, who grows angry; Rizzio pleads with the queen for mercy for the hapless consort. ("Clemenza, oh, Madonna") He is joined by the whole court in his plea for mercy. Mary agrees and the banquet continues, but soon the party is terrified by the growing storm. At the height of the storm, the banished Lennox himself bursts in, with armed men and Lord Ruthven, who has come from his deathbed to denounce Rizzio. He declares his innocence and denounces the court for its suspicion; he declares that he will leave the next morning to return to Italy. ("O gente, che state affondate") The lights in the room are blown out by a lightning bolt striking the tower, and Murray and Ruthven stab Rizzio. Mary curses the conspirators. Murray, terrified by the Queen's curse, runs off after the rest of the conspirators; Lady Argyle, repentant, weeps over Rizzio's body as Mary faints.