Ulissi made his move inside the final 300 meters, bursting out of the peloton to edge Robert Kiserlovski for a second stage win in the race.
Wilco Kelderman was third on the 179-kilometer (111-mile) leg from Foligno to Montecopiolo, which had two category one climbs, including the final ascent to the finish.
“It was a difficult race today, it was really too hard for my characteristics, but of course I am really happy,” Ulissi said.
Evans finished fifth to take the lead from fellow Australian Matthews, who had worn the pink jersey for six days.
Evans has a 57-second advantage over Rigoberto Uran, with Rafal Majka third.
TOUR OF CALIFORNIA: In Pasadena, Calif., Slovakia’s Peter Sagan won the seventh stage in a field sprint and Britain’s Bradley Wiggins closed in on the overall title.
Sagan, riding for Cannondale, jumped to the front just after the final corner to win the 88.7-mile Santa Clarita road race in 3 hours, 24 minutes, 33 seconds.
Wiggins, the 2012 Tour de France winner who rides for Sky, finished in the main field to maintain his 30-second lead over Australia’s Rohan Dennis of Garmin-Sharp. American Lawson Craddock, of Giant-Shimano, remained third overall, 1:48 back.
Norway’s Thor Hushovd, of BMC, was second the stage, followed by Dutchman Danny Van Poppel of the Trek.
There were no changes at the top of the overall standings, with all of the top 10 finishing in the front group.