The Senegalese defender left it desperately late to score the goal that fired Napoli to within a single point of reigning champions Juve ahead of the last four games of the season.
Massimiliano Allegri’s Bianconeri face a trickier run-in, too, with Champions League hopefuls Inter Milan and Roma providing league opposition in the next few weeks, while there is also the distraction of a Coppa Italia final against AC Milan.
Napoli will hope to capitalise on any slip-up by their modern rivals when they tackle the likes of Torino, Sampdoria and Crotone across their remaining fixtures.
Juve needed steel in defence to nullify Napoli’s forward line but a key figure was removed from the back three early on when Giorgio Chiellini suffered an injury during a clash with Lorenzo Insigne.
Stephan Lichtsteiner replaced him and despite the disappointment of the enforced change to the game plan, Juve came close to scoring with a little over a quarter of an hour on the clock in Turin.
Bosnian schemer Miralem Pjanic rattled the far post with a free-kick after Raul Albiol had fouled Gonzalo Higuain and the touchpaper had been lit.
Napoli soon found channels to run into given the modified Juve set-up and in the 21st minute Mario Rui let loose a twisting drive that Gianluigi Buffon did well to palm over the crossbar.
Another Azzurri raid down the left allowed Mario Rui to set up his captain Marek Hamsik, who grimaced after dragging his shot across Buffon’s goalmouth and wide of the far post.
Napoli had the ball in the back of the net as half-time approached, Insigne prodding home only for referee Gianluca Rocchi to rule that the Italy striker had been offside.
Allegri surprised many when he withdrew Paulo Dybala for Juan Cuadrado at the break, not only removing the X-factor of the Argentinian from the pitch but leaving himself with just one more substitute.
And with Napoli making up consistent ground in the Juve half as the second period progress, the boss rolled the dice and threw Mario Mandzukic into the fray when there were 20 minutes remaining.
Pjanic made it his business to try and pick out the tall Croatian with his pinpoint free-kicks – one in particular caused chaos in the Napoli box.
But the visitors were unruffled and roamed forward to test Buffon with a sighter from Piotr Zielinski.
Juve failed to heed the warning and in the 89th minute the unthinkable happened.
Increasing pressure resulted in a Napoli corner and, when Jose Callejon swung it in, Koulibaly strode forward before leaping and powering home an historic goal with his head.
JUVENTUS AND NAPOLI’S REMAINING LEAGUE GAMES
- April 28 – Inter Milan (A)
- May 5 – Bologna (H)
- May 13 – Roma (A)
- May 20 – Verona (H)
- April 29 – Fiorentina (A)
- May 6 – Torino (H)
- May 13 – Sampdoria (A)
- May 20 – Crotone (H)