Senegalese international Koulibaly, 29, returned a positive after 30-year-old Algerian Ghoulam's result was announced earlier in the day. The club said that Ghoulam was tested on Thursday.
The pair will miss Saturday's league trip to Genoa with Napoli due to host champions Juventus next weekend.
Napoli then face Atalanta in an Italian Cup semi-final, second leg, at Bergamo on Wednesday.
Napoli were already without the club's record scorer Dries Mertens to injury and midfielder Diego Demme is in doubt after taking a knock to the head during the goalless first leg this week.
Captain Lorenzo Insigne limped off during the game in Naples with a calf problem.