Earlier this week, a video of a phone call between Koulibaly and former incumbent John Terry went viral, the latter's legendary shirt having remained vacant since his departure.
However, having worn that number during his time at Napoli and in pre-season, the Senegal international will now take it over in West London.
A number of options were available to fellow new arrival Raheem Sterling, including the number nine shirt left vacant by Romelu Lukaku.
Instead, Sterling will take on the number 17 shirt, made available by Mateo Kovacic's recent switch to the number eight.
On Thursday morning, Chelsea announced that they had completed the signing of Carney Chukwuemeka on a six-year contract.
The 18-year-old, who has departed Aston Villa to join his boyhood club, will wear number 30, which was not occupied during last season.
Meanwhile, Conor Gallagher will wear the number 23 as he prepares to be involved in his first season at Chelsea after a number of loan spells.
Armando Broja, who has been linked with a move away from the West Londoners, has been provided with number 18, potentially a sign that Thomas Tuchel intends to retain his services.
Malang Sarr has kept the number 31 shirt, even if the defender has been heavily linked with a switch to Monaco over the coming days.
As it stands, the numbers two, four and 15 remain available for any new signings, with Chelsea known to want Leicester City defender Wesley Fofana and Brighton & Hove Albion left wing-back Marc Cucurella.
However, Levi Colwill is one of numerous players to be left without a number, the youngster expected to move to the Seagulls should Cucurella eventually make the switch in the opposite direction.
Ross Barkley, Kenedy, Ethan Ampadu, Billy Gilmour and Michy Batshuayi also miss out, an indication that they will depart Chelsea before the start of September.