The triple Olympic champion will headline the third Diamond League meeting of the season ahead of the defence of his titles in London.
The Jamaican won last year's meeting in 9.91 seconds before sensationally false-starting in the final at the World Championships in Daegu.
"I am looking forward to returning to Rome and to be back on the fast track of the Stadio Olimpico for the Golden Gala," Bolt said on www.iaaf.com.
"I have great memories of last year's edition. It was my first time running in Italy, and I enjoyed the fantastic crowd (of 47,732) at the stadium.
"Someone told me it was the biggest crowd for athletics in the last 10 years, something that made me really happy. I remember the huge coverage from the media there, the press conference I had two days before the race was one of the biggest I ever had outside the Olympics or World Championships.
"I didn't get to see much of the city, maybe I will get to see some more this year.
"I'm training hard, the off season has been good. Everything is going according to plan.
"The aim is to defend my Olympic titles in London. Rome's race will be a good early season test and tell me what I need to do on the way to the Olympic Games."
Bolt has already confirmed he will contest the 100m at the ExxonMobil Bislett Games in Oslo on June 7.