"I am disappointed to have to withdraw from Rome this year, but unfortunately I do not feel well enough to compete," world number four Rybakina said in a statement.
"I have such good memories from last year and was looking forward to defending my title."
Former Wimbledon champion Rybakina was supposed to open her title defence in the Italian capital on Friday against Romania's Irina-Camelia Begu.
Instead, the 24-year-old, who represents Kazakhstan, has been replaced by lucky loser Oceane Dodin, the world number 73 from France who was eliminated in the second round of qualifying.