Maria Sharapova endured a day to forget as the Russian retired from the Internazionali d'Italia soon after discovering she would not get a French Open wildcard, while Julia Goerges toppled the in-form Kristina Mladenovic in Rome.
French Tennis Federation president Bernard Giudicelli announced that Sharapova had been snubbed for the second major of the year less than an hour before she stepped out on court to face Mirjana Lucic-Baroni in the second round.
The five-time grand slam champion was a break up at 2-1 in the final set when she shook hands with Lucic-Baroni, unable to continue due to an injury to her left thigh.
Sharapova, who returned from a 15-month doping ban last month, will now need to come through Wimbledon qualifying or be handed a wildcard in order to play at SW19, but would have gained direct entry to the main draw by reaching the semi-finals this week.
Mladenovic reached the Madrid Open final last week, but was beaten 7-6 (8-6) 7-5 by the unseeded Goerges on day two in the Eternal City.
Daria Gavrilova battled back to stun Madison Keys, while fifth seed Johanna Konta eased to 6-3 6-0 win over Yulia Putintseva and will face Venus Williams in the next round if the American dumps Lesia Tsurenko out on Wednesday.
Sharapova was broken to love in the first game of the match and was unable to recover in the first set, but took advantage of errant Lucic-Baroni serving to level.
She was on top in the third, but went off court for a medical timeout and returned with strapping on her left thigh. Despite attempting to continue, she had to accept defeat on a miserable day for the two-time Roland Garros champion.
"I apologise for having to withdraw from my match today with a left thigh injury," said the 30-year-old.
"I will be getting all the necessary examinations to make sure it is not serious. I want to thank the tournament for giving me the opportunity to play in this special event again."
Mladenovic has been on a roll, progressing to the final in Madrid and Stuttgart but losing to Simona Halep and Laura Siegemund respectively in those showdowns.
The world number 14 from France fell at the first hurdle in Rome, as Georges dominated from the baseline and struck 29 winners to march into the second round.
German Georges came from a break down in the opening set and a mini-break down in the tie-break to take the opener and needed just the one break in the second to pull off the upset.
Gavrilova stunned 10th seed Keys, coming from behind to triumph 2-6 7-5 7-5.
Keys reached the final in Rome last year, but Australian qualifier Gavrilova came from a break down in the final set to win a contest which swung one way and another.
Konta had no such trouble, serving six aces and coming up with 36 winners as 32 unforced errors proved to be costly for Putintseva.
There were also wins for Anett Kontaveit, Anastasija Sevastova, Jelena Ostapenko, Kiki Bertens and CiCi Bellis. Lauren Davis, Ekatervina Makarova, Svetlana Kuznetsova and Timea Bacsinszky were also victorious.