Kerber hit 25 unforced errors in her 82-minute defeat to Makarova, who looked like her former top-10 self and was untroubled throughout the clash.
Kerber went down 6-2 6-2 in one hour and 22 minutes, leaving her number one status in the balance.
The German, who also lost in round one past year, has had a miserable season and is not a fan of clay so the result came as no real surprise.
The only seed she may have to face is recent Strasbourg champion Samantha Stosur, who will face Kristina Kucova in her first match tonight.
The 29-year-old German slumped to a dispiriting 6-2, 6-2 defeat to experienced Ekaterina Makarova of Russian Federation today.
Kerber now has just two wins from her past four tournaments.
Totti leaves future uncertain before final match with Roma
He was widely expected to quit playing last summer but signed a one-year extension to his contract. I don't imagine him going to another Italian club. "I'm ready for a new challenge", Totti added.
"I'm really glad to have made the decision to play here", said 27-year-old Kvitova, who was seriously hurt when she fought off a knife-wielding burglar at her home in the eastern Czech town of Prostejov in December. Clayking, Rafael Nadal will be looking to complete his La Decima (Tenth French Open title). I think I played well after six months off. So I think this is the most, yeah, thing that it's a little bit different than to the other surface. The 31st-seeded Vinci, most famous for ending Williams' hopes of a calendar year Grand Slam by beating her in the semifinals at Flushing Meadows in 2015, lost to Rio Olympics gold medalist Monica Puig 6-3, 3-6, 6-2. "And just, you know, going out and trying to find my game", she said.
But Sunday's setback - albeit on a surface for which she has no great affection, having made an opening-round exit in Paris a year ago - threatens to leave her season in tatters.