* World number one Serena upset by Spain's Muguruza
* Sister Venus falls to Slovak teenager Schmiedlova
* Djokovic eases past Frenchman Chardy in three sets
* Fourth seed Federer breezes into third round
On a chilly day in Paris, Serena never looked the force that has won 17 major titles. Her 20-year-old opponent broke the American's serve twice early in the first set and made the most of a series of unforced errors by a leaden-footed Williams.
The Spaniard, ranked 35 in the world, did not release the pressure and the match became a repeat of the nightmare of 2012, when Williams lost in the first round, not the glory of 2013 when she beat Maria Sharapova in the final here.
A netted return from Williams sent the Spaniard into ecstasy and even Williams's mother, Oracene, smiled as Muguruza leapt about the court. "Of course, it's amazing," Muguruza said. "I had to be really aggressive and focused.
"Today is a great day. I'm so happy," the Spaniard shouted.
Later Muguruza told a news conference: "She (Serena) said that if I continue playing like this, I can win the tournament. I said: 'I will try, I will try'.
Serena was philosophical, saying: "I don't think anything worked for me today. It was just one of those days, you can't be on (your game) every day.
"It's not the end of the world, there's always next year."
After the defeat of China's Li Na in the first round on Tuesday, it is the first time since tennis turned professional in 1968 that the top two women's seeds have failed to reach the third round at a grand slam.
It is also the third time, after 2004 and 2008, that both Williams sisters have lost on the same day at the French Open.
The big names on the men's side of the draw suffered no problems in reaching the third round.
Novak Djokovic, the second seed, despatched Jeremy Chardy of France 6-1 6-4 6-2 while Roger Federer, seeded fourth, won 6-3 6-4 6-4 against Diego Sebastian Schwartzman of Argentina, a player he had never seen before.
Chardy, who beat Federer at the Rome Masters earlier this month, spelled out why Djokovic is fancied to end eight-times champion Rafa Nadal's reign here.
"He was simply stronger than me, bigger than me in all compartments. You're always under pressure. He knows everything. He can do everything."
Czech Tomas Berdych, the sixth seed, started slowly before beating Aleksandr Nedovyesov of Kazakhstan 6-7(4) 6-4 7-5 6-3. Eighth seed Milos Raonic got past Jiri Vesely of the Czech Republic 7-6(4) 6-4 6-1.
The Williams sisters had been due to meet in the third round but that was knocked off the agenda when Venus became the first seed to lose in the second round, falling 2-6 6-3 6-4 to Anna Schmiedlova of Slovakia.
The 29th seed, wearing long sleeves in temperatures of 14 Celsius, has never won the claycourt grand slam in 16 visits and, now 32, she may have to accept she never will.
Schmiedlova, aged 19 and ranked 56 in the world, wasted two match points on Williams's serve before the American hit wide to set up a third, and the Slovak clinched the match with a fine backhand cross court shot.
"This is the biggest win of my life," said Schmiedlova, who broke down when expressing how much Venus had meant to her when she was growing up.
Venus was diagnosed with an auto-immune disease in September 2011 and since losing in the fourth round of Wimbledon that year, has exited in the first or second round of every grand slam she has taken part in.
"I think she just played so well and just kept getting so many balls in the court," said Venus. "It just wasn't my day in the last couple of sets."
Flavia Pennetta of Italy, seeded 12th, joined the Williams sisters in heading for the exit after going down 5-7 6-4 6-2 to Sweden's Johanna Larsson.
Eugenie Bouchard, the 18th seed, recovered from a slow start to beat Julia Goerges of Germany in three sets.
On the men's side, Russian Mikhail Youzhny, the 15th seed, lost 6-0 6-3 3-6 6-4 to Czech Radek Stepanek before Spain's Marcel Granollers overturned a two-set deficit to beat 20th seed Alexandr Dologopolov of Ukraine 1-6 3-6 6-3 6-0 6-2.
Gilles Simon, the 29th seed, eased past Alejandro Gonzales of Colombia 6-4 6-0 6-2 and there were also straight-set wins for Spain's Roberto Bautista Agut, the 27th seed, and Ernests Gulbis, the 18th seed from Latvia.