New York - French fifth seed Jo-Wilfried Tsonga became the first major casualty of the US Open on Thursday when he slumped to a 6-4, 1-6, 6-1, 6-3 defeat to unheralded Slovak Martin Klizan.
Tsonga, who made the quarter-finals in 2011, had come into the final Grand Slam event of the season in a fog of injury and form worries summed up by having to skip the Cincinnati event after cutting his knee on a fire hydrant.
His defeat was good news for British third seed Andy Murray, the Olympic champion, who had been due to face the Frenchman in the quarter-finals.
Left-hander Klizan, the world No 52, progressed to the third round of a Grand Slam for the first time and will tackle either 32nd seed Jeremy Chardy of France or Australia's Matthew Ebden.
Klizan only won his first career Grand Slam match this year at the French Open, spending most of his time on the second-tier Challenger tour where he won four titles this season.
But he shrugged off that inexperience on Thursday, firing 32 winners to help offset the 42 unforced errors he sent down in a daring display of attacking tennis.
Elsewhere on Thursday, Spanish 11th seed Nicolas Almagro fought back to beat Philipp Petzschner 6-3, 5-7, 5-7, 6-4, 6-4 with the German's challenge undone by 62 unforced errors.
Japan's 17th seed Kei Nishikori also made the third round with a 6-2, 6-2, 6-4 win over American qualifier Tim Smyczek.
Serbia's Ana Ivanovic reached the third round, but admitted she needs to take painkillers every morning to see her through the season's last Grand Slam tournament.
The 12th-seeded Serb, who has never got beyond the fourth round in New York, made the last 32 with a 6-2, 6-2 win over Sofia Arvidsson of Sweden.
But the 2008 French Open champion is still bothered by a right foot injury which forced her out of the Cincinnati tournament on the eve of the US Open.
"I still have to take a handful of pills every morning," said the 24-year-old.
"It was a little bit unfortunate but it's part of the sport. We play so much and changing surfaces and continents. My foot has been healing and at the moment I am feeling healthy."
Ivanovic goes on to face either America's Sloane Stephens or German qualifier Tatjana Malek for a place in the last 16.
Fellow Serb Jelena Jankovic, seeded 30th and the 2008 runner-up, also made the third round with a 6-4, 6-2 win over Spain's Lara Arruabarrena-Vecino.
She next meets either second seed Agnieszka Radwanska of Poland, the Wimbledon runner-up, or Spain's Carla Suarez Navarro.
Russian 14th seed Maria Kirilenko, watched by ice hockey superstar boy friend Alex Ovechkin, saw off Hungarian 33-year-old Greta Arn 6-3, 6-2.
Slovakian 13th seed Dominika Cibulkova, a quarter-finalist in 2010, edged Serbia's Bojana Jovanovski 7-6 (7/3), 7-6 (7/3).
Later on Thursday, five-time US Open champion Roger Federer, bidding to become the first six-time winner in the Open era, tackles 32-year-old German Bjorn Phau.
The two first met in 1999 in Washington, which was the first North American hardcourt event for either man - and Phau won in straight sets.
Federer won both their subsequent meetings, at the 2007 Australian Open and at the 2010 Estoril clay-court tournament.
World No 1 Federer is playing in his 52nd straight Grand Slam event while Phau, the world 83, has never gone beyond the second round of a major.
Both Williams sisters also take to the court looking to reach the third round.
Three-time champion Serena, who suffered an ankle injury scare playing doubles on Wednesday, goes for her 60th career win at the tournament when she tackles Spain's Maria Jose Martinez Sanchez.
That could be a spicy affair after Williams accused the Spaniard of cheating when they clashed in the 2009 French Open.
Two-time champion Venus faces sixth seed, and 2011 semi-finalist, Angelique Kerber, just a month after the German knocked her out of the Olympics.
Kerber has won a tour-leading 54 matches in 2012 and also ended Serena Williams' 19-match win streak with victory in the Cincinnati quarter-finals.
Latest results from the fourth day of the US Open on on Thursday (x denotes seeding):
Kei Nishikori (JPN x17) bt Tim Smyczek (USA) 6-2, 6-2, 6-4
Nicolas Almagro (ESP x11) bt Philipp Petzschner (GER) 6-3, 5-7, 5-7, 6-4, 6-4
Martin Klizan (SVK) bt Jo-Wilfried Tsonga (FRA x5) 6-4, 1-6, 6-1, 6-3
Feliciano Lopez (ESP x30) bt Pablo Andujar (ESP) 6-4, 6-1, 6-7 (5/7), 3-6, 7-5
Jack Sock (USA) bt Flavio Cipolla (ITA) 6-2, 6-2, 6-4
Ana Ivanovic (SRB x12) bt Sofia Arvidsson (SWE) 6-2, 6-2
Maria Kirilenko (RUS x14) bt Greta Arn (HUN) 6-3, 6-2
Jelena Jankovic (SRB x30) bt Lara Arruabarrena-Vecino (ESP) 6-4, 6-2
Dominika Cibulkova (SVK x13) bt Bojana Jovanovski (SRB) 7-6 (7/3), 7-6 (7/3)
Andrea Hlavackova (CZE) bt Galina Voskoboeva (KAZ) 6-2, 6-7 (3/7), 6-3
Tsvetana Pironkova (BUL) bt Ayumi Morita (JPN) 7-5, 6-2
Ekaterina Makarova (RUS) bt Elena Vesnina (RUS) 6-3, 6-4