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Fourth seed Agnieszka Radwanska became the first high-profile casualty of the US Open as she lost 6-3 6-4 to China’s world number 39 Peng Shuai.
The Pole has failed to advance beyond the last 16 in nine visits to New York.
Fifth seed and 2006 champion Maria Sharapova survived the loss of the first set to see off world number 95 Alexandra Dulgheru 4-6 6-3 6-2.
But American 21st seed Sloane Stephens also suffered a shock defeat, beaten 5-7 6-4 6-2 by Sweden’s Johanna Larsson.
Wimbledon semi-finalist, and Sharapova’s off-court partner, Grigor Dimitrov made easier work of his second-round tie, dispatching American Ryan Harrison 6-2 7-6 (7-4) 6-2 in a little over two hours.
Two-time champion Venus Williams, seeded 19th in this year’s draw, made the third round for the first time in four years as she beat Switzerland’s Timea Bacsinszky 6-1 6-4. The American will play Italian 13th seed Sara Errani next,
Former world number one Caroline Wozniacki eased aside Grand Slam debutant Aliaksandra Sasnovich of Belarus 6-3 6-4 to set up an intriguing meeting with Germany’s Andrea Petkovic, who she beat in the 2011 quarter-final.