Venus Williams was unable to win back-to-back WTA Tour matches for the first time since 2019 after letting a lead of a set and a break slip in Cincinnati.
On the day Williams received a wildcard for a record-extending 24th US Open, the 43-year-old American lost 1-6 6-2 6-1 to world number 24 Zheng Qinwen.
Williams, ranked 533rd, led 6-1 2-0 before losing 11 games in a row.
China’s Zheng, 20, served out victory and plays top seed Iga Swiatek in the Western and Southern Open last 16.
Williams will now continue her preparations for the US Open, where she will extend her own record of Open era singles appearances.
The seven-time major champion made her main-draw debut at Flushing Meadows in 1997, going on to win the 2000 and 2001 titles.
Two-time finalist Caroline Wozniacki, who is making a comeback after retiring in 2020, is also among the wildcards.
Dane Wozniacki, 33, will play in her third tournament since returning to the WTA Tour earlier this month.
The US Open, which is the final major event of the season, starts on 28 August.
Swiatek looking sharp before US Open defence
Polish world number one Swiatek, 22, continued her preparations for the defence of her title in New York with a ruthless 6-1 6-0 win over American wildcard Danielle Collins.
“I feel like I was really solid and I didn’t make a lot of unforced errors,” Swiatek said. “I really like playing that way because it gives me confidence.”
American world number three Jessica Pegula, who won the Canadian Open title last week, also reached the Cincinnati last 16 with a 6-7 (2-7) 6-2 6-3 victory over Italy’s Martina Trevisan.
Wimbledon champion Marketa Vondrousova of the Czech Republic won 6-4 6-2 against Russia’s Anastasia Popatova, while Russian 14th seed Daria Kasatkina also progressed by beating France’s Varvara Gracheva.
Greek eighth seed Maria Sakkari avoided a shock against Romania’s Sorana Cirstea, fighting back to win 2-6 6-3 7-5.
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