Jelena Jankovic is struggling for form ahead of the US Open
Top seed Jelena Jankovic was beaten 7-6 (7-3) 6-3 by qualifier Iveta Benesova in the second round of the Rogers Cup in Montreal.
Jankovic, who has an ankle problem, is struggling to find form ahead of the US Open which begins on 30 August.
"I didn't feel like a top-five player out there," said Jankovic, who lost to Serena Williams at the 2008 US Open in her only Grand Slam final appearance.
"My game hasn't been at the top level for the last couple of tournaments."
Benesova, the world number 75, enjoyed her sixth victory in five days, having won three times last weekend in qualifying plus first-round singles and doubles matches on Monday.
But beating world number three Jankovic was the biggest win of the Czech's 12-year professional career, surpassing her victory over then-world number five Mary Pierce in the second round of the 2006 Australian Open.
"It was almost a perfect match for me," Benesova said. "I was in the zone and focused on my game."
She will face either France's Marion Bartoli and Japan's Kimiko Date-Krumm in round three.
Jankovic got off to a strong start, winning her first two service games to love and breaking Benesova to go up 5-3 in the first, but she was unable to capitalise on her opportunity.
"I really wanted to play matches here, and at the last tournament, but I didn't do that and now I haven't played many matches before the US Open," she added.
"I'll just try my best over there, and hopefully I'll be better."
Meanwhile, Vera Zvonareva made quick work of Kazakhstan's Yaroslava Shvedova, winning their second-round match 6-2 6-1 in 67 minutes.
Svetlana Kuznetsova, however, needed three sets to defeat fellow Russian Anastasia Pavlyuchenkova 7-5 4-6 6-1.
Despite the struggle, Kuznetsova, a recent title winner in San Diego, said: "I think my game is better. I feel like I'm back, I'm back in my game and it all depends on me."
Victoria Azarenka of Belarus beat Kateryna Bondarenko of Ukraine 6-1 6-1, while 15th seed Flavia Pennetta of Italy ousted Slovakian Daniela Hantuchova 6-0 7-5.