Ennis, who is one of the top heptathletes in the world but also an accomplished hurdler, stormed to victory at the Great CityGames in Manchester on Sunday with a career-best time of 12.75 seconds, knocking 0.04s off her previous fastest set last year.
Not only was it her best time, but she had beaten Olympic champion Dawn Harper of the United States in the process.
But officials had only laid out nine of the 10 hurdles, meaning her time was rendered invalid.
"I am so annoyed," said Ennis, who is one of the favourites to win heptathlon Olympic gold in London this summer. "I still had a good competitive race, but I've just not got the result I wanted.
"It’s a massive, massive mess-up. I can’t believe it."
Organisers Nova International admitted to the error, promising a full investigation.
"There has been a technical error and there were nine sets of hurdles out on the course instead of 10," a spokesman for Nova said.
"We'll conduct a thorough investigation and find out what happened and why and by whom.
"It's a very unfortunate mistake, we're very unhappy about it."
Dwain Chambers, who was recently cleared to compete in the Olympics after the British Olympic Association's lifetime ban for convicted dopers was overturned by the Court of Arbitration for Sport, was also taking part in his first race in Europe since the decision went in his favour.
Chambers received a warm welcome from the fans before finishing second in a showpiece 150 metres in 15.27 seconds.
The Great CityGames is a special, free event featuring some of the world’s top athletes competing on a specially constructed street track.
It is affiliated with the Great Manchester Run 10,000m race. Haile Gebrselassie won the men’s race, with Linet Masai triumphing for the women.