Briton Mark Lewis-Francis was also disqualified for false a false start.
The Jamaican, with typical showmanship, had earlier arrived at the stadium in a faux-vintage dragster with sparks flying out of the back, and he did not disappoint on the track.
Bolt finished in a time of 9.79 seconds to edge out Asafa Powell, who recorded a season's best time of 9.85.
Lewis-Francis' false start came shortly after Ennis, who had finished third in her heat, was disqualified from the 100m hurdles for the same offence.
Australian Sally Pearson took the victory in a time of 12.49 ahead of American Kristi Catlin and Britain's Tiffany Porter.
"It wasn't the best execution as I had a good start so I have to go home and fix a couple of things," Porter told BBC Sport. "But it'll be okay."
World champion Pearson added: "I travelled 29 hours to get here so I'm a bit jet-lagged but doing well. I've put myself in the position I want to be in. I like being chased. It keeps me on my toes."
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Dai Greene could only manage fourth in the 400m hurdles with a time of 48.98, having coming in to the event off the back of an illness, with Javier Culson cruising to victory in 47.94 - the fastest time of the year.
"It's a bit disappointing to be honest, but given how things were last week with my illness it's a bit much to ask to be up to my game," he said.
"I haven't done much speed work yet. I'm a bit disappointed with the time but I hope to get that down over the next few weeks."
But there was better news for another Welshman, with Gareth Warburton winning the 800m in one minute 44.98 seconds, a Welsh record and inside the Olympic 'A' standard.
Abiodun Oyepitan finished second to Murielle Ahoure in the women's 200m, but was not happy with an apparently impressive time of 22.71.
"I always want to win," she said. "I didn't execute as well as I would like. I could have gone harder. It's Olympic year and I want to win all my races."
Milcah Chemos won the women's steeplechase in a blistering time of 9:07.15, the fourth fastest time in history, but Britain's Eilish McColgan could be happy with her ninth place, finishing in a new personal best of 9:38.45.