The charter bus from Austin, Texas, carrying 50 to 60 people, was heading north on Main Street when it was hit by an eastbound CSX train around 2:13 p.m., according to the spokesman, Vincent Creel.
Several people were injured, including some seriously, officials said. The train crew was not injured, said CSX spokesman Gary Sease.
Sease said the eastbound mixed-freight train, heading from New Orleans to Mobile, Alabama, had three locomotives and 52 cars, 27 loaded cars and 25 empty cars.
The crossing has flashing lights and crossing gates, according to Sease.
“This is a tragic event for our city,” Biloxi Mayor Andrew Gilich said.
Police are talking to the bus driver and the train operator.
Sease said CSX personnel are working with first responders at the scene.
Mississippi Gov. Phil Bryant tweeted: “Please pray for the injured and the families of the lost.”
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