In the year to April 2009, the number of residents shrunk by 58,294, largely due to a collapse in the local real estate sector, a report from the University of Florida said.
It is the first time the population of the "Sunshine State" has dropped since 1946, when troops left after the end of World War II.
"The population decline is really a reflection of how severe the national recession has been," said the university's Stan Smith, who led the research.
"Traditionally, Florida's growth has been spurred by both a booming economy and a booming housing market, and both have seen substantial losses over the last couple of years."
In February, US President Barack Obama selected Fort Myers, Florida, the epicenter of the mortgage foreclosure crisis, to highlight his government's response.
"This reflects a very abrupt change from three or four years ago, when Florida was experiencing some of its largest population increases ever," said Smith predicting long-term population growth would eventually return.