The snake, measuring at 18 feet 2 inches (5.5m), fell short of the state record by 6 inches, according to the Florida Fish and Wildlife Conservation Commission.
Last year, a snake collector in the state discovered the largest python on record there, measuring 18 feet 8 inches, commission spokeswoman Katie Johnson said.
The pythons, which can grow to more than 20 feet in their native habitat in Southeast Asia, are one of the most problematic invaders of Florida's sprawling Everglades wetlands.
They eat indigenous species and their food sources, fueling concerns that the predator snakes will fundamentally change the ecosystem.
The python was killed, and its corpse was taken to the University of Florida, where it will be measured and studied by scientists trying to combat the species, according to South Florida Water Management District spokesman Gabe Margasak.
Officials have said the python population is believed to have grown to as many as 150,000 in the Everglades. The cold-blooded reptiles are often found atop levees, where they lie for hours at a time to warm up under the Florida sun.
The snakes, one of the largest species in the world, found a home to their liking in the Everglades when pet owners started using the wetlands as a convenient dumping ground.
