Officials believe the ants were imported to Australia in a shipping container from overseas, rather than arriving from south-east Queensland, where Australia's main fire ant infestation is located.
The nest was destroyed, and specialist fire-ant-detection dog teams have surveyed the logistics hub without finding any other nests. The response will now focus on inspecting and treating properties within a 500-metre radius of the centre and searching for nests within a two-kilometre radius.
The NSW government said local businesses that deal in soil, turf, building materials or vegetation would be contacted to work through measures to identify and treat any fire ant incursion while minimising disruption.
"It's important to note that no further ants have been found, but we are focused on making sure any potential spread is identified and eradicated quickly," Agriculture Minister Tara Moriarty said. "While fire ants are one of the world's most invasive species, they can and have been successfully eradicated in NSW."
Fire ants were previously found at Port Botany in 2014 and were eradicated.
The Invasive Species Council describes fire ants as one of the world's worst super-pests, warning that if the ants became established and spread around Australia they would cost the economy up to $2 billion a year. Advocacy director Reece Pianta said the detection was a reminder of why investment in border biosecurity was so important, and used the news to call for more funding for the national eradication program.
"In south-east Queensland they invade backyards, parks and sporting fields, attack pets and wildlife and threaten agriculture and infrastructure. Sydney does not want to find out what living with fire ants looks like," Pianta said.