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The varroa destructor was first found at a port near Sydney last week but has since been spotted in hives 100km away.
The outbreak threatens to cost the honey and food production industries millions of dollars.
Keepers inside a new biosecurity zone will not be able to move hives, bees or honeycombs until further notice.
Australia was the only continent free of the varroa mites, which are the biggest threat to bees worldwide.
The pests - which are about the size of a sesame seed - weaken and kill colonies by feasting on them and transmitting viruses.
Authorities have introduced several biosecurity measures to limit the outbreak, after the mites were detected at seven sites across New South Wales.
Any hives within 10km of infested locations - about 400 so far - will also be destroyed, while colonies within 25km will be inspected and monitored.
A "bee lockdown", banning movement of bees across the state, is also being enforced.
That is critical to stop the outbreak devastating producers, says NSW Farmers' Ian McColl.
If the mites continue to spread, it could cost the honey production industry alone $70 million ($48m, £39m) a year, he says.
About a third of Australia's food production relies on bee pollination, including almonds, apples and avocadoes.
"Bees are an integral part of our production system," Mr McColl added. "This is not just a concern for industry, but it's a major community concern as well."
Previous detections of other kinds of varroa mite in Queensland and Victoria have been eradicated and farmers expect the next few days to be critical in tackling the outbreak.
