A total of 72 players are now isolating in their hotel rooms ahead of the tournament.
The fifth person tested positive on the night of Jan. 17 and they were on a charter flight from Doha that landed in Melbourne on Saturday.
It means three Australian Open charter flights have carried passengers who later tested positive, following infections on planes from Los Angeles and Abu Dhabi.
COVID-19 Quarantine Victoria on Sunday said four people in the Australian Open contingent and one airline crew member had now tested positive.
Despite some of the affected players complaining of being unable to leave their rooms for training, COVID-19 Quarantine Victoria commissioner Emma Cassar insisted the program would not be modified.
Cassar said that there had been cases of “challenging behaviour” from some confined players and support staff.
She cited two cases when they opened their doors to have a conversation or shout down the hallway.
“There is zero tolerance for breaches,” she said.
“I had a conversation with Victoria police to ensure we are increasing our compliance and enforcement efforts.
“It’s low level but dangerous acts that we just can’t tolerate.”
Cassar warned they could be fined up to $20,000 or repeat offenders transferred to the complex care hotel where they have a police officer stationed outside their door.
Australian Open boss Craig Tiley has ruled out delaying the tournament, which is due to begin on February 8.