Australia, fired by a game-high 30 points from Patty Mills, ended the USA's 78-game winning streak in competitions and exhibitions on Saturday, claiming a shock 98-94 win in a World Cup warm-up in Melbourne.
In front of a record crowd at Docklands stadium, the USA suffered their first loss in 13 years, while Australia secured their first ever win over the Americans after 26 attempts dating back to 1964.
Team USA had been impregnable since the 2006 World Championships, where they suffered a stunning loss to Greece in the semi-finals.
The Americans recovered to win the bronze medal game in that tournament, and then went unbeaten through all Olympics and major world tournaments since.
Then, on Saturday, they ran into Mills and a determined Australian outfit laden with their own NBA talents and desperate to land a blow before the FIBA World Cup in China.
Australia rallied from 10 points down at half-time to secure a famous win.
"They played well, that's a really good team over there," USA guard Kemba Walker said.
"We beat them the first night, they came out and wanted it more than us tonight. Lesson learned for us."
Though Australia lost Game One 102-86 at the same venue on Thursday, Mills had caused all sorts of headaches for USA coach Gregg Popovic, also his boss at the San Antonio Spurs.
He went up another level in Game Two, and could hardly miss in the fourth quarter as he tipped in 13 points and dominated the final minutes.
Although many of the United States' top NBA stars skipped the series and the World Cup, Australia's win thrilled the 52,079 fans - a record crowd for the sport at the stadium - and gave the Boomers hope of claiming a medal in China.
"It was awesome, we had no idea what it was going to be like and everyone came out in full force tonight," said Mills.
"It's a building block for us... This is just one step towards our progress and ultimate goal of creating history and winning a medal, so we're locked in on that focus, for sure."
The result was also a boost for organisers, who were under fire on Friday after hundreds of fans complained about seating arrangements during the opener the night before.
Australia, who finished fourth at the Rio Olympics, face Canada, Senegal and Lithuania at the World Cup which starts on 31 August.
The USA meet Canada in a third World Cup warm-up in Sydney on Monday.