Admiral Harry Harris, head of the US Pacific Command, warned Washington would not accept Chinese control of the region, despite Beijing's rapid development of artificial islands capable of hosting military planes.
He remarks come as Beijing reacted angrily after Trump, the US President-elect, broke convention by speaking directly to Taiwanese leader Tsai Ing-wen -- and suggested Washington could jettison Washington's decades-old "One China policy".
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"We will not allow the shared domains to be closed down unilaterally, no matter how many bases are built on artificial features in the South China Sea," Harris said.
"We will cooperate where we can but we will be ready to confront where we must."
China insists on sovereignty over virtually all the resource-endowed South China Sea, despite rival claims from its Southeast Asian neighbours.
But Washington has repeatedly said it does not recognise the claims, and has regularly sent warships into the strategically vital area to assert the right to freedom of navigation.
"Should others (countries) signal in this way in freedom of navigation operations? I think so, but that is again up to each individual country to make that decision," the admiral told Australian think tank the Lowy Institute in Sydney.
"The US fought its first war following our independence to ensure freedom of navigation, and we did that when we were weak and small," he added.
"This is an enduring principle, and one of the reasons our forces remain ready to fight tonight."
Harris added that Washington would not make Australia choose between its traditional ally, the United States, and rising world power China.
"Australia is more than capable of chewing gum and walking at the same time, and so we're not asking you to make a choice," he said.
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