It will be the final match Socceroos' coach Ange Postecoglou will have to give any fringe players an opportunity to push their claims for his squad at the tournament in Brazil before they re-assemble in May ahead of the June 12-July 13 showpiece.
"Were excited to have secured the match against Ecuador, which offers us a good test against South American opposition leading into the World Cup," Postecoglou said in a FFA statement on Monday.
"This is also the last match we will play before we get together in May for our final World Cup preparations so its important that we can build on the first positive steps we took in the match against Costa Rica late last year."
The clash will be the first between the two teams and will be at English Championship (second division) side Millwall's 'The Den' stadium in southeast London on March 5.
Australia are in Group B at the World Cup with defending champions Spain, the Netherlands and Chile, while Ecuador are in Group E with Switzerland, France and Honduras.
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- Ange Postecoglou