Prominent landmarks around Australia are being lit up in rainbow colours and the red, white and blue of the American flag.
In Melbourne, about 300 people gathered for a vigil where they bowed their heads and observed 50 seconds of silence.
World leaders, including the US President, have emphasised the massacre in Orlando was both an act of terror and an act of hate.
Australia's political leaders, Malcolm Turnbull and Bill Shorten, both recognised that gay people were deliberately targeted.
(Turnbull) "It was clearly directed by a murderous hatred of gay people."
(Shorten) "I particularly want to extend my sympathy to people in the LGBTI community who might be feeling additional pain."
At the Melbourne vigil, Michael Barnett was among those who bowed his head for 50 seconds - one second for each of those murdered.
"The pain of what's happened, it's a long way from Melbourne but it seems very close. As part of the community, feeling so many people have been killed so needlessly, so recklessly. I feel connected, it could have been me anywhere in the world that this happened to, my friends or my family."
This woman had been planning to go to work but decided she had to attend the vigil instead.
"And I just feel incredibly sad about what's happened and I felt like I needed to come together with my community, in some way to express that, so when I heard about the vigil I was meant to be going into work and I thought no, I need to come here and just spend some time with my community.
An American-born man addressed the vigil, extending a challenge to those who don't identify as heterosexual.
"And so today my challenge to all of you is have that patience that wasn't afforded (to) us; have that compassion that we have so seldom been shown; show people that we are better than how they portray us; we are better than how they perceive us; that we are loved. Our community is the embodiment of love and respect and hopefully, hopefully in the future these sorts of things won't happen again."
