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In an open letter, the 40 faith leaders said they feared a plebiscite could create division between faith communities and destabilise the place of religion in society.
"Polls have demonstrated that opinion on legalising same-sex marriage varies among people of faith. In fact most polls find that a majority favour change," the letter reads.
"Yet the negative case will be put by religious groups and leaders who claim to speak on behalf of people of faith generally, or religious institutions as a whole."
The Australian Christian Lobby is one of the lead campaigning groups against same-sex marriage. Earlier this week, it called for anti-discrimination laws to be suspended during the plebiscite, saying the vote would not be fair otherwise.
"A volatile, public and politically-charged debate could both distance leaders from lay people, marginalise faith communities from broader society and alienate LGBTI individuals within religious communities."
They also expressed worry that the plebiscite would cause harm to LGBTI Australians.
"After decades of legalised discrimination, and ongoing social stigma, LGBTI Australians will face an angry, drawn-out debate, one likely to multiply existing disadvantages and stigma," they wrote.
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Most of the signatories are Christians, including Catholic, Anglican and Uniting leaders. Gregg Heathcote, a Buddhist priest from Newcastle, is also on the list.
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