International research assessing differences in the prevalence of alcohol and other drug (AOD) use among Lesbian Gay Bisexual and Transgender (LGBTI) and heterosexual populations shows elevated prevalence rates of substance use among LGBTI people. To date no research has been published investigating these differences at a population level among both men and women in Australia.The 2013 National Drug Strategy Household Survey, a multistage stratified population sample collecting data on AOD use in the Australian population over 14 years of age, was analysed for differences between gay and bisexual (GB) men and lesbian/gay and bisexual (LGB) women and their heterosexual counterparts in: (1) the prevalence of lifetime and past year tobacco and AOD use; (2) age of initiation of tobacco and AOD use; and (3) frequency of alcohol and cannabis use, and history of AOD treatment.There were elevated rates of past year cannabis (22.4%), ecstasy (11.8%) and methamphetamine (9.7%) use among GB men compared to heterosexual men (12.4%, 2.9% and 2.5%). LGB women also reported elevated rates of past year use (tobacco - 23.7%; cannabis - 24.6%) compared to heterosexual women (10.6% and 7.1%). LGB women initiated tobacco (15.2 years) and alcohol (15.5 years) at an earlier age than heterosexual women (16.6 and 17.7 years), and were significantly more likely to report daily alcohol consumption (OR 3.2, 95% CI: 2.1, 5.1), and weekly or more frequent cannabis use (OR 1.7, 95%CI: 1.1, 3.1).These findings are indicative of the need for more responsive and targeted AOD harm reduction and treatment services for LGBTI communities in Australia. Of concern is the elevated risk among LGB women for earlier initiation of substance use, and the development of problematic consumption patterns. Further research, investigating the risk and protective factors for AOD use among LGB women is warranted.