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Objective: The aim of this study was to examine associations between nicotine, alcohol, and drug use, as well as key demographic variables, with barriers to antiretroviral therapy (ART) adherence among people living with HIV (PLHIV) in Guangxi, China. Method: Cross-sectional survey data were obtained from 2,095 PLHIV in Guangxi, China, among whom 61.2% were male (n = 1283) and 38.8% were female (n = 812). Participants provided information on substance use history and four types of barriers to ART adherence (symptomatic, emotional, forgetful, and time management barriers). Multivariate regression was used to analyze associations between substance use and perceived barriers to adherence. Results: A minority of participants reported current substance use, with 38.1% (n = 798) using nicotine, 30.7% (n = 644) using alcohol, and 6.6% (n = 138) using illicit drugs. A small but significant proportion of participants reported having quit nicotine, alcohol, and drug use following HIV diagnosis (10.0%, 8.5%, and 9.5%, respectively). Multivariate regression indicated that current drug use was associated with symptomatic, emotional, forgetful, and time management barriers to adherence; current alcohol use was associated with symptomatic, forgetful and time management barriers. Age, ethnicity, official residential status, employment status, duration of being on ART, ability to pay medical expenses, and perceived attitudes of healthcare providers were associated with one or more types of barriers. Conclusions: Psychosocial and behavioral health interventions designed to improve ART adherence in China may benefit from incorporating substance use treatment and care for current users. [ABSTRACT FROM AUTHOR]