BACKGROUND: Most of studies on the relationship between drug use and HIV have focused largely on people who inject drugs. Non-injecting drug use is much more common than injecting drug use, and although it can also predispose people to HIV infection, it is not widely explored. We therefore conducted this study to explore the prevalence of HIV and identify risk factors for HIV infection among people who use non-injecting drugs (PWUD) in Cambodia. METHODS: This cross-sectional study was conducted in 2017. The Respondent Driven Sampling method was used to recruit the study participants who were interviewed face-to-face using a structured questionnaire. Blood samples were collected for HIV and syphilis testing. A multivariable logistic regression analysis was conducted to identify risk factors associated with HIV infection. RESULTS: In total, 1367 PWUD were included in this study, whose mean age was 28.0 (SD = 7.7) years. The majority (95.1%) of the participants used methamphetamine. The prevalence of HIV was 5.7, and 35.2% of the identified HIV-positive PWUD were not aware of their status prior to the survey. After adjustment for other covariates, HIV infection remained significantly associated with being in the age group of ≥35 (AOR = 2.34, 95% CI = 1.04-6.11), having lower level of formal education of ≤ 6 years (AOR = 2.26, 95% CI = 1.04-5.15), living on the streets (AOR = 2.82, 95% CI = 1.10-7.23), perception that their HIV risk was higher as compared to that of the general population (AOR = 3.18, 95% CI = 1.27-8.62), having used injecting drugs in lifetime (AOR = 3.8, 95% CI = 1.36-4.56), and having cuts or sores around the genital area in the past 12 months (AOR = 3.42, 95% CI = 1.09-6.33). CONCLUSIONS: The prevalence of HIV among PWUD in this study was more than 10 times higher than the prevalence in the general adult population. The findings reveal a higher vulnerability to HIV infection among specific sub-populations of PWUD, such as those who are homeless, who may benefit from tailored interventions that respond to their specific needs. To enhance HIV case finding, stratification of PWUD to facilitate HIV risk profiling based on socio-economic profiles and drug injection history is recommended.