Abstract Aim and Background: This study aimed at studying the role of incarceration length and HIV/AIDS on severity of psychopathological symptoms in substance abusers. Methods and Materials: In a cross sectional-retrospective study, 43 substance abusers with HIV/AIDS and 49 ones without HIV/AIDS were selected by available and consecutive sampling methods that were under Methadone Maintenance Therapy (MMT) in counseling clinics of behavioral diseases and addiction abandonment. All of them were evaluated respecting matched confounding variables via symptom check list-90-Revised (SCL-90-R). Results were analyzed by chi-square (χ 2 ), independent t-test, two-variable regression, multivariate analysis of covariance (MANCOVA). Findings: Although the average of incarceration length was higher in substance abusers with HIV/AIDS, the prediction equations of psychological distress severity based on incarceration length indicated the significance and strength of this relationship in substance abusers without HIV/AIDS (P=0.004). However, this relationship was not significant in substance abusers with HIV/AIDS (P=0.061). The effect of history of incarceration and HIV/AIDS was significant on psychological distress severity in both groups (P=0.001, P=0.049, respectively). The interactive effect of each of these factors was not significant on psychological distress severity. While it was significant on depression levels and positive symptoms total (PST). Conclusions: The higher incarceration length has more detrimental role on psychological distress severity in substance abusers without HIV/AIDS. Having both a history of incarceration and HIV/AIDS endangers the mental health of substance abusers. Meanwhile, although substance abusers with HIV/AIDS without incarceration history had higher levels of depression, substance abusers with HIV/AIDS with history of incarceration suffered more from number or higher range of psychological symptoms.