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Substance abuse (SA) and mental illness (MI) commonly co-occur with HIV infection in the United States and have important implications for clinical management of HIV/AIDS. Yet SA/MI often go untreated due in part to a lack of practical, validated screening tools.HIV clinic in academic medical center.The 16-item SA/MI Symptoms Screener (SAMISS) targets SA/MI in HIV-positive patients. Consecutive consenting HIV-positive patients completed the SAMISS and then a reference standard diagnostic tool, SCID, the Structured Clinical Interview for DSM-IV (Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition).Twenty percent of participants (29/148) had an SA diagnosis and 41% (59/143) had an MI diagnosis in the past year on the SCID; 48% (68/143) had 1 or both. Thirty-seven percent (55/148) screened positive for SA and 69% (99/143) screened positive for MI on the SAMISS. The SAMISS had 86% (95% CI: 68%-96%) sensitivity and 75% (66%-82%) specificity for SA and 95% (86%-99%) sensitivity and 49% (38%-60%) specificity for MI. Patients with SA were likely to show up as false positives for MI and vice versa.The SAMISS functioned well as a first-line screening tool for SA/MI in this HIV clinic population. It missed few cases and was easily incorporated into a busy clinical setting. Persons screening positive require a more rigorous confirmatory psychiatric evaluation.