In the era of new antiretroviral treatments that have dramatically reduced both morbidity and mortality, a primary goal is to maximize function and wellbeing in the everyday life of HIV-infected patients. To be able to do so, it would be important for clinicians and policy makers to identify factors that influence health-related quality of life (HRQoL). The objective of this multicentre prospective cohort study was to identify determinants of HRQoL in a cohort of Italian HIV-infected patients, the majority of whom were taking highly active antiretroviral therapy (HAART). A total of 809 patients were enrolled. The MOS-HIV Health Survey (summarized using two scores, physical health (PHS) and mental health (MHS)), and an HIV-related symptom scale were administered at enrolment and six months later. At baseline, low CD4+ cell count, hospitalization during the three months before the enrollment and symptoms were independently related to poor PHS; hospitalization during the three months before the enrollment, symptoms and poor satisfaction with information from providers were independently related to MHS. Predictors of PHS at six months included the stage of HIV infection, baseline CD4+ cells count, PHS and symptom score; while age, baseline MHS, symptom score and education predicted six-month MHS. Among these factors, symptoms, recent hospitalization and satisfaction with information are most amenable to clinical intervention.