Background: Recent antiretroviral regimens are potent and better tolerated, resulting in a low prevalence of treatment failure. It is important to identify the drivers of virological failure, so that patients at risk can be identified early and prevention strategies implemented. Methods: We performed a retrospective case?control study of HIV-positive patients on antiretroviral therapy and managed at The Alfred Hospital during 2010 to evaluate the predictors of virological failure. Controls were matched 3 : 1 to cases by gender, and by clinical review in the same week as the diagnosis of virological failure in the case. Predictors of virological failure were identified by multivariate conditional logistic regression. Results: Thirty-nine patients were identified with treatment failure. In the multivariate model, characteristics associated with virological failure were missed clinic appointments in 1 year before virological failure (odds ratio (OR) = 13.1, 95 confidence interval (CI): 2.8?61.1), multiple previous combined antiretroviral therapy regimens (OR = 4.2, 95 CI:1.2?15.3), current hepatitis C infection (OR = 8.6, 95 CI: 1.9?38.7), older age at HIV diagnosis (OR = 1.1, 95 CI: 1.0?1.2), younger age at time of virological failure (OR = 0.9, 95 CI: 0.8 to 1.0), and CD4 cell count at virological failure (OR = 0.7, 95 CI: 0.5 to 0.9). Conclusions: Targeted and appropriate adherence support should be provided to treatment-experienced patients, particularly those who have missed clinical appointments and those with hepatitis C coinfection. Further elucidation of the barriers to clinic attendance may optimise linkage and retention in care.