Melioidosis is caused by the saprophytic organism Burkholderia pseudomallei. The use of the indirect hemagglutination assay (IHA) has found widespread use in areas endemic for this disease. Using this assay, we explored the serologic profile of 275 patients with culture-confirmed melioidosis in the Northern Territory of Australia. Based on a threshold titer of 1:40, the sensitivity of the IHA on admission was 56%. Female patients, those with positive blood cultures, and those with pneumonia independently predicted a negative IHA result. Most patients (68%) with negative admission IHA titers subsequently seroconverted. Most patients (92%) with positive admission IHA titers had persistently positive IHA titers. Relapses were not observed in 36 patients who had a negative IHA at least 1 month after admission, irrespective of initial admission IHA. The IHA has limited utility as a diagnostic test for acute disease, and most patients subsequently have persistently positive titers after recovery from illness.