The pathophysiology of post-treatment Lyme disease syndrome (PTLDS) may be linked to overactive immunity including aberrant activity of the brain’s resident immune cells, microglia. Here we used [11C]DPA-713 and positron emission tomography to quantify the 18 kDa translocator protein, a marker of activated microglia or reactive astrocytes, in the brains of patients with post-treatment Lyme disease symptoms of any duration compared to healthy controls. Genotyping for the TSPO rs6971 polymorphism was completed, and individuals with the rare, low affinity binding genotype were excluded. Data from eight brain regions demonstrated higher [11C]DPA-713 binding in 12 patients relative to 19 controls. [11C]DPA-713 PET is a promising tool to study cerebral glial activation in PTLDS and its link to cognitive symptoms.