Background <p>International guidelines for <i>Staphylococcus aureus</i> vertebral osteomyelitis recommend 6 weeks of treatment, including oral follow-up using antibiotics with high bioavailability such as a fluoroquinolone/rifampicin combination. Oral flucloxacillin is not recommended due to low bioavailability and scarce evidence. However, flucloxacillin as oral follow-up treatment is common practice in Sweden based on favourable clinical experience, good tolerability, few interactions, and low ecological impact. Our aim was to review a single-centre experience of <i>S. aureus</i> vertebral osteomyelitis, with focus on flucloxacillin treatment.</p> Methods <p>A single-centre retrospective cohort of patients with <i>Staphylococcus aureus</i> vertebral osteomyelitis (<i>n</i> = 40) was identified between 2010 and 2016. Patients were further stratified by antibiotic treatment strategy with focus on oral flucloxacillin therapy (<i>n</i> = 24). Primary outcomes were relapse or death within 12 months of treatment initiation, and antibiotic-related adverse effects during treatment.</p> Results <p>Methicillin-susceptible <i>S. aureus</i> (MSSA) caused 38 of the infections (95%), and five patients (13%) died, all in-hospital. Flucloxacillin was used for at least 75% of the oral treatment duration in 24 patients (60%). Median antibiotic treatment duration among these patients was 125.5 days (IQR 95–182), 109 days (IQR 76–149) of which comprised oral antibiotics. There were two relapses and two deaths among the patients treated predominantly with flucloxacillin, resulting in a composite clinical cure rate of 83% (20 of 24).</p> Conclusions <p>Prolonged oral flucloxacillin administration could be a potential treatment option for MSSA vertebral osteomyelitis. A prospective study of optimal treatment duration and dosing strategies for flucloxacillin in vertebral osteomyelitis is warranted.</p>