Mycobacterium arupense is a slow-growing, nontuberculous mycobacterium widely found in the environment and is known to cause tenosynovitis and osteomyelitis, mainly in the hands and wrists. We present the first case of vertebral osteomyelitis caused by M arupense in a 78-year-old man with renal cell carcinoma. The patient had a history of tuberculous pleuritis in childhood. Although the nucleic acid amplification test of the vertebral tissue for Mycobacterium tuberculosis was negative, we initiated tuberculosis treatment based on the history and pathological findings of auramine-rhodamine-positive organisms and epithelioid cell granulomas. Subsequently, the isolated mycobacterium was identified as M arupense by genome sequencing. Accordingly, the treatment regimen was changed to a combination of clarithromycin, ethambutol, and rifabutin. Owing to a subsequent adverse event, rifabutin was switched to faropenem, and the patient was treated for a total of 1 year. In previous literature, we found 15 reported cases of bone and soft tissue infections caused by M arupense, but none of them had vertebral lesions. Physicians should be aware that M arupense can cause vertebral osteomyelitis mimicking tuberculous spondylitis. In addition, molecular testing of isolated mycobacteria is essential for diagnosis, even if tuberculous spondylitis is suspected.