To explore the clinical features and imaging features of pyogenic vertebral osteomyelitis (PVO) caused by Staphylococcus, the clinical characteristics, laboratory examinations and imaging examinations of 20 cases of PVO caused by Staphylococcus admitted to Department of Infectious Diseases, Shanghai Sixth People’s Hospital, Shanghai Jiao Tong University from January 2010 to December 2019 were retrospectively analyzed. The results showed that 20 patients were mainly infected with Staphylococcus aureus, 85% occurred in the lumbar spine, 12 patients developed paravertebral abscess, 13 patients (65%) had fever, and 50% of the patients had elevated white blood cell counts. C­reactive protein, erythrocyte sedimentation rate, and ferritin elevated in all patients. Computed tomography (CT) plain scan showed that the infected vertebral bodies had bone destruction. In magnetic resonance imaging (MRI) vertebral bodies and intervertebral disc destruction area had abnormal signal focus. Emission computed tomography (ECT) showed uneven radioactive uptake of the diseased vertebral bodies. Over 80% of Staphylococcus aureus was not sensitive to penicillin. Twelve patients with paravertebral abscess recovered by CT ­guided drainage combined with sensitive antibiotic treatment. It is concluded that Staphylococcus aureus is the main pathogenic bacterium of pyogenic vertebral osteomyelitis caused by Staphylococcus, which is generally resistant to penicillin. Imaging examination is of great value in the diagnosis of this disease. The inflammatory indicators and MRI could be used in therapeutic effect evaluation and conservative treatment is effective.