Panton-Valentine leukocidin (PVL) gene (lukS/F-PV)-positive community-acquired methicillin-resistant Staphylococcus aureus (MRSA), particularly the USA300 clone, is a health concern worldwide. Recently, community-acquired MRSA clones have been found to spread and persist in hospital settings. We aimed to investigate the prevalence and molecular epidemiologic features of lukS/F-PV–positive MRSA in Japanese hospitals. A total of 3433 MRSA isolated from nine hospitals in 2011–2015 were assessed. Molecular epidemiologic analysis performed by staphylococcal cassette chromosome mec typing, PVL typing, arginine catabolic mobile element typing, detection of virulence determinants, multilocus sequence typing, pulsed-field gel electrophoresis and antimicrobial susceptibility testing. The detection rate of lukS/F-PV was increased annually from 0.6% (5/817) in 2011 to 3.1% (17/544) in 2015. Molecular epidemiologic analysis for 64 lukS/F-PV–positive MRSA isolates revealed that 42 isolates (65.6%) were the USA300 clone. Resistance rates of levofloxacin and gentamicin among lukS/F-PV–positive isolates increased annually as a result of increased prevalence of the multidrug-resistant USA300 clone. This is the first report on dissemination of the USA300 clone and PVL-positive MRSA in multiple Japanese hospitals. Our data strongly suggest that the USA300 clone may become epidemic in Japanese hospitals.