Pyomyositis is a severe bacterial infection of skeletal muscle, commonly affecting 35 children in tropical regions and predominantly caused by Staphylococcus aureus. To 36 understand the contribution of bacterial genomic factors to pyomyositis, we conducted a 37 genome-wide association study of S. aureus cultured from 101 children with pyomyositis and 38 417 children with asymptomatic nasal carriage attending the Angkor Hospital for Children in 39 Cambodia. We found a strong relationship between bacterial genetic variation and pyomyositis, 40 with estimated heritability 63.8% (95% CI 49.2-78.4%). The presence of the Panton-Valentine 41 leucocidin (PVL) locus increased the odds of pyomyositis 130-fold (p=10 -17.9 ). The signal of 42 association mapped both to the PVL-coding sequence and the sequence immediately upstream. 43 Together these regions explained >99.9% of heritability. Our results establish staphylococcal 44 pyomyositis, like tetanus and diphtheria, as critically dependent on expression of a single toxin 45 and demonstrate the potential for association studies to identify specific bacterial genes 46 promoting severe human disease. 47 48 49