Replicate populations of the social bacterium Myxococcus xanthus underwent extensive evolutionary adaptation to an asocial selective environment (liquid batch culture). All 12 populations showed partial or complete loss of their social (S) motility function after 1,000 generations of evolution. Mutations in the pil gene cluster (responsible for type IV pilus biogenesis and function) were found to be at least partially responsible for the loss of S motility in the majority of evolved lines. Restoration (partial or complete) of S motility in the evolved lines by genetic complementation with wild-type pil genes positively affected their fruiting body development and sporulation while negatively affecting their competitive fitness in the asocial regime. This genetic tradeoff indicates that mutations in the pil region were adaptive in the asocial selective environment. This finding was confirmed by experiments showing that defined deletions of pil gene regions conferred a competitive advantage under asocial conditions. Moreover, an amino acid substitution in an evolved genotype was located in a region predicted by genetic complementation analysis to bear an adaptive mutation.