IDENTIFICATION AND CHARACTERIZATION OF THE PUTP PROLINE PERMEASE THATCONTRIBUTES TO IN-VIVO SURVIVAL OF STAPHYLOCOCCUS-AUREUS IN ANIMAL-MODELS

Staphylococcus aureus is an important pathogen of humans and other ani mals, causing bacteremia, abscesses, endocarditis, and other infectiou s syndromes, A signature-tagged mutagenesis (STM) system was adapted f or use in studying the genes required for in vivo survival of S. aureu s. An STM library was ultimately created in S. aureus RN6390, with Tn9 17 being used to create the transposon mutations, Pools of S. aureus R N6390 mutants were screened in mouse abscess, bacteremia, and wound in fection models for growth attenuation after in vive passage, One of th e mutants that was identified displayed marked attenuation following l arge-pool screening in all three animal models, which was confirmed in bacteremia and endocarditis models of infection with a smaller pool o f mutants, Sequence analysis of the entire open reading frame showed a 99% identity to the high-affinity proline permease (putP) gene charac terized in another strain of S. aureus. In wound and murine abscess in fection models, the putP mutant was approximately 10-fold more attenua ted than was wild-type strain RN6390, Another S. aureus strain transdu ced with the putP mutation also displayed an attenuated phenotype afte r passage in the wound model, A [H-3]proline uptake assay showed that less proline was specifically transported into the putP mutant than in to strain RN6390. The reduced viability of the bacteria possessing the mutation in the S. aureus high-affinity proline permease suggests tha t proline scavenging by the bacteria is important for in vive growth a nd proliferation and that analogs of proline may serve as potential an tistaphylococcal therapeutic agents.