Staphylococcal species are a leading cause of nosocomial infections and a variety of infections and toxin mediated syndromes that affect patients of all ages. The glycopeptides (GP) vancomycin (Vm) and teicoplanin (Tco) are the most reliable alternatives for treatment of methicillin-resistant staphylococci, which are resistant to beta-lactams and a wide spectrum of antibiotics. Therefore, the emergence of clinical staphylococcal isolates with decreased susceptibility, heteroresistance, and true resistance to Gps has been alarming. The investigators' studies are aimed at identifying Vm and Tco resistance mechanisms in staphylococci. Resistance is likely to be complex and multifactorial and, thus, require a multifaceted investigative approach. In this regard, they propose to characterize a transposon (Tn) mutant of a Vm-resistant laboratory derived Staphylococcus aureus isolate and to perform Tn mutagenesis on other clinical Vm- and Tco-resistant S. aureus isolates and on a coagulase negative Vm-resistant S. haemolyticus isolate. They also plan to screen plasmid libraries prepared from selected glycopeptide- resistant clinical and laboratory isolate(s) for inserts capable of conferring Vm-resistance in Vm-susceptible isolates to determine the role of the plasmid insert in mediating resistance. They also propose to identify glycopeptide resistance determinants by global, random comparisons of genomic DNA and mRNA from resistant and susceptible isolates by restriction landmark genomic scanning and mRNA differential display aimed at comparing GP-resistant and -susceptible S. aureus isolates. The decision to undertake these will be based in part on what was learned with Tn mutagenesis and screening of the plasmid libraries. They will also determine whether there is a role for PBP2, found in increased abundance in many clinical and laboratory derived Vm-resistant isolates, by inactivating the pbpB gene and evaluating its effect on resistance and heteroresistance in staphylococci and employ an avian model of S. aureus bacteremia to begin to clarify some important issues regarding GP-resistance in S. aureus. These studies should lead to an understanding of the mechanisms by which staphylococci resist Gps and may provide novel protein targets against which antimicrobial agents could be designed to treat staphylococcal infections caused by GP resistant isolates.
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