title: Selection of Klebsiella pneumoniae mutants with high-level cefotaxime resistance during growth in serum containing therapeutic concentrations of cefotaxime.
abstract: BACKGROUND
  In a previous investigation of the genetic characterization of extended-spectrum beta-lactamases (ESBLs) in Klebsiella pneumoniae from Zagreb, Croatia, 20 strains were found to produce SHV-2 beta-lactamase. Those strains displayed varying degrees of beta-lactam resistance and a wide range of beta-lactamase activity. We concluded that more resistant isolates were hyperproducers of SHV-2 beta-lactamase.
  
  
  METHODS
  In this investigation, we tried to develop hyperproducing variants from 8 low-level SHV-2 beta-lactamase-producing Klebsiella strains by subculturing them in serum containing therapeutic concentrations of cefotaxime (CTX).
  
  
  RESULTS
  In most cases, there was a moderate increase in CTX resistance (twofold to threefold), except in one strain which displayed a 16-fold increase in the minimum inhibitory concentration (MIC) of CTX after incubation in the serum. That strain showed a marked increase in enzyme activity as well. The strains with a moderate increase in CTX MIC did not produce more enzyme after exposure to the serum, except for one strain which had a threefold rise in beta-lactamase activity after exposure to serum.
  
  
  CONCLUSIONS
  In this investigation, it was established that the mutants with high-level CTX resistance developed very quickly in the biological fluids containing therapeutic concentrations of CTX. It is reasonable to expect that a similar process occurs in patients infected with an ESBL-producing K. pneumoniae strain during antibiotic treatment. Since most of the high-level CTX-resistant mutants did not have a marked rise in beta-lactamase activity after exposure to serum, it is possible that the elevated resistance was due to some other mechanism, such as reduced penicillin-binding protein affinity, changes in outer membrane proteins or efflux by multidrug efflux pumps.
authors.affiliation: Department of Microbiology, A. Stampar School of Public Health, Medical School, University of Zagreb, Croatia. branka.bedenic@zg.hinet.hr
authors.forename: Branka
authors.initials: B
authors.lastname: Bedenić
mesh_terms: D002439:Cefotaxime / Q000494:pharmacology*; D018550:Cephalosporin Resistance; D002511:Cephalosporins / Q000494:pharmacology*; D003470:Culture Media; D007710:Klebsiella Infections / Q000382:microbiology*; D007711:Klebsiella pneumoniae / Q000187:drug effects* / Q000201:enzymology / Q000302:isolation & purification*; D008826:Microbial Sensitivity Tests; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010957:Plasmids; D001618:beta-Lactamases / Q000378:metabolism
chemical_list: D002511:Cephalosporins; D003470:Culture Media; C062760:beta-lactamase SHV-2; D001618:beta-Lactamases; D002439:Cefotaxime