Outer membrane profiles of clonally related Klebsiella pneumoniae isolates from clinical samples and activities of cephalosporins and carbapenems

Abstract

Fifteen isolates of Klebsiella pneumoniae producing extended-spectrum beta-lactamases (ESBLs) isolated during a nosocomial outbreak were studied. The strains belonged to the same clonal type, as shown by pulsed-field gel electrophoretic analysis of chromosomal DNA. All the isolates were resistant to extended-spectrum cephalosporins, aztreonam, gentamicin, and fluoroquinolones and were susceptible to carbapenems, tobramycin, netilmicin, and amikacin. None of the isolates expressed the OmpK36 porin. Eight isolates, for which the MICs of cefoxitin were > or = 64 micrograms/ml, showed a diminished level or no expression of a 35-kDa porin. The MICs of meropenem, cefotaxime, and cefpirome were three to eight times higher for porin-deficient isolates than for isolates expressing the 35-kDa porin, but the MICs of imipenem increased two times for porin-deficient isolates compared to those for isolates expressing the porin. This MIC increase reverted to a level similar to that for the parental strain when porin-deficient isolates were transformed with the gene coding for the K. pneumoniae porin OmpK36. It is concluded that the high level of resistance to cefoxitin and the increase in the MICs of meropenem, cefotaxime, and cefpirome for the ESBL-producing K. pneumoniae isolates studied are associated with porin deficiency.

FIG. 1

PFGE of total DNA from K. pneumoniae cut with XbaI. Lane 1, PFGE Molecular Weight Marker (New England Biolabs); Lanes 2 and 3 and lanes 6 and 7, porin-sufficient isolates; lanes 4 and 5, strains with diminished expression of porin; lanes 8 to 10, porin-deficient isolates; lanes 11 and 12, lanes 13 and 14, and lanes 15 and 16, pairs of porin-deficient and porin-sufficient isolates (in the respective pairs of lanes) from three patients.

FIG. 2

SDS-PAGE analysis of outer membrane proteins of K. pneumoniae isolates. Lane MW, molecular weight standard (in kilodaltons); lanes 1, 3, 6, and 7, porin-expressing isolates; lanes 4 and 5, isolates with diminished levels of porin expression; lane 2, porin-deficient isolate.

FIG. 3

SDS-PAGE analysis of outer membrane proteins of K. pneumoniae isolates. Lane MW, molecular mass standard (in kilodaltons); lanes 1 and 2, lanes 3 and 4, and lanes 5 and 6, porin-sufficient and porin-deficient isolates (in the respective pairs of lanes) from the same patients; lanes 7 and 8, porin-deficient isolates.

FIG. 4

SDS-PAGE analysis of OMPs and porins from strains CSUB10S (lanes 1 and 3) and CSUB10R (lanes 2 and 4). Lanes 1 and 2, OMP; lanes 3 and 4, porins. Numbers on the left side correspond to the approximate molecular masses of the proteins (in kilodaltons).