Characterization of a nosocomial outbreak caused by a multiresistant Acinetobacter baumannii strain with a carbapenem-hydrolyzing enzyme: high-level carbapenem resistance in A. baumannii is not due solely to the presence of beta-lactamases

Abstract

From February to November 1997, 29 inpatients at Ramón y Cajal Hospital, Madrid, Spain, were determined to be either colonized or infected with imipenem- and meropenem-resistant Acinetobacter baumannii (IMRAB) strains (MICs, 128 to 256 microg/ml). A wide antibiotic multiresistance profile was observed with IMRAB strains. For typing IMRAB isolates, pulsed-field gel electrophoresis was used. For comparative purposes, 30 imipenem- and meropenem-susceptible A. baumannii (IMSAB) strains isolated before, during, and after the outbreak were included in this study. The molecular-typing results showed that the outbreak was caused by a single IMRAB strain (genotype A). By cloning experiments we identified a class D beta-lactamase (OXA-24) encoded in the chromosomal DNA of this IMRAB strain which showed carbapenem hydrolysis. Moreover, the outer membrane profile of the IMRAB strain showed a reduction in the expression of two porins at 22 and 33 kDa when compared with genetically related IMSAB isolates. In addition no efflux mechanisms were identified in the IMRAB strains. In summary, we report here the molecular characterization of a nosocomial outbreak caused by one multiresistant A. baumannii epidemic strain that harbors a carbapenem-hydrolyzing enzyme. Although alterations in the penicillin-binding proteins cannot be ruled out, the reduction in the expression of two porins and the presence of this OXA-derived beta-lactamase are involved in the carbapenem resistance of the epidemic nosocomial IMRAB strain.

FIG. 1

Patterns obtained by PFGE with SmaI. Lanes 1, 10, 14, and 19, low-range PFG DNA marker (New England Biolabs, United Kingdom). Lanes 2 through 9, IMRAB strains 1, 3, 6, 8, 24, 25, 26, and 28, respectively, in Table 1. Lanes 11 through 13, IMSAB PRE strains 31 to 33, respectively; lanes 15 through 18, IMSAB AT strains 34 through 37, respectively; lanes 20 through 22, IMSAB POST strains 38 through 40, respectively.

FIG. 2

Patterns obtained by REP-PCR. The numbers above the figure indicate the corresponding strains (Table 1). Lanes λ_III and λ_V, DNA molecular weight markers III and V, respectively (Boehringer GmbH, Mannheim, Germany).

FIG. 3

(A) OMP profile of the IMSAB strain 34 (Table 1) (lane 1) and IMRAB strain RYC 52763/97 (lane 2). SDS—12.5% polyacrylamide gel electrophoresis and silver staining. Arrows, the different expression of porins at 22 and 33 kDa. (B) Densitometric analysis of the gel shown in panel A. 1, IMSAB protein pattern; 2, IMRAB protein pattern. Arrows indicate the different expression of porins at 22 and 33 kDa.

FIG. 4

Absence of the OXA-24 gene in different carbapenem-susceptible IMSAB strains by PCR amplification with P1 and P2 OXA primers. Lanes 1 and 11, molecular weight marker λ_III (Boehringer); lane 2, A. baumannii RYC 52763/97 as the positive control; lanes 3 and 4, two epidemic genetically related IMRAB strains; lanes 5 through 10, different genetically unrelated carbapenem-susceptible IMSAB strains.