Clonal spread of both oxyimino-cephalosporin- and cefoxitin-resistant Klebsiella pneumoniae isolates co-producing SHV-2a and DHA-1 β-lactamase at a burns intensive care unit

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Abstract

Over a 1-month period, a total of 16 ceftriaxone- and cefoxitin-resistant Klebsiella pneumoniae isolates were isolated from 15 patients hospitalised at a burns intensive care unit (ICU). These isolates showed negative results for extended-spectrum β-lactamase (ESBL) by the Vitek system and were highly resistant to ceftazidime, aztreonam and cefoxitin (minimum inhibitory concentrations ≥128 μg/mL). The blaSHV-2a and blaDHA-1 genes were detected by polymerase chain reaction and sequence analysis. Pulsed-field gel electrophoresis profiles of the isolates were identical. AmpC disk tests for AmpC enzymes as well as double-disk tests and Clinical and Laboratory Standards Institute (CLSI) confirmatory disk tests for ESBLs yielded positive results for all the isolates. However, only three isolates (18.8%) were shown to produce ESBL by CLSI confirmatory tests using broth microdilution. We report the first outbreak of colonisations and infections due to K. pneumoniae isolates co-producing an SHV-2a ESBL and a DHA-1 AmpC β-lactamase in a Korean hospital, which were suggested to represent a single clonal spread at a burns ICU. In addition, this report presents problems associated with ESBL detection using broth microdilution in isolates that co-produce an ESBL and an AmpC β-lactamase.

Introduction

β-Lactamase production is the predominant mechanism for resistance to β-lactams in Enterobacteriaceae. The majority of β-lactamases that confer resistance to extended-spectrum cephalosporins in Klebsiella pneumoniae are extended-spectrum β-lactamases (ESBLs) of Ambler's molecular class A [1]. β-Lactamases of molecular class C, also termed AmpC β-lactamases, provide a broader spectrum of resistance than ESBLs and have also been identified in K. pneumoniae [2]. Unlike ESBLs, AmpC enzymes are poorly inhibited by β-lactamase inhibitors and are less active against cefepime and cefpirome than ESBL enzymes. AmpC enzymes confer resistance to oxyimino- and 7-α-methoxy-cephalosporins. They occur naturally in the chromosomes of bacteria such as Enterobacter, Citrobacter, Serratia and Pseudomonas spp. [2]. Recently, genes coding for AmpC enzymes have made their way into plasmids and are increasingly detected in bacteria naturally lacking chromosomal AmpC such as Klebsiella spp., Salmonella spp. and Proteus mirabilis [3], [4], [5].

On the basis of the characteristics of these β-lactamases, various phenotypic detection methods have been proposed previously. However, coexistence of different classes of β-lactamases in a single bacterial isolate may pose diagnostic and therapeutic challenges [6], [7], [8].

Over a 1-month period in 2004, 16 K. pneumoniae isolates resistant to ceftriaxone, aztreonam and cefoxitin were isolated from 15 patients hospitalised at a burns intensive care unit (ICU) in Korea. These strains yielded negative ESBL results with the Vitek system (bioMérieux, Hazelwood, MO). The purpose of this study was to investigate the molecular epidemiology of these isolates and also to determine the mechanism of drug resistance.

Section snippets

Bacterial strains and patients

The burns centre is a 200-bed centre including 36 ICU beds and admits ca. 1500 patients per year. Over a 1-month period in 2004, a total of 16 ceftriaxone-, aztreonam- and cefoxitin-resistant isolates of K. pneumoniae were recovered from 15 patients hospitalised in the burns ICU. The ESBL test by the Vitek system was negative for all of the isolates. The first isolate from each source for each patient was included in the study.

Susceptibility testing and β-lactamase phenotype

Susceptibilities were determined by the broth microdilution minimum

Results

Of 16 K. pneumoniae isolates, 8 isolates were recovered from sputum, 3 isolates were from endotracheal tube samples, 3 isolates were from blood and 2 isolates were from burn wounds. Resistance to ceftazidime and cefoxitin of all 16 K. pneumoniae isolates was transferred to the E. coli J53 Azir recipients by conjugation. The isolates showed similar susceptibility profiles, characterised by elevated MICs of piperacillin (MICs > 256 μg/mL), cefoxitin (MICs > 128 μg/mL), ceftazidime (MICs  128 μg/mL) and

Discussion

This study began with the observation of K. pneumoniae isolates showing resistance to ceftriaxone, aztreonam and cefoxitin, but negative for ESBL by the Vitek system. During a 1-month period in 2004, a total of 16 isolates showing an identical antibiogram were isolated. These results strongly suggest that there are AmpC enzymes only or co-existence of ESBL and AmpC in these isolates. SHV-2a ESBL is increasingly described worldwide; particularly SHV-2a and SHV-12 are the most frequently

Acknowledgments

The authors are thankful to Tae-Jae Lee and Il Kwon Bae for molecular testing.

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