Shu-Huei Tsai

Prevalence of Plasmid-Mediated Quinolone Resistance Determinants QnrA, QnrB, and QnrS among Clinical Isolates of Enterobacter cloacae in a Taiwanese Hospital

ABSTRACT The prevalence of three plasmid-mediated quinolone resistance determinants, QnrA, QnrB, and QnrS, among 526 nonreplicate clinical isolates of Enterobacter cloacae collected at a Taiwanese university hospital in 2004 was determined by PCR and colony hybridization, and the association of Qnr with the IMP-8 metallo-β-lactamase was investigated. Eighty-six (16.3%) of all isolates were qnr positive, and the qnrA1-like, qnrB2-like, and qnrS1-like genes were detected alone or in combination in 3 (0.6%), 53 (10.1%), and 34 (6.5%) isolates, respectively. Among 149 putative extended-spectrum-β-lactamase-producing isolates, 59 (39.6%) isolates, all of which were SHV-12 producers, harbored qnrA (0.7%; 1 isolate), qnrB (28.9%; 43 isolates), or qnrS (12.1%; 18 isolates). Forty-four (78.6%) of 56 IMP-8 producers carried qnrB (58.9%; 33 isolates), qnrS (25.0%; 14 isolates), or both. PCR and sequence analysis revealed that qnrA1 was located in a complex sul1-type integron that contains dhr15, aadA2, qacEΔ1, sul1, orf513, qnrA1, ampR, and qacEΔ1. Conjugation experiments revealed the coexistence of qnrB and blaIMP-8 on the transferred plasmids and the absence of β-lactamase content on the transferred qnrS-positive plasmids. The transferred blaIMP-8-positive plasmids with and without qnrB had very similar restriction patterns, suggesting the horizontal mobility of qnrB. Pulsed-field gel electrophoresis showed six major patterns among the 44 qnr-positive IMP-8-producing isolates. Thus, the extremely high prevalence of qnr among the metallo-β-lactamase-producing E. cloacae isolates in the hospital may be due mainly to the intrahospital spread of a few clones and the dissemination of plasmids containing both qnrB and blaIMP-8.

Prevalence of SHV-12 among Clinical Isolates of Klebsiella pneumoniae Producing Extended-Spectrum β-Lactamases and Identification of a Novel AmpC Enzyme (CMY-8) in Southern Taiwan

ABSTRACT Twenty (8.5%) of 234 nonrepetitive clinical isolates ofKlebsiella pneumoniae from southern Taiwan were found to produce extended-spectrum β-lactamases (ESBLs): 10 strains produced SHV-12, 4 produced SHV-5, 2 produced a non-TEM non-SHV ESBL with a pI of 8.3, 3 produced a novel AmpC β-lactamase designated CMY-8 with a pI of 8.25, and 1 produced SHV-12 and an unidentified AmpC enzyme with a pI of 8.2. The CMY-8 enzyme confers a resistance phenotype similar to CMY-1 and MOX-1, and sequence comparisons showed high homologies (>95%) of nucleotide and amino acid sequences among these three enzymes. Plasmid and pulse-field gel electrophoresis analyses revealed that all isolates harboring an SHV-derived ESBL were genetically unrelated, indicating that dissemination of resistance plasmids is responsible for the spread of SHV ESBLs among K. pneumoniaein this area. All three isolates carrying CMY-8 had identical genotypic patterns, suggesting the presence of an epidemic strain.

Outbreak of Infection with Multidrug-Resistant Klebsiella pneumoniae Carrying bla IMP-8 in a University Medical Center in Taiwan

ABSTRACT Klebsiella pneumoniae strains with the transferable carbapenem-hydrolyzing metallo-β-lactamases, which include IMP- and VIM-type enzymes, remain extremely rare. To investigate whether IMP- or VIM-producing K. pneumoniaeisolates had spread at a university medical center in Taiwan, a total of 3,458 clinical isolates of K. pneumoniaeconsecutively collected in 1999 and 2000 were tested by the agar diffusion method, colony hybridization, PCR, and nucleotide sequencing. A total of 40 isolates (1.2%), or 17 nonrepetitive isolates, from 16 patients were found to carry blaIMP-8, a metallo-β-lactamase gene recently identified from a K. pneumoniae strain in Taiwan. Carriage ofblaVIM or otherblaIMP genes was detected in none of the remaining isolates. Of the 17 nonrepetitiveblaIMP-8-positive isolates, 15 isolates (88.2%) appeared susceptible to imipenem (MICs, ≤4 μg/ml) and meropenem (MICs, ≤1 μg/ml), indicating the difficulty in detecting blaIMP-8 in K. pneumoniae by routine susceptibility tests; 14 isolates (82.4%) produced SHV-12 as well; and 14 isolates (82.4%) were also resistant to fluoroquinolones. The organisms caused wound infections in eight patients and bloodstream infections in three patients. They were not directly associated with the death of nine patients. Before the recovery of the blaIMP-8-positive isolates, all 16 patients had undergone various surgical procedures, and 15 patients had been admitted to the surgical intensive care unit, suggesting a nosocomial outbreak. Two major patterns were observed by pulsed-field gel electrophoresis for 14 of the 17 nonrepetitive isolates, indicating that the clonal spread was mainly responsible for the outbreak.

Dissemination of blaCMY-2 among Escherichia coli Isolates from Food Animals, Retail Ground Meats, and Humans in Southern Taiwan

ABSTRACT Twenty-six Escherichia coli isolates recovered from food animal feces and retail ground meats and 14 urinary E. coli isolates from outpatients were shown to carry blaCMY-2. Similar CMY-2-encoding plasmids were found among seven human and three ground-pork isolates. These data indicate the community spread of blaCMY-2 in southern Taiwan.

Emergence of Ceftriaxone-Resistant Salmonella Isolates and Rapid Spread of Plasmid-Encoded CMY-2–Like Cephalosporinase, Taiwan

Of 384 Salmonella isolates collected from 1997 to 2000 in a university hospital in Taiwan, six ceftriaxone-resistant isolates of Salmonella enterica serovar Typhimurium were found in two patients in 2000. The resistance determinants were on conjugative plasmids that encoded a CMY-2–like cephalosporinase. During the study period, the proportion of CMY-2–like enzyme producers among Escherichia coli increased rapidly from 0.2% in early 1999 to >4.0% in late 2000. Klebsiella pneumoniae isolates producing a CMY-2–like β-lactamase did not emerge until 2000. The presence of blaCMY-containing plasmids with an identical restriction pattern from Salmonella, E. coli, and K. pneumoniae isolates was found, which suggests interspecies spread and horizontal transfer of the resistance determinant. Various nosocomial and community-acquired infections were associated with the CMY-2–like enzyme producers. Our study suggests that the spread of plasmid-mediated CMY-2–like β-lactamases is an emerging threat to hospitalized patients and the public in Taiwan.

Complexity of Klebsiella pneumoniae Isolates Resistant to Both Cephamycins and Extended-Spectrum Cephalosporins at a Teaching Hospital in Taiwan

ABSTRACT Among 99 clinical Klebsiella pneumoniae isolates resistant to cefoxitin and extended-spectrum cephalosporins, coexistence of AmpC (DHA-1, CMY-2, or CMY-8) and extended-spectrum β-lactamases (CTX-M and/or SHV) was detected in a total of 35. The remainder produced AmpC (n = 42), extended-spectrum β-lactamases (n = 9), metallo-β-lactamases (n = 2), or none of these enzymes (n = 11). Phenotypic characteristics of these isolates were demonstrated.

Characterization of carbapenem-non-susceptible Escherichia coli isolates from a university hospital in Taiwan

OBJECTIVES To investigate characteristics of nine carbapenem-non-susceptible (CP-NS) Escherichia coli isolates collected between 1999 and 2005 at a Taiwanese university hospital. METHODS Genetic relatedness was analysed by PFGE. beta-Lactamases were characterized by PCR and isoelectric focusing. Outer membrane proteins and transcripts were investigated by SDS-PAGE and northern blotting. Cloning experiments were performed to investigate the role of membrane permeability in carbapenem non-susceptibility. RESULTS The nine CP-NS isolates were found to produce the CMY-2 AmpC enzyme (n = 8), the CTX-M-14-type extended-spectrum beta-lactamase (ESBL) (n = 1), the SHV-12 ESBL (n = 1) and the IMP-8-type metallo-beta-lactamase (n = 1) alone or in combination. All CP-NS isolates revealed a decrease in the transcription and protein expression of ompC, and susceptibility to carbapenems was restored in one isolate by introducing the cloned ompC gene. PFGE revealed genetic diversity among the nine isolates. All patients with the CP-NS isolates had been treated with carbapenems (six patients) and/or extended-spectrum cephalosporins (five patients) before isolation. CONCLUSIONS Our study suggests that the decreased susceptibility to carbapenems in E. coli in the hospital might arise by the stepwise accumulations of multiple drug-resistance determinants in different clones.

Cephalosporin and Ciprofloxacin Resistance in Salmonella, Taiwan

We report the prevalence and characteristics of Salmonella strains resistant to ciprofloxacin and extended-spectrum cephalosporins in Taiwan from January to May 2004. All isolates resistant to extended-spectrum cephalosporins carried blaCMY-2, and all ciprofloxacin-resistant Salmonella enterica serotype Choleraesuis isolates were genetically related.

Comparison of the MB/BacT and BACTEC MGIT 960 system for recovery of mycobacteria from clinical specimens☆

A total of 543 specimens were cultured in parallel with the MB/BacT and BACTEC MGIT 960 systems and on the conventional solid media. Mycobacteria were identified from 95 (17.5%) specimens, including 63 (66.3%) Mycobacterium tuberculosis and 32 (33.7%) nontuberculous mycobacteria. The recovery rates for the MB/BacT, MGIT 960, and solid media were 91.6, 87.4, and 54.7%, respectively, for all mycobacteria; the recovery rates were 93.6, 88.9, and 63.4%, respectively, for M. tuberculosis complex alone, and 87.5, 84.4, and 37.5%, respectively, for all nontuberculous mycobacteria. The mean times to detection of all mycobacteria by individual systems were 13. 9, 8.7, 31.7 days for the MB/BacT, MGIT 960 and solid media, respectively, 13.9, 9.3, 32.9 days for M. tuberculosis alone, and 14. 1, 8.1, 27.2 days for all nontuberculous mycobacteria. The contamination rates of the MB/BacT and MGIT 960 were 10.2 and 5.4%, respectively. With regard to detection times and recovery rates, both automated systems are superior to the conventional media (all p < 0.005). As compared to the MB/BacT, the MGIT 960 detected mycobacterial growth more rapidly (p < 0.001), and had a lower contamination rate (p = 0.003); however, there was no statistically significant difference in recovery rates between these two systems. These results indicate that both MGIT 960 and MB/BacT systems are rapid, sensitive, and efficient methods for the recovery of mycobacteria from clinical specimens.

Epidemiological Investigation of Bloodstream Infections by Extended Spectrum Cephalosporin-Resistant Escherichia coli in a Taiwanese Teaching Hospital

ABSTRACT In an epidemiologic and case-control study including 30 case patients over a 3.5-year period in a Taiwanese university hospital, only β-lactamase inhibitor use and extended-spectrum cephalosporin use were identified as independent risk factors for nosocomial CMY-2-producing Escherichia coli bloodstream infection, and CMY-2 producers were found more prevalent than extended-spectrum β-lactamase-producing isolates.