Prevalence and evaluation of detection methods for plasmid-mediated AmpC beta-lactamases among Klebsiella pneumoniae and Escherichia coli in Hong Kong

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Prevalence and evaluation of detection methods for plasmid-mediated AmpC beta-lactamases among Klebsiella pneumoniae and Escherichia coli in Hong Kong


Author: Wong, Wing-kin
Title: Prevalence and evaluation of detection methods for plasmid-mediated AmpC beta-lactamases among Klebsiella pneumoniae and Escherichia coli in Hong Kong
Degree: M.Sc.
Year: 2012
Subject: Beta lactamases.
Escherichia coli.
Klebsiella pneumoniae.
Hong Kong Polytechnic University -- Dissertations
Department: Dept. of Health Technology and Informatics
Pages: xi, 71 leaves : ill. ; 30 cm.
Language: English
Abstract: Plasmid-mediated AmpCβ-lactamase has been reported since 1988. There are over 40 known plasmid-mediated AmpCβ-lactamases identified to date. They are commonly discovered in Klebsiella pneumoniae, Escherichia coli, Proteus mirabilis and Salmonella species. AmpCβ-lactamases confer resistance to a broad spectrum of beta-lactam drugs, but not to 4th generation cephalosporins and carbapenems. Besides, they are not inhibited by the commonly available beta-lactamases inhibitor (clavulanic acid). Moreover AmpCβ-lactamases producing pathogens may show in vitro susceptibility to some cephalosporins but it will not response to that therapy later because of its inducible resistance property. Unfortunately, there is no Clinical Laboratory Standard Institute (CLSI) recommended detection method for AmpCβ-lactamases; the data of worldwide prevalence of plasmid-mediated AmpCβ-lactamases is limited. This study aimed to find the prevalence of AmpCβ-lactamases in Klebsiella pneumoniae and Escherichia coli strains in Hong Kong, to evaluate different methods for detecting AmpCβ-lactamases and to study the associated antimicrobial resistance found in AmpC producing Klebsiella pneumoniae. A total of 881 non-duplicated and consecutive Klebsiella pneumoniae isolates and 1379 Escherichia coli isolates were recognized from blood culture in Princess Margaret Hospital (PMH) between 2004 and 2008. Any isolates showed augmentin zone diameter less than 18 mm which were regarded as screening positive strains. One hundred and sixty eight (19%) of the 881 Klebsiella pneumoniae clinical isolates and one hundred and thirteen (8%) of the 1379 Escherichia coli clinical isolates were recovered. The screening positive isolates were tested by cefoxitin non-susceptibility, disk approximation test, three-dimensional test, AmpC disk test, inhibitor-based test and multiplex PCR. For isolates positive of plasmid-mediated AmpCβ-lactamase, the minimal inhibitory concentrations (MIC) of cefuroxime, ceftriaxone, cefotaxime, ceftazidime, cefepime, imipenem, meropenem, ciprofloxacin and levofloxacin were determined by E tests.
Multiplex PCR was applied to detect six families of plasmid-mediated AmpCβ-lactamases (MOX, CIT, DHA, ACC, EBC, and FOX) and acted as a reference standard for evaluating the phenotypic tests. Plasmid-mediated AmpC genes were found in 52 (5.6%) of 881 K. pneumoniae isolates and 9 (0.65%) of 1376 E. coli isolates. Only DHA group AmpC beta-lactamase was harbored in K. pneumoniae isolates, DHA and CIT group AmpC beta-lactamases were present in 3 and 6 of E. coli isolates, respectively. No enzymes belonging to the ACC, FOX, MOX or EBC family were detected. By comparing the results of phenotypic methods against the genotypic method, the sensitivity of disk approximation test, AmpC disk test, inhibitor-based test and three-dimensional test were 75%, 97%, 95% and 92%, respectively. Their specificity approached 100%. In this study, there were 6.5%, 22.9% and 9.8% of blaDHA positive K. pneumoniae isolates classified as susceptible to cefotaxime, ceftriaxone and ceftazidime, respectively. Clinical failure of using cephalosporins to treat AmpC producing organisms has been reported. Reporting susceptibility to third generation cephalosporins for plasmid-mediated AmpC producing organisms is still controversial. Cefepime and carbapenems are still effective to treat AmpC beta-lactamases producing bacteria, 98.3% and 96.7% of plasmid-mediated blaAmpC positive isolates were susceptible to cefepime and imipenem, respectively. Moreover, around 30% of these isolates were susceptible to fluoroquinolone. In conclusion, overall prevalence of plasmid-mediated AmpC beta-lactamases is 2.7%, DHA type enzyme is the most prevalent type in Hong Kong. AmpC disk test, inhibitor-based test and three-dimensional test show good performance for AmpC beta-lactamase detection. AmpC disk test is the best among these phenotypic methods, it can be recommended in clinical laboratory. A few plasmid-mediated AmpC beta-lactamase producing organisms were falsely classified as susceptible to third generation cephalosporins. It reinforced highlights the need for detection and continuous surveillance of AmpC beta-lactamase in Hong Kong.

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