Ji Youn Sung

[Dissemination of IMP-1 and OXA type beta-lactamase in carbapenem-resistant Acinetobacter baumannii].

BACKGROUND Acinetobacter baumannii is an aerobic, gram-negative, glucose-nonfermenting bacterium, which has emerged as a serious opportunistic pathogen. In recent years, the increasing instance of carbapenem-resistant A. baumannii producing metallo-beta-lactamases (MBLs) or OXAtype beta-lactamases is causing a serious clinical problem. In this study, we investigated the prevalence of Ambler class A, B, and D beta-lactamases and their extended-spectrum derivatives in carbapenem-resistant A. baumannii isolates. METHODS A total of 31 consecutive, non-duplicate, carbapenem-resistant A. baumannii were isolated from three university hospitals in the Chungcheong province of Korea. The modified Hodge and inhibitor-potentiated disk diffusion tests were conducted for the screening of carbapenemase and MBL production, respectively. PCR and DNA sequencing were performed for the detection of beta-lactamase genes. We also employed the enterobacterial repetitive intergenic consensus (ERIC)-PCR method for the epidemiologic study. RESULTS Twenty-three of 31 isolates harbored bla(OXA-2)(51.6%), bla(OXA-23)(22.6%), bla(IMP-1)(48.4%),and bla(VIM-2)(3.2%). All of the OXA-2-producing strains also evidenced MBLs. The strains that harbored bla(OXA-23)were isolated only in hospital C, and only in a limited fashion. The ERIC-PCR pattern of the five OXA-23 strains indicated that the isolates were closely related in terms of clonality. The six strains producing IMP-1 isolated from hospital A were confirmed to be identical strains. CONCLUSIONS A. baumannii strains harboring IMP-1 or OXA-type beta-lactamases are currently widely distributed throughout the Chungcheong province of Korea. The most notable finding in this study was that a bla(OXA-2)-producing A. baumannii harboring MBL, which has not been previously reported, can also lead to outbreaks.

Evaluation of Two Automated Instruments for Pre-transfusion Testing: AutoVue Innova and Techno TwinStation

BACKGROUND Despite the advances in total laboratory automation, a considerable amount of work in blood banks is still done using outdated manual methods. Some automated pre-transfusion testing instruments have recently been developed. Of these, we evaluated and compared the AutoVue Innova (Ortho, USA) and the Techno TwinStation (DiaMed AG, Switzerland). METHODS Forward and reverse ABO/Rh typing and unexpected antibody screening and identification tests were performed on 4,628 samples using the manual method and the two automated instruments. Two different anticoagulants (EDTA and citrate) were compared in ABO/Rh typing and unexpected antibody screening tests. Titrating studies were conducted on the following 7 dilutions using 5 samples of irregular antibodies with anti-E, anti-E & -c, anti-D, and anti-Le(a) with anti-Fy(a): 1:2, 1:4, 1:8, 1:16, 1:32, 1:64, and 1:128. The test throughput per hour, the time required to perform 1 and 100 tests, and a simulation test for total events occurring in 1 day were also measured. RESULTS No erroneous results were reported between the two instruments and the manual method. Discrepancies observed in 10 cases (0.4%) of ABO/Rh typing were of higher intensity with AutoVue Innova than with the manual method. AutoVue Innova exhibited the highest sensitivity in the titrating study and throughput performance compared with the manual method and the Techno TwinStation. Especially in the throughput and time required to complete 100 antibody screening tests, AutoVue Innova had a 3.3- and 3.5-fold higher performance, respectively, than Techno TwinStation. CONCLUSIONS Because both of the two fully automated instruments (AutoVue Innova and Techno TwinStation) had high levels of accuracy and performance, it is expected that use of fully automated instruments will reduce human labor, turnaround time, and operator error in the blood bank.

Evaluation of the Vitek 2 Korean Antimicrobial Susceptibility Testing Cards AST N056 and AST N055

Background: The recently issued Korean version of antimicrobial susceptibility cards for Vitek 2 system uses an adjusted antimicrobial combination that reflects Korean clinical practice and CLSI guidelines. We evaluated the two Korean antimicrobial susceptibility testing cards for gram negative rods, AST N056 and AST N055. Methods: The results of susceptibility tests were compared between the original and Korean cards. A number of the same antimicrobials included in the both cards were 15 in AST N041-AST N056 and 17 in AST N022-AST N055. Susceptibilities to the newly added antimicrobials, aztreonam, tobramycin, and meropenem for AST N056; and cefotaxime, levofloxacin, and minocycline for AST N055 were compared with those obtained by disc diffusion test and, in case of discrepancy, by confirmative Etest or broth dilution method. Results: In comparison between AST N041 and AST N056 cards, the average discrepancy rate per strain was 0.34, minor error was 88.2%, and major error and very major error were both 5.9%. In comparison between AST N022 and AST055 cards, the average discrepancy rate per strain and very major error were 1.23 and 4.4%, respectively. The three antimicrobial agents added into AST N055 card showed highly discrepant results as a total of 49 items (44.1%) in 111 isolates were discrepant with very major error of 5.9% and major error of 2.0%. Conclusion: AST N056 showed acceptable results in most items including the newly added antimicrobial agents. However, in the case of AST N055 card that showed a relatively high discrepancy, other indicator antibiotics should be referred to for newly added three antimicrobials. For the antibiotics that showed a high discrepancy between the original and Korean cards, a comparison study should be performed using the standard method and clinical isolates collected in Korea. (Korean J Clin Microbiol 2008;11:2328)

Characteristics of the Molecular Epidemiology of CTX-M-Producing Escherichia coli Isolated from a Tertiary Hospital in Daejeon, Korea.

The aims of this study were to characterize the molecular epidemiological profiles of CTX-M-producing uropathogenic Escherichia coli isolates from a tertiary hospital in Daejeon, Korea, and to investigate the genetic diversity and compare the prevalence of sequence types (STs) in different areas. Extended spectrum β-lactamase-producing E. coli strains isolated from urine were analyzed for CTX-M, integrons, and insertion sequence common regions (ISCRs) by PCR and sequencing. Multilocus sequence typing (MLST), pulsed-field gel electrophoresis (PFGE), phylogenetic analysis, and rep-PCR were also used for molecular typing of the isolates. Of 80 CTX-M producers, 31 and 46 expressed CTX-M-15 and CTX-M-14, respectively. MLST analysis indicated that the most prevalent ST was ST131 (n = 34, 42.5%), followed by ST38 (n = 22, 27.5%), ST405 (n = 8, 10.0%), and ST69 (n = 6, 7.5%). Most CTX-M producers harbored class 1 integrons. ST131 strains belonged to phylogenetic group B2 and showed identical rep-PCR patterns, whereas ST69, ST38, and ST405 strains belonged to phylogenetic group D; the ST38 and ST405 strains displayed the same rep-PCR pattern, respectively. ST131 and ST38 isolates showed 21 and 19 distinct types, respectively, by PFGE. In Daejeon, D-ST38 CTX-M-14 producers were relatively more prevalent than in other countries and Korean cities. Our results indicate that CTX-M-producing E. coli isolates belonged mostly to ST131 or ST38 and were more related to hospital-onset than to community-onset infections and that the blaCTX-M gene may vary according to the ST.

Characterization of Class 1 Integrons in Metallo-β- lactamase-producing Pseudomonas aeruginosa

Background: The genes of metallo-β-lactamase (MBL), a powerful carbapenemase, are carried as a part of the mobile gene cassettes inserted into integrons playing an important role in rapid dissemination of antibiotic resistance genes among bacterial isolates. In this study, we investigated carbapenemase genes and class 1 integrons integrated into the gene cassettes in imipenem-non susceptible P. aeruginosa. Methods: From July 2006 to March 2008, 81 consecutive, non-duplicate, imipenem-non susceptible P. aeruginosa were isolated at Chungnam National University Hospital in Chungcheong province of Korea. The modified Hodge and double disk synergy tests were conducted for the screening of carbapenemase and MBL production, respectively, and PCR and DNA sequencing were performed for the detection of carbapenemase genes and class 1 integron gene cassettes. We also employed the repetitive element sequence-based (Rep)-PCR method for an epidemiologic study.

Characteristics of Acquired β-lactamase Gene in Clinical Isolates of Multidrug-resistant Pseudomonas aeruginosa

Background: Recently, there have been reports of infections with multidrug-resistant Pseudomonas aeruginosa. To determine the mechanism of the resistance, we investigated the prevalence of Ambler class A and D β-lactamases, their extended-spectrum derivatives, and class B and D carbapenemase in multidrug-resistant P. aeruginosa isolates. Methods: During the period of March 2006 to May 2007, clinical isolates of multidrug-resistant P. aeruginosa were collected from patients in Chungnam National University Hospital, Daejeon, Korea. Inhibitor-potentiated disk diffusion tests were used for the screening of metallo-β-lactamase (MBL) production. PCR and DNA sequencing were conducted for the detection of β-lactamase genes. We also employed the enterobacterial repetitive intergenic consensus (ERIC)PCR method for an epidemiologic study. Results: A total of 37 consecutive, non-duplicate, multidrug-resistant P. aeruginosa were isolated. Twentynine of 37 isolates harbored blaOXA-10 (56.8%), blaOXA-2 (18.9%), and blaOXA-1 (5.4%). Only one isolate produced IMP-1, and it also harbored blaOXA-1. None harbored Ambler class A β-lactamase or class D carbapenemase. The strains producing OXA type β-lactamases showed a significantly higher resistance to aminoglycoside compared to non-producers. The ERIC-PCR pattern of the 19 OXA-10 producing strains indicated that the isolates were closely related in terms of clonality. Conclusion: OXA type β-lactamases are the most prevalent among the acquired β-lactamases produced by multidrug-resistant P. aeruginosa isolated at a university hospital in Chungcheong Province. Besides β-lactam antibiotics, the strains harboring OXA type β-lactamase showed a significantly higher resistance to aminoglycoside and qunolone. (Korean J Clin Microbiol 2008;11:98-106)

Persistence of Multidrug-Resistant Acinetobacter baumannii Isolates Harboring blaOXA-23 and bap for 5 Years.

The emergence and dissemination of carbapenemase-producing Acinetobacter baumannii isolates have been reported worldwide, and A. baumannii isolates harboring blaOXA-23 are often resistant to various antimicrobial agents. Antimicrobial resistance can be particularly strong for biofilm-forming A. baumannii isolates. We investigated the genetic basis for carbapenem resistance and biofilm-forming ability of multidrug-resistant (MDR) clinical isolates. Ninety-two MDR A. baumannii isolates were collected from one university hospital located in the Chungcheong area of Korea over a 5-year period. Multiplex PCR and DNA sequencing were performed to characterize carbapenemase and bap genes. Clonal characteristics were analyzed using REP-PCR. In addition, imaging and quantification of biofilms were performed using a crystal violet assay. All 92 MDR A. baumannii isolates involved in our study contained the blaOXA-23 and bap genes. The average absorbance of biomass in Bap-producing strains was much greater than that in non-Bap-producing strains. In our study, only three REP-PCR types were found, and the isolates showing type A or type B were found more than 60 times among unique patients during the 5 years of surveillance. These results suggest that the isolates have persisted and colonized for 5 years, and biofilm formation ability has been responsible for their persistence and colonization.

Molecular Characterization of Salmonella Genomic Island 1 in Proteus mirabilis Isolates from Chungcheong Province, Korea

The emergence and dissemination of Salmonella genomic island 1 (SGI1) are strongly associated with the occurrence of multidrug-resistant (MDR) enterobacteria in humans and animals. Diverse SGI1s have been reported among Salmonella enterica and Proteus mirabilis in several countries. We aimed to characterize SGI1 in P. mirabilis isolates from humans and chickens in Chungcheong Province, Korea. A total of 44 P. mirabilis isolates were recovered from humans (n = 20) and chickens (n = 24). Antimicrobial susceptibility was determined by disk diffusion assay. To detect and characterize SGI1s, PCR amplification and PCR mapping experiments were performed. Repetitive extragenic palindromic-PCR (REP-PCR) was performed to assess the clonality of the isolates. The four P. mirabilis strains (16.7%) from chicken harbored a SGI1, whereas none of the isolates from clinical specimens contained SGI1. The SGI1s detected in our study were all confirmed as SGI1-PmABB harboring aminoglycoside-resistant genes (aacCA5 and aadA7). In P. mirabilis isolates, the presence of SGI1-PmABB was significantly correlated with high resistance rates of the isolates to antimicrobial agents, such as gentamicin, streptomycin, and spectinomycin. Moreover, the four SGI1-bearing isolates showed the same REP-PCR patterns and that suggested both horizontal and clonal spread of the isolates. This study is the first attempt to determine SGI1s in P. mirabilis isolates in Korea. We confirmed that P. mirabilis isolates carrying SGI1-PmABB were distributed at poultry farms in Korea. The present study emphasizes the need for continuous monitoring of SGI1s to prevent spreading of the MDR genomic islands among P. mirabilis isolates from humans and animals.