Yunsop Chong

Evaluation of the Hodge Test and the Imipenem-EDTA Double-Disk Synergy Test for Differentiating Metallo-β-Lactamase-Producing Isolates of Pseudomonas spp. and Acinetobacter spp.

ABSTRACT Gram-negative bacilli with acquired metallo-β-lactamase (MBL) production have been increasingly reported in some countries, necessitating their detection. The aim of this study was to evaluate the performance of the Hodge test and those of the imipenem (IPM)-EDTA, ceftazidime (CAZ)-mercaptopropionic acid (MPA), and CAZ-sodium mercaptoacetic acid (SMA) double-disk synergy tests (DDSTs). The efficiencies of testing CAZ-resistant and IPM-nonsusceptible isolates were also compared. Strains used for the evaluation were known IMP-1 and VIM-2 MBL-producing isolates and consecutive and CAZ-nonsusceptible isolates of pseudomonads and acinetobacters. The performance of the Hodge test was improved by addition of zinc sulfate (140 μg/disk) to an IPM disk. In DDSTs, EDTA (ca. 1,900 μg) disks were better at detecting MBL-producing strains among pseudomonads, while MPA (3 μl) and SMA (3 mg) disks performed better for acinetobacters. EDTA (ca. 750 μg)-plus-SMA (ca. 2 mg) disks performed better than EDTA, MPA, or SMA disks with both organisms. CAZ-SMA DDSTs failed to detect 22 of 80 (28%) MBL-producing acinetobacters. In conclusion, use of an IPM disk and an EDTA (750 μg)-plus-SMA (2 mg) disk improves performance, and testing IPM-nonsusceptible isolates rather than CAZ-resistant isolates could reduce screening work. Further evaluation of the test is required for the detection of other types of MBL-producing gram-negative bacilli.

Imipenem-EDTA Disk Method for Differentiation of Metallo-β-Lactamase-Producing Clinical Isolates of Pseudomonas spp. and Acinetobacter spp.

ABSTRACT Rapid detection of metallo-β-lactamase (MBL)-producing gram-negative bacilli is necessary to prevent their dissemination. The method using a disk with imipenem plus 750 μg of EDTA differentiated all MBL-producing pseudomonads, and the sensitivity and specificity for acinetobacters were 95.7 and 91.0%, respectively. The imipenem-EDTA disks were stable for 12 and 16 weeks at 4 and −20°C, respectively.

Novel Acquired Metallo-β-Lactamase Gene, blaSIM-1, in a Class 1 Integron from Acinetobacter baumannii Clinical Isolates from Korea

ABSTRACT Carbapenem resistance mediated by acquired carbapenemase genes has been increasingly reported, particularly for clinical isolates of Pseudomonas aeruginosa and Acinetobacter spp. Of 1,234 nonduplicate isolates of carbapenem-resistant Pseudomonas spp. and Acinetobacter spp. isolated at a tertiary-care hospital in Seoul, Korea, 211 (17%) were positive for metallo-β-lactamase (MBL). Of these, 204 (96%) had either the blaIMP-1 or blaVIM-2 allele. In addition, seven Acinetobacter baumannii isolates were found to have a novel MBL gene, which was designated blaSIM-1. The SIM-1 protein has a pI of 7.2, is a new member of subclass B1, and exhibits 64 to 69% identity with the IMP-type MBLs, which are its closest relatives. All SIM-1-producing isolates exhibited relatively low imipenem and meropenem MICs (8 to 16 μg/ml) and had a multidrug resistance phenotype. Expression of the cloned blaSIM-1 gene in Escherichia coli revealed that the encoded enzyme is capable of hydrolyzing a broad array of β-lactams, including penicillins, narrow- to expanded-spectrum cephalosporins, and carbapenems. The blaSIM-1 gene was carried on a gene cassette inserted into a class 1 integron, which included three additional cassettes (arr-3, catB3, and aadA1). The strains were isolated from sputum and urine specimens from patients with pneumonia and urinary tract infections, respectively. All patients had various underlying diseases. Pulsed-field gel electrophoresis of SmaI-digested genomic DNAs showed that the strains belonged to two different clonal lineages, indicating that horizontal transfer of this gene had occurred and suggesting the possibility of further spread of resistance in the future.

blaVIM-2 Cassette-Containing Novel Integrons in Metallo-β-Lactamase-Producing Pseudomonas aeruginosa and Pseudomonas putida Isolates Disseminated in a Korean Hospital

ABSTRACT We investigated the phenotypic and genetic properties of metallo-β-lactamase-producing Pseudomonas isolates collected at a tertiary-care hospital in Korea since 1995. The prevalence of imipenem resistance among Pseudomonas aeruginosa isolates reached 16% in 1997, when 9% of the resistant organisms were found to produce VIM-2 β-lactamase, a class B enzyme previously found only in P. aeruginosa isolates from Europe. VIM-2-producing isolates of Pseudomonas putida were also detected. Resistance was transferable from both these species to P. aeruginosa PAO4089Rp by filter mating, although the resistance determinant could not be found on any detectable plasmid. Serotyping showed that many of the VIM-2-producing P. aeruginosa isolates belonged to serotypes O:11 and O:12, and pulsed-field gel electrophoresis of XbaI-digested genomic DNA revealed that many had identical profiles, whereas the P. putida isolates were diverse. Sequencing showed that the blaVIM-2 genes resided as cassettes in class 1 integrons. In contrast to previous VIM-encoding integrons, the integron sequenced from a P. aeruginosa isolate had blaVIM located downstream of a variant of aacA4. blaVIM also lay in a class 1 integron in a representative P. putida strain, but the organization of this integron was different from that sequenced from the P. aeruginosa strain. In conclusion, the metallo-β-lactamase produced by these imipenem-resistant Pseudomonas isolates was VIM-2, and the accumulation of producers reflected clonal dissemination as well as horizontal spread. Strict measures are required in order to control a further spread of resistance.

VIM- and IMP-Type Metallo-β-lactamase–Producing Pseudomonas spp. and Acinetobacter spp. in Korean Hospitals

We determined the occurrence of acquired metallo-β-lactamase (MBL)–producing bacteria in Korean hospitals. Among the isolates nonsusceptible to imipenem that were collected from 28 hospitals from 2000 to 2001, 44 (11.4%) of 387 Pseudomonas spp. and 38 (14.2%) of 267 Acinetobacter spp. infections produced MBL and had alleles of blaVIM-2 or blaIMP-1. MBL-producing isolates were detected in 60.7% of the hospitals.

A two year global evaluation of the susceptibility of Candida species to fluconazole by disk diffusion

An improved fluconazole 25-mg disk diffusion method was used to test the susceptibility of 20,900 consecutively isolated clinical strains of Candida species from 40 hospital laboratories in 26 countries. The procedure is similar to the National Committee for Clinical Laboratory Standards (NCCLS) M2-A6 method for testing bacteria, except Mueller-Hinton agar is supplemented with 2% glucose and 0.5 mcg/mL methylene blue. Plates were incubated at 35 degrees C and read after 18 to 24h. Tentative zone interpretive criteria were based on the correlation by regression analysis with the NCCLS M27-A Reference Method for Broth Dilution Antifungal Susceptibility Testing of Yeasts: > =19mm Susceptible, < = 12mm Resistant, and 13-18 mm Susceptible-Dose Dependent. Of 14,368 isolates of Candida albicans, 2,073 C. glabrata, 869 C. tropicalis, 752 C. parapsilosis, and 351 C. krusei, 99%, 67%, 90%, 94%, and 26%, respectively, were susceptible in vitro to fluconazole. All study sites used the BIOMIC System to electronically read zones on plates, interpret, record test results and verify quality control data. This is the largest study to date that evaluated a broad range of sequentially collected yeasts from various infections and different types of hospitals. The modified disk testing procedure is facile and economical to perform and offers a reproducible and accurate means to assess the in vitro susceptibility of Candida species to fluconazole.

Multidrug-Resistant Acinetobacter spp.: Increasingly Problematic Nosocomial Pathogens

Pathogenic bacteria have increasingly been resisting to antimicrobial therapy. Recently, resistance problem has been relatively much worsened in Gram-negative bacilli. Acinetobacter spp. are typical nosocomial pathogens causing infections and high mortality, almost exclusively in compromised hospital patients. Acinetobacter spp. are intrinsically less susceptible to antibiotics than Enterobacteriaceae, and have propensity to acquire resistance. A surveillance study in Korea in 2009 showed that resistance rates of Acinetobacter spp. were very high: to fluoroquinolone 67%, to amikacin 48%, to ceftazidime 66% and to imipenem 51%. Carbapenem resistance was mostly due to OXA type carbapenemase production in A. baumannii isolates, whereas it was due to metallo-β-lactamase production in non-baumannii Acinetobacter isolates. Colistin-resistant isolates were rare but started to be isolated in Korea. Currently, the infection caused by multidrug-resistant A. baumannii is among the most difficult ones to treat. Analysis at tertiary care hospital in 2010 showed that among the 1,085 isolates of Acinetobacter spp., 14.9% and 41.8% were resistant to seven, and to all eight antimicrobial agents tested, respectively. It is known to be difficult to prevent Acinetobacter spp. infection in hospitalized patients, because the organisms are ubiquitous in hospital environment. Efforts to control resistant bacteria in Korea by hospitals, relevant scientific societies and government agencies have only partially been successful. We need concerted multidisciplinary efforts to preserve the efficacy of currently available antimicrobial agents, by following the principles of antimicrobial stewardship.

High Prevalence of PER-1 Extended-Spectrum β-Lactamase-Producing Acinetobacter spp. in Korea

ABSTRACT PER-1, an extended-spectrum β-lactamase, has been reported only in Europe. We detected PER-1 in 53 of 97 acinetobacters in Korea, mainly in the sputum of intensive care unit patients. Pulsed-field gel electrophoresis analysis suggested that clonal spread had occurred. Only PCR reliably detected PER-1 producers. PER-1 producers may also exist in other Asian countries.

Wide dissemination of OXA-type carbapenemases in clinical Acinetobacter spp. isolates from South Korea

Carbapenem-resistant Acinetobacter spp. are being increasingly reported worldwide, including in South Korea, where we examined 144 representative isolates collected in a nationwide hospital survey in 2005. Metallo-beta-lactamases were detected in only 19.4% of isolates, none of which were Acinetobacter baumannii, whereas 74.3% of isolates (mostly A. baumannii) expressed bla(OXA) carbapenemase genes. Among the latter, 47 had bla(OXA-23)-like genes and 56 had upregulated bla(OXA-51)-like variants, including bla(OXA-66), (-83), (-109) and (-115); bla(OXA-115) was a novel variant, detected in two isolates. bla(OXA-72) (bla(OXA-40)-like) was detected in only a single Acinetobacter baylyi isolate, whilst three Acinetobacter calcoaceticus isolates had both bla(VIM-2)-like and bla(OXA-58) genes. Pulsed-field gel electrophoresis (PFGE) suggested the spread of A. baumannii clones with OXA carbapenemases within and between hospitals. In conclusion, the recent increase in imipenem-resistant Acinetobacter spp. from South Korea is mostly due to OXA-type carbapenemases.

Further Increases in Carbapenem-, Amikacin-, and Fluoroquinolone-Resistant Isolates of Acinetobacter spp. and P. aeruginosa in Korea: KONSAR Study 2009

Purpose The increasing prevalence of antimicrobial resistant bacteria has become a serious worldwide problem. The aim of this study was to analyze antimicrobial resistance data generated in 2009 by hospitals and commercial laboratories participating in the Korean Nationwide Surveillance of Antimicrobial Resistance program. Materials and Methods Susceptibility data were collected from 24 hospitals and two commercial laboratories. In the analysis, resistance did not include intermediate susceptibility. Duplicate isolates were excluded from the analysis of hospital isolates, but not from the commercial laboratory isolates. Results Among the hospital isolates, methicillin-resistant Staphylococcus aureus, penicillin G-non-susceptible Streptococcus pneumoniae based on meningitis breakpoint, and ampicillin-resistant Enterococcus faecium remained highly prevalent. The proportion of vancomycin-resistant E. faecium gradually increased to 29%. Ceftazidime-resistant Escherichia coli and Klebsiella pneumoniae increased to 17% and 33%, respectively, and fluoroquinolone-resistant K. pneumoniae, Acinetobacter spp. and Pseudomonas aeruginosa increased to 33%, 67% and 39%, respectively. Amikacin-resistant Acinetobacter spp. increased to 48%. Imipenem-resistant Acinetobacter spp. and P. aeruginosa increased to 51% and 26%, respectively. Higher resistance rates were observed in intensive care unit (ICU) isolates than in non-ICU isolates among the isolates from hospitals. Resistance rates were higher in hospital isolates than in clinic isolates among the isolates from commercial laboratories. Conclusion Among the hospital isolates, ceftazidime-resistant K. pneumoniae and fluoroquinolone-resistant K. pneumoniae, Acinetobacter spp., and P. aeruginosa further increased. The increase in imipenem resistance was slight in P. aeruginosa, but drastic in Acinetobacter spp. The problematic antimicrobial-organism combinations were much more prevalent among ICU isolates.