Min Jeong Park

Boronic acid disk tests for identification of extended-spectrum β-lactamase production in clinical isolates of Enterobacteriaceae producing chromosomal AmpC β-lactamases

A study using boronic acid (BA) was designed to detect the extended-spectrum beta-lactamases (ESBLs) in Enterobacteriaceae producing chromosomal AmpC beta-lactamases. A total of 197 clinical isolates of Enterobacter spp. (n=100), Serratia marcescens (n=62) and Citrobacter freundii (n=35) were analysed. Genes encoding ESBLs were detected by polymerase chain reaction (PCR) amplification followed by direct sequencing of PCR products. The Clinical and Laboratory Standards Institute confirmatory test detected only 72.1% of the ESBL-producing isolates. When a > or =5mm increase in the zone diameter of either the cefotaxime/clavulanic acid and/or the ceftazidime/clavulanic acid disks tested in combination with BA versus cefotaxime and/or ceftazidime containing BA was considered to be a positive for ESBL, the method detected 60 (98.4%) of the 61 isolates that harboured ESBLs and showed no false-positive results for ESBL-non-producing isolates. In conclusion, the BA disk test is a highly sensitive and specific method for the detection of ESBLs in Enterobacteriaceae producing chromosomal AmpC beta-lactamases.

Case Report of Pediatric Gastroenteritis Due to CTX-M-15 Extended-Spectrum β-Lactamase-Producing Salmonella enterica Serotype Enteritidis

A clinical isolate of Salmonella enterica serotype Enteritidis in Korea was found to produce the extended-spectrum beta-lactamase CTX-M-15. The isolate was recovered in 2008 from the stool of a 3-yr-old boy with gastroenteritis. This isolate was found to be resistant to multiple drugs, including ampicillin, piperacillin, cefotaxime, ceftazidime, cefepime, and aztreonam. The resistance to cefotaxime was transferred by conjugation to recipient Escherichia coli J53. The patient was eventually successfully treated with trimethoprim-sulfamethoxazole. This is the first report of the bla (CTX-M-15) gene in S. enterica serotype Enteritidis in Korea.

An Increase in the Clinical Isolation of Acquired AmpC β-Lactamase-Producing Klebsiella pneumoniae in Korea from 2007 to 2010

We investigated the occurrence and genetic basis of AmpC β-lactamase (AmpC)-mediated antibiotic resistance, by examining Escherichia coli, Klebsiella pneumoniae, and Proteus mirabilis isolates at a university hospital, from 2007 to 2010. The ampC genes were detected by multiplex AmpC PCR, and AmpC-positive strains were subjected to DNA sequencing. Extended-spectrum β-lactamase (ESBL) production was assessed using the ESBL disk test based on the utilization of boronic acid. Carbapenem-resistant isolates were further investigated by the modified Hodge test, a carbapenemase inhibition test and SDS-PAGE experiments. AmpC expression was detected in 1.6% of E. coli (39 DHA-1, 45 CMY-2, and 1 CMY-1) isolates, 7.2% of K. pneumoniae (39 DHA-1, 45 CMY-2, and 1 CMY-1) isolates, and 2.5% of P. mirabilis (8 CMY-2 and 1 CMY-1) isolates. Of the 198 acquired AmpC producers, 58 isolates (29.3%) also produced an ESBL enzyme. Among the acquired AmpC-producing K. pneumoniae isolates, the minimum inhibitory concentration (MIC) MIC50/MIC90 values for cefoxitin, cefotaxime, cefepime, imipenem, and meropenem were >32/>32, 16/>32, 1/16, 0.25/0.5, and <0.125/0.125 µg/mL, respectively. The MIC values for carbapenem were ≥2 µg/mL for 2 K. pneumoniae isolates, both of which carried the blaDHA-1 gene with a loss of OmpK36 expression, but were negative for carbapenemase production. The acquisition of AmpC-mediated resistance in K. pneumoniae isolates increased, as did the proportion of AmpC and ESBL co-producers among the hospital isolates. The accurate identification of isolates producing AmpCs and ESBLs may aid in infection control and will assist physicians in selecting an appropriate antibiotic regimen.

Prevalence and Molecular Characteristics of Carbapenemase-Producing Enterobacteriaceae From Five Hospitals in Korea

Background The emergence of carbapenemase-producing Enterobacteriaceae (CPE) represents a major clinical problem because these bacteria are resistant to most antibiotics. CPE remain relatively uncommon in Korea. We report the prevalence, clinical characteristics, and molecular epidemiology of CPE isolates collected from five university hospitals in Korea. Methods Between January and December 2015, 393 non-duplicated isolates that were nonsusceptible to ertapenem were analyzed. Production of carbapenemase, extended-spectrum β-lactamase, and AmpC β-lactamase was determined by genotypic tests. Antimicrobial susceptibility profiles were determined by using an Etest. Clonality of Klebsiella pneumoniae carbapenemase (KPC)-2-producing and oxacillinase (OXA)-232-producing Klebsiella pneumoniae isolates was determined by pulsed-field gel electrophoresis (PFGE). Results Of the 393 isolates tested, 79 (20.1%) were CPE. Of these 79 isolates, 47 (59.5%) harbored the blaOXA-232 gene while the remaining isolates carried genes blaKPC-2 (n=27), blaIMP-1 (n=4), and blaNDM-1 (n=1). Among the 24 KPC-2 K. pneumoniae isolates from hospital B, 100% were resistant to carbapenems, 8% to colistin, and 0% to tigecycline. Among the 45 OXA-232 K. pneumoniae at hospital C, 95% were resistant to ertapenem, 68% to imipenem, 95% to meropenem, 10% to colistin, and 24% to tigecycline. PFGE analysis revealed a unique pattern for KPC-2 K. pneumoniae and identified 30 isolates belonging to the dominant pulsotypes (PT)1 and PT2 among 41 OXA-232 K. pneumoniae isolates. Conclusions CPE strains are present in Korea, with the majority of K. pneumoniae isolates producing OXA-232 and KPC-2. The prevalence and predominant genotypes of CPE show hospital-specific differences.

Clinical Usefulness of T-SPOT.TB Test for the Diagnosis of Tuberculosis

BACKGROUND Tuberculosis-specific ELISPOT assay (T-SPOT.TB, Oxford Immunotec, UK) is a test that detects interferon-gamma producing T-cells after stimulating patients lymphocytes with two kinds of tuberculosis-specific antigens (ESAT-6 and CFP-10). We evaluated clinical usefulness of T-SPOT.TB test in Korean adults with intermediate burden of tuberculosis and high rate of BCG vaccination at birth. METHODS T-SPOT.TB test was performed in 79 patients and 64 healthy volunteers and these patients and volunteers were classified into four groups: group 1, patients with active tuberculosis (n=30); group 2, patients with not-active tuberculosis (n=27); group 3, patients without tuberculosis (n=24); group 4, healthy volunteers without tuberculosis history (n=62). Positive rates and average spot counts of T-SPOT.TB test were obtained among these groups. RESULTS Positive rates of group 1 (96.4%) and group 2 (92.3%) were higher than those of group 3 (31.6%) and group 4 (27.4%) (P<0.0001). The sensitivity deduced from group 1 and specificity deduced from group 4 of T-SPOT.TB test were 96.4% and 72.6%, respectively. The average spot counts of group 1 and 2 were higher than those of group 3 and 4 (P<0.001). There was a tendency of increasing positive rate with increasing age. Overall agreement between T-SPOT.TB test and tuberculin skin test was 63.8% (kappa=0.29). CONCLUSIONS T-SPOT.TB test would be a very useful screening and supplementary test for diagnosis of tuberculosis due to its high sensitivity. However, positive results of T-SPOT.TB should be cautiously interpreted because of high positivity in treated tuberculosis patients and healthy volunteers in Korea.

Evaluation of feasible timing of elective noncardiac procedure after antiplatelet discontinuation in patients treated with antiplatelet agents.

Background The current standard of care is to delay noncardiac procedure (NCP) 5 to 7 days after discontinuation of antiplatelet agents (APAs) in patients with coronary stents. However, it is often difficult to follow because of concerns over stent thrombosis. The point-of-care aspirin/P2Y12 assay (VerifyNow; Accumetrics Inc, San Diego, CA) is useful to evaluate platelet reactivity in conjunction with APAs. In this study, we evaluated the feasible timing after discontinuation of APAs. Methods and Results Sixty-two patients taking APAs, who were scheduled to undergo elective NCP, were enrolled. All patients took either aspirin 100 mg or aspirin 100 mg plus clopidogrel 75 mg daily. The aspirin-reactivity unit (ARU) and P2Y12-reactivity unit (PRU) were measured from 0 days (day 0, no discontinuation) to as long as 5 days (day 5, 5 days after discontinuation) depending on each procedure schedule. For 15 patients, baseline ARU and PRU (592 and 288) before aspirin/clopidogrel loading at index percutaneous coronary intervention were collected as control. For ARU after discontinuation of APA, days 0 to 5 values progressively increased over time (489.4 ± 85.3, 512.6 ± 77.0, 589.9 ± 58.8, 613.6 ± 47.3, 632.6 ± 49.2, 662.0 ± 4.2). Likewise, for PRUs, days 0 to 5 values also increased over time (245.0 ± 96.9, 253.9 ± 80.9, 270.9 ± 45.8, 289.0 ± 68.6, 306.5 ± 29.2, 351.0 ± 8.5). The ARU and PRU well correlated with days after APA discontinuation by linear regression analysis (y = 490.897 + 39.238 * x, R 2 = 0.43, P < 0.001; y = 241.739 + 16.701 * x, R 2 = 0.092, P = 0.018, respectively). Assuming baseline ARU and PRU as 592 and 288, the mean days after complete reversal of platelet reactivity by APAs are 2.6 and 2.8, respectively. Conclusions The feasible timing of NCP after discontinuation of APAs showed less than 5 days. VerifyNow is useful in the evaluation of antiplatelet reversal after discontinuation of APAs.

Comparison of Clinical and Laboratory Standards Institute and European Committee on Antimicrobial Susceptibility Testing Breakpoints for beta-Lactams in Enterobacteriaceae Producing Extended-Spectrum beta-Lactamases and/or Plasmid-Mediated AmpC beta-Lacta.

Results: Among the 94 isolates containing ESBL and/ or PABL, the number of isolates that were susceptible to cefotaxime, ceftazidime, aztreonam, cefepime, and imipenem according to the CLSI 2010 vs. the EUCAST breakpoints were 4 (4.3%) vs. 4 (4.3%); 26 (27.7%) vs. 8 (8.5%); 37 (39.4%) vs. 14 (14.9%); 71 (75.5%) vs. 31 (33.0%); and 76 (80.9%) vs. 90 (95.7%), respectively. Of the 18 isolates that were not susceptible to imipenem according to the CLSI 2010 breakpoints, 13 isolates (72.2%) were P. mirabilis. Conclusion: The CLSI 2010 MIC breakpoints without tests to detect ESBL and/or PABL for Enterobacteriaceae could be unreliable. Thus, special tests for ESBLs and AmpC β-lactamases are required to detect the resistance mechanisms involved. (Korean J Clin Microbiol 2011;14:24-29)

Comparison of the BACTEC Peds Plus Pediatric Blood Culture Bottle to the BacT/Alert PF Pediatric Blood Culture Bottle for Culturing Blood from Pediatric Patients

Blood samples were collected for culture from pediatric patients who were hospitalized during 2010 at a university hospital. BACTEC Peds Plus and BacT/Alert PF bottles were placed in the BACTEC FX and BacT/Alert 3D blood culture system, respectively, and tested for 5 days. Bottles flagged by instruments as positive were removed from the instruments and the TTDs were recorded.

Emergence of multidrug-resistant Providencia rettgeri isolates co-producing NDM-1 carbapenemase and PER-1 extended-spectrum β-lactamase causing a first outbreak in Korea

BackgroundNosocomial outbreak due to carbapenem-resistant Enterobacteriaceae has become serious challenge to patient treatment and infection control. We describe an outbreak due to a multidrug-resistant Providencia rettgeri from January 2016 to January 2017 at a University Hospital in Seoul, Korea.MethodsA total of eight non-duplicate P. rettgeri isolates were discovered from urine samples from eight patients having a urinary catheter and admitted in a surgical intensive care unit. The β-lactamase genes were identified using polymerase chain reaction and direct sequencing, and strain typing was done with pulsed-field gel electrophoresis (PFGE).ResultsAll isolates showed high-level resistance to extended-spectrum cephalosporins, aztreonam, meropenem, ertapenem, ciprofloxacin, and amikacin. They harbored the blaNDM-1 carbapenemase and the blaPER-1 type extended-spectrum β-lactamases genes. PFGE revealed that all isolates from eight patients were closely related strains.ConclusionsThe 13-month outbreak ended following reinforcement of infection control measures, including contact isolation precautions and environmental disinfection. This is the first report of an outbreak of a P. rettgeri clinical isolates co-producing NDM-1 and PER-1 β-lactamase.

Carbapenem Inactivation Method: Accurate Detection and Easy Interpretation of Carbapenemase Production in Enterobacteriaceae and Pseudomonas spp.

Background: We evaluated the carbapenem inactivation method (CIM) compared with the modified Hodge test (MHT) for the detection of carbapenemase-producing Gram-negative bacilli. Methods: A total of 61 isolates of carbapenemaseproducing Enterobacteriaceae (CPE: 14 KPC, 7 GES5, 8 NDM-1, 9 VIM-2, 9 IMP-1, and 14 OXA-48-like), 34 isolates of metallo-β-lactamase (MBL)-producing Pseudomonas spp. (14 VIM-2 and 20 IMP-6), and 70 carbapenem-nonsusceptible carbapenemase-negative isolates were included. The CIM and MHT were performed for all of the isolates. To perform the CIM, a meropenem disk was incubated with a suspension of the isolate to be tested and then on Mueller-Hinton agar with the Escherichia coli ATCC 29522 strains. The absence of an inhibition zone indicates presence of a carbapenemase. The presence of a clearing zone indicates lack of a carbapenemase. Results: The total sensitivity and specificity of CIM (96% sensitivity and 100% specificity) in carbapenemnonsusceptible Enterobacteriaceae and Pseudomonas spp. were better than those of the MHT (77% sensitivity and 94% specificity). The interpretation of CIM results was easy, with no or <20 mm inhibition zones indicating positivity and >20 mm inhibition zones indicating negative carbapenemase activity. Conclusion: The CIM had excellent sensitivity and specificity for detection of CPE and MBL-producing Pseudomonas spp., and a positive result was easily determined, unlike the MHT. (Ann Clin Microbiol 2016;19:83-87)