Borum Suh

In vivo selection of carbapenem-resistant Klebsiella pneumoniae by OmpK36 loss during meropenem treatment

We recovered a carbapenem-resistant Klebsiella pneumoniae isolate H224 under in vivo meropenem selection pressure. Insertional inactivation of a major porin gene, ompK36, by IS5 element might play a role in acquiring carbapenem resistance in this strain harboring plasmid-borne DHA-1 AmpC beta-lactamase.

Outbreak of Meropenem-Resistant Serratia marcescens Comediated by Chromosomal AmpC β-Lactamase Overproduction and Outer Membrane Protein Loss

ABSTRACT The aim of this study was to investigate the mechanisms involved in the meropenem resistance of Serratia marcescens clinical isolates. Meropenem-resistant (MIC range, 16 to 32 μg/ml) S. marcescens isolates were recovered from nine patients in a tertiary hospital in Seoul, South Korea, from June to November 2005. All the isolates shared identical or similar (>85% similarity) SpeI macrorestriction patterns, indicating clonal spread. PCR experiments did not detect any carbapenemase in those isolates. They carried the blaCTX-M-22 gene located on a 150-kbp plasmid of the incompatibility group L/M; however, the addition of clavulanic acid exhibited few effects on meropenem MICs. Although meropenem MICs were reduced 4- to 16-fold with the addition of boronic acid, no plasmid-borne AmpC β-lactamase gene was detected in PCR experiments. Real-time quantitative PCR experiments showed that expression levels of the chromosomal ampC gene in those isolates were 87.06 to 155.76 times higher than that of the reference strain ATCC 8100. SDS-PAGE showed a lack of the 42-kDa outer membrane protein (OmpF). In combination with the overproduction of the chromosomal AmpC enzyme, the loss of OmpF may have played a role in the acquisition of meropenem resistance in our isolates.

Acute promyelocytic leukemia with insertion of PML exon 7a and partial deletion of exon 3 of RARA: a novel variant transcript related to aggressive course and not detected with real-time polymerase chain reaction analysis.

We report the case of a 38-year-old man with acute promyelocytic leukemia (APL) showing a distinct breakpoint cluster region 2 (bcr2) variant transcript. Findings from bone marrow, cytogenetic, fluorescence in situ hybridization, and qualitative reverse transcriptase-polymerase chain reaction (RT-PCR) analyses were consistent with the diagnosis of APL. Although PCR products of size 841 bp and 984 bp were amplified from bone marrow specimen, the quantitative PCR (RQ-PCR) findings were negative. Given the discrepancy in PCR results, sequencing of PCR products was performed to determine the detailed composition of these fusion transcripts. By cloning and sequencing, we discovered that these two bands were isoforms, in which one exon (exon 5, 144 bp) of the PML gene was spliced out of the smaller products (minor PCR products); one sequence (G) insertion and one base substitution (T-->C) of PML exon 4 generate a stop codon in the smaller fusion transcript. In addition, a search of the Ensembl database revealed that these variant PML/RARA fusion transcripts were composed of exon 7a insertion of the PML gene and partial deletion (46 bp) of exon 3 of the RARA gene, in addition to inserted sequences of intron 7 of PML and genomic sequence ATCT of unknown origin at the fusion junction site. Although the biological significance of most atypical transcripts remains unclear, sequence analysis of these atypical products should be performed, to reveal the composition of such a fusion transcript and elucidate the molecular mechanism.

JAK2 V617F/C618R mutation in a patient with polycythemia vera: a case study and review of the literature.

The acquired Janus kinase 2 (JAK2) V617F mutation shows a high frequency in diverse BCR/ABL-negative chronic myeloproliferative disorders (CMPD), and it is typically associated with polycythemia vera (PV). The frequency of JAK2 V617F mutation is about 90% in patients with PV, 50-60% in patients with essential thrombocythemia (ET), primary myelofibrosis (PMF), and less in patients with other myeloid neoplasms, while extremely rare in lymphoid malignancies. About 20 kinds of novel mutations of JAK2 other than V617F have been reported recently in the literature. Among these mutations, only one case of JAK2 V617F/C618R has been reported in a 67-year-old patient with PV. Here, we report a rare case of JAK2 V617F/C618R in a 41-year-old Korean male patient with review of the relevant literature on JAK2 mutations other than V617F. Although the frequency of JAK2 mutations other than the V617F is very low, this study emphasizes the need for assiduous analysis of the JAK2 gene to characterize new mutations, to determine their frequency, and to improve understanding of the clinical phenotypes as well as prognostic and biologic features associated with these mutations.

Molecular epidemiology of Pseudomonas aeruginosa clinical isolates from Korea producing β-lactamases with extended-spectrum activity

This study was performed to investigate the prevalence and molecular epidemiology of Pseudomonas aeruginosa isolates from Korea that produce enzymes with extended-spectrum (ES) activity to β-lactams. A total of 205 non-duplicate P. aeruginosa clinical isolates were collected from 18 university hospitals in Korea. PCR and sequencing experiments were performed to identify genes encoding β-lactamases. PCR mapping and sequencing of the regions surrounding the β-lactamase genes were performed. Multilocus sequence typing experiments were performed. The most common sequence type (ST) was ST235 (n = 96), and 2 single-locus variants of ST235, ST1015 (n = 1) and ST1162 (n = 1), were also identified. These 3 STs were grouped as a clonal complex (CC), CC235. The remaining 107 isolates were identified as 59 different STs. Isolates belonging to CC235 showed higher rates of non-susceptibility to imipenem (85.4% versus 47.7%) and meropenem (92.7% versus 52.3%) compared to non-CC235 isolates. All the metallo-β-lactamase (MBL)-producing isolates were identified as CC235, except for 1 ST591. Genes encoding OXA-17 and OXA-142 were detected in 1 isolate and 4 isolates of CC235, respectively; while the bla(SHV-12) gene was detected in 4 non-CC235 isolates. Class A and D β-lactamases with ES activity play a role in acquiring ceftazidime resistance in P. aeruginosa in Korea. Production of IMP-6 and VIM-2 MBLs is the main mechanisms in acquiring resistance to ceftazidime and carbapenems in P. aeruginosa isolates in Korea. Clonal spread of P. aeruginosa CC235 may be an important conduit for the dissemination of MBL genes in Korea.

Analysis of fluorescence in situ hybridization, mtDNA quantification, and mtDNA sequence for the detection of early bladder cancer

We designed this study to test the sensitivities of cytology, the nuclear matrix protein 22 (NMP22) assay, and fluorescence in situ hybridization (FISH) in the early detection of urothelial carcinoma, and to identify mtDNA alterations in urinary epithelial cells. We collected 41 urine samples and 26 corresponding peripheral blood samples from patients with clinically suspected urothelial carcinoma. The FISH and NMP22 assays detected 92.1% of the cancers, and cytology detected 60.5%. In the low-grade group, NMP22 and FISH analyses were more sensitive than cytology, but in the high-grade group, all three methods showed approximately 90% sensitivity. Overall, the FISH and NMP22, or FISH and cytology assays combined detected 97.4% of cancers, while cytology with NMP22 detected 92.1%. In the low-grade group, the sensitivity of the three methods combined was above 80%, but in high-grade group, the combined sensitivity was approximately 100%. In the mtDNA control region, we detected characteristic heteroplasmic mtDNA substitution mutations in 1 patient and a mtDNA length heteroplasmic mutation in 303 polyC or 16184 poly C in 20 patients. Overall, urothelial carcinoma-specific mtDNA mutations were observed in 20 of the 26 patients (76.9%). The average mtDNA copy numbers in urine samples and corresponding peripheral blood samples (83.45 +/- 60.36 and 39.0 +/- 24.38, respectively) (mean +/- standard deviation [SD]) differed significantly (P < 0.001). The mtDNA copy numbers in the urine samples from patients with high-grade and low-grade tumors (81.83 +/- 67.78 and 86.49 +/- 46.69, respectively) did not differ significantly (P = 0.589). In conclusion, the FISH assay showed the highest sensitivity for detecting low-grade urothelial carcinoma, and mtDNA copy numbers in urine samples were higher than those in the corresponding peripheral blood samples. The frequency of mtDNA mutations in the D-loop region in patients with cancer was approximately 80% in our study. This report further supports the significance of genetic alteration in urothelial carcinoma and the clinical utility of the FISH, mtDNA quantitation polymerase chain reaction, mtDNA sequencing, and capillary electrophoresis for this purpose.

Rare translocations involving chromosome band 8p11 in myeloid neoplasms.

Two types of distinct clinical syndromes are associated with abnormalities of chromosomal band 8p11 [1]. The first is 8p11 myeloproliferative syndrome (EMS)estem cell leukemia-lymphoma syndrome (SCLL), an aggressive chronic myeloproliferative disorder characterized by myeloid hyperplasia, eosinophilia, and lymphoblastic lymphoma. The second clinical syndrome is an acute myeloid leukemia (AML) of monocytoid phenotype (FAB type M4 or M5) with erythrophagocytosis. The abnormalities of chromosome band 8p11 target two different genes, FGFR1 in EMS and MYST3 (alias MOZ ) in AML. Rosati et al. [2] reported NSD3 (nuclear receptor-binding su(var)3-9, enhancer of zeste, trithorax [SET] domain containing gene 3) as the third translocation target in chromosome 8p11wp12 region, which is telomeric to FGFR1. (Note that NSD3 has since been classified as WHSC1L1; accession no. AF_332469.) Here, we report two rare translocations involving chromosome band 8p11 in myeloid neoplasms: t(8;22)(p11;q11) and t(8;11)(p11;p15). The first patient was a 50-year-old Korean woman, with no relevant past history, who presented with general weakness. An initial complete blood count showed a hemoglobin level of 9.5 g/dL, a platelet count of 596,000/mL, and a white blood cell count of 51,630/mL with 1% myeloblasts, 1% promyelocytes, 5% myelocytes, 6% metamyelocytes, 78% neutrophils, 4% lymphocytes, 1% monocytes, 2% eosinophils, and 2% basophils. A subsequent bone marrow biopsy revealed hypercellular marrow (O90%) with 18% blast cells and eosinophilia. An initial diagnosis of chronic myelogenous leukemia (CML), accelerated phase (AP), was made; however, the results of reverse transcriptasee polymerase chain reaction analysis (RT-PCR) for both major and minor BCReABL were negative. Three months later, transformation to blast phase (BP) occurred and the patient died of sepsis. The second patient was a 37-year-old Korean woman who was admitted with a complaint of abdominal pain and fever. She had been diagnosed with left-side breast cancer 11 years before and underwent a modified radical mastectomy. The patient was subsequently treated with chemotherapy and radiotherapy, when multiple metastases had developed. On presentation at our institution, many abnormal blasts (69%) were observed in the peripheral blood smear. An ensuing bone marrow aspiration revealed

A Report of Cat Scratch Disease in Korea Confirmed by PCR Amplification of the 16S-23S rRNA Intergenic Region of Bartonella henselae

We report a case of cat scratch disease in an 8-yr-old girl who presented with fever and enlargement of both axillary lymph nodes. Both aerobic and anaerobic cultures of the lymph node aspirate were negative for microbial growth. Gram staining and Warthin-Starry silver staining did not reveal any organism. Purified DNA from the PCR-amplicon of the 16S-23S rRNA intergenic region was sequenced and showed 99.7% identity with the corresponding sequence of Bartonella henselae strain Houston-1. Our findings suggest that the internal transcribed spacer is a reliable region for PCR identification of Bartonella species. In patients with lymphadenitis, a history of contact with cats or dogs necessitates the use of diagnostic approaches that employ not only the conventional staining and culture but also molecular methods to detect B. henselae.

Performance Characteristics of the UniCel DxI 800 Immunoassay for the Maternal Serum Quadruple Test, Including Median Values for Each Week of Gestation, in Korean Women

BACKGROUND Maternal serum prenatal quadruple screening includes testing for alpha-fetoprotein (AFP), human chorionic gonadotrophin (hCG), unconjugated estriol (uE3), and dimeric inhibin A (DIA). We evaluated quadruple screening using an automated platform and looked for any ethnic differences in the median values of each marker. METHODS We measured the concentrations of each quadruple test analyte using the UniCel DxI 800 system (Beckman Coulter, USA) in 788 Korean mid-trimester maternal serum samples and calculated their median values using Benetech software (Benetech, Canada). We also compared the results with those obtained using the Immulite 2000 assay (Siemens Healthcare Diagnostics, USA) or ELISA (DSL, USA) in 442 samples. RESULTS We obtained mid-trimester median values for each marker. The following are the comparative results for each test using the Immulite 2000 assay or ELISA (x) and the UniCel DxI 800 immunoassay (y): AFP, y=1.10x+0.01, r=0.925; uE3, y=0.28x+0.24, r=0.885; hCG, y=1.22x-3047.8, r=0.944; and DIA, y=0.86x+15.31, r=0.833. Assay results for each of the four markers showed good correlations. However, significant biases necessitated new median calculations of prenatal risk estimates in all four tests. CONCLUSIONS We established gestational age-specific second-trimester median values for four markers in Korean samples using the UniCel DxI 800 immunoassay system. Despite significant bias, there were good correlations between the results obtained using the UniCel DxI 800 immunoassay and those obtained using the Immulite 2000 assay.

MLL rearrangement with t(6;11)(q15;q23) as a sole abnormality in a patient with de novo acute myeloid leukemia: conventional cytogenetics, FISH, and multicolor FISH analyses for detection of rare MLL-related chromosome abnormalities

We report a rare case of acute myeloid leukemia (AML) with t(6;11)(q15;q23) in a 50-year-old female showing a poor prognosis. Bone marrow biopsy revealed markedly hypercellular marrow with infiltrates of myeloblasts, consistent with AML-M2 morphology. The karyotype of this patient was 46,XX,t(6;11)(q15;q23) in all analyzed cells, and the results of fluorescence in situ hybridization (FISH) and multi-color FISH analysis confirmed this unique MLL rearrangement as a sole abnormality. To our knowledge, t(6;11)(q13 approximately q15;q23) is the most rare type of MLL rearrangement involving the long arm of chromosome 6. Only two cases with t(6;11)(q13;q23) and three cases with t(6;11)(q15;q23) have been reported, but detailed clinical or laboratory data were not available. From this report, it is apparent that in a cytogenetic laboratory, the accurate detection of a rare type of MLL rearrangement is very important in the differential diagnosis, prompt treatment, and prediction of prognosis of leukemias.