Yunjung Cho

Evaluation of a novel real-time RT-PCR using TOCE technology compared with culture and Seeplex RV15 for simultaneous detection of respiratory viruses

n Abstractn n Backgroundn Various kinds of commercial molecular systems have been developed for fast and more accurate detection of respiratory viruses. Anyplex™ II RV16 [RV16] was designed for simultaneous detection of 16 respiratory viruses using multiplex PCR coupled with TOCE™ technology.n n n Objectivesn To compare the performance of RV16 with those of culture and Seeplex® RV15 ACE [RV15] by determining their sensitivity and specificity.n n n Study designn Seven hundred and thirty respiratory samples were tested by modified shell vial culture method, RV16, and RV15. For molecular tests, automated nucleic acid extraction and liquid handling system using MICROLAB Nimbus IVD (Hamilton, USA) was adopted to maximize the workflow and accuracy. Performance of each assay was determined against a composite reference standard.n n n Resultsn Two hundred and one samples (28%) out of 730 samples were positive by culture, while additional 281 (39%) were positive by RV16 or RV15. Sensitivities of RV16, RV15, and culture for virus tested were as follows: 100/93/63% for influenza A, 90/80/69% for influenza B, 98/94/63% for RSV, 98/52/23% for adenovirus, and 100/75/46% for PIV. For viruses not covered by culture, sensitivities of RV16 and RV15 were as follows: 99/81% for rhinovirus, 92/100% for coronavirus OC43, 100/56% for coronavirus 229E/NL63, 92/88% for metapneumovirus, 100/62% for bocavirus, and 91/91% for enterovirus. Overall, the specificities of culture, RV16, and RV15 (Seegene) were 100/99.9/99.9%.n n n Conclusionsn RV16 assay was superior to culture method and RV15 and will be a promising tool for patient management and public health epidemiology.n n

Validation of the Elecsys® HIV combi PT assay for screening and reliable early detection of HIV-1 infection in Asia

BACKGROUNDnThe Elecsys® HIV combi PT assay was developed to allow earlier detection of HIV infection with increased sensitivity and specificity.nnnOBJECTIVESnTo validate the assay for screening and reliable early detection of HIV-1 infection in Asia.nnnSTUDY DESIGNnSamples tested reflected those routinely screened in Asia and comprised: HIV-1 antigen lysate (25 samples) and antibody (20 samples) dilutions; seven HIV-1 seroconversion panels (46 samples); 39 patient samples from early infection; 183 known-positive sera; HIV-1 p24 antigen sensitivity panel (seven samples); >500 routine clinical samples per center. The Elecsys® HIV combi PT assay was compared with fourth- (ADVIA Centaur® HIV combo, ARCHITECT® HIV combo, Elecsys® HIV combi) and third-generation (VIRONOSTIKA® HIV Uni-Form II Plus O, Zhuhai Livzon Anti-HIV EIA, Serodia® Particle Agglutination) assays commonly used in the region.nnnRESULTSnOverall, the Elecsys® HIV combi PT showed superior or similar sensitivity to the comparators for detecting all subtypes. The assay correctly identified all positive samples, including those taken soon after infection, and detected seroconversion at a similar or shorter time interval than the comparators. The analytical sensitivity of Elecsys® HIV combi PT for HIV-1 p24 antigen was 0.90 IU/mL, which was lower than reported previously. The assay showed good specificity (99.86%) that was superior or equivalent to the other fourth-generation assays tested.nnnCONCLUSIONSnThese robust data demonstrate the good subtype inclusivity of the Elecsys® HIV combi PT assay and its suitability for screening and reliable early detection of HIV infection in Asia.

The Genomic Heterogeneity among Mycobacterium terrae Complex Displayed by Sequencing of 16S rRNA and hsp65 Genes

The species identification within Mycobacterium terrae complex has been known to be very difficult. In this study, the genomic diversity of M. terrae complex with eighteen clinical isolates, which were initially identified as M. terrae complex by phenotypic method, was investigated, including that of three type strains (M. terrae, M. nonchromogenicum, and M. triviale). 16S rRNA and 65‐kDa heat shock protein (hsp65) gene sequences of mycobacteria were determined and aligned with eleven other references for the comparison using similarity search against the GenBank and Ribosomal Database Project II (RDP) databases. 16S rRNA and hsp65 genes of M. terrae complex showed genomic heterogeneity. Amongst the eighteen clinical isolates, nine were identified as M. nonchromogenicum, eight as M. terrae, one as M. mucogenicum with the molecular characteristic of rapid growth. M. nonchromogenicum could be subdivided into three subgroups, while M. terrae could be subdivided into two subgroups using a 5 bp criterion (>1% difference). Seven isolates in two subgroups of M. nonchromogenicum were Mycobacterium sp. strain MCRO 6, which was closely related to M. nonchromogenicum. The hsp65 gene could not differentiate one M. nonchromogenicum from M. avium or one M. terrae from M. intracellulare. The nucleotide sequence analysis of 16S rRNA and hsp65 genes was shown to be useful in identifying the M. terrae complex, but hsp65 was less discriminating than 16S rRNA.

Evaluation of the nucleated red blood cell count in neonates using the Beckman Coulter UniCel DxH 800 analyzer

Introduction:u2002 Nucleated red blood cell (NRBC) count is closely associated with the prognosis of neonates. The analysis of NRBC has traditionally been measured manually. Recently, a newly developed automated hematology analyzer, the UniCel DxH 800 (DxH 800), was released. The goal of our study was to evaluate the performance of the DxH 800 NRBC method in neonates with the reference manual method and against previous generation hematology analyzers.

Carbapenem Resistance Mechanisms and Molecular Epidemiology of Acinetobacter spp. from Four Hospitals in Seoul and Gyeonggi Province in 2006

Background: Increasing numbers of Acinetobacter spp. resistant to multiple drugs, including carbape- nem, has been a serious problem. The aims of this study were to determine carbapenem resistance pat- terns and mechanisms, as well as to study the mo- lecular epidemiology of Acinetobacter spp. Methods: Clinical isolates of Acinetobacter spp. were collected from May to November in 2006. Antimicro- bial susceptibility testing was performed using CLSI disk diffusion and agar dilution methods. Metallo-β- lactamase- and OXA carbapenemase-producing iso- lates were detected by PCR. Carbapenem resistance and hydrolytic activities were compared according to OXA type and presence of ISAba1. Pulsed-field gel electrophoresis (PFGE) was performed to determine the epidemiologic features. Results: The imipenem non-susceptible rates were variable from 10% to 67%. Among 151 isolates car- rying blaOXA-51-like, 75 isolates carried both blaOXA-51-like and ISAba1, and 25 isolates had both blaOXA-51-like, blaOXA-23-like, and ISAba1. Carbapenem MICs of both blaOXA-51-like and ISAba1-carrying isolates were higher than those with blaOXA-51-like only. Carbapenem MICs of blaOXA-23-like-carrying isolates were higher than those with both blaOXA-51-like and ISAba1. Both blaOXA-51-like and ISAba1-carrying isolates and blaOXA-51-like, blaOXA-23-like, and ISAba1-carrying isolates demonstrated higher hydrolysis activities in oxacillin and carbapenems. Most of the tested isolates were susceptible to tige- cycline, and all of them were susceptible to colistin. Pulsed-field gel electrophoresis suggested that there had been several outbreaks of blaOXA-23-like and blaOXA-51-like-positive strains. Conclusion: Carbapenem non-susceptible Acineto- bacter isolates and OXA carbapenemase-producing isolates were prevalent. Dissemination of blaOXA-har- boring isolates may make it difficult to treat infections due to carbapenem-resistant Acinetobacter spp. Further surveillance studies are required to prevent the spread of carbapenem resistance. (Korean J Clin Microbiol 2010;13:27-33)

Concurrent MYC and MLL amplification on dmin and hsr in acute myeloid leukemia.

Genomic amplification is a frequent aberration in malignant proliferation that usually leads to an inappropriate expression of one or more oncogenes that are located within the amplicon. Cytogenetically, genomic amplification appears as a homogeneously staining region (hsr) or double minute chromosomes (dmin). In contrast to solid tumors [1], genomic amplification is rarely detected in hematological malignancies. The estimated incidence of cytogenetically detectable gene amplification in acute myeloid leukemia (AML) is *1% [2,3]. MYC is the most frequently amplified gene, but cases with MLL gene amplification have also been reported [2,4–6]. MLL is known as a regulator of growth of hematopoietic precursors [7], and MYC protein is known to be a nuclear transcription factor [8]. The genes are regarded as proto-oncogenes and amplification of them is associated with aggressive growth and poor prognosis. We report here a case of concurrent MYC and MLL amplification in AML on dmin and hsr, respectively, in a same clone. A 68-year-old male patient presented with a 2week history of progressive dyspnea and low back pain. His past medical history was notable only for a herniated nucleus pulposus that was treated by surgery 5 years prior, and there was no prior history of toxic or radiation exposure. Laboratory data on admission revealed an Hgb level of 8.6 g/dL, platelet count of 13 000/mL, and a WBC count of 1100/mL with 11% blasts. The bone marrow biopsy revealed hypercellular marrow, composed of blasts with a heterogenous morphology. The profound red cell abnormalities, hypogranular neutrophils and megakaryocytic atypia suggested the possibility of a preexisting myelodysplastic syndrome. Flow cytometry of the bone marrow identified a blast population (49.8%) that expressed myelomonocytic markers including CD13, CD14, CD33, CD45 and HLADR. Cytochemically, the blast cells were strongly positive for peroxidase and a-naphthyl-butyrate esterase. The patient was confirmed as FAB subtype M4 as determined by the morphological, cytochemical and immunophenotypical classification of a BM specimen. G-banding analysis showed that all of 20 metaphase cells had both numerical and structural abnormalities, including questionable regions in 1p32 (hsr) and 8q24.2 (deletion), trisomy 6 and 17, and monosomy 11 and 21. Sixteen of the 20 cells (80%) also had 1–14 dmin in each cells [Figure 1(A)]. Considering that MYC is located at 8q24.2 and that the hsr and dmin represent a form of gene amplification, we performed fluorescence in situ hybridization (FISH) using a CEP8/MYC/IGH probe set (Vysis, Downers Grove, IL). Furthermore, considering that the AML-M4 subtype should be evaluated whether 11q23/MLL rearrangement exists or not, and our patient showed monosomy11, we

Therapy-related acute megakaryoblastic leukemia in a lung cancer patient.

Therapy-related AML (t-AML) is one of the newly expanded disease entities in the 2008 WHO classification, accounting for 10-20% of all cases of AML, and its incidence is increasing worldwide because of improved survival rates following treatment for other primary cancers [1, 2]. Acute megakaryoblastic leukemia (AMKL) (M7) is the least common of the t-AML French-American-British (FAB) subtypes, and only two such cases have been reported to date [3, 4], neither of which was in Korea. AMKL accounts for about 7-10% of childhood AML cases (frequently associated with Down syndrome), but only about 1% of adult AML cases [5]. Here, we describe a rare case of therapy-related acute megakaryoblastic leukemia (t-AMKL) with chromosome 5 and 7 abnormalities that presented ten years after chemoradiotherapy in an elderly lung cancer patient.