Sook Young Bae

Enhanced detection of respiratory viruses using cryopreserved R-Mix ReadyCells

BACKGROUND R-Mix, which contains a fresh mixture of two cell lines, Mv1Lu (mink lung cells) and A549 cells, has shown good sensitivity and specificity for respiratory virus culture. However, it has until recently only been available in North America, in part due to the shipping constraints associated with cell aging and the difficulty in providing these cells to hard to reach regions. Recently, cryopreserved R-Mix ReadyCells for longer storage were developed. These cells, which are shipped on dry ice and have a shelf life as long as 6 months from date of manufacture, can be thawed and used as needed with minimal addition of refeeding media. OBJECTIVE Assess the potential for cryopreserved R-Mix ReadyCells to replace conventional culture. STUDY DESIGN Two hundred and twenty-three nasopharyngeal aspirates confirmed as respiratory virus-positive by conventional culture were inoculated into cryopreserved R-Mix ReadyCells and re-inoculated into conventional culture cells simultaneously. After 1 and 3 days of incubation cryopreserved R-Mix ReadyCells and conventional culture cells were screened using a respiratory virus fluorescent antibody pool for the detection of seven major respiratory viruses (influenza A and B viruses, parainfluenza 1, 2 and 3 viruses, respiratory syncytial virus and adenovirus). Positive pool results were further differentiated with specific monoclonal antibodies against the individual viruses. RESULTS After 1 day of incubation detection rates for conventional culture were 25%, 39%, 39%, 49%, and 10% for influenza A virus, influenza B virus, parainfluenza viruses, respiratory syncytial virus, and adenovirus, respectively. Corresponding detection rates for cryopreserved R-Mix ReadyCells were 78%, 91%, 72%, 81%, and 65%. Average detection rates of cryopreserved R-Mix ReadyCells for all respiratory viruses were 80% after 1 day incubation and 95% after 3 days incubation, compared to 35% and 70% by conventional culture. CONCLUSION The cryopreserved R-Mix ReadyCells system offers a highly sensitive and rapid method for detection of respiratory viruses that may allow it to replace conventional cell culture systems.

Absolute CD4+ cell count using a plastic microchip and a microscopic cell counter

We have designed and evaluated the performance of a simple, rapid, and affordable method for counting CD4+ T‐cells with the use of plastic microchips. This new system is an adaptation of a “no‐lyse, no‐wash,” volumetric single platform assay, and absolute CD4+ counts are determined with the use of a microscopic scanning cell counter. To assess the CD4+ count test precision and linearity of the system, measured CD4+ counts were compared with two other reference assays (single and dual platform flow cytometry) with the use of 123 clinical samples including samples obtained from 35 HIV‐infected patients, and artificially diluted samples. A correlation between the results from the use of the new method and from the use of the two other reference assays was r = 0.98 for the clinical samples. A dilution test of the new method demonstrated a linearity of r ≥ 0.99, with coefficients of variation ≤7.6% for all concentration levels. Our findings suggest that the new CD4+ counting device can be potentially be applied for other diagnostic procedures that measure quantities of characteristic antigens or other materials on cells.

Alteration of platelet counts and lipid profiles after treatment of acute Plasmodium vivax

During malaria infections, thrombocytopenia and low cholesterol levels are frequently observed changes. We compared these changes in patients admitted with fevers and infected with Plasmodium vivax, patients admitted with fevers with respiratory/urinary infections and afebrile normal (control) non-infected volunteers. Changes in the platelet count and lipid parameters are reported for malaria patients after treatment with hydroxychloroquine and primaquine for acute P. vivax malaria. Of a total 141 participants, 55 patients were diagnosed with malaria (positive blood smear) prior to treatment. Compared to the normal (n=52) and non-malaria fever groups (n=34), there was a significant decrease in five hematologic indices (white blood cell, red blood cell, hemoglobin, hematocrit and platelet) and three lipid parameters (total cholesterol, HDL-c and LDL-c) in the vivax malaria group at day 0 (pre-treatment). Following treatment, the platelet counts returned to normal limits (P<0.05) from 91,058/microL on day 0 to 246,833/microL by day 17 after treatment. However, changes in the lipid parameters of malaria patients showed a slow recovery to normal limits compared to the platelet counts. The HDL-c and LDL-c remained low for 1 month after treatment but increased at 3 and 6 months post-treatment. At 12 months after treatment, the levels of two lipid parameters had fully recovered to the normal limits. Thus, special attention should be applied when interpreting laboratory blood profiles of malaria patients, especially platelet and lipid based tests, until full recovery after treatment.

Acute myeloid leukemia (AML-M2) associated with variant t(8;21): report of three cases

Variants of the t(8;21)(q22;q22) involving chromosome 8, 21, and other chromosomes account for approximately 3% of all t(8;21)(q22;q22) found in patients with acute myeloid leukemia (AML). The clinicopathologic features of AML with the variant t(8;21) have not been well established. We report three cases of AML with variants of t(8;21) characterized, respectively, by derivative 8 with the interstitial inverted insertion of 21q and concurrent monosomy 21, t(8;18;21)(p22;q11.3;q22), and t(2;21;8)(q11.2;q22;q22). Fluorescence in situ hybridization or reverse transcriptase-polymerase chain reaction assay confirmed the presence of RUNX1-RUNX1T1 gene (previously AML1-ETO) rearrangements. Among these cases, three-way breakpoints 18p11.3 and 2q11.2 have not been previously reported. The present report deals with the results of hematologic, immunophenotypic, cytogenetic, fluorescence in situ hybridization, and molecular analyses of these variants. The possible role of the genes in this region in leukemogenesis, response to treatment, and clinical implications are discussed.

Hypereosinophilia in biphenotypic (B-cell/T-cell) acute lymphoblastic leukemia.

The association of acute lymphoblastic leukemia (ALL) and symptomatic eosinophilia is rare and only a few cases have been reported [1-5]. Biphenotypic acute leukaemias are also uncommon, and genera...

Identification of proteolipid protein 1 gene duplication by multiplex ligation-dependent probe amplification: First report of genetically confirmed family of pelizaeus-merzbacher disease in Korea

Pelizaeus-Merzbacher disease (PMD) is a rare X-linked recessive disorder with a prototype of a dysmyelinating leukodystrophy that is caused by a mutation in the proteolipid protein 1 (PLP1) gene on the long arm of the X chromosome in band Xq22. This mutation results in abnormal expression or production of PLP. We here present a Korean boy with spastic quadriplegia, horizontal nystagmus, saccadic gaze, intentional tremor, head titubation, ataxia, and developmental delay. The brain magnetic resonance imaging (MRI) showed abnormally high signal intensities in the white matter tract, including a subcortical U fiber on the T2-weighted and fluid attenuated inversion recovery (FLAIR) image. The chromosomal analysis was normal; however, duplication of the PLP1 gene in chromosome Xq22 was detected when the multiplex ligation-dependent probe amplification (MLPA) method was used. We also investigated the pedigree for a genetic study related to PMD. This case suggests that the duplication mutation of the PLP1 gene in patients with PMD results in a mild clinical form of the disorder that mimics the spastic quadriplegia of cerebral palsy.

Standardization of Terminology in Laboratory Medicine II

Standardization of medical terminology is essential in data transmission between health care institutes and in maximizing the benefits of information technology. The purpose of this study was to standardize medical terms for laboratory observations. During the second year of the study, a standard database of concept names for laboratory terms that covered those used in tertiary health care institutes and reference laboratories was developed. The laboratory terms in the Logical Observation Identifier Names and Codes (LOINC) database were adopted and matched with the electronic data interchange (EDI) codes in Korea. A public hearing and a workshop for clinical pathologists were held to collect the opinions of experts. The Korean standard laboratory terminology database containing six axial concept names, components, property, time aspect, system (specimen), scale type, and method type, was established for 29,340 test observations. Short names and mapping tables for EDI codes and UMLS were added. Synonym tables were prepared to help match concept names to common terms used in the fields. We herein described the Korean standard laboratory terminology database for test names, result description terms, and result units encompassing most of the laboratory tests in Korea.

A Case of Haemophilus parainfluenzae Endocarditis

The HACEK group of bacteria (Haemophilus parainfluenzae, H. aphrophilus, H. paraphrophilus, Actinobacilus actinomycetemcomitans, Cardiobacterium hominis, Eikenella corodens, and Kingella kingae) are the normal flora of the upper respiratory tract and oropharynx. The organisms infect abnormal cardiac valves, causing subacute native endocarditis or prosthetic valve endocarditis more than one year after valve surgery. Haemophilus species are responsible for only 0.5∼1% of all infective endocarditis cases. Embolization occurs in 60% and the mortality rate ranges from 16∼45% of cases of infective endocarditis caused by H. parainfluenzae. We experienced a case of infective endocarditis due to H. parainfluenzae in a 37-year-old male admitted with high fever, chills, nausea & vomiting, chest discomfort, and blurred vision. The organism was isolated from a blood culture and was identified as H. parainfluenzae by factor V requirement, negativity at urea, positivity at ornithine decarboxylase, and acid production from glucose and maltose. The patient was treated with antibiotics and symptoms and signs were improved. (Korean J Clin Microbiol 2009;12:78-81)

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

Development of correction methods for variable pinhole single-photon emission computed tomography

We propose a novel pinhole collimator in which the pinhole shape can be changed in real-time, and a new single-photon emission computed tomography (SPECT) system that utilizes this variable pinhole (VP) collimator. The acceptance angle and distance between the collimator and the object of VP SPECT are varied so that the optimum value of the region-of-interest (ROI) can be obtained for each rotation angle. Because of these geometrical variations, new correction methods are required for image reconstruction. In this study, we developed two correction methods. The first is the sensitivity-correction algorithm, which minimizes the variation of a system matrix caused by varying the acceptance angle for each rotation angle. The second is the acquisition-time-correction method, which reduces the variation of uniformity caused by varying the distance between the collimator and the object for each rotation angle. A 3D maximum likelihood expectation maximization (MLEM) algorithm was applied to image reconstruction, and two digital phantoms were studied to evaluate the resolution and sensitivity of the images obtained using the proposed methods. The images obtained by using the proposed correction methods show higher uniformity and resolution than those obtained without using these methods. In particular, the results of the resolution phantom study show that hot rods (0.8-mm-diameter) can be clearly distinguished using the proposed correction methods. A quantitative analysis of the ROI phantom revealed that the mean square error (MSE) was 0.42 without the acquisition-time-correction method, and 0.04 with the acquisition-time-correction method. The MSEs of the resolution phantom without and with the acquisition-time-correction method were calculated as 55.14 and 14.69, respectively.