Paradigm for diagnosing mycobacterial disease: direct detection and differentiation of complex and non-tuberculous mycobacteria in clinical specimens using multiplex real-time PCR.

Aims: and non-tuberculous mycobacteria (NTM) are clinically different, and the rapid detection and differentiation of complex (MTBC) and NTM is crucial for patient management and infection control. Given the slow growth of most pathogenic mycobacteria, nucleic acid amplification assays are excellent tools for direct identification of mycobacteria in clinical specimens. Recently, a multiplex real-time PCR assay was developed that can directly detect 20 mycobacterial species in clinical specimens.

Direct identification of mycobacteria from liquid media using a triplex real-time PCR coupled with pyrosequencing method.

Culture in enriched broth, as well as on a solid medium, is recommended for primary isolation of mycobacteria. With the introduction of liquid mycobacterial culture methods, a substantial workload regarding the identification of culture-recovered mycobacterial species, particularly Mycobacterium tuberculosis complex (MTC), has been imposed on our laboratory. We thus developed a triplex, real-time PCR coupled with pyrosequencing assay that can directly identify mycobacterial species from liquid media, which can reduce the workload.

A challenging diagnosis: crystal-storing histiocytosis in plasma cell myeloma.

Objectives: Crystal-storing histiocytosis (CSH) is an uncommon finding in plasma cell neoplasms. CSH is thought to be an intralysosomal deposition of secreted paraproteins or immunoglobulins, which usually express κ immunoglobulin light chains that finally aggregate in crystals. Because of its rarity, CSH in bone marrow often makes diagnosis difficult.

CD34 and p53 immunohistochemical stains differentiate hypocellular myelodysplastic syndrome (hMDS) from aplastic anemia and a CD34 immunohistochemical stain provides useful survival information for hMDS.

Background: The presence of significant dysplasia in bone marrow (BM) aspirates helps to distinguish between hypocellular myelodysplastic syndrome (hMDS) and aplastic anemia (AA). Occasionally, diluted BM aspirates make it difficult to recognize dysplastic changes and can also negatively affect the detection of cytogenetic abnormalities in hMDS. We evaluated the usefulness of CD34 and p53 immunoreactivity for discriminating between hMDS and AA and for estimating survival outcomes in hMDS patients.

Direct identification of mycobacteria from culture media using a multiplex real-time PCR assay: report on its application in a clinical laboratory in a region of high tuberculosis endemicity.

There are few commercial assays that easily and correctly identify the mycobacteria from culture in a clinical laboratory with a high workload. Thus, we developed and evaluated a scheme for the identification of mycobacteria using a multiplex real-time PCR assay and report on its application in our laboratory. The scheme consisted of 3 stepwise PCRs.

Male breast adenoid cystic carcinoma.

Introduction: Adenoid cystic carcinoma (ACC) of the breast is a rare condition, and cases in male patients are even less common.

Case: We describe a case of ACC of the breast with axillary lymph node metastasis, disseminated osteolytic bone metastasis and bone marrow involvement in a 41-year-old man.

Conclusion: Male breast ACC is an extremely rare malignancy; there can be difficulty in obtaining a final diagnosis.

Multiplex real-time PCR assay for detection of methicillin-resistant Staphylococcus aureus (MRSA) strains suitable in regions of high MRSA endemicity.

A multiplex real-time PCR assay that simultaneously detects the mecA, staphylococcal cassette chromosome (SCCmec)-open reading frame X (orfX) junction, and staphylococcal 16S rRNA genes was developed and evaluated using 444 staphylococcal strains. We demonstrated that this assay resulted in fewer false-positive results than a single-locus real-time PCR assay that amplified the SCCmec-orfX junction. This assay would be useful in a clinical laboratory in a region of high endemicity for methicillin-resistant Staphylococcus aureus (MRSA) infections.

A combination of CD15/CD10, CD64/CD33, CD16/CD13 or CD11b flow cytometric granulocyte panels is sensitive and specific for diagnosis of myelodysplastic syndrome.

Flow cytometry (FCM) is a reproducible and objective technique that may be useful in the diagnosis of myelodysplastic syndrome (MDS) by detecting abnormal immunophenotypes specific to MDS. We investigated 5 granulocyte/monocyte panels by FCM to find a sensitive and specific combination of panels in order to discriminate MDS from non-clonal hematologic disorders. Bone marrow aspirates from 35 patients with MDS and 25 patients with non-clonal hematologic disorders were studied.

Multiplex real-time PCR assay and melting curve analysis for identifying Mycobacterium tuberculosis complex and nontuberculous mycobacteria.

A multiplex real-time PCR assay and melting curve analysis for identifying 23 mycobacterial species was developed and evaluated using 77 reference strains and 369 clinical isolates. Concordant results were obtained for all 189 (100%) isolates of the Mycobacterium tuberculosis complex and 169 (93.9%) isolates of nontuberculous mycobacteria.

[Minimal residual disease detection in acute leukemia patients by flow cytometric assay of cross-lineage antigen expression].

Background: It has been demonstrated that flow cytometric detection of minimal residual disease (MRD) has a prognostic significance in the treatment of patients with acute leukemia. We investigated the significance of flow cytometric MRD detection for the first time in Korea.

Methods: We analyzed the results of MRD detection in morphologically complete remission bone marrow aspirates from 89 patients with newly-diagnosed or relapsed acute leukemia, in which leukemic cells had cross-lineage antigen expression.

Evaluation of the TEST 1 erythrocyte sedimentation rate system and intra- and inter-laboratory quality control using new latex control materials.

Background: The erythrocyte sedimentation rate (ESR) test has been considered to be a simple procedure, not requiring quality control (QC). However, QC is essential for accuracy and precision. We evaluated the TEST 1 ESR system and performed QC procedures using newly developed latex control materials in three hospitals.

[Detection of vancomycin-resistant enterococci using multiplex real-time PCR assay and melting curve analysis].

Background: We developed and evaluated the utility of a multiplex real-time PCR assay that uses melting curve analysis and allows simultaneous identification of vancomycin-resistant genotypes and clinically relevant enterococci.

Methods: The specificity of the assay was tested using 4 reference strains of vancomycin-resistant enterococci (VRE) and 2 reference strains of vancomycin-susceptible enterococci. Ninety-three clinical isolates of enterococci with different glycopeptide-resistant phenotypes were genotyped and identified using a multiplex real-time PCR assay and melting curve analysis.

[Subgenotype and serotype analysis of hepatitis B virus in Korean chronic hepatitis B patients under treatment].

Background: Hepatitis B virus (HBV) detected in Korean patients almost belongs to genotype C, which is subdivided into subgenotype C1 (or Cs) and C2 (or Ce). It was recently reported that the risk of hepatocellular carcinoma is different between subgenotype C1 and C2. Thus, we studied the distribution of subgenotypes of HBV in Korean chronic hepatitis B (CHB) patients.

Erythrocyte sedimentation rate measurements by TEST 1 better reflect inflammation than do those by the Westergren method in patients with malignancy, autoimmune disease, or infection.

We compared the TEST 1 (Alifax, Padova, Italy) and Westergren methods of measuring the erythrocyte sedimentation rate (ESR) to assess inflammation. The ESR was measured by both methods in 154 blood samples from patients with malignancy (n = 69), autoimmune disease (n = 44), or infection (n = 41). Total protein, albumin, and C-reactive protein (CRP) levels were measured in each plasma sample, and albumin and alpha(1)-, alpha(2)-, beta(1)-, beta(2)-, and gamma-globulin fractions were measured by capillary electrophoresis.

[Experience on identification of cross-reactive group specificity performed by anti-human globulin panel reactive antibody tests].

Background: Panel reactive antibody (PRA) is to screen and identify HLA antibody. Majority of antibody specificities in high-PRA are directed against cross reactive group (CREG). Thus, this study was to know the advantage of identifying CREG specificity and whether antibody specificities are changed according to CREG classification.

Identification of hepatitis C virus genotype 6 in Korean patients by analysis of 5' untranslated region using a matrix assisted laser desorption/ionization time of flight-based assay, restriction fragment mass polymorphism.

Previous surveys of the prevalence of hepatitis C virus (HCV) in Korea have identified types 1 and 2, but little has been said of other genotypes and viral subtypes. In this study, HCV genotypes in Korea were investigated using Restriction Fragment Mass Polymorphism (RFMP) assay, a sensitive and specific method for genotyping based on MALDI-TOF mass spectrometry. A total of 1,043 independent serum samples from HCV-infected patients were analyzed.

[Performance evaluation of Abbott RealTime HBV Quantification Kit for HBV viral load by real-time PCR].

Background: Hepatitis B virus (HBV) DNA quantification is necessary for starting and monitoring of antiviral therapy in patients with chronic hepatitis B. This study was intended to assess the clinical performance of Abbott RealTime HBV Quantification kit (Abbott Laboratories, USA).

Methods: The performance was evaluated in terms of precision, linearity, detection sensitivity, cross-reactivity, and carry-over.

[Development of a system for extracting the information of candidate tumor markers reported in biomedical literatures].

Background: Since the human genome project was completed in 2003, there have been numerous reports on cancer and related markers. This study was aimed to develop a system to extract automatically information regarding the relationship between cancer and tumor markers from biomedical literatures.

Methods: Named entities of tumor markers were recognized by both a dictionary-based method and machine learning technology of the support vector machine.

Meta-analysis of the association between HLA-DRB1 allele and rheumatoid arthritis susceptibility in Asian populations.

The aims of this study were to summarize results on the association of HLA-DRB1 with rheumatoid arthritis (RA) in Asians and to determine if the shared epitope (SE) hypothesis could explain the meta-analysis results. Among the papers published between January 1987 and July 2006 on RA susceptibility in Asian-Mongoloid populations (Korean, Japanese, Chinese, and Thai), 12 were selected for the metaanalysis. Mongoloid-Asian patients with RA had significantly higher frequencies of HLA-DRB1*0101, *0401, *0410, and *1001 than controls (OR 1.

[Development of a web-based program for the identification of human leukocyte antigen antibody specificities].

Background: Panel reactive antibody (PRA) test is used to determine whether a patient awaiting transplantation is previously sensitized. Tail analysis algorithm is widely used to identify antibody specificities, but it is very difficult to perform manually.

Methods: To develop a web-based program, PHP (5.

[Performance evaluation of real-q HBV quantification kit for HBV DNA by real-time PCR.].

Background: Hepatitis B virus (HBV) DNA quantification is important for the management of HBV infection and identification of the development of resistance. The susceptibility to contamination and more variable reproducibility of results with the conventional HBV DNA quantification method have raised the need of a more simple and accurate method for HBV DNA quantification. Real-time quantitative PCR assays recently introduced in the laboratory can meet these needs.

[Evaluation of BioSewoom HLA-A, -B, -C PCR/SSP kit].

Background: Human leukocyte antigen (HLA) typing based on polymerase chain reaction (PCR) is rapidly replacing the conventional serological method. This study was intended to evaluate BioSewoom HLA-A, -B, -C PCR/SSP kit (BioSewoom SSP) which had recently been developed in Korea.

Methods: A total of 158 samples from domestic (21) and international (137) HLA proficiency testing (PT) were genotyped with BioSewoom SSP, and its results were compared to consensus results.

Significantly better prognosis for patients with primary plasma cell leukemia than for patients with secondary plasma cell leukemia.

Plasma cell leukemia (PCL) is a rare variant of multiple myeloma (MM). Patients may either present de novo (primary PCL), or PCL may occur during the course of MM (secondary PCL). We compared the laboratory and clinical findings of both primary and secondary PCL and MM to elucidate their natural history and the relationship among these entities.