Choong-Hwan Cha

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 meta-analysis. Mongoloid-Asian patients with RA had significantly higher frequencies of HLA-DRB1*0101, *0401, *0410, and *1001 than controls (OR 1.5-2.1, p<0.05 for association). When analyses were restricted to more ethnically homogeneous populations, HLA-DRB1*0405 showed a significant susceptibility to RA in Koreans (OR 5.65, 95% CI 4.32-7.39), whereas the HLA-DRB1*0301, *0403, *0406, *0701, *1301, and *1405 alleles showed protective association with RA (OR 0.32-0.70, p<0.05 for association). In conclusion, it was found that HLA-DRB1 *0101, *0401, *0405, *0410, and *1001 are susceptible, while HLA-DRB1*0301, *0403, *0406, *0701, *1301, and *1405 are protective in Asian-Mongoloids. All the RA-associated alleles except DRB1*0301 could be explained by the structural model supporting the SE hypothesis that RA susceptibility is determined by the combination of amino acid residues at HLA-DR β71 and β74, not by β71 alone.

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. Of interest, 15 subjects (1.4%) were determined to contain HCV genotype 6 and 46 subjects (4.4%) contained mixed genotypes with the most prevalent genotypes being HCV 1b and 2a/c (45.0% and 35.4%, respectively). The 15 subjects with HCV genotype 6 comprised eight cases of subtype 6c, including one case of mixed infection with 1b, three cases of HCV 6a, and six cases of unassigned subtypes. Sequencing corroborated the identity of genotype 6 from 13 subjects, while the line probe assay (LiPA) mis‐identified them as genotype 1b. The majority (7/9) of the genotype 6 patients enrolled for interferon/ribavirin therapy, achieved a sustained virologic response. The ability of the RFMP assay to differentiate various HCV genotypes should enable better analysis of the relationship between HCV genotype and disease prognosis. J. Med. Virol. 80:1712–1719, 2008.

MICA polymorphisms and haplotypes with HLA-B and HLA-DRB1 in Koreans

Major histocompatibility complex (MHC) class I chain-related gene A (MICA) is located within the human MHC, centromeric to HLA-B and telomeric to HLA-DRB1. The location of MICA in the MHC indicates the presence of linkage disequilibrium with human leukocyte antigen (HLA). Like HLA, MICA is highly polymorphic; however, the information available for MICA polymorphisms is not as comprehensive as that for HLA polymorphisms. We estimated the allelic frequencies of MICA and haplotypes with HLA-B and HLA-DRB1 at high-resolution in a population of 139 unrelated Korean individuals by applying the newly developed method of sequence-based typing (SBT). A total of 17 MICA alleles were identified. The most frequent allele was MICA*010 (19.4%), followed by alleles *00201 (17.6%), *00801 (14.7%), *01201 (9.4%), *004 (8.3%) and *049 (7.9%). The most common two- and three-locus haplotypes were HLA-B*1501-MICA*010 (10.4%), MICA*010-HLA-DRB1*0406 (5.8%) and HLA-B*1501-MICA*010-HLA-DRB1*0406 (5.8%). This is the first study to provide such high-resolution information on the distribution of haplotypes comprising MICA, HLA-B and HLA-DRB1 in Korean individuals, a level of resolution made possible by use of the SBT method. The results of this study should help determine the mechanisms underlying diseases associated with MICA polymorphisms in Korean individuals.

Multiplex Real-Time PCR Assay and Melting Curve Analysis for Identifying Mycobacterium tuberculosis Complex and Nontuberculous Mycobacteria

ABSTRACT 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. Our results showed that this multiplex real-time PCR assay is an effective tool for the mycobacterial identification from cultures.

MICB polymorphisms and haplotypes with MICA and HLA alleles in Koreans

Major histocompatibility complex (MHC) class I chain-related gene B (MICB) is located within the human MHC class I region. The location of MICB in the MHC region may imply the presence of linkage disequilibrium with polymorphic MICA and human leukocyte antigen (HLA) loci. MICB is also polymorphic; however, MICB polymorphisms have not been investigated in Koreans. Using sequence-based typing (SBT), we estimated the allelic frequencies of MICB and haplotypes with MICA, HLA-B, and HLA-DRB1 at high resolution in a population of 139 unrelated Korean individuals. Eight MICB alleles were identified. The most frequent allele was MICB*005:02/*010 (57.2%), followed by *002 (11.5%), *004 (8.3%), *005:03 (8.3%), and *008 (6.8%). The most common two-locus haplotypes were MICB*005:02/*010-MICA*010 (19.4%), MICB*005:02/*010-DRB1*15:01 (6.5%), and MICB*005:02/*010-B*15:01 (10.4%); the most common three-locus haplotypes were B*15:01-MICA*010-MICB*005:02/*010 (5.8%) and MICA*010-MICB*005:02/*010-DRB1*04:06 (10.4%); and the most common four-locus haplotype was B*15:01-MICA*010-MICB*005:02/*010-DRB1*04:06 (5.8%). This is the first study to provide information about MICB allele frequencies and haplotypes with HLA in Koreans. These study results should help understand mechanisms of disease association between the MICB locus and neighboring loci in Koreans.

Multiplex Real-Time PCR Assay for Detection of Methicillin-Resistant Staphylococcus aureus (MRSA) Strains Suitable in Regions of High MRSA Endemicity

ABSTRACT 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.

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. RESULTS Representative melting curves were obtained for Enterococcus faecium, Enterococcus faecalis, vanA-containing E. faecium, vanB-containing E. faecalis, Enterococcus gallinarum, and Enterococcus casseliflavus. Phenotypic and genotypic analysis of the isolates revealed same results for 82 enterococcal isolates, while in 4 isolates, the glycopeptide-resistant phenotypes were inconsistent with the glycopeptide-resistant genotypes and in the 4 other isolates, species could not be accurately identified. Three isolates with mixed strains, which were detected by the PCR assay, could not be correctly identified using phenotypic methods. CONCLUSIONS VRE genotyping and identification of clinically relevant enterococci were rapidly and correctly performed using multiplex real-time PCR assay and melting curve analysis.

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. Mycobacterium tuberculosis complex (MTC) and nontuberculous mycobacteria (NTM) were differentially detected in the step 1 PCR, and the NTM species were identified in the step 2 and 3 PCRs. Over the 1.5-year study period, 1136 isolates of MTC and 618 isolates of NTM were detected, and the species of 608 (98.4%) of the 618 NTM isolates were identified. We conclude that the established scheme is a very useful diagnostic approach for the rapid and accurate identification of MTC and clinically relevant NTM in a clinical laboratory in a region where tuberculosis is endemic.

HLA‐C*03:93, a novel HLA‐C*03 allele identified by sequence‐based typing

The new allele C*03:93 showed one nucleotide difference with C*03:04:01 at codon 140 (GCT/ACT).

Direct identification of mycobacteria from clinical specimens by multiplex real-time PCR.

To directly identify clinically relevant mycobacteria from clinical specimens, we have developed a multiplex real‐time PCR assay with hydrolysis probes that can identify 20 mycobacterial species.