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Lee G.-C.,Water Analysis and Research Center | Jung G.S.,Chungbuk National University | Lee C.H.,Chungbuk National University
Virus Genes

The complete nucleotide and deduced amino acid sequences of the RNA genome of a recently isolated norovirus (NoV) from Korea, designated Hu/GII-4/CBNU2/ 2007/KR (CBNU2), were determined and characterized by phylogenetic comparison with several genetically diverse NoV sequences. The RNA genome of CBNU2 is 7,560 nucleotides in length, excluding the 30 poly (A) tract. It includes three open reading frames (ORFs): ORF1, which encodes the nonstructural polyprotein (5-5,104); ORF2, which encodes VP1 (5,085-6,707); and ORF3, which encodes VP2 (6,707-7,513). ORF2-based phylogenetic analysis revealed that CBNU2 belonged to the GII.4 genotype, the most prevalent genotype, and formed a cluster with NoVs isolated from Asian regions, between 2006 and 2008. Comparative analysis with the consensus sequence of 207 completely sequenced NoV genomes showed 47 mismatched nucleotides: 26 in ORF1, 14 in ORF2, and 7 in ORF3, resulting in 8 amino acid changes: 3 in ORF1, 2 in ORF2, and 3 in ORF3. Phylogenetic analysis with full genome ORF1, ORF2, and ORF3 nucleotide sequences obtained from CBNU2 and each of the other representative NoV genomes suggested that CBNU2 had not undergone recombination with any of the other NoVs. A SimPlot analysis further supported this finding. © Springer Science+Business Media, LLC 2012. Source

Lee G.-C.,Water Analysis and Research Center | Jeon E.-S.,Chungbuk National University | Kim W.-S.,Chungbuk National University | Le D.T.,Research Team for Vectorborne Diseases | And 2 more authors.
Virology Journal

Background. This study evaluated the clinical accuracy and analytical sensitivity of the NanoSign® Influenza A/B antigen kit in detecting 2009 pandemic influenza A/H1N1 viruses. The kit is one of the most popular rapid diagnostic tests for detecting influenza in Republic of Korea. Results. The NanoSign® Influenza A/B kit resulted in 79.4% sensitivity and 97.2% specificity compared to RT-PCR in the detection of the viruses from 1,023 specimens. In addition, the kit was able to detect two strains of novel influenza viruses, Influenza A/California/12/2009(H1N1) and clinically isolated wild-type novel influenza A/H1N1, both of which are spreading epidemically throughout the world. In addition, the correlation between NanoSign® Influenza A/B test and conventional RT-PCR was approximately 94%, indicating a high concordance rate. Analytical sensitivity of the kit was approximately 73 3.65 ng/mL of the purified viral proteins and 1.13 0.11 hemagglutination units for the cultured virus. Conclusions. As the NanoSign® Influenza A/B kit showed relatively high sensitivity and specificity and the good correlation with RT-PCR, it will be very useful in the early control of influenza infection and in helping physicians in making early treatment decisions. © 2010 Lee et al; licensee BioMed Central Ltd. Source

Lee G.-C.,Water Analysis and Research Center | Lee J.H.,Catholic Kwandong University | Kim B.Y.,Chungbuk National University | Lee C.H.,Chungbuk National University
Journal of Microbiology and Biotechnology

Mitochondria often play central roles in apoptotic pathways, and disruption of the mitochondrial transmembrane potential (ΔΨm) has been observed in various cells undergoing apoptosis. Human cytomegalovirus (HCMV) infection induces apoptosis in permissive cells; however, investigations of mitochondria-targeted apoptosis in HCMV-infected human foreskin fibroblast (HFF) cells have been limited. Here, we investigated the mitochondrial apoptosis pathway in HCMV-infected HFF cells. Flow cytometry analysis using JC-1 revealed that HCMV infection induces disruption of ΔΨm in HFF cells when administered 24 h postinfection (hpi), and this disruption was maximized at 48 hpi. Moreover, cytochrome c, normally a mitochondrial inner membrane protein, was detected in cytoplasmic extracts of HCMV-infected cells, but not mock-infected cells, by western blot analysis at 24 hpi. A caspase activity assay based on fluorescence spectrophotometry using a fluorogenic substrate revealed an increase in caspase-3 activity at 48 hpi in HCMV-infected cells. Caspase-8 activity was increased at 72 hpi in HCMV-infected cells. These results imply that HCMV infection induces mitochondria-mediated apoptosis in HFF cells. © 2013 by The Korean Society for Microbiology and Biotechnology. Source

Lee J.H.,Catholic Kwandong University | Lee G.-C.,Water Analysis and Research Center | Kim J.I.,Chungbuk National University | Yi H.A.,Chungbuk National University | Lee C.H.,Chungbuk National University
Journal of Virological Methods

The development of rapid and effective methods to detect water- and food-borne enteric viruses is important for the prevention and control of mass infection. This study represents an attempt to develop a reliable cell culture-based detection system and optimize an effective and rapid protocol for the assaying of environmental samples for the presence of infectious enteric viruses. Six enteric viruses were used in this study: poliovirus, Coxsackie virus A9, Coxsackie virus B5, human rotavirus G1, hepatitis A virus, and adenovirus type 41. Among the cell lines from humans (A549, HeLa, HEK293, and HFF) and other primates (Vero, BS-C-1, FRhK-4, BGMK, and MA104), a cytopathic effect (CPE) analysis indicated that the MA104 cell line was the most optimal for use in the detection of infectious enteric viruses. Both the sensitivity and specificity of virus detection in MA104 cells were similar to or higher than those in standard BGMK cells. Next, a method was developed for the determination of the infectiousness of enteric viruses using the colorimetric thiazolyl blue (MTT) assay. This assay utilizes 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide to yield % values based on colorimetric results. These results were compared with those from a conventional CPE-based TCID50 assay, revealing no statistically significant difference between the two methods. The MTT% values in MA104 cells were comparable to those in BGMK cells. This MA104 cell-based MTT assay could substitute for the classical BGMK cell-based CPE assay for infectious enteric viruses. © 2013 Elsevier B.V. Source

Nam S.,Chungnam National University | Kim M.-J.,Water Analysis and Research Center | Park C.,Catholic University of Korea | Park J.-G.,Daegu University | And 2 more authors.
International Journal of Hygiene and Environmental Health

The distribution characteristics of Enterococcus spp., which are indicators of fecal pollution, were investigated at 33 sites within the 3 major water systems of Korea. Enterococci were detected at concentrations ranging from 1 to 37 CFU/100. mL in 41 of 132 samples (31.1%) from the 3 major water systems. The overall average detected concentration was 1.2 CFU/100. mL, while the average concentration for all detection sites was 5.3 CFU/100. mL. After optimized multiplex polymerase chain reaction (PCR) was performed with newly developed VanA, VanB, VanC-1, and VanC-2/3 primers, concentrations of vancomycin-resistant Enterococcus spp. (VRE) ranging from 1 to 23 CFU/100. mL were detected in 17 of 132 samples (12.9%). Of 216 individual enterococcal colonies, 64 (29.6%) displayed the VanC genotype. The results of a susceptibility test to vancomycin showed that the range of the minimal inhibitory concentration (MIC), an indicator of bacterial resistance, was 4 to 24 μg/mL, with the average MIC at 9.2 ± 4.5 mu;g/mL. Of the bacterial isolates, 1 colony with the VanC-1 genotype was identified as E. gallinarum by 16S rDNA sequencing, whereas the other 63 colonies had the VanC-2/3 genotype and were identified as E. casseliflavus. Although these results imply that the major head bays of Korea are not contaminated with the highly vancomycin-resistant VanA- or VanB-type VREs, the misuse of antibiotics should be prohibited to minimize the presence of VREs and to maintain a safe water supply for protecting the health of the general population. Based on the study results, we also recommend the implementation of a continuous, broad-spectrum inspection program for Enterococcus spp. and VRE contamination in the major head bays. Furthermore, the multiplex PCR method described in this study can be used effectively for this purpose. © 2012 Elsevier GmbH. Source

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