• Journal of Food Hygiene and Safety
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• v.15 no.3
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• pp.241-247
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• 2000

The incidence of verotoxin-producing Escherichia coli(VTEC) was surveyed in domestic foods including hamburger, raw meats and vegetables from 1997 to 1999. The molecular biological characteristics of the isolates were analyzed. Three VTEC strain were isolated from 1,700 samples. Serotypes of those isolates were 0157 : H7, 026 H4, and 056 : Hl2, respectively. Serotype O26 : H4 produced VT I and VT II, and 055 Hl2 isolate produced VT I, however the 60 MDa plasmid DNA and eae gene were not found from both strains. One 0157 : H7 isolate produced VT II and harbour 60 MDa plasmid DNA, however eae gene was not found in the strain. Although they produced VT, it seemed that the virulence of two strains were relatively weak because of the lack of the eae gene. In addition, the serotype O157 : H7 isolate resistant to ampicillin and streptomycin, while isolates of serotype O26 : H4 and O55 : Hl2 were multi-resistant to antibiotics including ampicillin, carbenicillin , cephalothin, trimethoprim/sulfamethoxazole and tetracycline. Supernatants of cultures of all three isolates were showed cytotoxic effect to vero and HeLa cell

• Journal of Microbiology and Biotechnology
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• v.19 no.5
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• pp.525-529
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• 2009

Cytolethal distending toxins (CDTs) represent an emerging family of newly described bacterial products that are produced by a number of pathogens. The genes encoding these toxins have been identified as a cluster of three adjacent genes, cdtA, cdtB, and cdtC, plus 5 cdt genetic variants, designated as cdt-I, cdt-II, cdt-III, cdt-IV, and cdt-V, have been identified to date. In this study, a general multiplex PCR system designed to detect Escherichia coli cdts was applied to investigate the presence of cdt genes among isolates. As a result, among 366 E. coli strains, 2.7% were found to carry the cdtB gene. In addition, the use of type-specific primers revealed the presence of cdt-I, cdtIV, and cdt-V types of the cdt gene, yet no cdt-II or cdt-III strains. The presence of other virulence genes (stxl, stx2, eae, bfp, espA, espB, and espD) was also investigated using a PCR assay. Among the 10 cdtB gene-positive strains, 8 were identified as COT-producing typical enteropathogenic E. coli (EPEC) strains ($eae^+$, $bfp^+$), whereas 2 were identified as CDT-producing atypical EPEC strains ($eae^+$, $bfp^-$). When comparing the cytotoxic activity of the CDT-producing typical and atypical EPEC strains, the CDT-producing atypical EPEC strains appeared to be less toxic than the CDT-producing typical EPEC strains.

• Journal of Microbiology and Biotechnology
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• v.19 no.3
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• pp.238-249
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• 2009

The genus Listeria consists of six closely related species and forms three phylogenetic groups: L. monocytogenes-L. innocua, L. ivanovii-L. seeligeri-L. welshimeri, and L. grayi. In this report, we attempted to examine the evolutionary relationship in the L. monocytogenes-L. innocua group by probing the nucleotide sequences of 23S rRNA and 16S rRNA, and the gene clusters lmo0029-lmo0042, ascB-dapE, rplS-infC, and prs-ldh in L. monocytogenes serovars 1/2a, 4a, and 4b, and L. innocua. Additionally, we assessed the status of L. monocytogenes-specific inlA and inlB genes and 10 L. innocua-specific genes in these species/serovars, together with phenotypic characterization by using in vivo and in vitro procedures. The results indicate that L. monocytogenes serovar 4a strains are genetically similar to L. innocua in the lmo0035-lmo0042, ascB-dapE, and rplS-infC regions and also possess L. innocua-specific genes lin0372 and lin1073. Furthermore, both L. monocytogenes serovar 4a and L. innocua exhibit impaired intercellular spread ability and negligible pathogenicity in mouse model. On the other hand, despite resembling L. monocytogenes serovars 1/2a and 4b in having a nearly identical virulence gene cluster, and inlA and inlB genes, these serovar 4a strains differ from serovars 1/2a and 4b by harboring notably altered actA and plcB genes, displaying strong phospholipase activity and subdued in vivo and in vitro virulence. Thus, by possessing many genes common to L. monocytogenes serovars 1/2a and 4b, and sharing many similar gene deletions with L. innocua, L. monocytogenes serovar 4a represents a possible evolutionary intermediate between L. monocytogenes serovars 1/2a and 4b and L. innocua.

• Microbiology and Biotechnology Letters
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• v.33 no.3
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• pp.207-214
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• 2005

To characterize the genotypic traits of clinical methicillin-resistant Staphyiococus aureus (MRSA) isolates (n=49), major virulence-associated genes were detected by using PCR-based methods. All the MRSA isolates possessed coagulase gene and showed four polymorphism types [500bp ($6{\%}$),580bp ($27{\%}$), 660bp ($65{\%}$) and 740bp ($2{\%}$)] due to variable numbers of tandem repeats present within the gene. The four or five different loci of hemolysin gene family were dominant in the MRSA isolates,25 of which($51{\%}$) possessed a combination of hla / hlb / hld/ hlg / hlg-2 genes as the most prevalent type. The prevalence of enterotoxin genes was varied among the MRSA isolates. sea and seb genes were detected from all the MRSA isolates. But sei, tsst-1, seg, sec, and seh genes were detected from 31 ($63{\%}$), 16 ($33{\%}$), 14 ($29{\%}$), 8 ($16{\%}$), and 5 ($10{\%}$) isolates, respectively. sed and sej genes were detected from 1 ($2{\%}$) isolate, respectively. see, eta, and etb genes were not detected at all. sea / seb genes were co-detected from 11 ($23{\%}$) isolates, sea / seb / sei genes from 9 ($19{\%}$) isolates, and sea / seb / seg / sei / tsst-1 genes from 5 ($10{\%}$) isolates. Other genes were co-detected with below $10{\%}$ frequencies.

• 한국균학회소식:학술대회논문집
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• 2014.10a
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• pp.15-15
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• 2014

Fusarium graminearum (teleomorph Gibberella zeae) is an important plant pathogen that causes head blight of major cereal crops such as wheat, barley, and rice, as well as causing ear and stalk rot on maize worldwide. Plant diseases caused by this fungus lead to severe yield losses and accumulation of harmful mycotoxins in infected cereals [1]. Fungi utilize spore production as a mean to rapidly avoid unfavorable environmental conditions and to amplify their population. Spores are produced sexually and asexually and their production is precisely controlled. Upstream developmental activators consist of fluffy genes have been known to orchestrate early induction of condiogenesis in a model filamentous fungus Aspergillus nidulans. To understand the molecular mechanisms underlying conidiogenesis in F. graminearum, we characterized functions of the F. graminearum fluffy gene homologs [2]. We found that FlbD is conserved regulatory function for conidiogenesis in both A. nidulans and F. graminearum among five fluffy gene homologs. flbD deletion abolished conidia and perithecia production, suggesting that FlbD have global roles in hyphal differentiation processes in F. graminearum. We further identified and functionally characterized the ortholog of AbaA, which is involved in differentiation from vegetative hyphae to conidia and known to be absent in F. graminearum [3]. Deletion of abaA did not affect vegetative growth, sexual development, or virulence, but conidium production was completely abolished and thin hyphae grew from abnormally shaped phialides in abaA deletion mutants. Overexpression of abaA resulted in pleiotropic defects such as impaired sexual and asexual development, retarded conidium germination, and reduced trichothecene production. AbaA localized to the nuclei of phialides and terminal cells of mature conidia. Successful interspecies complementation using A. nidulans AbaA and the conserved AbaA-WetA pathway demonstrated that the molecular mechanisms responsible for AbaA activity are conserved in F. graminearum as they are in A. nidulans. F. graminearum ortholog of Aspergillus nidulans wetA has been shown to be involved in conidiogenesis and conidium maturation [4]. Deletion of F. graminearum wetA did not alter mycelial growth, sexual development, or virulence, but the wetA deletion mutants produced longer conidia with fewer septa, and the conidia were sensitive to acute stresses, such as oxidative stress and heat stress. Furthermore, the survival rate of aged conidia from the F. graminearum wetA deletion mutants was reduced. The wetA deletion resulted in vigorous generation of single-celled conidia through autophagy-dependent microcycle conidiation, indicating that WetA functions to maintain conidia dormancy by suppressing microcycle conidiation in F. graminearum. In A. nidulans, FlbB physically interacts with FlbD and FlbE, and the resulting FlbB/FlbE and FlbB/FlbD complexes induce the expression of flbD and brlA, respectively. BrlA is an activator of the AbaA-WetA pathway. AbaA and WetA are required for phialide formation and conidia maturation, respectively [5]. In F. graminearum, the AbaA-WetA pathway is similar to that of A. nidulans, except a brlA ortholog does not exist. Amongst the fluffy genes, only fgflbD has a conserved role for regulation of the AbaA-WetA pathway.

• Korean Journal of Microbiology
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• v.41 no.4
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• pp.275-280
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• 2005

Anthrax is an infectious disease caused by the gram-positive bacterium, Bacillus anthracis. Anthrax toxin is a tripartite toxin comprising of protective antigen (PA), lethal factor (LF) and edema factor (EF). PA is the receptor-binding component, which facilitates the entry of LF or EF onto the cytosol. LF is a zinc-dependent metalloprotease, which is a critical virulence factor in cytotoxicity of infected animals. Therefore, it is of interest to develop its potent inhibitors for the neutralization of anthrax toxin. The first step to identify the inhibitors is the development of a rapid, sensitive, and simple assay method with a high-throughput ability. Much efforts have been concentrated on the preparation of powerful assays and on the screening of inhibitors using these system. In the present study, we have tried to construct anthrax lethal factor in yeast expression system to prepare cell-based high-throughput assay system. Here, we have shown the results covering the construction of a new vector system, subcloning of LF gene, and the expression of target gene. Our results are first trial to express LF gene in eukaryote and provide the basic steps in design of cell-based assay system.

• Microbiology and Biotechnology Letters
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• v.34 no.4
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• pp.289-298
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• 2006

3',5'-cyclic adenosine monophosphate (cAMP) is an important molecule, which mediates diverse cellular processes. For example, it is involved in regulation of sugar uptake/catabolism, DNA replication, cell division, and motility in various acterial species. In addition, cAMP is one of the critical regulators for syntheses of virulence factors in many pathogenic bacteria. It is believed that cAMP acts as a signal for environmental changes as well as a regulatory factor for gene expressions. Therefore, intracellular concentration of cAMP is finely modulated by according to its rates of synthesis (by adenylate cyclase), excretion, and degradation (by cAMP phosphodiesterase). In the present review, we discuss the bacterial physiological characteristics governed by CAMP and the molecular mechanisms for gene regulation by cAMP. Furthermore, the effect of cAMP on phosphotransferase system is addressed.

• BMB Reports
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• v.43 no.6
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• pp.419-426
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• 2010

Aeromonas hydrophila is a bacterial pathogen that infects a large number of eukaryotes, including humans. The UDP-galactose 4'-epimerase (GalE) catalyzes interconversion of UDP-galactose to UDP-glucose and plays a key role in lipopolysaccharide biosynthesis. This makes it an important virulence determinant, and therefore a potential drug target. Our earlier studies revealed that unlike other GalEs, GalE of A. hydrophila exists as a monomer. This uniqueness necessitated elucidation of its structure and active site. Chemical modification of the 6xHis-rGalE demonstrated the role of histidine residue in catalysis and that it did not constitute the substrate binding pocket. Loss of the 6xHis-rGalE activity and coenzyme fluorescence with thiol modifying reagents established the role of two distinct vicinal thiols in catalysis. Chemical modification studies revealed arginine to be essential for catalysis. Site-directed mutagenesis indicated Tyr149 and Lys153 to be involved in catalysis. Use of glycerol as a cosolvent enhanced the GalE thermostability significantly.

• Journal of Microbiology and Biotechnology
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• v.14 no.1
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• pp.74-80
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• 2004

Aspergillus fumigatus is one of the most common fungi in the human environment, both in-doors and out-doors. It is the main causative agent of invasive aspergillosis, a life-threatening mycosis among immunocompromised patients. The genome has been sequenced by an international consortium, including the Wellcome Trust Sanger Institute (U.K.) and The Institute for Genomic Research (TIGR, U.S.A.), and a ten times whole genome shotgun sequence assembly has been made publicly available. In this study, we identified tricarboxylic acid (TCA) cycle enzymes of A. fumigatus by comparative analysis with four other fungal species. The open reading frames showed high amino acid sequence similarity with the other fungal citric acid enzymes and well-conserved functional domains. All genes present in Saccharomyces cerevisiae, Schizosaccharomyces pombe, Candida albicans, and Neurospora crassa were also found in A. fumigatus. In addition, we identified four A. fumigatus genes coding for enzymes in the glyoxylate shunt, which may be required for fungal virulence. The architecture of multi-gene encoded enzymes, such as isocitrate dehydrogenase, 2-ketoglutarate, succinyl-CoA synthetase, and succinate dehydrogenase was well conserved in A. fumigatus. Furthermore, our results show that genes of A. fumigatus can be detected reliably using GlimmerM.

• Journal of Microbiology and Biotechnology
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• v.26 no.2
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• pp.263-269
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• 2016

Temperate phages have been suggested to carry virulence factors and other lysogenic conversion genes that play important roles in pathogenicity. In this study, phage TEM123 in wild-type Staphylococcus aureus from food sources was analyzed with respect to its morphology, genome sequence, and antibiotic resistance conversion ability. Phage TEM123 from a mitomycin C-induced lysate of S. aureus was isolated from foods. Morphological analysis under a transmission electron microscope revealed that it belonged to the family Siphoviridae. The genome of phage TEM123 consisted of a double-stranded DNA of 43,786 bp with a G+C content of 34.06%. A bioinformatics analysis of the phage genome identified 43 putative open reading frames (ORFs). ORF1 encoded a protein that was nearly identical to the metallo-β-lactamase enzymes that degrade β-lactam antibiotics. After transduction to S. aureus with phage TEM123, the metallo-β-lactamase gene was confirmed in the transductant by PCR and sequencing analyses. In a β-lactam antibiotic susceptibility test, the transductant was more highly resistant to β-lactam antibiotics than S. aureus S133. Phage TEM123 might play a role in the transfer of β-lactam antibiotic resistance determinants in S. aureus. Therefore, we suggest that the prophage of S. aureus with its exotoxin is a risk factor for food safety in the food chain through lateral gene transfer.