• The Journal of the Korean Society for Microbiology
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• v.35 no.2
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• pp.109-115
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• 2000

FtsH is a membrane-bound, ATP-dependent metalloprotease that is involved in a variety of cellular functions including the regulation of responses to heat and stress shock. Previously, we had cloned and sequenced pneumococcal ftsH gene whose deduced amino acid sequence was very similar to those of several gram-positive bacteria and Escherichia coli, except for the N-terminal domain that was responsible for membrane anchoring. In order to better understand the role of Streptococcus pneumoniae FtsH, we expressed pneumococcal ftsH gene in Escherichia coli. When it was expressed from a strong promoter, $P_{tac}$, a considerable amount of the recombinant FtsH was produced, although the prolonged induction resulted in not only accumulation of breakdown products but also ceasing of the further growth of E. coli host. This indicated that the expression of the exogenous ftsH gene was tightly regulated since the excessive FtsH appeared detrimental to bacterial cells. In Western blotting, the pneumococcal FtsH protein, whether native or recombinant, was reactive to anti-E. coli FtsH serum. The observation that FtsH proteins were well conserved throughout the bacterial kingdom and its expression level was fine-tuned suggests an important role for this protein in the stress adaptation which may be related to infecting process by pneumococci.

• Journal of fish pathology
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• v.18 no.3
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• pp.287-292
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• 2005

In the present study, immune gene expression of rainbow trout, Oncorhynchus mykiss, against bacterial endotoxin (LPS) was studied in vivo. The expression of prointflammatory cytokine genes (IL-1$\beta$ and TNF-$\alpha$) and IFN-related genes (IRF-I, Mx-3) at gill was assessed by RT-PCR at different time point of day1 and day 3 post-injection. It was shown that the proinflammatory cytokine gene expression at gill was induced 1 day after LPS injection but the expression was not sustained until day 3. Meanwhile upregulated expression of IFN-related genes was found to be only at day 3 post injection, indicating indirect effect of LPS on these genes.

• Korean Journal of Veterinary Research
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• v.52 no.4
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• pp.231-238
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• 2012

The expression profiles of inflammatory cytokines viz. interleukins (IL)-6, IL-8, IL-12, granulocyte macrophage-colony stimulating factor, interferon-${\gamma}$ and tumor necrosis factor-${\alpha}$ in response to subclinical mastitis in indigenous cattle breed Kankrej (n = 6), Gir (Bos indicus) (n = 12) and crossbred (Bos taurus${\times}$Bos indicus) (n = 7) were investigated using quantitative real time PCR. Significant correlation (p < 0.05) was observed between total bacterial load and somatic cell count (SCC) in all three breeds of cattle. All the cytokines were observed to be up-regulated compared to cows with healthy quarters, however, level of their expression varied among three breeds of cattle. In Kankrej most cytokines were found to be transcribed to higher levels than in other two breeds; the milk had higher load of bacteria but not so high SCC, implying that Kankrej has a higher inherent resistance against mastitis. The results of present study indicated that mammary glands of crossbred cattle are more sensitive to bacterial infection than indigenous breed of cattle as they elicit immune response at lower bacterial load and result into higher SCC. Research on identification of factors responsible for differentially expressed cytokines profiles and use of cytokines as immunomodulatory tools can pave way for formulating control strategies against bovine mastitis.

• The Plant Pathology Journal
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• v.25 no.2
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• pp.136-141
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• 2009

Marking biocontrol bacteria is an essential step to monitor bacterial behavior in natural environments before application in agricultural ecosystem. In this study, we presented the simple green fluorescent protein (GFP) reporter system driven by the promoter active in Bacillus species for tagging of the biocontrol bacteria. A constitutive promoter P43 from Bacillus subtilis was fused to an enhanced promoterless gfp gene by overlap extension PCR. The GFP expression was demonstrated by the high fluorescence intensity detected in B. subtilis and Escherichia coli transformed with the P43-gfp fusion construct, respectively. The GFP reporter system was further investigated in two bacterial biocontrol strains B. licheniformis and Pseudomonas fluorescens. When the reconstructed plasmid pWH34G was introduced into B. licheniformis, GFP level measured with the fluorescence intensity in B. licheniformis was almost equivalent to that in B. subtilis. However, GFP expression level was extremely low in other biocontrol bacteria P. fluorescens by transposon based stable insertion of the P43-gfp construct into the bacterial chromosome. This study provides information regarding to the efficient biomarker P43-gfp fusion construct for bio-control Bacillus species.

• Journal of Microbiology and Biotechnology
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• v.20 no.5
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• pp.917-924
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• 2010

The practicability of transgenic tobacco engineered to express bacterial native mercuric reductase (MerA), responsible for the transport of $Hg^{2+}$ ions into the cell and their reduction to elemental mercury ($Hg^0$), without any codon modification, for phytoremediation of mercury pollution was evaluated. Transgenic tobacco plants reduce mercury ions to the metallic form; take up metallic mercury through their roots; and evolve the less toxic elemental mercury. Transformed tobacco produced a large amount of merA protein in leaves and showed a relatively higher resistance phenotype to $HgCl_2$ than wild type. Results suggest that the integrated merA gene, encoding mercuric reductase, a key enzyme of the bacterial mer operon, was stably integrated into the tobacco genome and translated to active MerA, which catalyzes the bioconversion of toxic $Hg^{2+}$ to the least toxic elemental $Hg^0$, and suggest that MerA is capable of reducing the $Hg^{2+}$, probably via NADPH as an electron donor. The transgenic tobacco expressing merA volatilized significantly more mercury than wild-type plants. This is first time we are reporting the expression of a bacterial native merA gene via the nuclear genome of Nicotiana tabacum, and enhanced mercury volatilization from tobacco transgenics. The study clearly indicates that transgenic tobacco plants are reasonable candidates for the remediation of mercurycontaminated areas.

• The Korean Journal of Physiology and Pharmacology
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• v.2 no.3
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• pp.331-343
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• 1998

Sublethal dose of bacterial lipopolysaccharide (LPS) would induce protection against cardiac ischemic/reperfusion (I/R) injury. This study examines the following areas: 1) the temporal induction of the cardio-protection produced by LPS; and 2) the relations between a degree of protection and the myocardial prostacyclin ($PGI_2$) production. Rats were administered LPS (2 mg/kg, i.v.), and hearts were removed 1, 4, 8, 14, 24, 48, 72,and 96 h later. Using Langendorff apparatus, haemodynamic differences during 25 min of global ischemia/30 min reperfusion were investigated. The concentration of $PGI_2$ in aliquots of the coronary effluent was determined by radioimmunoassay as its stable hydrolysis product $6-keto-PGF1_{\alpha}$ and lactate dehydrogenase release were measured as an indicative of cellular injury. LPS-induced cardiac protection against I/R injury appeared 4 h after LPS treatment and remained until 96 h after treatment. $PGI_2$ release increased 2-3 fold at the beginning of reperfusion compared to basal level except in hearts treated with LPS for 48 and 72 h. In hearts removed 48 and 72 h after LPS treatment, basal $PGI_2$ was increased. To determine the enzymatic step in relation to LPS-induced basal $PGI_2$ production, we examined prostaglandin H synthase (PGHS) protein expression, a rate limiting enzyme of prostaglandin production, by using Western blot analysis. LPS increased PGHS protein expression in hearts at 24, 48, 72, 96 h after LPS treatment. Induction of PGHS expression appeared in both isotypes of PGHS, a constitutive PGHS-1 and an inducible PGHS-2. To identify the correlationship between $PGI_2$ production and the cardioprotective effect against I/R injury, indomethacin was administered in vivo or in vitro. Indomethacin did not inhibit LPS-induced cardioprotection, which was not affected by the duration of LPS treatment. Taken together, our results suggest that $PGI_2$ might not be the major endogenous mediator of LPS-induced cardioprotection.

• Journal of Microbiology and Biotechnology
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• v.10 no.1
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• pp.86-90
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• 2000

We have earlier reported on the cloning and identification of bft-k from an enterotoxigenic strain of Bacteroides fragilis 419, which was isolated from the blood of a Korean patient who suffered from systemic infections [4,5]. The bft-k gene encodes a 397-amino-acids metalloprotease enterotoxin, and the protein has been identified as a new isoform of B. fragilis enterotoxins (BFTs), which are cytopathic to intestinal epithelial cells to induce fluid secretion and tissue damage in ligated intestinal loops [4, 6, 18, 20]. This report describes the cloning and sequencing of the enterotoxin pahogenicity islet of B. fragilis 419 which contains the bft-k gene. the cloned enterotoxin pathogenicity islet was found to have 6,045 bp in length and to contain 120bp direct repeats near its end. In the pathogenicity islet, in addition to the BFR-K, two putative open reading frames (ORFs) were identified; (1) the t-3 gene encoding a 396-amino-acids protein of a putative metalloprotease; (2) the third gene encoding an ORF of a 59-amino-acids protein, whose function has not yet beenn characterized. The expression of the t-3 gene in B. fragilis 419 was verified by western blot analysis.

• Journal of Korean Biological Nursing Science
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• v.10 no.2
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• pp.131-140
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• 2008

Purpose: This study is about safety of the expressed human milk by investigating its bacterial contamination and antibiotic resistance of the microbes in the expressed milk. Methods: The data are collected from the 156 mothers and their newborn infants in NICU of U University Hospital from January 2004 to September 2005. Results: 1) The expressed human milk was contaminated by Staphylococcus epidermidis for 66.7%, Two mixed bacterial strain for 11.5%, Acinetobacter species for 8.0%, Klebsiella species for 4.6%, Staphylococcus aureus for 4.6%. 2) The microbes in the human milk had high resistance to the Antimicrobial agents: 77.5% for Penicillin-G, 66.6% for Oxacillin, and 63.7% for Cephalothin. 3) The distribution of microbes showed a significant depending on the place of the milk expression (p=.020). In particular, mixed bacterial strain was found more in the milks expressed at home than the milk expressed at the hospital. Conclusion: This study shows the importance of systematic education of feeding process in expressed milk: poor management of a breast pump, inadequate hand washing and imperfect breast cleaning explain the reasons of contamination 156 cases.

• Proceedings of the Korean Society for Applied Microbiology Conference
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• 2000.04a
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• pp.94-99
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• 2000

Bioluminescence is being used as a prevailing reporter of gene expression in microorganisms and mammalian cells. Bacterial bioluminescence draws special attention from environmental biotechnologists since it has many advantageous characteristics, such as no requirement of extra substractes, highly sensitive, and on-line measurability. Using bacterial bioluminescence as a reporter of toxicity has replaced the classical toxicity monitoring technology of using fish or daphnia with a cutting-edge technology. Fusion of bacterial stress promoters, which control the transcription of stress genes corresponding to heat-shock, DNA-, or oxidative-damaging stress, to the bacterial lux operon has resulted in the development of novel toxicity biosensors with a short measurement time, enhanced sensitivity, and ease and convenient usage. Therefore, these recombinant bioluminescent bacteria are expected to induce bacterial bioluminescence when the cells are exposed to stressful conditions, including toxic chemicals. We have used these recombinant bioluminescent bacteria in order to develop toxicity biosensors in a continuous, portable, or in-situ measurement from for air, water, and soil environments. All the data obtained from these toxicity biosensors for these environments were found to be repeatable and reproducible, and the minimum detection level of toxicity was found to be ppb (part per billion) levels for specific chemicals.

• Microbiology and Biotechnology Letters
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• v.25 no.6
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• pp.566-570
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• 1997

During sporulation, Bacillus thuringiensis strains produce crystals consist of toxin proteins highly specific against insect pests. Their host specificities are desirable from a standpoint of environmental safety, but also limit market potential. Thus, development of improved Bacillus thuringiensis strains having broad host spectrum will contribute to increase its use. For the construction of Bacillus thuringiensis strain having broad host spectrum, we cloned cry1C gene encoding a toxin protein highly toxic against Spodoptera exigua from a B. thuringiensis isolate and constructed two recombinant plasmids, pUBClC and plC60. The plasmid PUBC1C has a replication origin of the natural plasmid pBC16 from B. cereus which is closely related species to B. thuringiensis, and the pBC16 was known to be replicated by rolling-circle mechanism. The plasmid pIC60 has a replication origin of a resident 60 MDa plasmid from B. thuringiensis subsp. kurstaki HD263, and it is believed that the pIC60 is replicated in a theta mode. The two plasmids were introduced into B. thuringiensis subsp. kurstaki cryB strain, and the transformed strains produced well-shaped bipyramidal crystals. We confirmed the expression of the cry1C gene by SDS-PAGE, and Western blotting. By investigating the segregational stability, it was found that the plasmid pIC60 is more stable than the pUBC1C.