Keun-Ki Kim

Increased production of bacterial cellulose by Acetobacter sp. V6 in synthetic media under shaking culture conditions

Acetobacter strains are bacteria that can synthesize cellulose when grown in a complex medium containing glucose. The effect of the components of a synthetic medium on bacterial cellulose (BC) production by a newly isolated Acetobacter sp. V6 in shaking cultures was investigated. BC production was dependent on the presence of MgSO4 x 7H2O and cosubstrates such as ethanol and lactic acid in the medium. The optimal synthetic medium contained 1.5% glucose, 0.2% (NH4)2SO4, 0.3% KH2PO4, 0.3% Na2HPO4 x 12H2O, 0.08% MgSO4 x 7H2O, 0.0005% FeSO4 x 7H2O, 0.0003% H3BO3, 0.00005% nicotinamide, and 0.6% ethanol. A maximum BC concentration of 4.16 g/l was achieved after 8 days of cultivation at 200 rpm. The production of BC by Acetobacter sp. V6 was higher in synthetic medium than complex medium (Hestrin and Schramm medium) traditionally used for Acetobacter strains.

Overexpression of rice isoflavone reductase-like gene (OsIRL) confers tolerance to reactive oxygen species.

Isoflavone reductase is an enzyme involved in isoflavonoid biosynthesis in plants. However, rice isoflavone reductase-like gene (OsIRL, accession no. AY071920) has not been unraveled so far. Here, we have characterized its behavior in response to oxidizing agents. Using Northern and Western blot analyses, the OsIRL gene and protein were shown to be down-regulated in young seedling roots treated with reduced glutathione (GSH) and diphenyleneiodonium (DPI), known quenchers of reactive oxygen species (ROS). The OsIRL transcript level in rice suspension-cultured cells was also found to be induced by oxidants such as hydrogen peroxide (H(2)O(2)), ferric chloride (FeCl(3)), methyl viologen (MV) and glucose/glucose oxidase (G/GO), but down-regulated when co-treated with GSH. Furthermore, to investigate whether overexpression of OsIRL in transgenic rice plants promotes resistance to ROS, we generated transgenic rice lines overexpressing the OsIRL gene under an abscisic acid (ABA) inducible promoter. Results showed that the OsIRL transgenic rice line activated by ABA treatment was tolerant against MV and G/GO-induced stress in rice leave and suspension-cultured cells. Our results strongly suggest the involvement of OsIRL in homeostasis of ROS.

Influence of glycerol on production and structural-physical properties of cellulose from Acetobacter sp. V6 cultured in shake flasks.

Cost-effective production of bacterial cellulose (BC) by Acetobacter sp. V6 was investigated in shake culture using glycerol as carbon source and its structural and physical properties were determined. In medium containing 3% (w/v) glycerol, BC production was 4.98+/-0.03g/l after 7 days. This value was 3.8-fold higher than the yield in the glucose medium. FT-IR spectra revealed that all the BC samples were highly crystalline and were cellulose type capital I, Ukrainian. The crystallinity index value of the BC produced was 9% higher in the glycerol medium than in the glucose medium. Scanning electron micrographs showed that BC from the glycerol medium was more compact than that from the glucose medium. Water-holding capacity and viscosity of BC from the glycerol medium had 61.3% and 22.4% lower values than those from the glucose medium. These results suggest that glycerol could be a potential low-cost substrate for BC production by Acetobacter sp. V6, leading to the reduction in the production cost.

Antimicrobial and Anticancer Activity of Korean Traditional Soy Sauce and Paste with Chopi

The fruits of Zanthoxylum piperitum are known as having various physiology vitality, and the abstraction ingredient of the pericarp is also known as having strong antibiotic activities against various bacteria. Therefore, this study was carried out to estimate the effect of physiology vitality when the abstraction ingredient of Z. piperitum was added in soy sauce(Chopi-kanjang) and soybean paste(Chopi-doenjang). For the antibiotic activity against the pathogens of sitotoxism such as Staphylococcus aureus, Salmonella typhimurium, Vibrio parahemolyticus, Escherichia coli 0157:H7, the extracts of the Chopi-kanjang was added 1%, 2%, 4% pericarp of Z. piperitum in the manufacturing process of soy source. According to the results, the growth of E. coli 0157:H7 and V. parahemolyticus were respectively inhibited as 70% and 50% by the Chopi-kanjang added 2% of the ingredient. For the antibiotic effects of the aforementioned Chopi-kanjang against Sal. typhimurium and Sta. aureus, the growth of those pathogens was also inhibited between 40% and 60% according to the manufacturing period of Chopi-kanjang. It was confirmed that the antibiotic activity using the mixture of the abstraction ingredient and Chopi-doenjang was lower than those of Chopi-kanjang. In order to estimate the anticancer activity using by caspase-3 activity, the mixture of the abstraction ingredient of the pericarp of Z. piperitum and Chopi-kanjang was treated to leukemia cells. According to the results, the activities of caspase-3 using the mixture added 1%, 2% and 4% of the abstraction ingredient were respectively increased as much as 4, 12, 15 times comparing with the control which was treated with the soy source only. It could be that the mixture of the abstraction ingredient of the pericarp of Z. piperitum and soy source induced apoptosis, and the mixture of the abstraction ingredient and soybean paste had no effect on the activity of caspase-3. In order to find out the death of the aforementioned cells caused by necrosis or apoptosis, DNA fragmentation in the cell was examined. U-937 cells showed apoptotic DNA fragmentation in the incubation with Chopi-kanjang extract.

Effects of Dietary Flammulina velutipes Mycelium on Broiler Chick Performance, Pathogenic Bacterial Counts in Caecal Contents and Amount of NH 3 in Excreta

The purpose of this study was to evaluate the effects of dietary Flammulina velutipes mycelium (FVM) on broiler chick performance, pathogenic bacterial (E. coli, Salmonella) counts in caecal contents and amount of NH3 in excreta. Ninety-six broiler chicks (HanHyup No. 3, Korea) were divided into four groups: 1) Control (basal diet), 2) T1 (supplemented with 1% FVM), 3) T2 (3% FVM), and T3 (5% FVM), and rose for 7 weeks. In results, there were no significant differences among treatments in weight gain, feed intake, feed efficiency and carcass yield. However, giblets were significantly increased in FVM treatments compared with control group (p<0.05). The number of Salmonella in caecum was significantly decreased in FVM treatments compared with control group (p<0.05). The number of E. coli was decreased in T2 but increased in T1 and T3 compared with control group. The emission of fecal NH3 gas was significantly decreased in accordance with increasing the feeding level of FVM (p<0.05). In conclusion, our data indicated that the supplementation of high level of FVM could inhibit the Salmonella in caecum and reduce the emission of fecal NH3 gas. Therefore, FVM at 5% level could be added in the diet of broiler chicks. (Key words : Flammulina velutipes mycelium, Broiler chick, Pathogenic bacterial, Fecal NH3 gas)

Isolation and Characteristics of Bacteria Showing Biocontrol and Biofertilizing Activities

To develop multifunctional microbial inoculant, microorganisms with antagonistic activity and biofertilizing activity were screened. Pantoea agglomerans and Bacillus megaterium from our laboratory culture collection, and strain MF12 from soil near poultry farm in Miryang were selected. On the basis of morphological, physiological studies and 16S rDNA sequence analysis, isolate MF12 was identified as the Bacillus pumilis. Three strains were studied for insoluble phosphate solubilization, indole-3-acetic acid (IAA) and siderophore production, ammonification ability, hydrolytic enzyme production and antifungal activity against phytopathogenic fungi. P. agglomerans did not produce any visible clear zone on agar plate containing 0.5% as a sole phosphorus source. However, this strain could solubilize insoluble phosphate in liquid medium. All strains produced IAA ranged from depending on culture time and had ammonification ability. Among three strains, only P. agglomerans produced siderophore. P. agglomerans produced pectinase and lipase, B. megaterium produced amylase, protease and lipase while B. pumilis produced protease and lipase. P. agglomerans showed antifungal activities against phytopathogenic fungi, Fusarium oxysporum and Colletotrichum gloeosporioides. B. pumilis showed antifungal activities against Botrytis cinerea, Sclerotinia sclerotiorum and Phythium ultimum.

Effect of Phytohormones and Chemical Inhibitors on Pathogenesis-related Genes Identified by Differential Hybridization in Rice Suspension Culture Cells

In order to study disease resistance mechanisms in rice against the rice blast fungus Magnaporthe grisea, we screened fungal elicitor-responsive genes from rice suspension-cultured cells treated with fungal elicitors employing differential hybridization (DH). By DH screening, 31 distinct rice clones were isolated and a majority of them were full-length cDNAs encoding pathogenesisrelated (PR) genes. Sixteen of the 31 genes were upregulated at 4, 8, and 12 h following fungal elicitor treatment. To elucidate the effect of signal molecules and biotic elicitors on the regulation of rice defense genes, we further characterized the transcriptional expression patterns of representative isolated PR genes; OsGlu1, OsGlu2, OsTLP, OsRLK, and OsPR-10, following treatment with fungal elicitor, phytohormones, cycloheximide, and inhibitors of protein phosphorylation. Jasmonic acid (JA) induced transcriptional expression of OsGlu1, OsTLP, and OsRLK, but not of OsGlu2 and OsPR-10 at any of the tested time points. Salicylic acid (SA) and abscisic acid weakly induced the expression of OsTLP and OsRLK. SA showed an antagonistic effect with fungal elicitor and JA. Cycloheximide suppressed all these genes upon elicitor treatment, except for OsGlu2. Staurosporine only induced the expression of OsRLK. Application of calyculin A strongly induced OsRLK expression, but suppressed the expression of OsGlu2. Our study yielded a number of PR genes that play a role in defense mechanisms against the rice blast fungus, as well as contribute towards the elucidation of crosstalk between phytohormones and other modifications during defense signaling.

Fermentation Characteristics of Chungkookjang Prepared Using Different Soybean

Abstract This study was carried out to investigate changes of protease and amylase activities and nitrogen content in Chungkookjang prepared by Bacillus subtilis S8 and different soybean. Amino-type nitrogen and ammonia-type nitrogen contents increased with an increase in fermentation time and was the highest in black soybean Chungkookjang. The number of viable cells increased up to 24 h of fermentation at all temperatures tested; especially, their levels were the highest at 40. Protease activity was the highest in black soybean Chungkookjang. α-amylase activity increased significantly up to 6 h of fermentation at 30and 40and then maintained constantly. It also increased up to 30-36 h of fermentation at 45and then decreased. β-amylase activity was the highest in black soybean Chungkookjang at 35and 40and in yellow soybean Chungkookjang at 45. Production pattern of reducing sugar was similar to that of β-amylase. Amino-type nitrogen, viable cell number and reducing sugar content and β-amylase activity was the highest in Chungkookjang fermented at 40. Considering amino-type and ammonia-type nitrogen contents, Chungkookjang fermentation using yellow soybean was favorable. However, the fermentation using black soybean was favorable, considering protease and amylase activities and reducing sugar content.Key words: Bacillus subtilis, Chungkookjang, Fermentation, Soybean cultivar

Biological Control of Plant Pathogens by Bacillus sp. AB02.

In the greenhouse fields for fruits and vegetables during the winter in Korea, there are serious damages by the sclerotium diseases due to the low temperature and humidity. This study was carried out to select an antagonic agent for the biological control of the sclerotium diseases. The 55 antagonic agents were selected from the rhizosphere in soil where the fruits and vegetables were cultivated in the green house fields, and strain AB02 among the tested isolates was estimated to be the strongest antagonist against the sclerotium disease. Using strain AB02, the antifungal spectrum was tested against 5 different plant pathogens. According to the results of the test, strain AB02 showed the high antagonistic effect against Botrytis cinerea and Sclerotinia sclerotiorum. For the experiment of biological control against the sclerotium disease, it was estimated the suppression effect and the control effect by the strain in the pot experiment using the green perilla. According to the result of the pot experiments, the suppression effect was 40% and the control effect was 62%, respectively. For the stimulation effect of the tested plant growth by strain AB02 compared to the control, it was improved as 120% for the total length, 141% for the liveweight, 121% for the total number of leaves, 185% for the leaf area, and 327% for the liveweight of the root, respectively. Strain AB02 showing the antagonistic effect against the sclerotium disease and the stimulation effect for the plant growth was identified as Bacillus sp.

Effects of Deletion of Ca Supplement (limestone) on Growth and Beef Quality in Hanwoo Finishing Steers

A study was conducted to determine the effects of feeding a diet containing no Ca supplement (limestone) during the late finishing period on growth, marbling and serum 1,25-dihydroxy vitamin level in Korean native cattle. Twenty-four steers (2024 mo of age) were divided into two groups of 12 each: one group assigned to a control diet (concentrates containing 2.5% limestone) and the other to a diet containing no calcium supplement. They were allowed to have free access to diets (concentrates and orchard grass hay) and water during the entire feeding period (223 d). Serum , Ca and P concentrations were not influenced by diets, but serum 1,25-dihydroxy vitamin concentrations determined 2 or 6 mo after the beginning of feeding the experimental diets were higher (P) with the diet without Ca supplement, compared to the control diet (P synthesis that may increase intracellular concentration and in turn fat synthesis.