• Journal of Microbiology and Biotechnology
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• v.13 no.6
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• pp.960-968
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• 2003

A bacterial isolate showing a strong endochitosanase activity was isolated from soil and then characterized. The isolate was identified and designated as Bacillus cereus P16, based on morphological and biochemical properties, assimilation tests, cellular fatty acids pattern, along with 16S rRNA gene sequence. The optimized medium for producing extracellular chitosanase in a batch culture contained 1% tryptone, 0.5% chitosan, and 1% NaCl (pH 7.0). Powder chitosan and tryptone served the best as carbon and nitrogen sources, respectively, for the chitosanase production. Chitosanase activity was the highest when culture was completed at $37^{\circ}C$ among various temperatures ($20-42^{\circ}C$) tested in a shaking incubator (200 rpm). The levels of chitosanase activity in the culture fluid were 2.0 U/ml and 3.8 U/ml, respectively, when incubated in a flask for 60 h and in a jar fermenter for 24 h. The culture supernatant showed a strong liquefying activity on the soluble chitosan. The viscosity of 1% chitosan solution, that was incubated with the culture supernatant, was rapidly decreased, suggesting the secretion of endochitosanolytic enzymes by P16. The culture fluid revealed six endo-type chitosanase isozymes, two major (38 and 45 kD), and four minor (54, 65, 82, and 96 kD) forms by staining profile. The crude enzymes were very stable, and full activity was maintained for 4 weeks at $4^{\circ}C\;or\;-20^{\circ}C$ in the culture supernatant, suggesting a highly desirable stability rate for making an industrial application of the crude enzymes. The supernatant also cleaved the insoluble chitosan powder, but the hydrolysis rate was much lower. The enzymic degradation products of chitosan contained $(GlcN)_n$ (n=2-8). The concentration of chitosan in the reaction mixture of the crude enzyme affected the chitooligosaccharides composition of the hydrolysis products. When the higher concentration of chitosan was used, the higher degree of polymerized chitooligosaccharides were produced. By comparison with other commercial chitosanase preparations, P16 was indeed found to be a valuable enzyme source for industrial production of chitooligosaccharides from chitosan.

• Microbiology and Biotechnology Letters
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• v.47 no.4
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• pp.596-602
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• 2019

In September 2017, the rhizospheric soil of Tetragonia tetragonoides (Pall.) Kuntze was further sampled. One hundred and thirty eight species of microorganisms were isolated from the soil. Indole-3-acetic acid (IAA) production, siderophore production, and phosphate degradation were examined in order to confirm bacterial growth from isolated microorganisms. As a result, most strains were able to produce auxins or siderophores and to solubilize phosphate. In addition, 138 isolated strains were treated with tobacco extract and conferred pathogen resistance to host plants upon treatment. As a result, 35 strains that were able to reduce pathophysiology by more the 60% were selected. Among them, 6 strains with high induced systemic resistance (ISR) activity were found. All of these strains belong to the genus Bacillus according to the 16S rDNA sequence analysis. Bacillus aryabhattai KUDC6619 showed outstanding effects with reduced infection in tobacco and pepper plants. Probably, these Bacillus species play a beneficial role by association with T. tetragonoides for its survival in the harsh conditions found on the island of Dokdo.

• The Korean Journal of Pesticide Science
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• v.20 no.4
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• pp.349-354
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• 2016

Triazole fungicides occupy an important portion in the global fungicide market and are relatively persistent in soil compared to the other fungicides, suggesting possible adverse effects of the fungicides on human health and environment. In this study, we tried to isolate microorganisms from orchard soils, which can decompose the triazole fungicides, tebuconazole, fluquinconazole, and difenoconazole. Only difenoconazole was completely degraded in the enrichment culture, from which several difenoconazole-degrading bacteria were isolated. They showed the same rep-PCR pattern thus only one strain, C8-2, was further studied. The strain was identified as Sphingomonas sp. C8-2 based on its 16S rRNA gene sequence and decomposed 100 mg/L of difenoconazole in a minimum medium to an unknown metabolite with a molecular weight of 296 within 24 hours. The inhibition effect of the metabolite against representative soil microorganisms significantly decreased compared to that of difenoconazole thus the bacterial strain is expected to be used for the detoxification of difenoconazole in soil and crop.

• Asian-Australasian Journal of Animal Sciences
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• v.14 no.6
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• pp.880-884
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• 2001

Rumen microbiology research has undergone several evolutionary steps: the isolation and nutritional characterization of readily cultivated microbes; followed by the cloning and sequence analysis of individual genes relevant to key digestive processes; through to the use of small subunit ribosomal RNA (SSU rRNA) sequences for a cultivation-independent examination of microbial diversity. Our knowledge of rumen microbiology has expanded as a result, but the translation of this information into productive alterations of ruminal function has been rather limited. For instance, the cloning and characterization of cellulase genes in Escherichia coli has yielded some valuable information about this complex enzyme system in ruminal bacteria. SSU rRNA analyses have also confirmed that a considerable amount of the microbial diversity in the rumen is not represented in existing culture collections. However, we still have little idea of whether the key, and potentially rate-limiting, gene products and (or) microbial interactions have been identified. Technologies allowing high throughput nucleotide and protein sequence analysis have led to the emergence of two new fields of investigation, genomics and proteomics. Both disciplines can be further subdivided into functional and comparative lines of investigation. The massive accumulation of microbial DNA and protein sequence data, including complete genome sequences, is revolutionizing the way we examine microbial physiology and diversity. We describe here some examples of our use of genomics- and proteomics-based methods, to analyze the cellulase system of Ruminococcus flavefaciens FD-1 and explore the genome of Ruminococcus albus 8. At Illinois, we are using bacterial artificial chromosome (BAC) vectors to create libraries containing large (>75 kbases), contiguous segments of DNA from R. flavefaciens FD-1. Considering that every bacterium is not a candidate for whole genome sequencing, BAC libraries offer an attractive, alternative method to perform physical and functional analyses of a bacterium's genome. Our first plan is to use these BAC clones to determine whether or not cellulases and accessory genes in R. flavefaciens exist in clusters of orthologous genes (COGs). Proteomics is also being used to complement the BAC library/DNA sequencing approach. Proteins differentially expressed in response to carbon source are being identified by 2-D SDS-PAGE, followed by in-gel-digests and peptide mass mapping by MALDI-TOF Mass Spectrometry, as well as peptide sequencing by Edman degradation. At Ohio State, we have used a combination of functional proteomics, mutational analysis and differential display RT-PCR to obtain evidence suggesting that in addition to a cellulosome-like mechanism, R. albus 8 possesses other mechanisms for adhesion to plant surfaces. Genome walking on either side of these differentially expressed transcripts has also resulted in two interesting observations: i) a relatively large number of genes with no matches in the current databases and; ii) the identification of genes with a high level of sequence identity to those identified, until now, in the archaebacteria. Genomics and proteomics will also accelerate our understanding of microbial interactions, and allow a greater degree of in situ analyses in the future. The challenge is to utilize genomics and proteomics to improve our fundamental understanding of microbial physiology, diversity and ecology, and overcome constraints to ruminal function.

• Journal of Korean Society of Environmental Engineers
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• v.22 no.3
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• pp.463-473
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• 2000

This research was carried out to investigate chemical pretreatment using limestone in treating abandoned metallic mine drainage with anaerobic biological treatment, and to estimate application of papermill and livestock sludge as carbon sources for SRB (Sulfate Reducing Bacterial. Capacity of anaerobic limestone bed was steeply decreased. But if limestone was utilized as pretreatment process in treating them with anaerobic biological treatment. it could look forward to stabilize system because it did initally neutralize them. Effluent SCOD in R-4 was lower than R-l~R-3 in inital HRT 5day but its concentration was high in HRT 1day after passed time. Therefore in point of durability and supply of organic matter. it seemed that R-4 was useful became organic matter in R-4 was not consumed by excessive degradation within short period. In all reactors, pH was suitable for SRB growth in whole HRT, but on the evidence of ORP, SRB was active after HRT 2day. Fixation trend of heavy meta s showed high as $SO_4{^{2-}}$ reduction efficiency increased, and $SO_4{^{2-}}$ reduction and fixation of heavy metals were relatively high in HET 2day.

• The Korean Journal of Pesticide Science
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• v.20 no.2
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• pp.152-158
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• 2016

The cause of death was investigated with several dead cabbage moth larvae in breeding box. Bacterial strains were isolated and selected from the dead larvae by bioassay. One of them was identified as Serratia marcescens used by morphological characteristics and gene sequencing. S. marcescens were cultured by Luria Bertani (LB) media broth for bioassay. When 100-fold dilution of culture broth to third larvae of diamondback moth, Plutella xylostella, it was showed a 100% mortality at 2 days after treatment, and only 10-fold dilution of supernatant liquid was showed 86.6% mortality. When the culture broth of S. marcescens was applied to the larvae of beet armyworm, Spodoptera exigua, contact and feeding toxicity were 20 and 8% of mortality, respectively. Otherwise, when the culture broth of S. marcescens was applied to 5 major plant pathogens, antibacterial activities against Fusarium oxysporum, Rhizoctonia solani, Colletotrichum gloeosporioides, Phytophthora capsici and Sclerotinias clerotiorum were 4.7, 11.3, 20, 15.7 and 42.6%, respectively. Also, degradation ability of S. marcescens against protein and chitin were examined.

• Journal of Life Science
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• v.25 no.11
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• pp.1273-1279
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• 2015

Agarase from a novel agar-degrading bacterium isolated from seawater in Namhae at Gyeongsangnamdo province of Korea was characterized. The SH-1 strain was selected from thousands of colonies on Marine agar 2216 media. Almost full 16S rRNA gene sequence of the agarolytic SH-1 strain showed 99% similarity with that of bacteria of Simiduia genus and named as Simiduia sp. SH-1. Agarase production was growth related, and activity was declined from stationary phase. Secreted agarase was prepared from culture media and characterized. It showed maximum activity of 698.6 units/L at pH 7.0 and 30℃ in 20 mM Tris-HCl buffer. Agarase activity decreased as the temperature increased from an optimum of 30℃, with 90% and 75% activity at 40℃ and 50℃, respectively. Agarase was not heat resistant. Slightly lower agarase activity was observed at pH 6.0 than at pH 7.0, without statistical difference, and 80% and 75% activity were observed at pH 5.0 and 8.0, respectively. Neoagarotetraose and neoagarobiose were the main final products of agarose, indicating that it is β-agarase. Simiduia sp. SH-1 and its β-agarase would be useful for the industrial production of neoagarotetraose and neoagarobiose, which have a whitening effect on skin, delaying starch degradation, and inhibiting bacterial growth.

• Asian-Australasian Journal of Animal Sciences
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• v.19 no.3
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• pp.376-380
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• 2006

An experiment was conducted to study the effect of temperature and pH on in vitro nutrient degradability, volatile fatty acid profile and methane production. The fermenter used was the semi-continuous system, known as the rumen simulation technique (RUSITEC). Sixteen cylinders were used at one time with a volume of 800 ml, the dilution rate was set at 3.5%/hour, the infused buffer being McDougall's artificial saliva. Basal diet (9.6 g DM) used in RUSITEC consisted of (DM) 6.40 g Timothy hay, 1.86 g crushed corn and 1.34 g soybean meal. The food for the fermentation vessel was provided in nylon bags, which were gently agitated in the liquid phase. The experiment lasted for 17 d with all the samples taken during the last 5 d. Treatments were allocated at random to four vessels each and were (1) two temperature levels of $39^{\circ}C$ and $41^{\circ}C$ (2) two pH levels of 6.0 and 7.0. The total diet contained ($g\;kg^{-1}$ DM) 957 OM, 115 CP and $167MJ\;kg^{-1}$ (DM) GE. Although increase in temperature from $39^{\circ}C$ to $41^{\circ}C$ reduced degradation of major nutrients in vitro, it was non-significant. Interaction effect of temperature with pH also reflected a similar trend. However, pH showed a significant (p<0.05) negative effect on the degradability of all the nutrients in vitro. Altering the in vitro pH from 7 to 6 caused marked reduction in DMD from 60.2 to 41.8, CPD from 76.3 to 55.3 and GED from 55.3 to 35.1, respectively. Low pH (6) depressed total VFA production (61.9 vs. 34.9 mM) as well as acetate to propionate ratio in vitro (from 2.0 to 1.5) when compared to pH 7. Compared to pH 7, total gas production decreased from 1,841 ml to 1,148 ml at pH 6, $CO_2$ and $CH_4$ production also reduced from 639 to 260 ml and 138 to 45 ml, respectively. This study supported the premise that pH is one of the principal factors affecting the microbial production of volatile fatty acids and gas. Regulating the ruminal pH to increase bacterial activity may be one of the methods to optimize VFA production, reduce methane and, possibly, improve animal performance.

• Fisheries and Aquatic Sciences
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• v.14 no.4
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• pp.267-274
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• 2011

Bacterial lipopolysaccharide (LPS) induces expression of pro-inflammatory cytokines and enzymes producing nitric oxide (NO) and prostaglandins (PGs) in immune cells. This process is mediated by the activation of nuclear factor kappaB (NF-${\kappa}B$). In this study, we investigated the anti-inflammatory characteristics of Codium fragile ethanolic extract (CFE) mediated by the regulation of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) using LPS-stimulated murine macrophage RAW 264.7 cells. CFE significantly inhibited LPS-induced NO and $PGE_2$ production in a dose-dependent manner and suppressed the expression of iNOS and COX-2 proteins in LPS-stimulated RAW 264.7 cells with no cytotoxicity. Pro-inflammatory cytokines, such as interleukin (IL)-$1{\beta}$, IL-6, and tumor necrosis factor-${\alpha}$, were significantly reduced by treatment of CFE in LPS-stimulated RAW 264.7 cells. CFE inhibited the promoter activity of (NF)-${\kappa}B$ in LPS-stimulated macrophages. Treatment with CFE suppressed translocation of the NF-${\kappa}B$ p65 subunit by preventing proteolytic degradation of inhibitor of ${\kappa}B-{\alpha}$. These results indicate that the CFE-mediated inhibition of NO and $PGE_2$ production in LPS-stimulated RAW 264.7 cells is mediated through the NF-${\kappa}B$-dependent transcriptional downregulation of iNOS and COX-2, suggesting the potential of CFE as a nutraceutical with anti-inflammatory activity.

• Microbiology and Biotechnology Letters
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• v.17 no.3
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• pp.236-240
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• 1989

The isolated bacterial strain 1K1 able to grow on extremely high concentration of toluene was morphologically and physiologically best described as Pseudomonas putida. This strain could grow on at least eight aromatic compounds, e.g., benzene, benzoate, phenol, o-cresol, m-cresol, toluene, m-tolunte, and xylene, but did not Brow on alkanes, such as hexane, octane, decane, and cyclohexane. Strain 1K1 could grow on above 95% toluene, but it could not grow on above 1% of other aromatic compounds. In the point of survival, strain 1K1 was resistant to high concentration of alkanes, appreciably resistant to toluene and xylene, and damaged by to other aromatic compounds. Strain 1K1 which grew on high concentration of toluene had irregular cell shape in comparing with normal cell shape of the genus Pseudomonas. Strain 1K1 was shown to have at least two aromatic compound dissimilation pathway, one for benzoate and the other for toluene.