• The Korean Journal of Pesticide Science
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• v.10 no.1
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• pp.56-65
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• 2006

An antagonistic bacterial isolate, that inhibits the growth of plant pathogens, was selected and identified from 5,000 isolates screened from the rhizosphere of various crop plants. An isolate Bacillus sp. N32, tested against Colletotrichum gloeosporioides causing anthracnose disease in hot pepper, produced both a heat resistant antifungal protein and a heat sensitive antifungal protein. The heat resistant protein was partially purified by Ammonium sulfate fractionation and gel filtration chromatography. The bioautography showed that the proteins possessed high antifungal activity. The biosynthetic gene cluster responsible for the heat resistant antifungal protein was cloned from cosmid library using DNA probe obtained from PCR product with the primers targeting the conserved nucleotide sequence of the synthetic genes reported earlier, Most of the clones obtained showed higher homology to fengycin antibiotic synthetic gene family reported earlier. On the other hand, the heat sensitive protein was isolated from SDS-PAGE and electroblotting to determine the N-terminal amino acid sequences. The heat sensitive antifungal protein gene was cloned from the ${\lambda}-ZAP$ libraries using a DNA probe based on the N-terminal amino acid sequences of the heat sensitive protein. We are contemplating to clone and sequence the whole gene cluster encoding the heat sensitive protein for further analysis.

• Korean Journal of Microbiology
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• v.51 no.4
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• pp.419-426
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• 2015

The aims of this study were to isolate spore-forming Bacillus strains that exhibit high digestibility and anti-pathogenic bacteria toward feed for calves. Total 136 spore-forming strains were isolated from finished feeds and their ingredients. Among them, 93 strains were identified as Bacillus species when analyzed by 16S rRNA sequencing. For industrial use, three strains named as Bacillus licheniformis SHS14, B. subtilis LCB7, B. amyloliquefaciens LCB10 were selected after evaluating the industrial standards that are related with heat and acid resistance, enzyme activities, and anti-pathogenic activities against Samonella dublin ATCC15480 and E. coli K99. After each culture, 3 selected strains were mixed together at 1:1:1 (v/v/v) ratio and then prepared as the mixed starter culture for feeding. The changes in microbial community were analyzed via 16S rRNA metagenomics. The initial community ratio among three strains was maintained even after manufacturing into final products. Also, in vitro, enzymatic and anti-pathogenic activities were almost same as those when cultured in single culture, and results of anti-pathogenic activities conducted with calves showed 90% activities against lincomycin, which would be indicative of a promising feed starter.

• Journal of Ginseng Research
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• v.41 no.3
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• pp.353-360
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• 2017

Background: Endophytic fungi play an important role in balancing the ecosystem and boosting host growth. In the present study, we investigated the endophytic fungal diversity of healthy Panax notoginseng and evaluated its potential antimicrobial activity against five major phytopathogens causing root-rot of P. notoginseng. Methods: A culture-dependent technique, combining morphological and molecular methods, was used to analyze endophytic fungal diversity. A double-layer agar technique was used to challenge the phytopathogens of P. notoginseng. Results: A total of 89 fungi were obtained from the roots, stems, leaves, and seeds of P. notoginseng, and 41 isolates representing different morphotypes were selected for taxonomic characterization. The fungal isolates belonged to Ascomycota (96.6%) and Zygomycota (3.4%). All isolates were classified to 23 genera and an unknown taxon belonging to Sordariomycetes. The number of isolates obtained from different tissues ranged from 12 to 42 for leaves and roots, respectively. The selected endophytic fungal isolates were challenged by the root-rot pathogens Alternaria panax, Fusarium oxysporum, Fusarium solani, Phoma herbarum, and Mycocentrospora acerina. Twenty-six of the 41 isolates (63.4%) exhibited activity against at least one of the pathogens tested. Conclusion: Our results suggested that P. notoginseng harbors diversified endophytic fungi that would provide a basis for the identification of new bioactive compounds, and for effective biocontrol of notoginseng root rot.

• Korean Journal of Pharmacognosy
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• v.24 no.4
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• pp.267-281
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• 1993

To find antitumor components from higher fungi, the mycelia of Collybia confluens (Pers. ex Fr.) Kummer were cultured in artificial media. For efficient production of the mycelia, the influences of various modifications of culture conditions were examined. A water-soluble protein-bound polysaccharide fraction, Fr. A, was obtained from the mycelia by hot water extraction. When Fr. A was purified and fractionated by DEAE-cellulose and Sepbadex G-200 gel filtration chromatographies into four fractions which were designated B, C, C-I and C-II. The tumor inhibition ratios of these fractions ranged from 46% to 75% against the solid forms of sarcoma 180 in ICR mice at doses of 20 and 50 mg/kg/day when given intraperitoneally. Especially, Fr. C which was named Collyban(CB) exhibited a marked life-prolonging effect of the mice against ascitic forms of sarcoma 180 at a dose of 50 mg via i.p. administration. To extend spectra of the antitumor activities and eliminate the effects of allograft rejection, the characterization of antitumor effects of CB was performed in syngeneic host-tumor systems. It did not show any antitumor activity against L1210 murine leukemia in $CD_2Fl$ mice but prolonged their life span against ascitic forms of $MM_{46}$ carcinoma in $C_3H/He$ mice. Also it exhibited antitumor activity against human cervical cancer HeLa cells that were xenografted into nude mice having BALB/c genetic backgrounds by the i.p. injection at a dose of 100 mg/kg/day. In order to characterize the antitumor components, CB was examined by chemical analysis. It was acidic protein-bound polysaccharides composed of 31% polysaccharide, 27% protein and 3% hexosamine. CB was fractionated into two fractions, Fr. C-I(M.W.: 500 Kd) and Fr. C-II(M.W.:30 and 8 Kd) by Sephadex G-200 gel filtration chromatography.

• Journal of Microbiology and Biotechnology
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• v.32 no.3
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• pp.307-316
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• 2022

Yeasts play an important role in spontaneous fermentation of traditional alcoholic beverages. Our previous study revealed that a mixed-consortia of both Saccharomyces and non-Saccharomyces yeasts were responsible for fermentation of khadi, a popular, non-standardized traditional beverage with an immense potential for commercialization in Botswana. Functional characterization of isolated fermenting yeasts from mixed consortia is an indispensable step towards the selection of potential starter cultures for commercialization of khadi. In this study, we report the characterization of 13 khadi isolates for the presence of brewing-relevant phenotypes such as their fermentative capacity, ability to utilize a range of carbon sources and their ability to withstand brewing-associated stresses, as a principal step towards selection of starter cultures. Khadi isolates such as Saccharomyces cerevisiae, Saccharomycodes ludwigii and Candida ethanolica showed good brewing credentials but Lachancea fermentati emerged as the isolate with the best brewing attributes with a potential as a starter culture. However, we were then prompted to investigate the potential of L. fermentati to influence the fruity aromatic flavor, characteristic of khadi. The aroma components of 18 khadi samples were extracted using headspace solid phase micro-extraction (HS-SPME) and identified using a GC-MS. We detected esters as the majority of volatile compounds in khadi, typical of the aromatic signature of both khadi and L. fermentati associated fermentations. This work shows that L. fermentati has potential for commercial production of khadi.

• Mycobiology
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• v.36 no.2
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• pp.121-133
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• 2008

Aspergillus niger isolated from Allium sativum was used at large scale fermentation (150 mg flavone/200ml medium) to obtain suitable amounts of the products, efficient for identification. Then spectral analysis (UV, IR, $^1H$-NMR, $^{13}C$-NMR) and mass spectrometry were performed for the two products, which contributed to the identification process. The metabolite (1) was identified as 2'-hydroxydihydrochalcone, and the metabolite (2) was identified as 2'-hydroxyphenylmethylketone, which were more active than flavone itself. Antioxidant activities of the two isolated metabolites were tested compared with ascorbic acid. Antioxidant activity of metabolite (1) was recorded 64.58% which represented 79% of the antioxidant activity of ascorbic acid, and metabolite (2) was recorded 54.16% (67% of ascorbic acid activity). However, the antioxidant activity of flavone was recorded 37.50% which represented 46% of ascorbic acid activity. The transformed products of flavone have anti-microbial activity against Pseudomonas aeruginosa, Aspergillus flavus and Candida albicans, with MIC was recorded $250{\mu}g/ml$ for metabolite (2) against all three organism and 500, 300, and $300{\mu}g/ml$ for metabolite (1) against tested microorganisms (P. aeruginosa, Escherichia coli, Bacillus subtilis, and Klebsiella pneumonia, Fusarium moniliforme, A. flavus, Saccharomyces cerviceae, Kluveromyces lactis and C. albicans) at this order.

• Journal of Microbiology and Biotechnology
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• v.26 no.1
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• pp.120-129
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• 2016

An industrial complex in Wonju, contaminated with trichloroethene (TCE), was one of the most problematic sites in Korea. Despite repeated remedial trials for decades, chlorinated ethenes remained as sources of down-gradient groundwater contamination. Recent efforts were being made to remove the contaminants of the area, but knowledge of the indigenous microbial communities and their dechlorination abilities were unknown. Thus, the objectives of the present study were (i) to evaluate the dechlorination abilities of indigenous microbes at the contaminated site, (ii) to characterize which microbes and reductive dehalogenase genes were responsible for the dechlorination reactions, and (iii) to develop a PCE-to-ethene dechlorinating microbial consortium. An enrichment culture that dechlorinates PCE to ethene was obtained from Wonju stream, nearby a trichloroethene (TCE)-contaminated industrial complex. The community profiling revealed that known organohalide-respiring microbes, such as Geobacter, Desulfuromonas, and Dehalococcoides grew during the incubation with chlorinated ethenes. Although Chloroflexi populations (i.e., Longilinea and Bellilinea) were the most enriched in the sediment microcosms, those were not found in the transfer cultures. Based upon the results from pyrosequencing of 16S rRNA gene amplicons and qPCR using TaqMan chemistry, close relatives of Dehalococcoides mccartyi strains FL2 and GT seemed to be dominant and responsible for the complete detoxification of chlorinated ethenes in the transfer cultures. This study also demonstrated that the contaminated site harbors indigenous microbes that can convert PCE to ethene, and the developed consortium can be an important resource for future bioremediation efforts.

• Molecular & Cellular Toxicology
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• v.1 no.3
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• pp.189-195
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• 2005

Industrial development has increase consumption of crude oil and environmental pollution. A large number of microbial lipolytic enzymes have been identified and characterized to date. To development for a new lipase with catalytic activity in degradation of crude oil as a microbial enzyme, Acinetobactor sp. B2 was isolated from soil samples that were contaminated with oil in Daejon area. Acinetobactor sp. B2 showed high resistance up to 10 mg/mL unit to heavy metals such as Ba, Li, Al, Cr, Pb and Mn. Optimal growth condition of Acinetobactor sp. B2 was confirmed $30^{\circ}C$. Lipase was purified from the supernatant by Acinetobactor sp. B2. Its molecular mass was determined to the 60 kDa and the optimal activity was shown at $40^{\circ}C$ and pH 10. The activation energies for the hydrolysis of p-nitrophenyl palmitate were determined to be 2.7 kcal/mol in the temperature range 4 to $37^{\circ}C$. The enzyme was unstable at temperatures higher than $60^{\circ}C$. The Michaelis constant $(K_{m})\;and\;V_{max}$ for p-nitrophenyl palmitate were $21.8{\mu}M\;and\;270.3{\mu}M\;min^{-1}mg\;of\;protein^{-1}$, respectively. The enzyme was strongly inhibited by $Cd{2+},\;Co^{2+},\;Fe^{2+},\;Hg^{2+},\;EDTA$, 2-Mercaptoethalol. From these results, we suggested that lipase purified from Acinetobactor sp. B2 should be able to be used as a new enzyme for degradation of crude oil, one of the environmental contaminants.

• Biotechnology and Bioprocess Engineering:BBE
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• v.10 no.5
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• pp.408-417
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• 2005

Increasing numbers of value added chemicals are being produced using microbial fermentation strategies. Computational modeling and simulation of microbial metabolism is rapidly becoming an enabling technology that is driving a new paradigm to accelerate the bioprocess development cycle. In particular, constraint-based modeling and the development of genome-scale models of industrial microbes are finding increasing utility across many phases of the bioprocess development workflow. Herein, we review and discuss the requirements and trends in the industrial application of this technology as we build toward integrated computational/experimental platforms for bioprocess engineering. Specifically we cover the following topics: (1) genome-scale models as genetically and biochemically consistent representations of metabolic networks; (2) the ability of these models to predict, assess, and interpret metabolic physiology and flux states of metabolism; (3) the model-guided integrative analysis of high throughput 'omics' data; (4) the reconciliation and analysis of on- and off-line fermentation data as well as flux tracing data; (5) model-aided strain design strategies and the integration of calculated biotransformation routes; and (6) control and optimization of the fermentation processes. Collectively, constraint-based modeling strategies are impacting the iterative characterization of metabolic flux states throughout the bioprocess development cycle, while also driving metabolic engineering strategies and fermentation optimization.

• Journal of Microbiology
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• v.33 no.2
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• pp.115-119
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• 1995

The alkaline protease of Xanthomonas sp. YL-37 has been purified, and the properties of the enzyme investigated. The alkaline protease of Xanthomonas sp. YL-37 was purified form crude enzyme by ammonium sulfate fractionation, CM-cellulose ion exchange chromatography, and Sephadex G-100 gel filtration. Through the series of chromatographies, the enzyme was purified to homogenecity with specific activity of 4.23 fold higher than that of the crude broth. The molecular weight of the purified protease has been estimated to be 62 KDa on SDS-polyacrylamide gel electrophoresis. The optimal pH and temperature for alkaline protease activity were 11.0 and 50.deg.C, respectively. The enzyme was stable between pH 5.0 and 10.0 and up to 50.deg.C. Enzyme activity was lost up to 50% on heating at 70.deg.C for 30 minutes. The activity of alkaline protease was inhibited by Cu$\^$2+/, Zn$\^$2+/, Hg$\^$2+/, PMSF, and activated by Mn$\^$2+/ and Ca$\^$2+/. The $K_{m}$ value for casein as a substrate was 4.0 mg/ml.