[Show abstract][Hide abstract] ABSTRACT: A bacterium producing antimicrobial substance was isolated from cheonggukjang. The bacterium was identified as a strain of Bacillus subtilis by 16S rDNA sequencing and designated as Bacillus subtilis HH28. The antimicrobial substance produced from Bacillus subtilis HH28 was purified by 0-80% ammonium sulfate precipitation, DEAE-sepharose FF column chromatography, and Sephacryl S-200 HR gel chromatography The molecular weight of the purified antimicrobial substance was estimated to be approximately 3,500 Da using Tricine sodium dodecyl sulfate-polyacrylamide gel electrophoresis and direct detection analysis. Antimicrobial substance from B. subtilis HH28 not only inhibited B cereus, but also Listeria monocytogenes and Vibrio parahaemolyticus. The purified antimicrobial substance was stable at 40-80°C, and between pH 2 and 8. Antimicrobial activity of the purified substance was completely destroyed by treatment of protease, proteinase K, and pronase E, indicating that it is proteinaceous.
Korean Journal of Microbiology and Biotechnology 12/2014; 42(4):393-401. DOI:10.4014/kjmb.1411.11003
[Show abstract][Hide abstract] ABSTRACT: A Gram-positive, aerobic, endospore forming, and moderately halophilic rod, designated strain R1T, was isolated from the rice husks and was subjected to a polyphasic taxonomic study. Strain R1T produced spherical or ellipsoidal endospores at a subterminal position in swollen sporangia, and was catalase and oxidase positive. The isolate grew optimally at 37 °C and pH 6.0-7.0, and could grow with up to 9% (w/v) NaCl. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain R1T belongs to the genus Bacillus. The species most closely related to strain R1T were Bacillus subtilis subsp. subtilis NCIB3610T, Bacillus aquimaris TF12T, and Bacillus marisflavi TF11T, with 16S rRNA similarities of 96.0%, 98.4%, and 98.7%, respectively. DNA-DNA relatedness values between the isolate and the reference strains were ≤ 42±3%. The predominant menaquinones were MK-5 (50%) and MK-7 (50%). The major polar lipids were phosphatidylglycerol, diphosphatidylglycerol, and phosphatidylethanolamine. The major cellular fatty acids were iso-C15:0 (48.6%) and anteiso-C15:0 (20.6%), and the cell-wall diamino acid was meso-diaminopimelic acid. On the basis of the 16S rRNA gene sequence and the chemotaxonomic and phenotypic characteristics, it is concluded that strain R1T represents a novel species of the genus Bacillus, for which we propose the name Bacillus oryzaecorticis sp. nov. The type strain is R1T (=KACC 17217T =KCCM 90231T=JCM 19602T).
International Journal of Systematic and Evolutionary Microbiology 05/2014; 64(Pt 8). DOI:10.1099/ijs.0.058768-0 · 2.51 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: For the development of hardy kiwi wine, we arranged for the post-maturity of hardy kiwi fruit, treated them with calcium carbonate and a pectinase enzyme complex, investigated the resulting physicochemical properties and conducted a sensory evaluation. The period determined for creating post-maturity in the hardy kiwi fruit was determined as 5 days storage at room temperature following maturity. During this time the yield of fruit juice was increased from 22.1% to 53.5% using 0.1% (v/v) cytolase PCL5 for 2 h at room temperature. 0.1% (w/v) calcium carbonate was also added during the process of aging, for the reduction of the sour taste. The fermentation trial of the hardy kiwi wine was prepared using water (25% or 50%), sugar (), 0.1% (w/v) , 0.1% (v/v) cytolase PCL5, (200 ppm), and yeast ( cell/ml). Fermentation then occurred for 2 weeks at . The pH value, total acidity, alcohol, and reducing sugar content of the resulting hardy kiwi wines of 25% (v/w) and 50% (v/w) water, were in a range of pH 3.4-3.7, 1.12-1.21%, 14.3-14.4%, and 15-16 g/l, respectively. Citric acid and fructose constituted the major organic acids and the free sugar of the 25% and 50% hardy kiwi wine, respectively. Volatile flavor components, including 10 kinds of esters, 8 kinds of alcohols, 5 kinds of acids, 3 kinds of others and aldehydes, were determined by GC analysis. The results of sensory evaluation demonstrated that 50% hardy kiwi wine is more palatable than 25% hardy kiwi wine.
Korean Journal of Microbiology and Biotechnology 09/2013; 41(3). DOI:10.4014/kjmb.1301.01002
[Show abstract][Hide abstract] ABSTRACT: Bacterial communities derived from cheonggukjang and raw rice straw collected from a Mireuksan farm and a Heungup cheonggukjang in Gangwon province were investigated using both culture-based method and denaturing gradient gel electrophoresis (DGGE) analysis. Pure cultures, which were isolated from raw rice straw and cheonggukjang and cultured on tryptic soy agar plates (53-76 colonies per plate), were identified by analysis of 16S rRNA sequences. The traditional culture-based method and analysis of PCR-amplified 16S rRNA by DGGE revealed that for samples collected from the Mireuksan farm, Pantoea agglomerans and Bacillus subtilis were the predominant species in the raw rice-straw and cheonggukjang, respectively. For samples collected from the Heungup cheonggukjang, Bacillus amyloliquefaciens was the predominant species in both raw rice straw and cheonggukjang. Other microorganisms, including members of Pantoea, Bacillus, Enterococcus, Enterobacter, Pseudomonas, Rhodococcus, and Acinetobacter, were also present in the raw rice-straw and cheonggukjang, as were bacteria that could not be cultured.
Korean Journal of Food Science and Technology 08/2013; 45(4). DOI:10.9721/KJFST.2013.45.4.515
[Show abstract][Hide abstract] ABSTRACT: A bacterium producing a fibrinolytic enzyme was isolated from Cheonggukjang. The bacterium was identified as a strain of Bacillus amyloliquefaciens by 16S rDNA analysis and designated as B. amyloliquefaciens HC188. The optimum culture medium appeared to be one containing 0.5% (w/v) maltose and 0.5% (w/v) soytone. Bacterial growth in the optimal medium at reached the stationary phase after 27 h of incubation and the fibrinolytic enzyme showed optimum activity at 24 h. The enzyme was purified by 20-80% ammonium sulfate precipitation, CM Sepharose fast flow ion exchange chromatography, and Sephacryl S-200HR column chromatography. Its specific activity was 38359.3 units/mg protein and the yield was 5.5% of the total activity of the crude extracts. The molecular weight was 24.7 kDa and the amino acids of the N-terminal sequence were AQSVPYGVSQIKAPA. The fibrinolytic enzyme activity had an optimum temperature of and an optimum pH of 8.0, and the enzyme was stable in the ranges and pH 6.0-8.0. Enzyme activity was increased by and but inhibited by , EDTA, and PMSF. It is suggested that the purified enzyme is a metallo-serine protease.
Korean Journal of Microbiology and Biotechnology 03/2013; 41(1). DOI:10.4014/kjmb.1208.08010
[Show abstract][Hide abstract] ABSTRACT: The bacterial community of Chungkookjang and raw rice-straw collected from various areas in South Korea was investigated using both culture-dependent and culture-independent methods. Pure cultures were isolated from Chungkookjang and raw rice-straw on tryptic soy agar plates with 72 to 121 colonies and identified by 16S rDNA gene sequence analysis, respectively. The traditional culture-based method and denaturing gradient gel electrophoresis analysis of PCR-amplified 16S rDNA confirmed that Pantoea agglomerans and B. subtilis were identified as predominant in the raw rice-straw and Chungkookjang, respectively, from Iljuk district of Gyeonggi province, P. ananatis and B. licheniformis were identified as predominant in the raw rice-straw and Chungkookjang from Wonju district of Gangwon province, and Microbacterium sp. and B. licheniformis were identified as predominant in the raw rice-straw and Chungkookjang from Sunchang district of Jeolla province. Other strains, such as Bacillus, Enterococcus, Pseudomonas, Rhodococcus, and uncultured bacteria were also present in raw rice-straw and Chungkookjang.
Practical Application: A comprehensive analysis of these microorganisms would provide a more detailed understanding of the biologically active components of Chungkookjang and help improve its quality. Polymerase chain reaction-denaturing gradient gel electrophoresis analysis can be successfully applied to a fermented food to detect unculturable or more species than the culture-dependent method. This technique is an effective and convenient culture-independent method for studying the bacterial community in Chungkookjang. In this study, the bacterial community of Chungkookjang collected from various areas in South Korea was investigated using both culture-dependent and culture-independent methods.
[Show abstract][Hide abstract] ABSTRACT: A swarming and moderately halotolerant bacterium, designated strain A1-2(T), was isolated from the intestinal tract of the earthworm Eisenia fetida L. Cells were endospore-forming rods that were facultatively anaerobic, catalase-positive, oxidase-negative and motile by peritrichous flagella. The isolate grew optimally at 30 °C and pH 7.0, and could grow with up to 9 % (w/v) NaCl. Phylogenetic analysis of 16S rRNA gene sequences indicated that strain A1-2(T) belonged to the genus Bacillus and exhibited 16S rRNA gene sequence similarities of 96.8, 96.0, 96.0, 96.4 and 96.7 % with Bacillus drentensis LMG 21831(T), B. horneckiae PT-45(T), B. niacini BAC 1015, B. infantis SMC 4352-1(T) and B. shackletonii LMG 18435(T), respectively. DNA-DNA relatedness values between the isolate and the reference strains were ≤38.3 %. The DNA G+C content of strain A1-2(T) was 38.5 mol%. The predominant menaquinone was MK-7 and the major polar lipids were diphosphatidylglycerol and phosphatidylethanolamine. The major cellular fatty acids were iso-C(15 : 0) (51.5 %) and anteiso-C(15 : 0) (29.6 %) and the cell-wall diamino acid was meso-diaminopimelic acid. On the basis of 16S rRNA gene sequence analysis and chemotaxonomic and phenotypic characteristics, it is concluded that strain A1-2(T) represents a novel species of the genus Bacillus, for which we propose the name Bacillus eiseniae sp. nov. The type strain is A1-2(T) ( = KCCM 90092(T) = JCM 16993(T)).
International Journal of Systematic and Evolutionary Microbiology 10/2011; 62(Pt 9):2077-83. DOI:10.1099/ijs.0.034892-0 · 2.51 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The bacterial communities in the intestinal tracts of earthworm were investigated by culture-dependent and - independent approaches. In total, 72 and 55 pure cultures were isolated from the intestinal tracts of earthworms under aerobic and anaerobic conditions, respectively. Aerobic bacteria were classified as Aeromonas (40%), Bacillus (37%), Photobacterium (10%), Pseudomonas (7%), and Shewanella (6%). Anaerobic bacteria were classified as Aeromonas (52%), Bacillus (27%), Shewanella (12%), Paenibacillus (5%), Clostridium (2%), and Cellulosimicrobium (2%). The dominant microorganisms were Aeromonas and Bacillus species under both aerobic and anaerobic conditions. In all, 39 DNA fragments were identified by polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) analysis. Aeromonas sp. was the dominant microorganism in feeds, intestinal tracts, and casts of earthworms. The DGGE band intensity of Aeromonas from feeds, intestinal tracts, and casts of earthworms was 12.8%, 14.7%, and 15.1%, respectively. The other strains identified were Bacillus, Clostridium, Enterobacter, Photobacterium, Pseudomonas, Shewanella, Streptomyces, uncultured Chloroflexi bacterium, and uncultured bacterium. These results suggest that PCR-DGGE analysis was more efficient than the culture-dependent approach for the investigation of bacterial diversity and the identification of unculturable microorganisms.
Journal of Microbiology and Biotechnology 09/2011; 21(9):885-92. · 1.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The bacterial communities in the food, intestines, and feces of earthworms were investigated by PCR-denaturing Gradient gel electrophoresis (DGGE). In this study, PCR-DGGE was optimized by testing 6 universal primer sets for microbial 16S rRNA in 6 pure culture strains of intestinal microbes in earthworms. One primer set effectively amplified 16S rRNA from bacterial populations that were found in the food, intestines, and feces of earthworms. Compared with the reference markers from the pure culture strains, the resulting DGGE profiles contained 28 unique DNA fragments. The dominant microorganisms in the food, intestines, and feces of earthworms included Rhodobacterales bacterium, Fusobacteria, Ferrimonas marina, Aeromonas popoffii, and soil bacteria. Other straisn, such as Acinetobacter, Clostridium, and Veillonella, as well as rumen bacteria and uncultured bacteria also were present. These results demonstrated that PCR-DGGE analysis can be used to elucidate bacterial diversity and identify unculturable microorganisms.
The Journal of Microbiology 08/2011; 49(4):544-50. DOI:10.1007/s12275-011-0423-8 · 1.44 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We analyzed the bacterial community structure of the intestines of earthworms and determined the effect of enzyme producing microorganisms on the biomass of earthworms in vermicompost. Fifty-seven bacterial 16S rDNA clones were identified in the intestines of earthworms by using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) analysis. Entomoplasma somnilux and Bacillus licheniformis were the dominant microorganisms; other strains included Aeromonas, Bacillus, Clostridium, Ferrimonas, and uncultured bacteria. Among these strains, Photobacterium ganghwense, Aeromonas hydrophila, and Paenibacillus motobuensis were enzyme-producing microorganisms. In the mixtures that were inoculated with pure cultures of A. hydrophila WA40 and P. motobuensis WN9, the highest survival rate was 100% and the average number of earthworms, young earthworms, and cocoons were 10, 4.00-4.33, and 3.00-3.33, respectively. In addition, P. motobuensis WN9 increased the growth of earthworms and production of casts in the vermicompost. These results show that earthworms and microorganisms have a symbiotic relationship.
[Show abstract][Hide abstract] ABSTRACT: Bacillus subtilis CF92, an isolate from cattle feces, produces phytase, which catalyzes the hydrolysis of phytic acid into myo-inositol and inorganic phosphates. Phytase from B. subtilis CF92 was purified via ethanol precipitation, anion-exchange chromatography, and gel filtration chromatography. Molecular weight of the purified phytase was estimated to be 46 kDa by SDS-PAGE. Purified phytase exhibited optimal activity at 60°C. The enzyme retained 40% of its original activity after 30 min incubation at 80°C. Optimum pH was 7.0, although activity remained fairly stable over pH range of 4.0 to 8.0. The enzyme was activated in the presence of EDTA and significantly inhibited by metal ions. Phytase exhibited substrate-specificity on polyphosphate compounds such as adenosine triphosphate, sodium tripolyphosphate, and sodium phytate. Km and Vmax values for sodium phytate were 0.42 mM and 4.35 μmol/min, respectively.
Journal of the Korean Society for Applied Biological Chemistry 02/2011; 54(1). DOI:10.3839/jksabc.2011.012 · 0.69 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A B cell-specific growth substance (BGS) was isolated from the slime layer of Bacillus licheniformis E1. Unlike LPS, the BGS was not affected by polymixin B, an inhibitor of LPS, or by TLR4, and resulted in the growth of B cells. When BALB/c mice were treated with the BGS, the B cell population was found to increase in both the bone marrow and the spleen, with a marked increase after 24 h in the bone marrow and after 48 h in the spleen. When using antibodies to B cell lineage-restricted surface molecules to analyze the B cell population changes resulting from treatment with the BGS, an increase in immature B cells (IgM(+) and AA4.1(+)) and mature B cells (IgM(+) and IgD(+)) was found in the bone marrow 24 h after treatment with the BGS, whereas a decrease in mature B cells and increase in IgG(+) B cells were found in the spleen. When the BGS and OVA antigen were injected into the peritoneal cavity of BALB/c mice, this resulted in a high OVA-specific antibody titer in the sera, similar to that induced by aluminum hydroxide. Therefore, it is anticipated that the mass production of the BGS by B. licheniformis E1 could be used for studies of B cells in immunology, and contribute to the development of a new adjuvant for vaccine manufacture.
Journal of Microbiology and Biotechnology 02/2009; 19(1):55-64. DOI:10.4014/jmb.0806.396 · 1.53 Impact Factor