• Korean Journal of Food Science and Technology
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• v.44 no.5
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• pp.600-606
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• 2012

A Bacillus sp. that produces fibrinolytic enzyme was isolated from Cheonggukjang, a traditional Korean soybean-fermented food. According to 16S rRNA gene base sequencing, the bacillus was identified as a variety of Bacillus subtilis, and named Bacillus subtilis IDCC 9204. Fibrinolytic enzyme NK-IL9204 was stable up to $60^{\circ}C$ and within pH range of 5-10. Purified NK-IL9204 was detected through fibrin zymography. The molecular weight and isoelectric point of the enzyme were estimated to be 27.7 kDa and 6.7 by SDS-PAGE and 2D electrophoresis, respectively. Its amino acid sequence was similar to that of nattokinase (identities 99.5%) and different from that of nattokinase BPN (identities 86.4%). The plasma fibrinolytic activity of NK-IL9204 was measured by euglobulin clot lysis times (ECLT). The NK-IL9204 was orally administered to SD rats for 3 weeks (1,000 FU/rat/day). The ECLT was significantly shortened by supplementation of NK-IL9204.

• The Korean Journal of Mycology
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• v.42 no.3
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• pp.225-230
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• 2014

Sclerotinia sclerotiorum, a plant pathogenic fungus, can cause serious yield and quality losses in the winter lettuce field. For biological control of S. sclerotiorum, soil-born microorganisms that inhibit the mycelia growth of S. sclerotiorum and Fusarium oxysporum were isolated from diseased soil. Among the isolates, bacterial isolate, GG95, which was identified as Bacillus subtilis according to the morphological, physiological characteristics and by 16S rRNA similarity, showed the highest level of inhibitory activity. The growth conditions for B. subtilis GG95 were optimized in TSB media (pH 7) by culturing at $28^{\circ}C$ for 24 hrs. Maltose or fructose and peptone were selected as the best carbon and nitrogen sources, respectively. Greenhouse experiment was performed to test effectiveness of B. subtilis GG95 in the control sclerotinia rot. Drench application ($1{\times}10^8cfu/mL$, 3 times) of the bacterial culture broth to lettuce showed an effectiveness value of 88%, suggesting that B. subtilis GG95 would be a promising biocontrol agent for control of sclerotinia rot.

• Journal of the Korean Society of Food Science and Nutrition
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• v.40 no.12
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• pp.1694-1699
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• 2011

The quality characteristics and physiological effects of chunggukjang fermented naturally (NF-c), with Bacillus subtilis-SKm (BS-c), with Bacillus subtilis HJ18-4 (BH-c), and with Bacillus subtilis KCCM 42923 (BK-c) were investigated. The characteristics of fermentation were determined by protease, ${\alpha}$-amylase and ${\gamma}$-GTP activities, and additionally the amounts of amino-type and ammonia-type nitrogens. BS-c showed the highest protease, ${\alpha}$-amylase, and ${\gamma}$-GTP activities, and also amino-type nitrogen content among the four types of chunggukjang. The ammonia-type nitrogen content in BS-c was similar to that of BK-c and NF-c. BH-c showed the lowest enzyme activities and amino-type and ammonia-type nitrogen content. BS-c, BH-c, BK-c, and NF-c showed a similar overall acceptability during sensory evaluation. BS-c also showed the strongest DPPH free radical scavenging and anti-proliferative activities in HT-29 human colon carcinoma cells. These results suggested that B. subtilis-SKm was suitable to be used as a starter to enhance the quality and effects of chunggukjang.

• Journal of Animal Science and Technology
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• v.64 no.2
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• pp.291-301
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• 2022

The objective of this study was to evaluate the effects of different mixing ratios of Bacillus licheniformis and Bacillus subtilis in diets on nutrient digestibility, fecal microflora, and odor gas emissions of growing pigs. A total of four crossbred ([Landrace × Yorkshire] × Duroc) barrows with average body weight (BW) of 41.2 ± 0.7 kg were randomly allotted four diets over four periods in a 4 × 4 Latin square design. Treatments were as follows: Control (CON, basal diet), CON + 0.2% probiotic complex (L4S6, B. licheniformis and B. subtilis at a 4:6 ratio), CON + 0.2% probiotic complex (L5S5, B. licheniformis and B. subtilis at a 5:5 ratio), CON + 0.2% probiotic complex (L6S4, B. licheniformis and B. subtilis at a 6:4 ratio). Dietary probiotic supplementation showed higher crude protein (CP) digestibility values and lower Escherichia coli counts in fecal samples than the CON group (p < 0.05). There was no significant difference in NH₃ or H₂S emission until day 3. The positive effect of H₂S and NH₃ emissions was detected earlier with the L4S6 and L5S5 compared to the L6S4, which had a lower ratio of B. subtilis. Both the L4S6 and L5S5 probiotic complexes significantly decreased the fecal H₂S and NH₃ emission in days 4 and 6 (p < 0.05). On day 7, all probiotic complexes decreased (p < 0.05) H₂S and NH₃ emissions than the CON group. Our results agreed that the dietary supplementation of Bacillus licheniformis and Bacillus subtilis complexes in growing pigs can significantly improve CP digestibility and reduce fecal E. coli counts, NH₃ and H₂S emissions. Notably, the higher mixing ratio of Bacillus subtilis in probiotic supplementation is more effective in reducing the odor of manure.

• Korean Journal of Environmental Biology
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• v.22 no.4
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• pp.487-493
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• 2004

To investigate the antifungal activity related protein in pesticidal bacteria, a bacterial strain LTD was isolated from soil collected at Gimje in Jeonbuk province, Korea, and identified as Bacillus subtilis LTD based on a API50 CHB kit and 168 rDNA sequencing. It has an antifungal activity against 9 plant pathogenic fungi in a paper disc assay. The antifungal activity- deficient mutant, B. subtilis mLTD was induced at a 5 kGy dose of $^{60}Co$ gamma radiation. Using the two-dimensional electrophoresis and the matrix assisted laser desorption ionization time-of-flight mass spectrometry, the comparison analysis of proteins between the wild and mutant were performed. A major intracellular serine proteinase IspA (MW: 32.5 kDa), a NAD (P) H dehydrogenase (MW: 20.0 kDa), and a stage II sporulation protein AA, SpoIIAA (MW: 14.3kDa) were detected only in the B. subtilis LTD. These results suggested that the functions of these proteins found only in the B. subtilis LTD could. be closely related to the antifungal activity against plant pathogenic fungi.

• Journal of Microbiology and Biotechnology
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• v.20 no.1
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• pp.138-145
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• 2010

A bacterial strain isolated from soil for its potential to control the anthracnose disease caused by Colletotrichum gloeosporioides was identified as a Bacillus subtilis. Bacillus subtilis CMB32 produced antifungal agents on M9 broth at $30^{\circ}C$. Biosurfactant lipopeptides produced by Bacillus subtilis CMB32 were precipitated by adjusting to pH 2 and extracting using chloroform/methanol, and then were purified using column chromatography and reverse-phase HPLC. The molecular masses of the lipopeptides were estimated by MALDI-TOF mass spectrometry as (a) 1,080, (b) 1,486, and (c) 1,044 Da, respectively. They had cyclic structures and amino acid compositions of (a) Pro, Asx, Ser, Tyr, Glx, (b) Glx, Tyr, Thr, Ala, Pro, lie, and (c) Glx, Leu, Val, Asx, respectively. Further analysis revealed that Bacillus subtilis CMB32 produced three antifungal lipopeptides: (a) iturin A, (b) fengycin, and (c) surfactin A.

• Journal of Microbiology and Biotechnology
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• v.9 no.2
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• pp.230-233
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• 1999

A plasmid vector was constructed for the expression and secretion of Bacillus macerans cyclodextrin glucanotransferase (CGTase) in Bacillus subtilis. The vector, pUBACGT, was composed of the ribosome-binding sequence, signal sequence, and cgt gene from B. macerans under the control of amyR2, the promoter of amyE gene coding for $\alpha$-amylase from B. subtilis var. natto. Bacillus subtilis LKS88, a mutant strain lacking genes for an amylase and two proteases, was used as a host for the transformation of the plasmid vector. The transformants were selected on kanamycin-containing Luria-Bertani plates. The starch hydrolyzing activity was observed on the starch-containing plates by the iodine method and cyclodextrin-forming activity was detected in the culture medium. A SDS-PAGE analysis showed that most of the expressed CGTase in the recombinant B. subtilis was secreted into the medium at a high expression level.

• Journal of Applied Biological Chemistry
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• v.57 no.2
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• pp.183-188
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• 2014

In order to increase the nutritional quality of soybean meal (SBM) as an animal feed, Bacillus subtilis TP6, a previously isolated strain from an Indonesian traditional fermented soybean food, Tempeh, was used as a starter organism for solid-state fermentation. In the pre-treated SBM with water content of 60% (v/w), B. subtilis TP6 was grown to a maximum viable cell number of $3.5{\times}10^9CFU/g$. Compared to control, crude protein in Bacillus fermented SBM was increased by 16%, while raffinose, stachyose, and trypsin inhibitors were reduced by 31, 37, and 90%, respectively. The Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis showed that proteins in the fermented SBM were remarkably hydrolyzed into smaller molecular masses, resulting in a decrease in large sized proteins. Our data suggested that B. subtilis fermentation could increase the nutritive value of SBM through reduction of anti-nutritive factors and improvement of protein quality by hydrolysis of soy protein. In addition, B. subtilis TP6 produced a functional ingredient, poly-${\gamma}$-glutamic acid which has various health benefits.

• Journal of Animal Science and Technology
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• v.64 no.3
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• pp.443-461
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• 2022

This study was conducted to assess the effects of the dietary supplementation of riboflavin (as a bile salt hydrolase [BSH] inhibitor) and Bacillus subtilis on growth performance and woody breast of male broilers challenged with Eimeria spp. Intestinal bacteria, including supplemented probiotics, can produce BSH enzymes that deconjugate conjugated bile salts and reduce fat digestion. A 3 × 2 × 2 (riboflavin × Bacillus subtilis × Eimeria spp. challenge) factorial arrangement of treatments in randomized complete block design was used. On d 14, birds were gavaged with 20× doses of commercial cocci vaccine (Coccivac^R -B52, Merck Animal Health, Omaha, NE). Dietary treatment of riboflavin and B. subtilis did not affect body weight (BW), body weight gain (BWG), and feed conversion (FCR) d 0 to 14 and overall d 0 to 41. Eimeria spp challenge reduced BWG, feed intake (FI), and increased FCR between d 14 to 28, but increased BWG and lowered FCR between d 28 to 35. There were no effects of the Eimeria spp. challenge on the overall d 0 to 41 FCR and FI, but BWG was reduced. Eimeria spp. challenge increased the abdominal fat pad weight and slight woody breast incidences on processed birds on d 42. Dietary inclusion of B. subtilis and riboflavin at tested levels did not help birds to mitigate the negative impact of Eimeria spp. challenge to enhance the growth performance.

• Food Science of Animal Resources
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• v.28 no.2
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• pp.181-186
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• 2008

This study was conducted to investigate the effects of supplementation with transgenic Bacillus subtilis secreting chitinase on the growth performance, nutrient digestibility, blood characteristics and carcass traits in finishing pigs. A total of sixty-four pigs ($50.82{\pm}0.82kg$, average initial body weight) were assessed over a period of 84 days. Dietary treatments included: 1) CON (basal diet without antibiotics), 2) AD (basal diet + 0.1% Virginiamycin), 3) CD0.5 (basal diet + 0.5% transgenic B. subtilis), 4) CD1.0 (basal diet + 1.0% transgenic B. subtilis). Each dietary treatment had 4 replicates of 4 pigs per pen in a randomized complete block design. In terms of growth performance, the ADG (average daily gain) and gain/feed ratio were significantly increased with the CD1.0 diet compared to the AD diet during weeks 0-4 (p<0.05). During weeks 4-8, the ADG was significantly increased with the AD diet compared to the CON and CD0.5 diets (p<0.05). The ADFI was also significantly increased with the AD diet compared to the other diets (p<0.05). During weeks $8{\sim}12$, the ADFI was significantly increased with the CON diet compared to the other diets (p<0.05), and the gain/feed ratio was significantly increased with the CD0.5 and CD1.0 diets compared to the AD diet (p<0.05). Over the entire 84 day test period, the ADFI was significantly increased with the AD diet compared to the CD0.5 and CD1.0 diets (p<0.05). The gain/feed ratio was significantly increased with the CD0.5 and CD1.0 diets compared to the CON diet (p<0.05). In terms of meat color, the L value was significantly increased with the CD0.5 diet compared to the CON and AD diets (p<0.05), and the a value was significantly increased with the CON diet compared to the other diets (p<0.05). In terms of sensory evaluation, meat color was significantly improved with the CON, CD0.5 and CD1.0 diets compared to the AD diet (p<0.05). Marbling was significantly increased with the CON diet compared to the other diets (p<0.05). Firmness was significantly increased with the CD0.5 diet compared to the AD diet (p<0.05). In conclusion, supplementation with transformed B. subtilis secreting chitinase improved gain/feed ratios and influenced meat color. Thus, we suggest that transformed B. subtilis secreting chitinase can partially substitute for antibiotics.