• 한국미생물·생명공학회지
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• 제51권3호
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• pp.289-292
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• 2023

Thermotolerant Bacillus subtilis NIB353 was isolated from Nuruk, a traditional Korean fermentation starter. The complete B. subtilis NIB353 genome sequence was obtained using MinION and Illumina (MiSeq) platforms. The B. subtilis NIB353 genome sequence was 4,247,447 bp with a GC content of 43%. The B. subtilis NIB353 strain exhibited orthologous average nucleotide identity values of 98.39% and 98.38% with B. subtilis 168 and B. subtilis ATCC6051a, respectively. The genome has been deposited in GenBank under the accession number NZ_CP089148.1.

• Journal of Animal Science and Technology
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• 제64권2호
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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.

• 한국균학회지
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• 제49권4호
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• pp.441-454
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• 2021

Fungal diseases including anthracnose, stem rot, blight, wilting, and root rot of crops are caused by phytopathogens such as Colletotrichum species, Sclerotinia sclerotiorum, Phytophthora species, and Fusarium oxysporum and F. solani which threaten the production of chili pepper. In this study, to identify biological control agents (BCAs) of phytopathogenic fungi, potentially useful Bacillus species were isolated from the field soils. We screened out five Bacillus strains with antagonistic capacity that are efficiently inhibiting the growth of phytopathogenic fungi. Bacillus species were characterized by the production of extracellular enzymes, siderophores, and indole-3-acetic acid (IAA). Furthermore, the influence of bacterial strains on the plant growth promoting activity and seedling vigor index were assessed using Brassica juncea as a model plant. Inoculation with Bacillus subtilis SRCM 121379 significantly increased the length of B. juncea shoots and roots by 45.6% and 52.0%, respectively. Among the bacterial isolates, Bacillus subtilis SRCM 121379 showed the superior enzyme activities, antagonistic capacity and plant growth promoting effects. Based on the experimental results, Bacillus subtilis SRCM 121379 (GenBank accession no. NR027552) was finally selected as a BCA candidate.

• 한국버섯학회지
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• 제21권3호
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• pp.140-144
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• 2023

본 연구는 길항성 미생물, Bacillus subtilis BSM320을 이용한 표고 푸른곰팡이병의 생물학적 방제를 목적으로 하였다. B. subtilis BSM320는 표고 톱밥배지 에서 분리된 T. hazianum을 포함한 3 Trichoderma 분리균주 대해 균사 생장을 억제하는 항균 활성을 나타냈다. B. subtilis BSM320은 1% glucose를 포함하는 LeCSMS물 추출물(LeCSMS WE)에서 4.2 × 10⁹±113.7 cfu/mL 세균수로 고밀도 배양 되었으며 상용배지 Lurina-Bertani broth 배지보다 높은 세균 밀도를 보였다. B. subtilis BSM320의 LeCSMS WE 배양액은 표고 자실체에 약해 없이 표고 톱밥 배지에 발생하는 푸른곰팡이병에 대하여 75%의 방제효과를 보였다. 그 결과는 LeCSMS 물 추출물 배양 B. subtilis BSM320는 표고 푸른곰팡이병의 생물학적 방제에 활용할 수 있는 것을 시사한다.

• Journal of Microbiology and Biotechnology
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• 제25권7호
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• pp.1015-1025
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• 2015

The development of diverse polymyxin derivatives is needed to solve the toxicity and resistance problems of polymyxins. However, no platform has generated polymyxin derivatives by genetically engineering a polymyxin synthetase, which is a nonribosomal peptide synthetase. In this study, we present a two-step approach for the construction of engineered polymyxin synthetases by substituting the adenylation (A) domains of polymyxin A synthetase, which is encoded by the pmxABCDE gene cluster of Paenibacillus polymyxa E681. First, the seventh L-threonine-specific A-domain region in pmxA was substituted with the L-leucine-specific A-domain region obtained from P. polymyxa ATCC21830 to make polymyxin E synthetase, and then the sixth D-leucine-specific A-domain region (A_6-D-Leu-domain) was substituted with the D-phenylalanine-specific A-domain region (A_6-D-Phe-domain) obtained from P. polymyxa F4 to make polymyxin B synthetase. This step was performed in Escherichia coli on a pmxA-containing fosmid, using the lambda Red recombination system and the sacB gene as a counter-selectable marker. Next, the modified pmxA gene was fused to pmxBCDE on the chromosome of Bacillus subtilis BSK4dA, and the resulting recombinant strains BSK4-PB and BSK4-PE were confirmed to produce polymyxins B and E, respectively. We also succeeded in constructing the B. subtilis BSK4-PP strain, which produces polymyxin P, by singly substituting the A_6-D-Leu-domain with the A_6-D-Phe-domain. This is the first report in which polymyxin derivatives were generated by genetically engineering polymyxin synthetases. The two recombinant B. subtilis strains will be useful for improving the commercial production of polymyxins B and E, and they will facilitate the generation of novel polymyxin derivatives.

• 한국식품과학회지
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• 제44권3호
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• pp.312-316
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• 2012

막걸리에서 분리된 B. subtilis의 특성을 파악하고자 영양유무(buffer와 TSB), pH(pH 7과 4, 막걸리 pH 4), adaptation 조건에 따라 초고압에 의한 저감 효과를 살펴보았다. B. subtilis 포자는 영양유무, pH에 관계없이 환경 적응력이 있었고, 초고압에 의한 저감효과도 미미하게 관찰되었다. B. subtilis 영양세포는 영양유무와 pH에 따라 대조구(non-treatment)의 균수 차이가 관찰되었고, 초고압에 의한 저감 효과도 다르게 나타났다. B. subtilis 영양세포는 저온에서 영양과 pH가 불리한 환경에서 endospore를 형성하여 적응을 하게 되고 초고압에 의한 저감효과는 감소하는 것으로 나타났다. 특히 막걸리에 접종한 경우는 buffer(pH 4)와 TSB(pH 4)보다 적응력이 약한 것으로 관찰되었다. B. subtilis 포자를 접종한 막걸리의 저장 실험 결과, 내열성이 있으며 균의 증식은 관찰되지 않았다. 결론적으로, 막걸리에서 B. subtilis는 대부분 포자상태로 존재하며, 포자는 초고압에 대한 저항력이 큰 것을 알 수 있었다. 그러나 초고압은 영양세포 사멸에 효과적이었다.

• 한국미생물·생명공학회지
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• 제48권4호
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• pp.439-446
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• 2020

Two Bacillus strains, K3 and K208, both demonstrating strong fibrinolytic activities were isolated from Kimchi, a traditional Korean preparation of fermented vegetables. Isolates were subjected to various molecular biology based identification methods including RAPD-PCR and identified as B. subtilis and B. velezensis, respectively. Tryptic soy broth (TSB) was found to best maintain both the growth and the fibrinolytic activity of these strains. Culture supernatants were analyzed by SDS-PAGE and fibrin zymography, and the results indicate that a 40 and 27 kDa band seem to be responsible for the fibrinolytic activities of these two isolates and the 27 kDa band was subsequently identified as the mature form of AprE, the major fibrinolytic enzyme. Thus the aprE genes were cloned and the translated amino acid sequences demonstrated 99.3% identity with each other, and 86.5% identity with BsfA, a fibrinolytic enzyme from B. subtilis ZA400 also isolated from Kimchi, and AprE2, a fibrinolytic enzyme from B. subtilis CH3-5 isolated from Cheonggukjang, a traditional Korean fermented soy. Given this B. subtilis K3 and B. velezensis K208 may be promising starter cultures in the production of fermented foods.

• 한국식품영양과학회지
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• 제37권12호
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• pp.1576-1582
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• 2008

기존 칼슘보충용식품의 성장촉진효능을 증진시키기 위하여 칼슘흡수 관여 물질인 $\gamma$-PGA를 다량 생산하는 Bacillus subtilis SE4 균주를 선발하였다. 선발된 B. subtilis SE4의 발효대사물을 기존 칼슘보충용식품에 첨가하여 성장기 수컷 흰쥐에 28일간 경구투여 한 결과 발효대사물이 첨가된 신규칼슘보충용식품군에서 혈청 내 IGF-I 함량이 대조군이나 기존칼슘보충용식품군에 비해 유의적으로 높게 나타났다(p<0.05). 일일증체량, 일일사료섭취량, 장골길이, 장골 무게, 장골폭 등의 성장관련 지표에서도 장골의 길이와 일부 부위별 두께에서의 유의적 결과를 포함하여 전반적으로 신규칼슘보충용식품군이 대조군과 기존칼슘보충용식품군에 비해 양호한 성적을 나타내었다. 이러한 결과는 기존칼슘 보충용 식품에 발효대사물을 첨가함으로써 나타난 칼슘의 생체흡수율 증대 및 성장호르몬의 분비활성화 등 여러 성장 관련요인들이 흰쥐 성장에 긍정적으로 작용한 때문으로 생각되어진다.

• Mycobiology
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• 제40권1호
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• pp.59-65
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• 2012

Antagonistic microorganisms against $Rhizoctonia$ $solani$ were isolated and their antifungal activities were investigated. Two hundred sixteen bacterial isolates were isolated from various soil samples and 19 isolates were found to antagonize the selected plant pathogenic fungi with varying degrees. Among them, isolate C9 was selected as an antagonistic microorganism with potential for use in further studies. Treatment with the selected isolate C9 resulted in significantly reduced incidence of stem-segment colonization by $R.$ $solani$ AG2-2(IV) in Zoysia grass and enhanced growth of grass. Through its biochemical, physiological, and 16S rDNA characteristics, the selected bacterium was identified as $Bacillus$ $subtilis$ subsp. $subtilis$. Mannitol (1%) and soytone (1%) were found to be the best carbon and nitrogen sources, respectively, for use in antibiotic production. An antibiotic compound, designated as DG4, was separated and purified from ethyl acetate extract of the culture broth of isolate C9. On the basis of spectral data, including proton nuclear magneric resonance ($^1H$ NMR), carbon nuclear magneric resonance ($^{13}C$ NMR), and mass analyses, its chemical structure was established as a stereoisomer of acetylbutanediol. Application of the ethyl acetate extract of isolate C9 to several plant pathogens resulted in dose-dependent inhibition. Treatment with the purified compound (an isomer of acetylbuanediol) resulted in significantly inhibited growth of tested pathogens. The cell free culture supernatant of isolate C9 showed a chitinase effect on chitin medium. Results from the present study demonstrated the significant potential of the purified compound from isolate C9 for use as a biocontrol agent as well as a plant growth promoter with the ability to trigger induced systemic resistance of plants.

• Journal of Microbiology and Biotechnology
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• 제30권11호
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• pp.1720-1728
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• 2020

We have previously characterized AprESJ4, the major fibrinolytic enzyme from Bacillus subtilis SJ4 (Yao et al., 2019). During that study, we observed a 68 kDa protein with fibrinolytic activity. In this study, we cloned the gene (vprSJ4) encoding the 68 kDa protein, a mature Vpr and minor protease secreted by Bacillus species. vprSJ4 encodes a preproenzyme consisting of 810 amino acids (aa) including signal sequence (28 aa) and prosequence (132 aa). The mature enzyme (650 aa) has a predicted molecular weight of 68,467.35. Unlike Vprs from other B. subtilis strains, VprSJ4 has 4 additional amino acids (DEFA) at the C-terminus. vprSJ4 was overexpressed in Escherichia coli. PreproVprSJ4 was localized in inclusion bodies, and subjected to in vitro renaturation and purification by an affinity column. SDS-PAGE and western blot showed that autoprocessing of preproVprSJ4 occurred and 68 kDa and smaller proteins were produced. The optimum pH and temperature of the recombinant VprSJ4 were pH 7.0 and 40℃, respectively. Kinetic parameters of recombinant VprSJ4 were measured by using an artificial substrate, N-succinyl-ala-ala-pro-phe-p-nitroanilide. Coexpression of vprSJ4 and aprESJ4 using pHY300PLK increased the fibrinolytic activity a further 117% when compared with aprESJ4 single expression using the same vector in B. subtilis WB600.