• Preventive Nutrition and Food Science
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• v.17 no.1
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• pp.64-70
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• 2012

Bacilli with fibrinolytic activities were isolated from traditionally-prepared Meju and some of these strains showed strong antifungal activities. One isolate, MJ1-4, showed the strongest antifungal activity. MJ1-4 and other isolates were identified as B. amyloliquefaciens strains by recA gene sequencing and RAPD-PCR results. B. amyloliqufaciens MJ1-4 efficiently inhibited an Aspergillus spp.-producing aflatoxin B1 ($AFB_1$) and a Penicillium spp.-producing ochratoxin (OTA) in addition to other fungi. Antifungal activity of B. amyloliquefaciens MJ1-4 culture reached its maximum (40 AU/mg protein) in LB or TSB medium around 48 hr at $37^{\circ}C$. Antifungal activity of the concentrated culture supernatant was not decreased significantly by protease treatments, implying that the antifungal substance might not be a simple peptide or protein. Considering its antifungal and fibrinolytic activities together, B. amyloliquefaciens MJ1-4 can serve as a starter for fermented soyfoods such as Cheonggukjang and Doenjang.

• Journal of Microbiology and Biotechnology
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• v.28 no.5
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• pp.784-795
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• 2018

Bacillus amyloliquefaciens Pc3 was isolated from Antarctic seawater with antifungal activity. In order to investigate the metabolic regulation mechanism in the biosynthesis of lipopeptides in B. amyloliquefaciens Pc3, GC/MS-based metabolomics was used when exogenous indole was added. The intracellular metabolite profiles showed decreased asparagine, aspartic acid, glutamine, glutamic acid, threonine, valine, isoleucine, hexadecanoic acid, and octadecanoic acid in the indole-treated groups, which were involved in the biosynthesis of lipopeptides. B. amyloliquefaciens Pc3 exhibited a growth promotion, bacterial total protein increase, and lipopeptide biosynthesis inhibition upon the addition of indole. Besides this, real-time PCR analysis further revealed that the transcription of lipopeptide biosynthesis genes ituD, fenA, and srfA-A were downregulated by indole with 22.4-, 21.98-, and 26.0-fold, respectively. It therefore was speculated that as the metabolic flux of most of the amino acids and fatty acids were transferred to the synthesis of proteins and biomass, lipopeptide biosynthesis was weakened owing to the lack of precursor amino acids and fatty acids.

• Research in Plant Disease
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• v.23 no.2
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• pp.177-185
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• 2017

Bacillus amyloliquefaciens LM11 was isolated from the feces of larvae of the rhino beetle and showed strong antifungal activities against various phytopathogenic fungi by producing biosurfactants. In this study, our overall goal was to determine relationship between biosurfactants produced from the LM11 strain and its role in growth inhibition of phytopathogenic fungi. Production and expression levels of B. amyloliquefaciens LM11 biosurfactants were significantly differed depending on growth phases. Transcriptional and biochemical analysis indicated that the biosurfactants of the LM11 strain were greatly enhanced in late log-phase to stationary phase. Inhibitions of phytopathogenic mycelial growth and spore germination were directly correlated (P<0.001, R=0.761) with concentrations of the LM11 cell-free culture filtrates. The minimum inhibitory surface tension of the culture filtrate of the B. amyloliquefaciens LM11 grown in stationary phase to inhibit mycelial growth of the phytopathogenic fungi was 38.5 mN/m (P<0.001, R=0.951-0.977). Our results indicated that the biosurfactants of B. amyloliquefaciens LM11 act as key antifungal metabolites in biocontrol of plant diseases, and measuring surface tension of the cell-free culture fluids can be used as an easy indicator for optimal usage of the biocontrol agents.

• Journal of Microbiology and Biotechnology
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• v.20 no.2
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• pp.370-374
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• 2010

A gene encoding the major secreted fibrinolytic protein of Bacillus amyloliquefaciens CH86-1 was cloned from genomic DNAs. DNA sequencing showed that the gene, aprE86-1, could direct the synthesis of a mature protein 275 amino acids in length after processing. When aprE86-1 was introduced into B. subtilis, a mature 27-kDa protein was produced as expected. The fibrinolytic activity of the B. subtilis transformant (TF) was higher than that of B. amyloliquefaciens CH86-1, showing the possibility of increasing the fibrinolytic activity of Bacillus strains through genetic engineering.

• Korean Journal of Microbiology
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• v.48 no.3
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• pp.220-224
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• 2012

In order to reduce harmful biogenic amines in the traditionally fermented soybean products, five isolates with biogenic amines-degrading property were obtained from 83 traditionally fermented soybean products. The strains were found to reduce biogenic amines including histamine, tyramine, putrescine, and cadaverine by 27 to 92% in the cooked soybean containing 5.3% of each biogenic amine over 10 days of incubation. The morphological and biochemical tests and the phylogenetic relationships based on 16S rRNA gene sequences indicated that the five isolates were most closely related to Bacillus subtilis or B. amyloliquefaciens. The use of selected strains would be a potential control measure in manufacturing traditionally fermented soybean products that are difficult to control biogenic amine levels.

• Korean Journal of Microbiology
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• v.50 no.1
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• pp.84-86
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• 2014

Mass production of biocontrol agent is an essential step for its commercial use. Media composition and culture conditions for production of Bacillus amyloliquefaciens SKU-78, a potential biocontrol agent against bacterial wilts, were optimized by a flask culture. Low cost media combining nitrogen and carbon sources were tested. Maximum cell growth (> $2{\times}10^9$ CFU/ml) was obtained in a medium of 5% soy flour combined with 3% corn starch after 24 h cultivation. The optimum initial pH, temperature and shaking speed was 5.5, $30^{\circ}C$ and 150-250 rpm, respectively. Fermentation of SKU-78 was scaled up in 30 L fermenter and the profiles of cell density, pH, dissolved oxygen and spore formation were recorded. After 8 h lag phase, exponential growth occurred and reached at maximum viable cell number ($1.2{\times}10^{11}$ CFU/ml) after 20 h. The SKU-78 strain grown in a low cost medium exhibited the high suppression of bacterial wilts. The results indicate that SKU-78 strain can be produced in a low cost medium and provide a basis for scaling up to industrial level.

• Journal of Microbiology and Biotechnology
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• v.23 no.7
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• pp.974-983
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• 2013

Bacillus amyloliquefaciens CB1 was isolated from cheonggukjang, a Korean fermented soy food. B. amyloliquefaciens CB1 secretes proteases with fibrinolytic activities. A gene homologous to aprE of Bacillus subtilis, aprECB1, was cloned from B. amyloliquefaciens CB1, and DNA sequencing showed that aprECB1 can encode a prepro-type serine protease consisting of 382 amino acids. When aprECB1 was introduced into B. subtilis WB600 using an E. coli-Bacillus shuttle vector, pHY300PLK, transformants showed fibrinolytic activity and produced a 28 kDa protein, the size expected for the mature enzyme. The 28 kDa fibrinolytic enzyme was purified from the culture supernatant of B. subtilis WB600 transformant. AprECB1 was completely inhibited by phenylmethylsulfonyl fluoride and almost completely inhibited by EDTA and EGTA, indicating that it is a serine metalloprotease. AprECB1 exhibited the highest specificity for N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide, a known substrate for ${\alpha}$-chymotrypsin. $A{\alpha}$ and $B{\beta}$ chains of fibrinogen were quickly degraded by AprECB1, but the ${\gamma}$-chain was resistant.

• Journal of Microbiology
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• v.40 no.1
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• pp.26-32
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• 2002

From the culture supernatant of the psychrotrophic strain of Bacillus amyloliquefaciens an extracellular serine protease was purified to apparent homogeneity by successive purification steps using QAE-Sephadex, SP-Sephadex and Sephacryl S-100 column chromatography. The pretense is monomeric, with a relative molecular mass of 23,000. It is inhibited by the serine protease inhibitor phenylmethylsulfonyl fluoride, but not by EDTA. The enzyme is most active at pH 9-10 and at $45^{\circ}C$, although it is unstable at $60^{\circ}C$.

• The Korean Journal of Mycology
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• v.41 no.4
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• pp.268-273
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• 2013

Bacillus amyloliquefaciens M27 was isolated from the cotton-waste compost for cultivation of oyster mushroom (Pleurotus ostreatus). B. amyloliquefaciens M27 is a biocontrol agent with antagonistic activities against a wide range of fungal pathogens. The aim of this work was to evaluate the possibility of exploiting antagonistic bacteria, B. amyloliquefaciens M27, in the biological control of the cucumber powdery mildew fungus, Podosphaera fusca. In greenhouse tests, the isolate was found to be very effective to control powdery mildew on cucumber leaves showing 4.0% diseased area, whereas diseased area in the control was 80.5%. The filtrate of the isolate cultured on MH and LB media were more effective for control of the disease than those cultured on TSB, NB, and KB media. When two, five, ten, 20, 50 and 100-fold diluted culture broth of isolate on LB media were treated, disease areas were 0%, 0%, 0%, 1.3%, 3.1% and 5.0%, respectively, whereas diseased area in the control was 60.0%. The filtrate of the isolate cultured on LB media was treated to cucumber plants on July, October and December just before the outbreak of the powdery mildew occurred. When 10-fold diluted filtrate of the isolate was treated, control efficacy was 88.9~98.9% in the treated seasons. The results showed that the culture filtrate of B. amyloliquefaciens M27 was very effective to control powdery mildew of cucumber.

• The Korean Journal of Mycology
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• v.37 no.1
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• pp.96-103
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• 2009

For the control of pathogenic microorganisms, Bacillus spp. were isolated from diseased pepper fruits in Korea. Among them, Bacillus sp. IUB158-03 showed high inhibitory effect on mycelial growth and spore germination of C. gloeosporioides and Botrytis cinerea. The strain was identified as B. amyloliquefaciens IUB158-03 based on its physiological, biochemical characteristics and Microlog analysis. The highest level of antifungal substances by B. amyloliquefaciens IUB158-03 were obtained when the bacterium was cultured in medium containing 2% soluble starch, 3% yeast extract, 0.5% tryptone, 0.5% $NH_4H_2PO_4$, and 1% NaCl (pH 6.0) at $25^{\circ}C$ for 72 hrs. The antifungal substances were purified by butanol extraction, silica gel column chromatography, preparative thin layer chromatography, and high performance liquid chromatography. The purified antifungal substance was confirmed $R_f$ 0.27 by TLC. This substance exhibited antifungal activity against Fusarium solani, Rhizoctonia solani, Botrytis cineria, Alternata alternaria of plant pathogenic fungi and Trichophyton mentagrophytes, Epidermophyton floccosum, Cryptococcus neoformans of human pathogenic fungi.