• Microbiology and Biotechnology Letters
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• v.26 no.2
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• pp.122-129
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• 1998

It were reported that antifungal mechanism of Enterobacter cloacae is a volatile ammonia that produced by the strain in soil, and the production of ammonia is related to the bacterial urease activity. A powerful bacterium SH14 against soil-borne pathogen Fusarium solani, which cause root rot of many important crops, was selected from a ginseng pathogen suppressive soil. The strain SH14 was identified as Bacillus subtilis by cultural, biochemical, morphological method, and $API^{circledR}$ test. From several in vitro tests, the antifungal substance that is produced from B. subtilis SH14 was revealed as heat-stable and low-molecular weight antibiotic substance. In order to construct the multifunctional biocontrol agent, the urease gene of Bacillus pasteurii which can produce pathogenes-suppressive ammonia transferred into antifungal bacterium. First, a partial BamH I digestion fragment of plasmid pBU11 containing the alkalophilic B. pasteurii l1859 urease gene was inserted into the BamH I site of pEB203 and expressed in Escherichia coli JM109. The recombinant plasmid was designated as pGU366. The plasmid pGU366 containing urease gene was introduced into the B. subtilis SH14 with PEG-induced protoplast transformation (PIP) method. The urease gene was very stably expressed in the transformant of B. subtilis SH14. Also, the optimal conditions for transformation were established and the highest transformation frequency was obtained by treatment of lysozyme for 90 min, and then addition of 1.5 ${mu}g$/ml DNA and 40% PEG4000. From the in vitro antifungal test against F. solani, antifungal activity of B. subtilis SH14(pGu366) containing urease gene was much higher than that of the host strain. Genetical development of B. subtilis SH14 by transfer of urease gene can be responsible for enhanced biocontrol efficacy with its antibiotic action.

• ALGAE
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• v.24 no.2
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• pp.85-91
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• 2009

The filamentous cyanobacterium Anabaena sp. Strain PCC 7120 produces a developmental patten of single hete- rocysts separated by approximately 10 vegetative cells. Heterocysts differentiate from vegetative cells and are spe- cialized for nitrogen fixation. The patS gene, which encodes a small peptide that inhibits heterocyst differentiation, is expressed in proheterocysts and plays a critical role in establishing the heterocyst pattem. Another key regulator of heterocyst development is the hetR gene. hetR mutants fail to produce heterocysts and extra copies of hetR on a plas- mid cause a multiple contiguous heterocyst phenotype. To elucidate the relationship between these two counter act- ing factors in the genetic regulatory pathway during heterocyst differentiation, the expression patterns of a patS-gfp and a hetR-gfp fusion were examined in a patS deletion and a hetR deletion strain. The results, in combination with the result from a hetR and patS double deletion strain, suggest patS and hetR are mutually antagonistic and the bal- ance between these two factors in tow different cell types (heterocysts and vegetative cells) may be critical during the decision making process on their cell fates.

• Mycobiology
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• v.49 no.5
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• pp.498-506
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• 2021

An endophytic fungus strain DYSJ3 was isolated from a stem of Aphanamixis grandifolia Blume, which was identified as Aspergillus versicolor based on the morphological characteristics, internal transcribed spacer (ITS) and calmodulin gene sequences analyses. A. versicolor DYSJ3 exhibited strong antagonistic activity against Colletotrichum musae, C. gloeosporioides and Fusarium oxysporum f. sp. cubense with the inhibition rates of 61.9, 51.2 and 55.3% respectively. The antifungal metabolites mainly existed in the mycelium of A. versicolor DYSJ3, and its mycelial crude extract (CE) had broad-spectrum antifungal activities against plant pathogenic fungi. The CE had a good thermal stability, and the inhibition rate of 100 mg/mL CE against C. musae was above 70.0% after disposing at 120 ℃ for 1 h. Five secondary metabolites were isolated from the CE and identified as averufanin, ergosterol peroxide, versicolorin B, averythrin and sterigmatocystin. Activity evaluation showed versicolorin B exhibited inhibitory effects on the mycelial growth and conidial germination of C. musae, and sterigmatocystin had a weak inhibitory effect on the mycelial growth of C. musae.

• The Plant Pathology Journal
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• v.27 no.4
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• pp.333-341
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• 2011

Two antifungal bacteria were selected from forest soils during the screening of microorganisms antagonistic to Cylindrocarpon destructans, a cause of ginseng root rot. The antifungal bacteria were identified as Bacillus subtilis (I4) and B. amyloliquefaciens (yD16) based on physiological and cultural characteristics, the Biolog program, and 16S rRNA gene sequencing analyses. Antagonistic activity of both bacterial isolates to C. destructans increased with increasing temperature. More rapid starch hydrolytic activity of the bacteria was seen on starch agar at higher temperatures than at lower temperatures, and in the higher density inoculum treatment than in the lower density inoculum treatment. The bacterial isolates failed to colonize ginseng root the root tissues inoculated with the bacteria alone at an inoculum density of $1{\times}10^6$ cfu/ml, but succeeded in colonizing the root tissues co-inoculated with the bacteria and C. destructans. Scanning electron microscopy showed that the pathogen was damaged by the low-density inoculum treatment with the bacterial isolates as much as by the high-density inoculum treatment. Both bacterial isolates were more effective in reducing root rot when they were treated at a concentration of $1{\times}10^6$ cfu/ml than at $1{\times}10^8$ cfu/ml. Also, only the former treatment induced prominent wound periderm formation, related to structural defense against pathogen infection. The results suggest that the bacterial antagonists may have high potential as biocontrol agents against ginseng root rot at relatively low-inoculum concentrations.

• Weed & Turfgrass Science
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• v.6 no.2
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• pp.157-164
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• 2017

Functional microorganisms decompose various organic matter by enzyme activity and suppress plant disease caused by pathogen. This study was conducted to isolate and select functional microorganisms with protein or carbohydrate degradation activities and antagonistic activity against turfgrass fungal pathogens for eco-friendly turfgrass management in golf course from compost containing livestock manure of poultry or swine. Totally 68 isolates collected from livestock manure compost strains were isolated and tested for their activities of amylase, protease and lipase and antagonistic activities against Rhizoctonia solani AG2-2, R. solani AG1-1, and Sclerotinia homoeocarpa. Among the isolates, 34 strains were selected as functional microbes showing higher activities of amylase and protease. Three isolates of ASC-14, ASC-18, and ASC-35 among the 34 strains were selected as antifungal bacterial strains repressing the above 3 turfgrass fungal pathogens. Analysis results of 16s rRNA gene sequence and phylogenic cluster indicated that ASC-14 and ASC-18 belonged to Bacillus amyloliquefaciens, while ASC-35 was B. subtilis, respectively.

• The Plant Pathology Journal
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• v.29 no.1
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• pp.52-58
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• 2013

Fusarium head blight (FHB) caused by the filamentous fungus Fusarium graminearum is one of the most severe diseases threatening the production of small grains. Infected grains are often contaminated with mycotoxins such as zearalenone and trichothecences. During survey of contamination by FHB in rice grains, we found a bacterial isolate, designated as BN1, antagonistic to F. graminearum. The strain BN1 had branching vegetative hyphae and spores, and its aerial hyphae often had long, straight filaments bearing spores. The 16S rRNA gene of BN1 had 100% sequence identity with those found in several Streptomyces species. Phylogenetic analysis of ITS regions showed that BN1 grouped with S. sampsonii with 77% bootstrap value, suggesting that BN1 was not a known Streptomyces species. In addition, the efficacy of the BN1 strain against F. graminearum strains was tested both in vitro and in vivo. Wheat seedling length was significantly decreased by F. graminearum infection. However, this effect was mitigated when wheat seeds were treated with BN1 spore suspension prior to F. graminearum infection. BN1 also significantly decreased FHB severity when it was sprayed onto wheat heads, whereas BN1 was not effective when wheat heads were point inoculated. These results suggest that spraying of BN1 spores onto wheat heads during the wheat flowering season can be efficient for plant protection. Mechanistic studies on the antagonistic effect of BN1 against F. graminearum remain to be analyzed.

• The Plant Pathology Journal
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• v.36 no.3
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• pp.244-254
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• 2020

Gom-chwi (Ligularia fischeri) is severely infected with Phytophthora drechsleri, the causal organism of Phytophthora root rot, an economically important crop disease that needs management throughout the cultivation period. In the present study, Phytophthora root rot was controlled by using bacterial isolates from rhizosphere soils collected from various plants and screened for antagonistic activity against P. drechsleri. A total of 172 bacterial strains were isolated, of which, 49 strains showed antagonistic activities by dual culture assay. In the seedling assay, six out of the 49 strains showed a predominant effect on suppressing P. drechsleri. Among the six strains, the ObRS-5 strain showed remarkable against P. drechsleri when treated with seed dipping or soil drenching. The ObRS-5 strain was identified as Enterobacter asburiae based on 16S ribosomal RNA gene sequences analysis. The bacterial cells of E. asburiae ObRS-5 significantly suppressed sporangium formation and zoospore germination in P. drechsleri by 87.4% and 66.7%, respectively. In addition, culture filtrate of E. asburiae ObRS-5 also significantly inhibited sporangium formation and zoospore germination by 97.0% and 67.6%, respectively. Soil drenched bacterial cells, filtrate, and culture solution of E. asburiae ObRS-5 effectively suppressed Phytophthora root rot by 63.2%, 57.9%, and 81.1%, respectively. Thus, E. asburiae ObRS-5 could be used as a potential agent for the biological control of Phytophthora root rot infecting gom-chwi.

• Journal of Microbiology and Biotechnology
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• v.33 no.5
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• pp.607-620
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• 2023

The biocontrol approach using beneficial microorganisms to control crop diseases is becoming an essential alternative to chemical fungicides. Therefore, new and efficient biocontrol agents (BCA) are needed. In this study, a rhizospheric actinomycete isolate showed unique and promising antagonistic activity against three of the most common phytopathogenic fungi, Fusarium oxysporum MH105, Rhizoctonia solani To18, and Alternaria brassicicola CBS107. Identification of the antagonistic strain, which was performed according to spore morphology and cell wall chemotype, suggested that it belongs to the Nocardiopsaceae. Furthermore, cultural, physiological, and biochemical characteristics, together with phylogenetic analysis of the 16S rRNA gene (OP869859.1), indicated the identity of this strain to Nocardiopsis alba. The cell-free filtrate (CFF) of the strain was evaluated for its antifungal potency, and the resultant inhibition zone diameters ranged from 17.0 ± 0.92 to 19.5 ± 0.28 mm for the tested fungal species. Additionally, the CFF was evaluated in vitro to control Fusarium wilt disease in Vicia faba using the spraying method under greenhouse conditions, and the results showed marked differences in virulence between the control and treatment plants, indicating the biocontrol efficacy of this actinomycete. A promising plant-growth promoting (PGP) ability in seed germination and seedling growth of V. faba was also recorded in vitro for the CFF, which displayed PGP traits of phosphate solubilization (48 mg/100 ml) as well as production of indole acetic acid (34 ㎍/ml) and ammonia (20 ㎍/ml). This study provided scientific validation that the new rhizobacterium Nocardiopsis alba strain BH35 could be further utilized in bioformulation and possesses biocontrol and plant growth-promoting capabilities.

• Journal of Microbiology and Biotechnology
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• v.34 no.5
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• pp.1040-1050
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• 2024

To isolate and analyze bacteria with Verticillium wilt-resistant properties from the fermentation residue of kitchen wastes, as well as explore their potential for new applications of the residue. A total of six bacterial strains exhibiting Verticillium wilt-resistant capabilities were isolated from the biogas residue of kitchen waste fermentation. Using a polyphasic approach, strain ZL6, which displayed the highest antagonistic activity against cotton Verticillium wilt, was identified as belonging to the Pseudomonas aeruginosa. Bioassay results demonstrated that this strain possessed robust antagonistic abilities, effectively inhibiting V. dahliae spore germination and mycelial growth. Furthermore, P. aeruginosa ZL6 exhibited high temperature resistance (42℃), nitrogen fixation, and phosphorus removal activities. Pot experiments revealed that P. aeruginosa ZL6 fermentation broth treatment achieved a 47.72% biological control effect compared to the control group. Through activity tracking and protein mass spectrometry identification, a neutral metalloproteinase (Nml) was hypothesized as the main virulence factor. The mutant strain ZL6ߡNml exhibited a significant reduction in its ability to inhibit cotton Verticillium wilt compared to the strain P. aeruginosa ZL6. While the inhibitory activities could be partially restored by a complementation of nml gene in the mutant strain ZL6CMߡNml. This research provides a theoretical foundation for the future development and application of biogas residue as biocontrol agents against Verticillium wilt and as biological preservatives for agricultural products. Additionally, this study presents a novel approach for mitigating the substantial amount of biogas residue generated from kitchen waste fermentation.

• Horticultural Science & Technology
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• v.32 no.6
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• pp.879-886
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• 2014

This study was conducted to test in vitro the antagonistic effect of composted liquid manure (CLM) against soil-borne turfgrass pathogenic fungi, Rhizoctonia solani AG-2-2 (IIIB) (brown patch), R. solani AG-2-2 (IV) (large patch), and Sclerotinia homoeocarpa (dollar spot) for environmentally friendly turfgrass management. CLMs were collected from 9 livestock excretion treatment facilities around the country including Gunwi (GW), Hapcheon (HC), Hoengseong (HS), Icheon (IC), Iksan (IS), Muan (MA), Nonsan (NS), and Yeoju (YJ). CLMs of IC, GW, and IS showed s ignificant (p < 0.05) mycelium growth inhibition that was 17.8%, 20.4%, and 48.0% against R. solani AG-2-2 (IIIB), R. solani AG-2-2 (IV), and S. homoeocarpa, respectively. A t otal of 110 bacterial isolates were obtained from the CLMs that showed antagonistic effects. Among them, 5, 4, and 10 microbe isolates showed promising antifungal activity against mycelium growth of R. solani AG-2-2 (IIIB), R. solani AG-2-2 (IV), and S. homoeocarpa, respectively. The bacterial isolates ICIIIB60, GWIV70, and ISSH20 effectively inhibited the mycelial growth of three soil-borne turfgrass pathogens. Selected bacterial isolates were identified as Alcaligenes sp., Bacillus licheniformis Ab2, and B. subtilis C7-3 through 16s rDNA gene sequence analysis. Among 5 fungicides, the most compatible fungicide with ICIIIB60, GWIV70, and ISSH20 was tebuconazol, toclofos-methyl and toclofos-methyl, respectively. These findings suggested that CLMs could be effectively used not only as organic liquid fertilizer sources but also as biological control agents for soil-borne turfgrass diseases such as brown patch, large patch, and dollar spot.