• 한국균학회지
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• 제42권3호
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• pp.219-224
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• 2014

P. tolaasii에 의해 발생하는 세균갈색무늬병은 버섯재배에서 문제가 되는 대표적인 병해이다. 본 연구에서는 세균갈색무늬병의 생물학적 방제법에 이용할 수 있는 길항미생물의 항균활성과 선발된 길항미생물에 대해 폿트 수준의 생물검정 실험을 실시하였다. 재배중인 느타리 폐면배지와 양송이 퇴비에서 세균갈색무늬병원균을 강하게 억제하는 길항세균 HC5를 선발하였으며, 생리 생화학적 실험과 유전적 실험결과 HC5균주는 P. azotoformans로 동정되었다. P. azotoformans HC5를 양송이, 팽이버섯, 느타리에 처리한 결과 각각 78%, 73%, 71%의 방제효과를 보였다. 따라서 P. azotoformans HC5가 버섯 세균갈색무늬병 방제를 위해 합성농약을 대체할 수 있는 친환경 방제제가 될 수 있을 것으로 생각된다.

• 한국유기농업학회지
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• 제22권4호
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• pp.855-868
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• 2014

파프리카에 발생하는 병들을 방제하기 위하여 합성농약이 광범위하게 사용되어왔지만 최근에 수많은 농약사용의 부작용에 대한 관심이 증가 하고 있다. 파프리카 주요병인 잿빛곰팡이병, 줄기 및 과실썩음병, 역병, 균핵병, 시들음병을 방제하기 위한 미생물을 분리하고 특성을 파악하기 위하여 본 연구를 실시하였다. 지방산분석과 16S rDNA 염기배열은 이 연구에서 분리한 YKJ1가 Serratia marcescens 그룹에 속하는 것을 밝혔다. 특히, YKJ1의 16S rDNA 염기배열은 S. marcescens의 염기서열과 99% 상동성을 보였다. 광학현미경을 통해 YKJ1처리에 의해 병원균의 포자 발아 및 균사 생장이 저해됨을 확인 하였다. YKJ1처리는 팽윤균사와 같은 현저한 형태적 변화와 세포벽의 분해를 유발하였다. 역병균의 경우 유주자낭의 형성이 억제되었다. 본 연구에서 동정한 S. marcescens는 S. marcescens-YKJ1으로 부르고자 한다. 포장실험등과 같은 시험이 차후 더 요구되어지나 파프리카의 주요 병관리를 위한 생물적 방제제의 하나로 가치가 있을 것으로 생각된다.

• 한국미생물·생명공학회지
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• 제34권4호
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• pp.311-317
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• 2006

식물생육을 촉진하고 고추역병균을 방제하는 다기능 PGPR 균주 Bacillus subtilis AH18 항진균성 cellulase 유전자를 PCR을 이용해 pUC18과 재조합 후 E. coli DH5$\alpha$에 cloning하여 E. coli내에 발현시켰으며, 그 형질전환 균주를 E. coli DH5$\alpha$(pCM 41)이라 명명하였고, 발현된 cellulase를 ce/H라 하였다. E. coli DH5$\alpha$(pCM 41)의 inset 부위는 B. subtilis AH18의 1,582 bp 유전자를 포함하며 cellulase의 유전자는 1,524 bp로 508개의 amino acid가 암호화된 것으로 추정되었고, CMC를 함유한 SDS-PAGE의 방법으로 약 55 kDa의 분자량을 확인하였다. B. subtilis AH18이 가지는 ce/H는 3종의 대표적인 Bacillus spp.들의 cellulase 유전자의 DNA와 아미노산 배열이 98% 이상 유사하였으며, CMC(carboxymethyl-cellulose) 뿐만 아니라, 불용성 섬유소인 Avicel, filter paper(Whatman No. 1) 특히 고추역병균인 Phytophthora capsici의 건조 cell wall도 분해하였다. 또한 colH의 cellulase는 $50^{\circ}C$에서 효소활성이 가장 높았으며, 최적 pH는 pH 6.0이었다. 그리고 $AgNO_3$ 또는 $CoCl_2$ 첨가시 활성이 1.7배, 2배 정도 증가하였고 $HgC1_2$ 첨가시는 활성이 20%까지 떨어졌다. 또한 여러 화학 저해제들 중 Sodium azide 또는 Hydroxy urea는 효소 활성을 증가시켰으며, CDTA 또는 EDIA는 섬유소분해능을 감소시켰다. 이들의 결과는 고추역병균 P. capsici의 생육을 억제하는 B. subtilis AM18의 진균세포벽 용해성 cellula의 효소학적 특성을 구명한 것이라고 할 수 있다.

• 농약과학회지
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• 제10권1호
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• pp.56-65
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• 2006

작물 근권 토양으로부터 분리한 5,000여 길항 균주로부터 Erwinia 및 Pseudomonas등의 세균과 Trichoderma, Colletotrichum 등 곰팡이의 성장을 동시에 억제하는 Bacillus sp. N 32 균주를 선발 동정 하였다. 특히 Bacillus sp. N32 균주는 고추 탄저병균인 Colletotrichum gloeosporioides에 대하여 열에 저항성이 있는 단백질과 열에 민감한 단백질의 2종류의 항균 단백질을 동시에 생산함을 단백질 침전과 활성 검정을 통하여 확인하였다. 이 항균 단백질들을 FPLC를 이용한 gel filtration chromatography방법으로 분리한 후 SDS-PAGE와 bioautography로 항균력을 확인하였다. 또한 이 항균 단백질의 유전자들을 선발하기 위하여 기존의 알려진 그람양성 세균의 대표적인 열 저항성 항균 펩타이드 생합성 유전자 서열을 primer로 이용한 PCR 방법으로 fengycin의 생합성 유전자 단편을 분리하고 이 PCR 산물을 이용하여 Bacillus sp. N32 균주의 cosmid library로부터 fengycin의 생합성 유전자 cluster중 일부를 분리하여 염기서열을 분석하였다. 또한 열에 민감한 항균 단백질 생산 유전자는 이 항균 단백질을 SDS-PAGE 및 electroblotting으로 분리한 뒤 N-terminal 부위의 15개의 아미노산 서열을 분석하고 이를 DNA 염기배열로 치환한 다음 probe로 이용하여 ${\lambda}-ZAP$ library로부터 항균 단백질 생산 유전자가 포함된 다수의 clone을 선발하였다.

• The Plant Pathology Journal
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• 제22권3호
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• pp.289-294
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• 2006

Two isolates of mucous bacteria, mc75 and pc78, were isolated from fungal culture plate as culture contaminants with an interesting swarming motility. Both isolates were identified as Pseudomonas fluorescens based on microscopy, biochemical analysis, Biolog test and DNA sequence analysis of the 16S rRNA gene. Both strains have the exactly the same 16S rRNA gene sequences, and yet their biological control activity were not identical each other. In vitro analysis of antagonistic activity of two isolates against several plant pathogenic fungi indicated that both produced diffusible and volatile antifungal compounds of unknown identities. Treatment of the bacterial culture of P. fluorescens pc78 and its culture filtrate exhibited a strong biological control activity against rice sheath blight in vivo among six plant diseases tested. More effective disease control activity was obtained from treatment of bacterial culture than that of culture filtrate. Therefore, in addition to antifungal compound and siderophore production, other traits such as biofilm formation and swarming motility on plant surface may contribute to the biological control activity of P.fluorescens pc78 and mc75.

• The Plant Pathology Journal
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• 제25권1호
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• pp.62-69
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• 2009

In this study, we characterized the bacterial strain KJ2C12 in relation with its biocontrol activity against Phytophthora capsici on pepper, and identified this strain using morphological, physiological, biochemical, fatty acid methyl ester, and 16S rRNA gene sequence analyses. Strain KJ2C12 significantly (P=0.05) reduced both final disease severity and areas under the disease progress curves of 5-week-old pepper plants inoculated with P. capsici compared to buffer-treated controls. As for the production of antibiotics, biofilms, biosurfactant, extracellular enzyme, HCN, and swarming activity, strain KJ2C12 produced an extracellular enzyme with protease activity, but no other productions or swarming activity. However, Escherichia coli produced weak biofilm only. Strain KJ2C12 could colonize pepper roots more effectively in a gnotobiotic system using sterile quartz sand compared to E. coli over 4 weeks after treatments. However, no bacterial populations were detected in 10 mM $MgSO_4$ buffer-treated controls. Strain KJ2C12 produced significantly higher microbial activity than the $MgSO_4$-treated control or E. coli over 4 weeks after treatments. Bacterial strain KJ2C12 was identified as Bacillus luciferensis based on morphological, physiological, and biochemical characteristics as well as FAME and 16S rRNA gene sequence analyses. In addition, these results suggested that B. luciferensis strain KJ2C12 could reduce Phytophthora blight of pepper by protecting infection courts through enhanced effective root colonization with protease production and an increase of soil microbial activity.

• Journal of Microbiology and Biotechnology
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• 제26권4호
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• pp.748-756
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• 2016

In the present study, a lipopeptide (named AXLP14) antagonistic to Xanthomonas oryzae pv. oryzae (Xoo) was obtained from the culture supernatant of Bacillus amyloliquefaciens B014. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis demonstrated that AXLP14 consisted of surfactin homologs. The minimum inhibition concentration and minimum bactericidal concentration of AXLP14 against Xoo were determined to be 1.25 and 2.50 mg/ml, respectively. At a concentration of 0.613 mg/ml, AXLP14 strongly inhibited the formation of Xoo biofilm. AXLP14 also inhibited the motility of Xoo in a concentration-dependent manner. Applying AXLP14 to rice seedlings significantly reduced the incidence and severity of disease caused by Xoo. In Xoo-infected rice seedlings, AXLP14 strongly and continuously up-regulated the expression of both OsNPR1 and OsWRKY45. In addition, AXLP14 effectively inhibited the Xoo-induced up-regulation of the expression of the abscisic acid biosynthesis gene OsNECD3 and the abscisic acid signalingresponsive gene OsLip9, indicating that AXLP14 may protect rice against Xoo-induced disease by enhancing salicylic acid defense and interfering with the abscisic acid response to virulence.

• Journal of Microbiology and Biotechnology
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• 제20권2호
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• pp.281-286
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• 2010

A Pseudomonas fluorescens strain was isolated and found to show antagonistic activity against phytopathogenic fungi and to possess a gene responsible for production of antibiotic 2,4-diacetylphloroglucinol. For the extension of biocontrol range, a gene for an Androetonus australis Hector insect toxin 1 (AaHIT1), one of the most known toxic insect-selective peptides, was designed and synthesized according to the preferred codon usage of Pseudomonas fluorescens, cloned, and transformed into the strain by pSUP106 vector, a broad-host-range plasmid. Bioassays indicated that the engineered strain was able to produce AaHIT1 with insecticidal activity, and at the same time retain the activity against plant pathogen. The experiments for nonplanted soil and rhizosphere colonization showed that, similar to the population of the wild-type strain, that of the engineered strain remained relatively constant in the first 10 days, and the subsequent 50 days, suggesting that AaHIT1 expression in the bacterial cell does not substantially impair its long-term colonization. It is first reported that a Pseudomonas fluorescens strain expressing an active scorpion neurotoxin has dual activity against phytopathogenic fungi and insects, making at attractive for agronomic applications.

• The Plant Pathology Journal
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• 제35권3호
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• pp.234-242
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• 2019

Thirty-four strains of bacteria were isolated from Phellodendron amurense. Using Nectria haematococca as an indicator strain, the best strain, B18, was obtained by the growth rate method. The morphological, physiological and biochemical characteristics of strain B18 and its 16S DNA gene sequence were identified, and the biocontrol effect of strain B18 was assessed in pot and field tests, as well as in a field-control test. Drilling methods were used to determine the antibacterial activity of metabolites from strain B18 and their effects on the growth of pathogen mycelia and spores. The best bacteriostatic rate was 85.4%. B18 can hydrolyse starch and oxidize glucose but does not produce gas; a positive result was obtained in a gelatine liquefaction test. According to 16S DNA gene sequencing, strain B18 is Bacillus methylotrophicus (GenBank accession number: MG457759). The results of pot and field-control trials showed 98% disease control when inoculating $10^8cfu/ml$ of the strain. The disease control effect of the B18 culture liquid (concentrations of $10^8$, $2{\times}10^6$, $10^6$, $5{\times}10^5$ and $2.5{\times}10^5cfu/ml$) in the field-control test was higher than 80%, and the cure rate of the original delivery solution was 96%. Therefore, in the practical forestry production, a $2.5{\times}10^5cfu/ml$ culture liquidshould be applied in advance to achieve good control effects.

• Journal of Microbiology and Biotechnology
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• 제30권3호
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• pp.368-377
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• 2020

Enterotoxigenic Bacteroides fragilis (ETBF) is the main pathogen causing severe inflammatory diseases and colorectal cancer. Its biofilm plays a key role in the development of colorectal cancer. The objective of this study was to determine the antagonistic effects of cell-free supernatants (CFS) derived from Clostridium butyricum against the growth and biofilm of ETBF. Our data showed that C. butyricum CFS inhibited the growth of B. fragilis in planktonic culture. In addition, C. butyricum CFS exhibited an antibiofilm effect by inhibiting biofilm development, disassembling preformed biofilms and reducing the metabolic activity of cells in biofilms. Using confocal laser scanning microscopy, we found that C. butyricum CFS significantly suppressed the proteins and extracellular nucleic acids among the basic biofilm components. Furthermore, C. butyricum CFS significantly downregulated the expression of virulence- and efflux pump-related genes including ompA and bmeB3 in B. fragilis. Our findings suggest that C. butyricum can be used as biotherapeutic agent by inhibiting the growth and biofilm of ETBF.