• Journal of Microbiology and Biotechnology
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• v.27 no.10
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• pp.1763-1772
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• 2017

This study aimed to isolate and characterize antibacterial metabolites from Pharbitis nil seeds and investigate their antibacterial activity against various plant pathogenic bacteria. The methanol extract of P. nil seeds showed the strongest activity against Xanthomonas arboricola pv. pruni (Xap) with a minimum inhibition concentration (MIC) value of $250{\mu}g/ml$. Among the three solvent layers obtained from the methanol extract of P. nil seeds, only the butanol layer displayed the activity with an MIC value of $125{\mu}g/ml$ against Xap. An antibacterial fraction was obtained from P. nil seeds by repeated column chromatography and identified as pharbitin, a crude resin glycoside, by instrumental analysis. The antibacterial activity of pharbitin was tested in vitro against 14 phytopathogenic bacteria, and it was found to inhibit Ralstonia solanacearum and four Xanthomonas species. The minimum inhibitory concentration values against the five bacteria were $125-500{\mu}g/ml$ for the n-butanol layer and $31.25-125{\mu}g/ml$ for pharbitin. In a detached peach leaf assay, it effectively suppressed the development of bacterial leaf spot, with a control value of 87.5% at $500{\mu}g/ml$. In addition, pharbitin strongly reduced the development of bacterial wilt on tomato seedlings by 97.4% at $250{\mu}g/ml$, 7 days after inoculation. These findings suggest that the crude extract of P. nil seeds can be used as an alternative biopesticide for the control of plant diseases caused by R. solanacearum and Xanthomonas spp. This is the first report on the antibacterial activity of pharbitin against phytopathogenic bacteria.

• Korean Journal of Medicinal Crop Science
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• v.19 no.6
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• pp.464-471
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• 2011

Conventional Thiamine Dilauryl Sulfate (TDS) powder has a low stability. In order to solve this problem, this study was performed to improve the solubility of TDS. The process for enhance solubility of TDS was nano grinding mill and ultrasonic dispersion process. TDS paticle was manufactured to nano size through nano grinding mill process. The size of TDS nanoparticle was measured as average 220 nm by DLS. And The TDS nanoparticle in water solution manufactured through ultrasonic dispersion process. The TDS nanoparticle in water solution was showed the highest solubility with 40% ethanol. These results was increased the concentration of TDS from 200 ppm to 240 ppm in water solution. The TDS nanoparticle in water solution showed diameter of Colletotrichum gloeosporioides growth with smaller than about 1.56 cm compared to the TDS paticle in water solution at same concentration. Also, TDS nanoparticle in water solution showed growth inhibition activity as 59.2% with higher than about 10% compared to the TDS paticle water solution in same concentration. Finally, TDS nanoparticle in water solution was increased solubility through nano grinding mill and ultrasonic dispersion process. Also, the increase of concentration in TDS nanopaticle in water solution according to solubility enhancement lead to an result enhancement of antifungal activity. Consequently, we suggested that the TDS nanoparticle in water solution was more effective than TDS particle in water solution owing to the sub-cellular particle size, ability to persistence and targeting to cell membrane of Colletotrichum gloeosporioides. Furthermore we expected the applicating possibility with bio pesticide.

• Research in Plant Disease
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• v.18 no.3
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• pp.186-193
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• 2012

Biotransformation is an eco-friendly and efficient method for enhancing the bioavailability of biopesticide. To increase the antifungal activity of the crude extract of Cynanchum wilfordii roots against barely powdery mildew, we performed biotransformation of wilfoside C1G using ${\beta}$-glucosidase (cellobiase from Aspergillus niger). The mixture (G sample) of partially purified wilfoside C1G and cynauricuoside A (K1G) was treated with ${\beta}$-glucosidase to remove a glucopyranosyl moiety. The enzyme completely converted C1G to C1N and K1G to K1N. Optimal conditions for enzymatic biotransformation of G sample were determined to be 10% ethanol, 1,555 ${\mu}U$ ${\beta}$-glucosidase/ml, pH 5, and $45^{\circ}C$. In in vivo experiment, the G sample transformed by ${\beta}$-glucosidase showed stronger antifungal activity against barley powdery mildew than the non-treated G sample. These results suggest that ${\beta}$-glucosidase biotransformation can be applied to increase the antifungal activity of the crude extract of C. wilfordii roots against powdery mildews.

• The Korean Journal of Pesticide Science
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• v.18 no.1
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• pp.21-25
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• 2014

Bukholderia pyrrocinia CAB08106-4 has an anti-fungal effect on Garlic White Rot caused by Sclereotium cepivorum and Sclereotium sp. It is environmentally friendly microbial product that prevents and controls a variety of phytopathogens including Garlic White Rot caused by Sclereotium cepivorum and Sclereotium sp. The aim of this study was to assess the environmental toxicity using Cyprinus carpio and Daphnia magna. Bukholderia pyrrocinia CAB08106-4 ($1.0{\times}10^9cfu/mL$) was adminatrated to Cyprinus carpio and Daphnia magna according to the toxicity test guideline for peciticide. $LC_{50}$ of Bukholderia pyrrocinia CAB08106-4 is over $6.67{\times}10^4cfu/mL$ in Cyprinus carpio and Daphnia magna and no adverse effect was observed. Based on these results, we concluded that Bukholderia pyrrocinia CAB08106-4 has no toxiciy for Cyprinus carpio and Daphnia magna.

• The Korean Journal of Pesticide Science
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• v.20 no.3
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• pp.181-188
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• 2016

BtPlus is a group of biopesticides that are made of Bacillus thuringiensis and immunosuppressant. A new BtPlus that exhibits high insecticidal activity against mosquito larvae has been investigated in control efficacy in field conditions and its environmental safety against aquatic system. This study assessed the control efficacy of BtPlus against mosquito larvae with two different application methods. In aerial spraying application (100 mL per $3.3m^2$), BtPlus was effective at 50% or above formulation concentrations to control mosquito larvae. For a direct application to aqueous mosquito habitat, a semi-field mimicking paddy rice field was constructed. In this condition, BtPlus showed 80% and 100% control efficacies at 0.1% and 0.2% concentrations, respectively. BtPlus also showed 40% mortality against adults at 0.1% concentration in 10% sugar bait. However, its control efficacies against adults were much less than against larvae. Safety assessment of BtPlus against ecosystem was evaluated using young carp (Cyprinus carpio), a water flea (Daphnia magna), and a honey bee (Apis mellifera). BtPlus did not give any adverse effects on these nontarget organisms. Based on these results, BtPlus can be applied to control mosquitoes by direct aqueous application to paddy rice field.

• Korean journal of applied entomology
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• v.59 no.1
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• pp.41-54
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• 2020

Two mosquito species were collected in still-water near farming area in Andong, Korea. Based on morphological characters, these two mosquitoes were identified as Aedes koreicus and Culex vagans, respectively. DNA barcode analyses supported the identification. An entomopathogenic bacterium, Bacillus thuringiensis subsp. israelensis (BtI), exhibited insecticidal activities against the two mosquito species and its virulence was more potent than that of B. thuringiensis subsp. kurstaki. It has been known that the bacterial metabolites of Xenorhabdus spp. suppress insect immunity and enhance pathogenicity of B. thuringiensis. This study tested the effect of the bacterial culture broth of Xenorhabdus spp. on enhancing BtI pathogenicity. Among three Xenorhabdus spp., culture broth of X. ehlersii (Xe) was relatively effective to enhance BtI pathogenicity against both mosquito species. Indeed, organic extracts of Xe culture broth suppressed the hemocyte-spreading behavior, suggesting the presence of immunosuppressant in the culture broth. These results suggest a formulation of BtPlus by mixing BtI spore and Xe culture broth to be applied to control the two mosquito species.

• KSBB Journal
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• v.14 no.3
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• pp.303-308
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• 1999

An in vitro culture method for entomopathogenic nematode Steinernema glaseri was developed. A symbiotic bacterium was isolated from Steinernema glaseri and identified as Xenorhabdus nematophilus. Phase variation that differed in some biochemical characteristics of symbiotic bacterium was observed. Entomopathogenic nematodes carried only phase I bacterium in their guts. Phase I bacterium could be converted into phase II form in in vitro culture medium consisting of 5% yeast extract, 0.5% NaCl, 0.05% $K_2HPO_4$, $0.02% MgSO_4$.$7H_2O$. The optimum temperature for bacterial growth was $28^{\circ}C$. The pH of the culture medium increased up to 9.0-9.5 during the exponential growth period of the culture, regardless of initial pH 6-7. Various culture media such as chicken offal, dog food, bovine liver, peanut, and so on were tested for in vitro culture of the nematodes. The best medium for Steinernema glaseri production was obtained from concentrated homogenate of bovine liver and the nematode growth was highest at 80% bovine liver. In the co-culture of entomopathogenic nematode with its symbiont, the growth rate of nematodes was 2 times faster than that without its symbiont and the nematode concentration reached about $5.5\times10^4$/mL within 15 days.

• Research in Plant Disease
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• v.21 no.3
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• pp.208-214
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• 2015

To develop an effective biopesticide to control pepper anthracnose disease, an isolate which showed strong inhibitory effect on the mycelial growth and conidial germination of Colletotrichum acutatum was selected among the antagonistic bacterial isolates collected from pepper grown soil. The bacterial isolate was identified as Bacillus sp. KBC1004 using 16S rRNA sequence analysis. The liquid culture of KBC1004 was freeze-dried and formulated as a wettable powder(WP). The wettable powder form of KBC1004 required at least 24 hours to activate and to inhibit the conidial germination of C. acutatum. In vitro bioassay using the detached green pepper fruits, biocontrol activity of the WP was not recognizable in simultaneous inoculation, but significant disease suppression was observed pre-treatment (24 hr) of the WP before pathogen inoculation. In field experiment, 4 times foliar applications of the 1/500 diluted wettable powder from the end of June showed great control efficacy similar to that of the chemical fungicide application. These results suggest that the formulated WP product could be an alternative mean to control of pepper anthracnose disease in environmentally friendly farming practices.

• The Korean Journal of Pesticide Science
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• v.18 no.3
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• pp.156-160
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• 2014

The stabilities of four limonoidal substances including azadirachtin A, azadirachtin B, deacetylsalannin and salannin were investigated both in controlled aquatic and soil conditions. The half-life of the total limonoid for neem extracts and its two commercial biopesticides was estimated 86.6-173 days in water under air, while degradation of the compounds was detected below 10% after eight weeks in deoxygenated water. The half-life in dry soil was estimated 43.3-57.7 days, and there was a similar degradation pattern with in aerobic water condition. In case of wet soil condition, the total bacteria of the soils ranged 6-8 log CFU/g soil for during the experiment, and the half-life of the total limonoid was 6.4-12.3 days. From the result, the fast limonoid degradation in wet soil environment was the result of both chemical oxidation and microbial degradation.

• The Korean Journal of Pesticide Science
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• v.12 no.2
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• pp.184-191
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• 2008

Entomopathogenic bacteria (Xenorhabdus nematophila, X. sp. and Photorhabdus temperata subsp. temperata) isolated from entomopathogenic nematodes express potent insecticidal activity in insect hemocoel. They are also known to suppress insect immune mediation by inhibiting phospholipase $A_2$, leading to host immunosuppression. This study analyzed effects of their cultured broths on inhibiting insect immunosuppression. For this, we removed all bacterial cells using $0.2\;{\mu}m$ pore sized membrane from the bacteria-cultured broth. All three sterilized cultured media, in dose-dependent manners, significantly inhibited hemocyte-spreading behavior of 5th instar larvae of Spodoptera exigua. However, they showed differential inhibitory activities among different bacterial species, in which X. nematophila showed the most potent inhibitory activity. This immunosuppressive effect was applied to increase the pathogenicity of Bacillus thuringiensis (Bt). All three bacterial cultured broths including bacterial cells significantly potentiated Bt pathogenicity against young S. exigua larvae when each of them was orally administered in a mixture of low dose of Bt. Finally, we tested the effect of oral administration of the cultured media containing the immunosuppressive compound(s) secreted by the bacteria. The membrane-sterilized cultured broths were mixed with the low dose of Bt and then orally administered to the young S. exigua. Only the cultured medium of X. nematophila showed increase of Bt pathogenicity. These results indicated that the; cultured media of the three bacteria possessed immunosuppressive factor(s), which may act to potentiate Bt toxicity to young S. exigua larvae.