• Journal of Microbiology and Biotechnology
• /
• v.21 no.8
• /
• pp.869-873
• /
• 2011

Most medium formulations for improving culture of entomopathogenic nematodes (EPN) based on protein sources have used enriched media like animal feed such as dried egg yolk, lactalbumin, and liver extract, among other ingredients. Most results, however, showed unstable yields and longer production time. Many of the results do not show the detailed parameters of fermentation. Soy flour, cotton seed flour, corn gluten meal, casein powder, soytone, peptone, casein hydrolysates, and lactalbumin hydrolysate as protein sources were tested to determine the source to support optimal symbiotic bacteria and nematode growth. The protein hydrolysates selected did not improve bacterial cell mass compared with the yeast extract control, but soy flour was the best, showing 75.1% recovery and producing more bacterial cell number ($1.4{\times}10^9$/ml) than all other sources. The highest yield ($1.85{\times}10^5$ IJs/ml), yield coefficient ($1.67{\times}10^6$ IJs/g medium), and productivity ($1.32{\times}10^7$ IJs/l/day) were also achieved at enriched medium with soybean protein.

• KSBB Journal
• /
• v.15 no.1
• /
• pp.100-105
• /
• 2000

Optimal medium composition, culture conditions, characteristics of phase variation and activity of insecticidal toxin by Xenorhabdus nematophilus isolated and identified from Korean entomopathogenic nematode Steinernema carpocapsae were examined. Optimal medium composition of this strain was 50-70 g/L yeast extract, 3 g/L $K_{2}HPO_{4}$, 1g/L $NH_{4}H_{2}PO_{4}$, 2g/L ${MgSO}_4$$\cdot$${7H}_{2}O$, 10g/L NaCl and, these, yeast extract was found as a limiting nutrient for cell growth. When Monod equation was applied, maxmum specific growth rate and Monod constant were estimated as 0.13 $hr^{-1}$ and 20g/L, respectively. The pH of culture medium increased up to 8.5-9.5 regardless of initial pH 6-7 as the cells continued to grow. The specific growth rate in a 7 L fermentor was 0.18 $hr^{-1}$, which was enhancement 1.4 fold compared to a flask culture. In case of phase variation, phase I fraction was maintained above 90% at the stationary phase for both flask and fermentor cultures. According to oral toxicity test of Gallena mellonella by Xenorhabdus nematophilus, the addition of cell pellets into feed inhibited normal growth of insect larvae and killed completely then after 20 days cultivation. When culture supernatant of this strain was injected into hemolymph of insect larva, the toxicity was strongest at 24hr cultivation in the early exponential phase and gradually decreased as the culture time proceeded.

• Journal of Applied Biological Chemistry
• /
• v.55 no.2
• /
• pp.123-127
• /
• 2012

In order to develop eco-friendly biopesticide, an entomopathogenic bacterium Photorhabdus temperata M1021 has been lyophilized via freeze-drying along with protective agents such as skim milk, starch, sodium alginate, glucose and sodium glutamate to protect cells from lysis. Freeze-drying powder of P. temperata M1021 containing 7% skim milk (w/v) showed highest survival rate of 63% among all the protective agents used in trials. Furthermore, the freeze-dried microbial powder showed 75% of survival rate after stored at $4^{\circ}C$ for 4 weeks at air contact conditions. Injection toxicity of the freeze-dried sample was tested against larvae of Galleria mellonella. A dose of $2.0{\times}10^1$ cells of P. temperata M1021 killed 100% of the G. mellonella larvae within 4 days after injection. Moreover, $2.0{\times}10^0$ cells caused 50% mortality within the 4 days after injection. Freeze-dried P. temperata M1021 strains exhibited effective insecticidal activity and could be a better candidate for being used as a biopesticide.

• Korean journal of applied entomology
• /
• v.47 no.4
• /
• pp.435-445
• /
• 2008

Due to internal feeding behavior, the oriental tobacco budworm, Helicoverpa assulta ($Guen\acute{e}e$), infesting hot pepper has been regarded to be effectively controlled by targeting egg and neonate larval stages just before entering the fruits. This study aimed to develop an efficient biological control method focusing on these susceptible stages of H. assulta. An egg parasitoid wasp, Trichogramma evanescens Westwood, was confirmed to parasitize the eggs of H. assulta. A mixture of Gram-positive soil bacterium, Bacillus thuringiensis subsp. kurstaki, and Gram-negative entomopathogenic bacterium, Xenorhabdus nematophila ANU101, could effectively kill neonate larvae of H. assulta. A sex pheromone trap monitored the occurrence of field H. assulta adults. The microbial insecticide mixture was proved to give no detrimental effects on immature development and adult survival of the wasp by both feeding and contact toxicity tests. A combined treatment of egg parasitoid and microbial pesticide was applied to hot pepper fields infested by H. assulta. The mixture treatment of both biological control agents significantly decreased the fruit damage, which was comparable to the chemical insecticide treatment, though either single biological control agent did not show any significant control efficacy. This study also provides morphological and genetic characters of T. evanescens.

• Journal of Microbiology and Biotechnology
• /
• v.28 no.1
• /
• pp.12-21
• /
• 2018

Photorhabdus temperata (PT), a gram-negative bacterium, lives symbiotically within entomopathogenic nematodes. The insecticidal compounds derived from Photorhabdus are used as biopesticides in agriculture. However, the physiological properties are not well characterized. In the course of our screening for neuroprotective and anti-neuroinflammatory substances from natural products, the culture broth of PT showed considerable activities. By activity-guided purification, five anthraquinones, namely, 3-methoxychrysazine (1), 1,3-dimethoxy-8-hydroxy-9,10-anthraquinone (2), 1,3,8-trihydroxy-9,10-anthraquinone (3), 3,8-dihydroxy-1-methoxy-9,10-anthraquinone (4), and 1,3,4-trimethoxy-8-hydroxy-9,10-anthraquinone (5), were isolated from the ethyl acetate fraction of the PT culture broth. Among the isolated compounds, $75{\mu}M$ 3 significantly protected mouse hippocampal neuronal cells (HT22) against 5 mM glutamate-induced cell death via the inhibition of reactive oxygen species production, $Ca^{2+}$ influx, and lipid peroxidation. Additionally, 3 and 4 effectively suppressed the interferon-${\gamma}$-induced neuroinflammation of mouse-derived microglial cells (BV2) at 10 ng/ml, via the reduction of nitric oxide, interleukin-6, and tumor necrosis factor-${\alpha}$. Anthraquinones 3 and 4 derived from the PT culture broth are a potential starting point to discover neuroprotective and anti-neuroinflammatory drug leads. The novel compound 5 is reported for the first time in this study.

• Journal of Microbiology and Biotechnology
• /
• v.14 no.5
• /
• pp.1057-1062
• /
• 2004

The entomopathogenic bacterium Bacillus thuringiensis is the most widely used biopesticide. Insecticidal proteins, coded by genes located in plasmids, form typical parasporal, crystalline inclusions during sporulation. We isolated a Bacillus thuringiensis strain having insecticidal activity against larvae of the house fly (M. domestica) from the soils at a pig farm in Korea, and named it Bacillus thuringiensis SM. The culture filtrate from Bacillus thuringiensis SM showed strong lethality (83.3%) against M. domestica larvae. The parasporal crystal is enclosed within the spores' outermost envelope, as determined by transmission electron microscopy, and exhibited a bipyramidal form. The crystal proteins of strain SM consisted of five proteins with molecular weights of approximately ~130, ~80, ~68, ~42, and ~27 kDa on a 10% SDS-PAGE (major band, a size characteristic of Cry protein). Examination of antibiotic resistance revealed that the strain SM showed multiple resistant. The strain SM had at least three different plasmids with sizes of 6.6, 9.3, and 54 kb. Polymerase chain reactions (PCRs) revealed the presence of cry1, cry4A2, and cry11A1 genes in the strain SM. The cry1 gene profile of the strain SM appeared in the three respective products of 487 bp [cry1A(c)], 414 bp [cry1D], and 238 bp [cry1A(b)]. However, the strain SM has not shown the cry4A2 md cry11A1 genes. In in vivo toxicity assays, the strain SM showed high toxicity on fly larvae (M. domestic) [with $LC_{50}$ of 4.2 mg/ml, $LC_{90}$ of 8.2 mg/ml].

• International Journal of Industrial Entomology and Biomaterials
• /
• v.36 no.1
• /
• pp.1-9
• /
• 2018

The purpose of this study was to determine the best method for minimizing the occurrence of Metarhizium anisopliae infection of Protaetia brevitarsis seluensis during mass breeding on agricultural farms. There is a high demand for the use of P. b. seluensis larvae in animal feed and as food for humans. However, mass breeding results in the entomopathogenic fungal (usually M. anisopliae) infection of P. b. seluensis. A mixture of microorganisms (Bacillus subtilis, Lactobacillus plantarum, and Saccharomyces cerevisiae) delayed fungal infection by M. anisopliae, which infected fewer P. b. seluensis when the microorganism mixture was added to sawdust as feed for P. b. seluensis. When sawdust with the effective microorganisms (EM) was given to P. b. seluensis for 30 d, their mortality rate was approximately 35 % less than that of the control group, which was fed sawdust without the EM. In addition, the growth of M. anisopliae on agar media spread with each bacterium as inhibited by up to 80 % more than those spread with 4 % sodium hypochlorite, which is a harmless fungal inhibitor generally used in agricultural farms for disinfection.

• Research in Plant Disease
• /
• v.19 no.3
• /
• pp.208-215
• /
• 2013

3-(4-Hydroxyphenyl)-propionic acid (HPP) is a bacterial metabolite synthesized and released by an entomopathogenic bacterium Xenorhabdus nematophila K1. In this study, the control efficacy of HPP was tested against Phytophthora blight and anthracnose of red pepper plants. HPP suppressed mycelial growth of Phytophthora blight and anthracnose pathogens. Under natural sunlight condition, HPP maintained the antifungal activity on the diseases for at least twenty five days. The antifungal activity was not decreased even in the condition of soil-water. It was proved that HPP was able to penetrate the roots and travel upward of the red pepper plants. When HPP suspension was applied to soil rhizosphere before transplanting the red pepper seedlings or was regularly sprayed to the foliage of the plants with ten days interval, it resulted in significant reduction of the disease occurrences (Phytophthora blight and anthracnose) without any phytotoxicity. These results suggested that HPP can be developed to a systemic agrochemical against Phytophthora blight and anthracnose of red pepper plants.

• Research in Plant Disease
• /
• v.16 no.2
• /
• pp.170-175
• /
• 2010

A monoterpenoid benzylideneacetone (BZA) is a bacterial metabolite isolated from culture broth of an entomopathogenic bacterium, Xenorhabdus nematophila K1. It was tested in this study the control efficacy of the metabolite against two major fungal diseases occurring in red-pepper plants. BZA exhibited significant antifungal activities against Phytophthora capsici and Colletotrichum acutatum. Under natural light conditions, the antifungal activity of BZA was maintained for more than sixty days. The antifungal activity of BZA was not lost even in soil because the incidence of Phytophthora blight against red-pepper plants was significantly reduced when the suspensions of P. capsici were poured to the rhizosphere soils mixed with BZA. Application of the BZA suspension spray to the fruit surface infected with C. acutatum significantly suppressed the disease occurrence of anthracnose on the red-pepper plants. These results suggest that BZA can be used to develop a promising agrochemical to control phytophthora blight and anthracnose of redpepper plants.

• The Korean Journal of Pesticide Science
• /
• v.18 no.3
• /
• pp.181-190
• /
• 2014

An entomopathogenic bacterium, Bacillus thuringiensis (Bt), can sporulate along with production of insecticidal Cry toxins. Bt Cry toxins exhibit relatively narrow spectrum to target insects due to their specific interactions with midgut receptors. This study designed several strategies to enhance Bt efficacy in target insect spectrum and insecticidal activity. Four Cry toxins were purified from four different Bt strains and showed relatively narrow target insect spectrum. However, the Cry mixtures significantly expanded their target insect spectra. The additional effect of baculovirus to Cry toxin was tested with recombinant baculoviruses expressing Cry1Ac or Cry1Ca. However, the baculovirus was little effective to expand target insect spectrum. Bacterial culture broth of Xenorhabdus nematophila (Xn) significantly suppressed insect cellular immune response and increased Cry toxicity. The addition of Xn culture broth to Cry mixture significantly enhanced Bt efficacy in target insect spectrum and insecticidal activity.