• International Journal of Industrial Entomology and Biomaterials
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• v.8 no.1
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• pp.77-80
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• 2004

For the development of the alternative control system against the major forest pests, Beauveria spp. F-101, isolated from a dead larva of Thecodiplosis japonensis, was selected because this isolate showed high pathogenicities against T. japonensis and Acantholyda parki. Beauveria spp. F-101 had irregular clustered conidio-phores and conidia borne on a distinctive apical zigzag extension, and it showed typical characteristic of the genus, Beauveria in morphology. For molecular based-identification, the ribosomal ITS region of Beauveria spp. F-101 was amplified with ITS1 and ITS4 primers, and cloned into pGEM- T Easy vector. The amplified PCR product was 569 bp in size and completely sequenced. The similarities of the cloned ITS sequence were 99 ％ and 97％ to those of B. bassiana and B. brongniartii, respectively. In comparison to other species among the genus Beauveria, the ITS region of Beauveria spp. F-101 showed a similarity of 95％ to B. amorpha, 95％ to B. tenella, 89％ to B. vermiconia and 69％ to B. alba, respectively. In addition, in comparison to different genus, it had 95％ similarities to Cordyceps militaris and 91％ to Paecilomyces tenuipes. Accordingly, the current result suggests that Beauveria spp. F-101 was a variant of B. bassiana and it seems to be a new isolate considering sequence variation in ITS region.

• International Journal of Industrial Entomology and Biomaterials
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• v.11 no.1
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• pp.37-42
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• 2005

We cloned the $\beta$-tubulin genes from the wild type strain and two benomyl-resistant mutants of Metahizium anisopliae and determined their nucleotide sequences. A $\beta$-tubulin encoding 448-residue protein from wild type M. anisopliae shows strong homology to other $\beta$-tubulins. The coding region is interrupted by four introns. Comparisons of intron position between the M. anisopliae gene and other fungal $\beta$-tubulin genes show considerable positional conservation. The mutations responsible for benomyl resistance were determined in two spontaneous mutants, 8-18 and 8­19. One mutant 8-18 substituted glutamate for aspar­agine at position 33 and lysine for glutamine at position 134. The other mutant 8-19 showed alterations at three positions of $\beta$-tubulin arginine for tryptophan at position 21, lysine for asparagine at position 33, and phenylalanine for leucine at position 240. These data suggest that regions of $\beta$-tubulin containing amino acids 21, 33,134, and 240 interact to form the binding site of benomyl.

• Journal of Microbiology and Biotechnology
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• v.27 no.3
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• pp.439-449
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• 2017

Beauveria bassiana infects a number of pest species and is known to produce insecticidal substances, such as the nonribosomal peptides (NRPs) beauvericin and bassianolide. However, most NRPs and their biological roles in B. bassiana remain undiscovered. To identify NRPs that potentially contribute to pathogenesis, the 21 predicted NRP synthetases (NRPSs) or NRPS-like proteins of B. bassiana ARSEF 2860 were primarily ranked into three functional groups: basic metabolism (7 NRPSs), pathogenicity (12 NRPSs), and unknown function (2 NRPSs). Based on the transcript levels during in vivo growth on diamondback moth (Plutella xylostella (Linnaeus)), half of the Group II NRPSs were likely to be involved in infection. Given that the metabolites biosynthesized by these NRPSs remain to be determined, our result underlines the importance of the NRPSome in fungal pathogenesis, and will serve as a guide for future genomic mining projects to discover functionally essential and structurally diverse NRPs in fungal genomes.

• Applied Biological Chemistry
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• v.41 no.3
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• pp.219-223
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• 1998

We have established a transformation system for entomopathogenic fungus, Metarhizium anisopliae, in order to develop mycoinsecticide by recombinant DNA techniques. Protoplasts of M. anisopliae would be transformed to a benomyl-resistant by introducing pBRG-4 plasmid DNA, which contains a ${\beta}-tubulin$ gene of Aspergillus flavus conferring resistance to benomyl and a pyr4 gene of Neurospora crassa, in the presence of 5% polyethylene glycol and 10 mM calcium chloride. Transformants occuring at a frequency of 10 colonies per $50\;{\mu}g$ pBRG-4 DNA grew on the $5\;{\mu}g/ml$ concentrations of benamyl, while the wild type was inhibited by $2.5\;{\mu}g/ml$. From the Southern analysis using genomic DNAs isolated from M. anisopliae transformants, the positive signals suggested that the ${\beta}-tubulin$ gene had integrated in the M. anisopliae genome by homologous recombination.

• Journal of Microbiology and Biotechnology
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• v.20 no.7
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• pp.1053-1060
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• 2010

Cordyceps militaris, an entomopathogenic fungus belonging to the class Ascomycetes, has been reported to have beneficial biological activities such as hypoglycemic, anti-inflammatory, antitumor, antimetastatic, hypolipidemic, immunomodulatory, and antioxidant effects. In this study, the crude water-soluble polysaccharide CMP, which was obtained from the fruiting body of C. militaris by hot water extraction and ethanol precipitation, was fractionated by DEAE-cellulose and Sepharose CL-6B column chromatographies. This process resulted in three polysaccharide fractions, termed CMP Fr I, CMP Fr II, and CMP Fr III. Of these fractions, CMP Fr II, with an average molecular mass of 127 kDa, was able to upregulate effectively the phenotypic functions of macrophages such as NO production and cytokine expression. The chemical property of the stimulatory polysaccharide, CMP Fr II, was determined based on its monosaccharide composition, which consisted of glucose (56.4%), galactose (26.4%), and mannose (17.2%). Its structural characteristics were investigated by a combination of chemical and instrumental analyses, including methylation, reductive cleavage, acetylation, Fourier transform infrared spectroscopy (FTIR), and gas chromatography-mass spectrometry (GCMS). Results indicated that CMP Fr II consisted of the (1${\rightarrow}$4) or (1${\rightarrow}$2) linked glucopyranosyl or galactopyranosyl residue with a (1${\rightarrow}$2) or (1${\rightarrow}$6) linked mannopyranosyl, glucopyranosyl, or galactopyranosyl residue as a side chain. The configuration of the ${\beta}$-linkage and random coil conformation of CMP Fr II were confirmed using a Fungi-Fluor kit and Congo red reagent, respectively.

• Journal of Life Science
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• v.15 no.1 s.68
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• pp.15-20
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• 2005

The unstructured model was tested to describe mycelial growth, exopolysaccharide formation, and substrate consumption in submerged mycelial culture of Paeeiliomyees tenuipes C240. The Logistic equation for mycelial growth, the Luedeking-Piret equation for exopolysaccharide formation, and Luedeking­Piret-like equations for glucose consumptions were successfully incorporated into the model. The value of the key kinetic constants were: maximum specific growth rate ${\mu}m,\;0.7281\;h^{-1};$ growth­associated constant for exopolysaccharide production $(\alpha),\;0.1743g(g\;cells)^{-1}$; non-growth associated constant for exopolysaccharide production $(\beta),\;0.0019g(g\;cells)^{-1}\;;$ maintenance coefficient $(m_s),\;0.0572g\;(g\;cells)^{-1}$. When compared with batch experimental data, the model successfully provided a reasonable description for each parameter during the entire growth phase. The model showed that the production of exopolysaccharide in P. tenuipes C240 was growth-associated. The model tested in the present study can be applied to the design, scale-up, and control of fermentation process for other kinds of basidiomycetes or ascomycetes.

• 한국생물공학회:학술대회논문집
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• 2002.04a
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• pp.469-472
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• 2002

Solid state fermentation was performed for the production of entomopathogenic fungus Metarhizium anisopliae HY-2 using wheat bran media containing rice bran. Fungal growth in a solid state fermentation system was estimated by viable cell count, spore count, and mycelial biomass. It was used chemical method measuring N-acetyl-glucosamine (chitin) content for estimating of mycelial biomass. In static flask culture, viable cell reached 2.40 ${\times}$ $10^8$ cfu/g at 23 days of culture at $27^{\circ}C$ and then mycelial biomass was 41.59 mg/g. Specific growth rate(${\mu}$ max) was 0.0418 $h^{-1}$ between 3 and 9 days when estimated by viable cell count and was 0.00976 $h^{-1}$ between 9 and 17 days when N-acetylglucosamine content was measured. Viable cells reached 1.12 ${\times}$ $10^8$ cfu/g in polypropylene-bag at 28 days of culture at $27^{\circ}C$. Formulated microbial pesticide containing M. anisopliae HY-2 were tested their bio-activity against Chestnut Brown Chafer (Adoretus tenuimaculatus). The protection rate of the liquid culture showed 13 ${\sim}$ 26 % with 1st to 3rd instar, and spore suspension of M. anisopliae HY-2 showed 56 ${\sim}$ 64%. Conidia produced by large scale solid-state fermentation showed 20 ${\sim}$ 27 % activity 60 ${\sim}$ 64 % with M. anisopliae HY-2.

• Mycobiology
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• v.46 no.4
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• pp.382-387
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• 2018

The entomopathogenic fungus Cordyceps militaris is a valuable medicinal ascomycete, which degenerates frequently during subsequent culture. To avoid economic losses during industrialized production, scratching stimuli of mycelia was introduced to improve the fruiting body production. The present results indicated that higher yields and biological efficiency were obtained from two degenerate strains (YN1-14 and YN2-7) but not from g38 (an insertional mutant in Rhf1 gene with higher yields and shorter growth periods). Furthermore, the growth periods of the fruiting bodies were at least 5 days earlier when the mycelia were scratched before stromata differentiation. Three ROS-scavenging genes including Cu/Zn superoxide dismutase (CmSod1), Glutathione peroxidase (CmGpx), and Catalase A (CmCat A) were isolated and their expression profiles against scratching were determined in degenerate strain YN1-14 and mutant strain g38. At day 5 after scratching, the expression level of CmGpx significantly decreased for strain g38, but that of CmSod1 significantly increased for YN1-14. These results indicated that scratching is an effective way to promote fruiting body production of degenerate strain, which may be related at least with Rhf1 and active oxygen scavenging genes.

• The Korean Journal of Mycology
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• v.29 no.1
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• pp.61-66
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• 2001

Entomopathogenic fungus of Cordyceps militaris is famous of its medicinal efficacies. An antitumor compound was purified from the n-hexane extract of artificially cultivated fruiting bodies of C. militaris and identified as ergosterol peroxide $(5{\alpha},\;8{\alpha}-epidioxy-24(R)-methylcholesta-6,22-dien-3{\beta}-o1,\; C_{28}H_{44}O_3)$ mainly by $^1H\;and\;^{13}C-NMR$ spectroscopic techniques. When the antitumor activity of ergosterol peroxide was measured against 3 tumor cell lines from Korean cancer patients, it showed the most strong activity against gastric cancer SNU-l cell line 3 days after treatment. The 50% inhibitory concentrations $(IC_{50})$ of ergosterol peroxide 6 days after treatment were $75.8{\mu}g/ml$ for human gastric SNU-1 tumor cell line, $39.7{\mu}g/ml$ for human colorectal SNU-C4 tumor cell line and $32.7{\mu}g/ml$ for human hepatoma SNU-354 cell line.

• Korean journal of applied entomology
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• v.35 no.3
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• pp.221-227
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• 1996

Four strains of Beauveria bassiana F101, F587, F9 and FJ8, were received from Forestry Research Institute,Seoul. The strain, B. bassiana F101, was the most active in the enzymatic activity and spore production. Whenspores of B. bassiana FlOl were sprayed on the female larva of the black tipped sawfly with various concentrationsof 6 xl 0'~-10s~p ores/ml, insects started to die from 5-7 days, and were covered with mycelia andspores in 24-28 days at 25"C, while the insect did not show visible symptoms even after 50 days at 4'C. Theinsect injected with 5 pl of spore solution (3X l0'~-10s~p oreslml) died within 30-98 and 38-218 hours at25$^{\circ}$C and 4"C, respectively. About 3 days (60 hours) after the injection with a concentration of 3 x lo9 spores1ml, at the point of the insect's death, lots of proteins started to disappear '||'&'||' the hemolymph, fat body and carcaseat 25'C. Esterase activity in the tissues was gone suddenly after that time. Six days after the spray, manyprotein and esterase bands were lost in the hemolymph, but not those in the fat body and carcase. When thefungi growing in the host were exposed in the air, they put energy for spore production, while numerous longand thin mycelia branched out from the host body in the soil.e host body in the soil.