• Journal of Microbiology and Biotechnology
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• v.26 no.5
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• pp.909-917
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• 2016

The shiitake mushroom industry has suffered from Camptomyia (gall midges) pest, which feeds on the mycelium of shiitake mushroom during its cultivation. It has been postulated that fungal damage of shiitake bed-logs is associated with infestation by the insect pest, but this is not well understood. To understand the fungal damage associated with Camptomyia pest, various Trichoderma species were isolated, identified, and characterized. In addition to two previously known Trichoderma species, T. citrinoviride and T. deliquescens, two other Trichoderma species, T. harzianum and T. atroviride, were newly identified from the pestinfested bed-log samples obtained at three mushroom farms in Cheonan, Korea. Among these four species, T. harzianum was the most evident. The results of a chromogenic media-based assay for extracellular enzymes showed that these four species have the ability to produce amylase, carboxyl-methyl cellulase, avicelase, pectinase, and ß-glucosidase, thus indicating that they can degrade wood components. A dual culture assay on PDA indicated that T. harzianum, T. atroviride, and T. citrinoviride were antagonistic against the mycelial growth of a shiitake strain (Lentinula edodes). Inoculation tests on shiitake bed-logs revealed that all four species were able to damage the wood of bed-logs. Our results provide evidence that the four green mold species are the causal agents involved in fungal damage of shiitake bed-logs infested by Camptomyia pest.

• Mycobiology
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• v.43 no.1
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• pp.81-86
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• 2015

To promote the selection of promising monokaryotic strains of button mushroom (Agaricus bisporus) during breeding, 61 progeny strains derived from basidiospores of two different lines of dikaryotic parental strains, ASI1038 and ASI1346, were analyzed by nucleotide sequencing of the intergenic spacer I (IGS I) region in their rDNA and by extracellular enzyme assays. Nineteen different sizes of IGS I, which ranged from 1,301 to 1,348 bp, were present among twenty ASI1346-derived progeny strains, while 15 different sizes of IGS I, which ranged from 700 to 1,347 bp, were present among twenty ASI1038-derived progeny strains. Phylogenetic analysis of the IGS sequences revealed that different clades were present in both the ASI10388- and ASI1346-derived progeny strains. Plating assays of seven kinds of extracellular enzymes (${\beta}$-glucosidase, avicelase, CM-cellulase, amylase, pectinase, xylanase, and protease) also revealed apparent variation in the ability to produce extracellular enzymes among the 40 tested progeny strains from both parental A. bisporus strains. Overall, this study demonstrates that characterization of IGS I regions and extracellular enzymes is useful for the assessment of the substrate-degrading ability and heterogenicity of A. bisporus monokaryotic strains.

• Proceedings of the Korea Technical Association of the Pulp and Paper Industry Conference
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• 2011.04a
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• pp.81-91
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• 2011

Current fuel ethanol research and development deals with process engineering trends for improving biotechnological production of ethanol. Recently, a large amount of studies regarding the utilization of lignocellulosic biomass as a good feedstock for producing fuel ethanol is being carried out worldwide. The plant biomass is mainly composed of cellulose, hemicellulose and lignin. The main challenge in the conversion of biomass into ethanol is the complex, rigid and harsh structures which require efficient process and cost effective to break down. The isolation of microorganisms is one of the means for obtaining enzymes with properties suitable for industrial applications. For these reasons, crude cultures containing cellulosic biomass degrading microorganisms were isolated from rice field soil, cow farm soil and rotten rice straw from cow farm. Carboxymethyl cellulose (CMC), xylan and Avicel (microcrystalline cellulose) degradation zone of clearance on agar platefrom rice field soil resulted approximately at 25 mm, 24 mm and 22 mm respectively. As for cow farm soil, CMC, xylan and Avicel degradation clearancezone on agar plate resulted around at 24mm, 23mm and 21 mm respectively. Rotten rice straw from cow farm also resulted for CMC, xylan and Avicel degradation zone almost at 24 mm, 23 mm and 22 mm respectively. The objective of this study is to isolatebiomass degrading microbial strains having good efficiency in cellulose hydrolysis and observed the effects of different substrates (CMC, xylan and Avicel) on the production of cellulase enzymes (endo-glucanase, exo-glucanase, cellobiase, xylanase and avicelase) for producing low cost biofuel from cellulosic materials.

• The Plant Pathology Journal
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• v.35 no.6
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• pp.674-683
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• 2019

Some species of the Trichoderma genus are reported as the major problem in oak wood mushroom production in Korea. In spite of economic loss by the fungi, scientific information on airborne Trichoderma species is not much available. To generate information for disease management development we analyzed airborne Trichoderma. A total of 1,063 fungal isolates were purely obtained from indoor air sampling of cultivation houses used for oak wood mushroom using sawdust media. Among the obtained isolates, 248 isolates were identified as Trichoderma fungi including T. harzianum, T. atroviride, T. citrinoviride, and T. pseudokoningii, by morphological and molecular analysis. T. harzianum was dominant among the four identified species. All the four Trichoderma species grew fast on solid nutrient media tested (potato dextrose agar [PDA], malt extract agar [MEA], Czapek's Dox + yeast extract agar [CYA] and cornmeal dextrose agar). Compact mycelia growth and mass spore production were better on PDA and CYA. In addition, T. harzianum and T. citrinoviride formed greenish and yellowish mycelium and spores on PDA and CYA. Greenish and yellowish pigment was saturated into PDA only by T. pseudokoningii. These four Trichoderma species could produce extracellular enzymes of sawdust substrate degradation such as β-glucosidase, avicelase, CM-cellulase, amylase, pectinase, xylanase, and protease. Their mycelia inhibited the growth of oak wood mushroom mycelia of two tested cultivars on dual culture assay. Among of eleven antifungal agents tested, benomyl was the best to inhibit the growth of the four Trichoderma species. Our results demonstrate that the airborne Trichoderma fungi need to be properly managed in the cultivation houses for safe mushroom production.

• Asian-Australasian Journal of Animal Sciences
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• v.24 no.3
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• pp.364-368
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• 2011

An experiment was conducted to examine the effects of Cordyceps militaris mycelia on microbial populations and fibrolytic enzyme activities in vitro. C. militaris mycelia was added to buffered rumen fluid with final concentrations of 0.00, 0.10, 0.15, 0.20, 0.25 and 0.30 g/L and incubation times were for 3, 6, 9, 12, 24, 36, 48 and 72 h. At all incubation times, the supplementation of C. militaris mycelia linearly increased the number of total viable and celluloytic bacteria; maximum responses were seen with 0.25 g/L supplementation of C. militaris mycelia. The addition of C. militaris mycelia above the level of 0.20 g/L significantly (p<0.01) increased the number of total and cellulolytic bacteria compared with the control. On the other hand, the response of fungal counts to the supplementation of C. militaris mycelia showed a linear decrease; the lowest response was seen with 0.30 g/L supplementation of C. militaris mycelia. It would seem that C. militaris mycelia possess a strong negative effect on rumen fungi since the lowest level of C. militaris mycelia supplementation markedly decreased fungal counts. Carboxylmethyl cellulase activities were linearly increased by the addition of C. militaris mycelia except at 3 and 9 h incubation times. At all incubation times, the supplementation of C. militaris mycelia linearly increased the activities of xylanase and avicelase. In conclusion, the supplementation of C. militaris mycelia to the culture of mixed rumen microorganisms showed a positive effect on cellulolytic bacteria and cellulolytic enzyme activities but a negative effect on fungi.

• Korean Journal of Organic Agriculture
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• v.20 no.3
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• pp.327-343
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• 2012

In order to develop a high cellulolytic direct-fed microorganism (DFM) for ruminant productivity improvement, this study isolated cellulolytic bacteria from the rumen of Holstein dairy cows, and compared their cellulolytic abilities via DM degradability, gas production and cellulolytic enzyme activities. Twenty six bacteria were isolated from colonies grown in Dehority's artificial (DA) medium with 2% agar and cultured in DA medium containing filter paper at $39^{\circ}C$ for 24h. 16s rDNA gene sequencing of four strains from isolated bacteria showed that H8, H20 and H25 strains identified as Ruminococcus flavefaciens, and H23 strain identified as Fibrobacter succinogenes. H20 strain had higher degradability of filter paper compared with others during the incubation. H8 (R. flavefaciens), H20 (R. flavefaciens), H23 (F. succinogenes), H25 (R. flavefaciens) and RF (R. flavefaciens sijpesteijn, ATCC 19208) were cultured in DA medium with filter paper as a single carbon source for 0, 1, 2, 3, 4 and 6 days without shaking at $39^{\circ}C$, respectively. Dry matter degradability rates of H20, H23 and H25 were relatively higher than those of H8 and RF since 2 d incubation. The cumulative gas production of isolated cellulolytic bacteria increased with incubation time. At every incubation time, the gas production was highest in H20 strain. The activities of carboxymethylcellulase (CMCase) and Avicelase in the culture supernatant were significantly higher in H20 strain compared with others at every incubation time (p<0.05). Therefore, although further researches are required, the present results suggest that H20 strain could be a candidate of DFM in animal feed due to high cellulolytic ability.

• The Korean Journal of Mycology
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• v.10 no.2
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• pp.67-73
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• 1982

The $(NH_4)_2\;SO_4$ (70%) treated crude enzymes from the culture filtrates of the$C-7^{+t}$ strain of Pyricularia oryzae which was grown on 2% CMC (carboxymethyl cellulose) for 8 days at $28^{\circ}C$ were chromatographied on Sephadex G-150 and DEAE-Sephadex A-25 columns. From the chromatography, three fractions of CMCase$(C_x)$ was examined using Na-CMC as substrate. The $C_x$ enzyme activity was optimal at pH 6.0 and $40^{\circ}C$, stable up to $40^{\circ}C$. The values of Km and Vmax of the enzyme were $2.8{\times}10\;mM$ and 5.9m moles/hour, respectively. The molecular weight determined by Sephadex G-150 column chromatography was around 80,000. Approximately sevenfold purified $C_x$ enzyme gave a single protein band on the polyacrylamide gel electrophoresis.

• KSBB Journal
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• v.22 no.2
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• pp.114-118
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• 2007

In this study, an amylase-producing fungus, strain 15 was isolated from soil in order to saccharify food wastes with cellulolytic and amylolytic enzymes. The amylase production cultures were performed in Mandel's medium with 1% rice straw and 1% paper wastes as carbon sources. The strain produced various cellulolytic (FPase 0.25, xylanase 20.09, CMCase 3.15 U/mL-supernatant) and amylolytic ($\alpha$-amylase 1.20, gluco-amylase 0.70, $\beta$-amylase 2.40 U/mL-supernatant) enzymes in Mandel's medium. In 10 L jar fermenter, maximum amylase and FPase activities, 3.25 and 0.23 U/mL, were obtained when the culture was grown at 30$^{\circ}C$, 200 rpm and 0.6 vvm for 3 days. In 100 mL flask level and 10 L jar fermenter, amylase produced by the strain 15 showed similar cellulolytic and amylolytic enzyme activities with Trichoderma inhamatum KSJ1 isolated from rotten woods by previous researcher. The ability of saccharification to food wastes also showed similar degree. However, the isolate 15 appeared to be yellowish in YMEA plate comparing to Trichoderma inhamatum KSJ1 in greenish.

• Mycobiology
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• v.43 no.4
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• pp.458-466
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• 2015

Oak tree death caused by symbiosis of an ambrosia beetle, Platypus koryoensis, and an ophiostomatoid filamentous fungus, Raffaelea quercus-mongolicae, has been a nationwide problem in Korea since 2004. In this study, we surveyed the yeast species associated with P. koryoensis to better understand the diversity of fungal associates of the beetle pest. In 2009, a total of 195 yeast isolates were sampled from larvae and adult beetles (female and male) of P. koryoensis in Cheonan, Goyang, and Paju; 8 species were identified by based on their morphological, biochemical and molecular analyses. Meyerozyma guilliermondii and Candida kashinagacola were found to be the two dominant species. Among the 8 species, Candida homilentoma was a newly recorded yeast species in Korea, and thus, its mycological characteristics were described. The P. koryoensis symbiont R. quercusmongolicae did not show extracelluar CM-cellulase, xylanase and avicelase activity that are responsible for degradation of wood structure; however, C. kashinagacola and M. guilliermondii did show the three extracellular enzymatic activities. Extracelluar CM-cellulase activity was also found in Ambrosiozyma sp., C. homilentoma, C. kashinagacola, and Candida sp. Extracelluar pectinase activity was detected in Ambrosiozyma sp., C. homilentoma, Candida sp., and M. guilliermondii. All the 8 yeast species displayed compatible relationships with R. quercus-mongolicae when they were co-cultivated on yeast extract-malt extract plates. Overall, our results demonstrated that P. koryoensis carries the yeast species as a symbiotic fungal associate. This is first report of yeast diversity associated with P. koryoensis.

• Korean Journal of Microbiology
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• v.36 no.3
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• pp.181-186
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• 2000

The cellulosome of Clo.~tr~rlil~m tl\ulcornererfnocellum consistmg of 26 dfferent polypeptides contains calcium. The polypeptides dissociated when calcium was removed. Most of dockerill region in the catalytic polypeptides cleavcd during dmociation. The dissociated polypeptides were well separated by MonoQ column chromatography into CipA containing fraction, a fraction still complexed wit11 91 kDa (CelK-a). 60 IiDa and 57 kDa polypeptides, and fractious contailling mainly single polypeptide of 46 kDa (CelA-a) or 71 1d)a polypeptide (CelS-trj Most or the fractions hydrolyzed c~ystalliue cellulose The purified 71 kDa polypeptide was strictly dependent on calcium for crystalline cellulose hydvolyzing activities a1 $60^{\circ}C$~$70^{\circ}C$ but 46 kDa polypeptide was not. 46 M)a polypeptide digested cellodextri~~ as cellobiose or cellotriose unit, and glucose was produced together with cellobiose and cellotriose froln cellotetraosc. It seems that cellulosome produces final product, cellobiose, through coordinated ~qulation of activities of vannus subunits.