• Horticultural Science & Technology
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• v.31 no.6
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• pp.657-665
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• 2013

A large patch disease caused by Rhizoctonia solani AG2-2 (IV) is a serious problem in Korean lawngrass (Zoysia japonica) sites including golf courses and sports fields in Korea. Antagonistic microorganisms against R. solani AG2-2 (IV) were isolated from various forest and crop soil sources in Southern Korea. Among the 61 isolates, I-009, FRIN-001-1, and YPIN-022 strains showing dramatic inhibition of the mycelial growth of R. solani AG2-2 (IV) in the pairing culture were selected as the most potential antagonistic microorganisms for this study. Based on the 16s RNA sequence comparison, I-009 and FRIN-001-1 isolates were identified as Bacillus spp., while YPIN-022 isolate belongs to the genus Pseudomonas. The greater inhibition (clear) zone between two edges of the selected and pathogenic microbes ranged from 11 to 15 mm in three selections, but the others averaged to 7 mm out of 30 mm distance. In another antifungal test using culture filtrate, those three isolates represented a range of 51.7 to 63.5% suppression potential. The selected isolates also inhibited significantly the stem-segment colonization by R. solani AG2-2 (IV) in vivo test by 28.1%, 43.0%, and 23.7% when inoculated with I-009, FRIN-001-1, and YPIN-022, respectively. The highest antagonistic activity for the large patch disease was demonstrated by the isolate FRIN-001-1, which will be useful for developing a bio-pesticide against Rhizoctonia.

• The Plant Pathology Journal
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• v.20 no.4
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• pp.283-288
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• 2004

Four Erwinia carotovora strains isolated from potatoes showing blackleg symptoms and rotted Chinese cabbage were analysed by biochemical tests and sequence analysis of 16S rDNA and 16S-23S rRNA intergenic spacer (IGS) regions, and the data were compared to related E. carotovora strains. Based on the results of the biochemical tests and sequence analysis, 2 of the 4 strains were identified as E. carotovora subsp. carotovora (Ecc), whereas the rest strains were distinct from Ecc. The last two strains, HCC3 and JEJU, were biochemically similar to E, carotovora subsp. atroseptica (Eca). However, the results of sequence analysis and Eca-specific PCR assays showed that the strains were distinct from Eca. On the basis of 16S rDNA sequence analysis, HCC3 and JEJU strains were placed in E. carotovora subsp. odorifera and E. carotovora subsp. wasabiae, respectively. The results of sequence analysis and specific PCR assay for Eca indicated that Asian Eca strains were distinct from European Eca strains, although they were phenotycally homogeneous.

• Biomedical Science Letters
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• v.18 no.1
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• pp.79-82
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• 2012

Acinetobacter infections are of great concern in clinical settings because of multi-drug resistance (MDR) and high mortality of the infected patients. The MDR Acinetobacter baumannii has emerged as a significant infectious agent in hospitals worldwide. The purpose of this study was to determine for molecular characterization of MDR A. baumannii clinical isolates obtained from the Wonju Christian Hospital in Gangwon province of Korea. A total of seventy nonduplicate A. baumannii isolates were collected from the Wonju Christian Hospital in Korea from March to April in 2011. All of the MDR A. baumannii isolates were encoded by $bla_{OXA-23-like}$ gene and all isolates with the $bla_{OXA-23-like}$ gene had the upstream element ISAba1 to promote increased gene expression and subsequent resistance to carbapenem. 16S rRNA methylase gene (armA) was detected in 44 clinical isolates which were resistant to amikacin, and phosphotransferase genes encoding aac(3)-Ia and aac(6')-Ib were the most prevalent. A combination of 16S rRNA methylase and aminoglycoside-modifying enzyme genes (armA, aac(3)-Ia, aac(6')-Ib, and aph(3')-Ia) were found in 31 isolates. The sequencing results for the quinolone resistance-determining region (QRDR) of gyrA and parC revealed the presence of Ser (TCA) 83 Leu (TTA) and Ser (TCG) 80 Leu (TTG) substitutions in the respective enzymes for all MDR. Molecular typing for MDR A. baumannii could be helpful in confirming the identification of a common source or cross-contamination. This is an important step in enabling epidemiological tracing of these strains.

• Journal of Life Science
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• v.23 no.12
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• pp.1486-1494
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• 2013

A high protease-producing strain was isolated and identified from the digestive tract of octopus vulgaris by detecting a hydrolysis circle of protease and its activity. The strain was identified by morphology observation, biochemical experiments, and 16S rRNA sequence analysis. The protease obtained from the strain was purified by a three-step process involving ammonium sulfate precipitation, carboxy methyl-cellulose (CM-52) cation-exchange chromatography, and DEAE-Sephadex A50 anion-exchange chromatography. The properties of protease were characterized as well. The strain Bacillus sp. QDV-3, which produced the highest activity of protease, was isolated. On the basis of the phenotypic and biochemical characterization and 16S rRNA gene-sequencing studies, the isolate was identified as follows: domain: Bacteria; phylum: Firmicutes; class: Bacilli; order: Bacillales; family: Bacillaceae; and genus: Bacillus. The isolate was shown to have a 99.2% similarity with Bacillus flexus. A high active protease designated as QDV-E, with a molecular weight of 61.6 kDa, was obtained. The enzyme was found to be active in the pH range of 9.0-9.5 and its optimum temperature was $40^{\circ}C$. The protease activity retained more than 96% at the temperature of $50^{\circ}C$ for 60 min. Phenylmethylsulfonyl fluoride (PMSF) inhibited the enzyme activity, thus confirming that this protease isolated from Bacillus sp. QDV-3 is an alkaline serine protease. Metal ions, $Mn^{2+}$ and $Mg^{2+}$, were determined to enhance the protease activity, whereas $Ba^{2+}$, $Zn^{2+}$, and $Cu^{2+}$ were found to inactivate the enzyme.

• Asian-Australasian Journal of Animal Sciences
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• v.25 no.7
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• pp.1045-1054
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• 2012

The phylogenetic diversity of the bacteria in hot composting samples collected from three spatial locations was investigated by molecular tools in order to determine the influence of gradient effect on bacterial communities during the thermophilic phase of composting swine manure with rice straw. Total microbial DNA was extracted and bacterial near full-length 16S rRNA genes were subsequently amplified, cloned, restriction fragment length polymorphism-screened and sequenced. The superstratum sample had the highest microbial diversity among the three samples which was possibly related to the surrounding conditions of the sample resulting from the location. The results showed that the sequences related to Bacillus sp. were most common in the composts. In superstratum sample, 45 clones (33%) and 36 clones (27%) were affiliated with the Bacillus sp. and Clostridium sp., respectively; 74 clones (58%) were affiliated with the Clostridium sp. in the middle-level sample; 52 clones (40%) and 29 clones (23%) were affiliated with the Clostridium sp. and Bacillus sp. in substrate sample, respectively. It indicated that the microbial diversity and community in the samples were different for each sampling site, and different locations of the same pile often contained distinct and different microbial communities.

• Journal of Microbiology
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• v.34 no.3
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• pp.229-235
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• 1996

Microgram quantities of DNA per gram soil were recovered with SDS- based and freeze-and thaw procedures. The average DNA fragment size was > 23 Kb. This method generated minimal shearing of extracted DNA. However, the DNA extracts still contained considerable amounts of humic impurities sufficient to inhibit PCR. Several approaches were used to reduce the interferences with the PCR (use of CTAF in extraction step, Elutip-d column purification, addition of BSA to PCR buffer) to accomplish PCR with DNA extract as a template. Most of the DNA extracts were not digested completely by restriction endonuclease, and CTAB-TREATED ane Elutip-d column purified DNA extracts were partially digested. Regarding as restriction enzyme digestion, all PCRs failed to amplify 16S rRNA gene fragments in the DNA extracts. In the case of DNA extracts only where BSA was added to PCR buffer, PCR was successfully conducted whether the DNA extracts were treated with CTAB or purified with columns. However, these two treatments were indispensable for humic impurity-rich DNA extracts to generate the PCR-compatible DNA samples. Direct extraction of DNA, coupled with these procedures to remove and relieve interferences by humic impurities and followed by the PCR, can be rapid and simple method for molecular microbiological study on soil microorganisms.

• Asian-Australasian Journal of Animal Sciences
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• v.20 no.12
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• pp.1887-1894
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• 2007

In this paper, a rapid and reliable gene-targeted species-specific polymerase chain reaction (PCR) technique based on a two-step process was established to identify bifidobacteria in dairy products. The first step was the PCR assay for genus Bifidobacterium with genus specific primers followed by the second step, which identified the species level with species-specific primer mixtures. Ten specific primer pairs, designed from nucleotide sequences of the 16-23S rRNA region, were developed for the Bifidobacterium species including B. angulatum, B. animalis, B. bifidum, B. breve, B. catenulatum, B. infantis, B. longum, B. minimum, B. subtile, and B. thermophilum. This technique was applied to the identification of Bifidobacterium species isolated from 6 probiotic products, and four different Bifidobacterium spp. (B. bifidum, B. longum, B. infantis, and B. breve) were identified. The findings indicated that the 16S-23S rDNA gene-targeted species-specific PCR technique is a simple and reliable method for identification of bifidobacteria in probiotic products. PCR combined with Denaturing Gradient Gel Electrophoresis (DGGE) for identification of the bifidobacteria was also evaluated and compared with the gene-targeted species-specific technique. Results indicated that for fermented milk products consistency was found for both species-specific PCR and PCR-DGGE in detecting species. However, in some lyophilized products, the bands corresponding to these species were not visualized in the DGGE profile but the specific PCR gave a positive result.

• Journal of Life Science
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• v.23 no.6
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• pp.770-778
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• 2013

To determine the seasonal variation of bacterial community in the seawater of Gwangyang Bay, three hundred thirty six bacterial strains were isolated on February, May, July and October 2011. Amplified Ribosomal DNA Restriction Analysis (ARDRA) was used to construct the phylotyes of the isolates using the restriction endonuclease, Hae III. Diversity indices of ARDRA patterns were calculated. One hundred and one phylotypes including 40 unique pylotypes were found at the 80% similarity level. Partial 16S rRNA genes of one hundred thirty nine strains representing each phylotypes were sequenced and compared. Bacterial community composed of 4 different phyla which include Proteobacteria, Actinobacteria, Bacteroidetes and Firmicutes. Proteobacteria was the prevailing phylum in all seasons, followed by Bacteroidetes in winter, spring and autumn while Actinobacteria in summer. At the family level, Flavobacteriaceae dominated in winter and spring and Pseudoalteromonadaceae did in summer and autumn. Genera Altererythrobacter, Loktanella, Pseudoalteromonas and Vibrio were encountered in all seasons. The most diverse bacterial community was found in autumn followed by the order of spring, winter and summer.

• Journal of Life Science
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• v.31 no.2
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• pp.175-182
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• 2021

Saccharomyces lysate has the well-known function of soothing the skin in various ways: it is an anti-irritant and can treat skin care conditions, such as skin whitening and antioxidative activity. However, data on the safety for use of Saccharomyces lysate in cosmetics and skin care products are still limited. To design a new cosmetic material with antioxidant and skin-whitening effects, 80 yeast strains were isolated from berries grown in Sunchang. Among the isolates, the FT4-4 strain, which exhibited superior biological activities, was selected for further experiments. The FT4-4 strain was identified as Saccharomyces cerevisiae by 18S rRNA gene sequencing analysis. S. cerevisiae FT4-4 showed higher DPPH radical-scavenging (51.41%), superoxide dismutase (62.23%), and tyrosinase inhibition (64.75%) activities. The highest yield of biomass (3.16 g/l) and maximum growth rate of S. cerevisiae FT4-4 were observed within 16 h. Furthermore, the cytotoxicity potential of S. cerevisiae FT4-4 on B16F10 melanoma cells was measured by an MTT assay, and the results indicated that S. cerevisiae FT4-4 had a capacity to inhibit melanin up to 72.02% at an initial 10 mg/ml concentration. These results suggest that S. cerevisiae FT4-4 could be a promising candidate as a multi-functional material for application in the cosmetic industry, especially because of its antioxidant and skin-whitening effects.

• Journal of Microbiology and Biotechnology
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• v.19 no.3
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• pp.323-330
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• 2009

Nontuberculous mycobacteria (NTM) are a major cause of opportunistic infections in immunocompromised patients, making the reliable and rapid identification of NTM to the species level very important for the treatment of such patients. Therefore, this study evaluated the usefulness of the novel target genes tuf and tmRNA for the identification of NTM to the species level, using a PCRrestriction fragment length polymorphism analysis (PRA). A total of 44 reference strains and 17 clinical isolates of the genus Mycobacterium were used. The 741 bp or 744 bp tuf genes were amplified, restricted with two restriction enzymes (HaeIII/MboI), and sequenced. The tuf gene-PRA patterns were compared with those for the tmRNA (AvaII), hsp65 (HaeIII/HphI), rpoB (MspI/HaeIII), and 16S rRNA (HaeIII) genes. For the reference strains, the tuf gene-PRA yielded 43 HaeIII patterns, of which 35 (81.4%) showed unique patterns on the species level, whereas the tmRNA, hsp65, rpoB, and 16S rRNA-PRAs only showed 10 (23.3%), 32 (74.4%), 19 (44.2%), and 3 (7%) unique patterns after single digestion, respectively. The tuf gene-PRA produced a clear distinction between closely related NTM species, such as M. abscessus (557-84-58) and M. chelonae (477-84-80-58), and M. kansasii (141-136-80-63-58-54-51) and M. gastri (141-136-117-80-58-51). No difference was observed between the tuf-PRA patterns for the reference strains and clinical isolates. Thus, a diagnostic algorithm using a tuf gene-targeting PRA is a promising tool with more advantages than the previously used hsp65, rpoB, and 16S rRNA genes for the identification of NTM to the species level.