Journal of Microbiology

Volume 42(1); March 2004

Research Support, Non-U.S. Gov'ts

Denaturing Gradient Gel Electrophoresis Analysis of Bacterial Populations in 5-Stage Biological Nutrient Removal Process with Step Feed System for Wastewater Treatment: Soo-Youn Lee , Hyeon-Guk Kim , Jong Bok Park , Yong Keun Park; J. Microbiol. 2004;42(1):1-8.; DOI: https://doi.org/2009 [pii]

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Abstract: Changes in the bacterial populations of a 5-stage biological nutrient removal (BNR) process, with a step feed system for wastewater treatment, were monitored by denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S ribosomal DNA fragments. DGGE analysis indicated seasonal community changes were observed, however, community profiles of the total bacteria of each reactor showed only minor differences in the samples obtained from the same season. The number of major bands was higher in the summer samples, and decreased during the winter period, indicating that the microbial community structure became simpler at low temperatures. Since the nitrogen and phosphate removal efficiencies were highly maintained throughout the winter operation period, the bacteria which still remaining in the winter sample can be considered important, playing a key role in the present 5-stage BNR sludge. The prominent DGGE bands were excised, and sequenced to gain insight into the identities of the predominant bacterial populations present, and most were found to not be closely related to previously characterized bacteria. These data suggest the importance of culture independent methods for the quality control of wastewater treatment

A Method for Comparing Multiple Bacterial Community Structures from 16S rDNA Clone Library Sequences: Inae Hur , Jongsik Chun; J. Microbiol. 2004;42(1):9-13.; DOI: https://doi.org/2008 [pii]

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Abstract: Culture-independent approaches, based on 16S rDNA sequences, are extensively used in modern microbial ecology. Sequencing of the clone library generated from environmental DNA has advantages over fingerprint-based methods, such as denaturing gradient gel electrophoresis, as it provides precise identification and quantification of the phylotypes present in samples. However, to date, no method exists for comparing multiple bacterial community structures using clone library sequences. In this study, an automated method to achieve this has been developed, by applying pair wise alignment, hierarchical clustering and principle component analysis. The method has been demonstrated to be successful in comparing samples from various environments. The program, named CommCluster, was written in JAVA, and is now freely available, at http://chunlab.snu.ac.kr/commcluster/.

Journal Article

Genomic Relationship of Salmonella enterica Serovar Typhimurium DT104 Isolates from Korea and the United States: Shukho Kim , Sung Guen Chun , Ok Young Lim , Mi Sun Park , Yeon Ho Kang , Yong Ho Park , Bok Kwon Lee; J. Microbiol. 2004;42(1):14-19.; DOI: https://doi.org/2007 [pii]

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Abstract: Salmonella enterica serovar Typhimurium DT104 (Salmonella Typhimurium DT104 or DT104) hasbeen emerging as a common pathogen for human in Korea since 1997. In order to compare the genomic relationship and to search for the dominant strains in Korea, we conducted pulsed-field gel electrophoresis (PFGE) and IS200 fingerprinting of 25 epidemiological unrelated isolates from human and animals from Korea and cattle from America. Two Salmonella Typhimurium DT104 isolates from human in Korea and all 8 isolates from American cattle had indistinguishable patterns from the PFGE and IS200 fingerprinting but multidrug-resistant Salmonella Typhimurium isolates, including DT104, from Korean animals had diverse genetic patterns. The data suggest that a dominant DT104 strain might have circulated between Korean and American cattle and that it had a high level of clonality.

Research Support, Non-U.S. Gov't

Characterization of Recombinant Drosophila melanogaster Myo-inositol-1-phosphate Synthase Expressed in Escherichia coli: Sang-Hee Park , Jong-Il Kim; J. Microbiol. 2004;42(1):20-24.; DOI: https://doi.org/2006 [pii]

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Abstract: Cloned myo-inositol-1-phpsphate synthase (INOS) of Drosophila melanogaster was expressed in Escherichia coli, and purified using a His affinity column. The purified INOS required NAD^+ for the conversion of glucose-6-phosphate to inositol-1-phosphate. The optimum pH for myo-inositol-1-phosphate synthase is 7.5, and the maximum activity was measured at 40^oC. The molecular weight of the native enzyme, as determined by gel filtration, was approximately M_r 271,000±15,000. A single subunit of approximately M_r 62,000±5,000 was detected upon SDS-polyacrylamide gel electrophoresis. The Michaelis (K_m) and dissociation constants for glucose-6-phosphate were 3.5 and 3.7 mM, whereas for the cofactor NAD^+ these were 0.42 and 0.4 mM, respectively.

Journal Articles

Study on Persistent Infection of Japanese Encephalitis Virus Beijing-1 Strain in Serum-free Sf9 Cell Cultures: Hun Kim , Su Jeen Lee , Jin Yong Park , Yong Wook Park , Hyun Sung Kim , Heui-Yun Kang , Byung-Ki Hur , Yeon-Woo Ryu , Sang In Han , Jong Su Kim; J. Microbiol. 2004;42(1):25-31.; DOI: https://doi.org/2005 [pii]

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Abstract: Sf9 cells have obvious advantages for the conventional production technology of vaccine. They are useful tools for high concentration and large-scale cultures. Sf9 cells were grown to maximal concentration, 8×10^6 cells/ml in a 500ml spinner flask, with a doubling time at the exponentially growing phase of 24.5 hours, using serum-free media. To explore the ability of Sf9 cells to be infected by the Japanese encephalitis (JE) virus Beijing-1 strain, Sf9 cells were infected with the virus. By 4-5 days post-infection, 10-15% of the Sf9 cells showed cytopathic effect (CPE), from granularity to the formation of syncytia and multinucleated giant cells continuously observed over a period of 35 days. Positive fluorescent reactions were detected in 30-40% of cells infected with the JE virus Beijing-1 strain, and the uninfected Sf9 cells were completely negative. Virus particles, propagated in Sf9 and Vero cells, were concentrated by sedimentation on 40% trehalose cushions by ultracentrifugation, and showed identical patterns of viral morphogenesis. Complete virus particles, 40 to 50 nm in diameter, were observed, and JE virus envelope (E) proteins, at 53 kDa, were found in the western blot analysis to the anti-JE virus E protein monoclonal antibody and reacted as a magenta band in the same position to the glycoprotein staining. To evaluate whether the infectious virus was produced in Sf9 cells inoculated with the JE virus Beijing-1 stain, Sf9 cells were inoculated with the virus, and sample harvested every 5 days. The titers of the JE virus Beijing-1 strain rose from 1.0×10^5 to 1.5×10^6 pfu/ml. The infected Sf9 cells could be subcultured in serum-free medium, with no change in the plaque sizes formed by the JE virus Beijing-1 strain in the plaque assay. It is suggested that the ability of the JE virus Beijing-1 strain to infect Sf9 cells in serum-free media will provide a useful insect cell system, where the JE virus replication, cytopathogenicity and vaccine immunogen can be studied.

Schizosaccharomyces pombe rsm1 Genetically Interacts with spmex67, Which Is Involved in mRNA Export: Jin Ho Yoon; J. Microbiol. 2004;42(1):32-36.; DOI: https://doi.org/2004 [pii]

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Abstract: We have previously isolated three synthetic lethal mutants from Schizosaccharomyces pombe in order to identify mutations in the genes that are functionally linked to spmex67 with respect to mRNA export. A novel rsm1 gene was isolated by complementation of the growth defect in one of the synthetic lethal mutants, SLMex1. The rsm1 gene contains no introns and encodes a 296 amino-acid-long protein with the RING finger domain, a C3HC4 in the N-terminal half. The [delta]rsm1 null mutant is viable, but it showed a slight poly(A)^+ RNA accumulation in the nucleus. Also, the combination of [delta]rsm1 and [delta]spmex67 mutations confers synthetic lethality that is accompanied by the severe poly(A)^+ RNA export defect. These results suggest that rsm1 is involved in mRNA export from the nucleus.

The Role of Enzymes Produced by White-Rot Fungus Irpex lacteus in the Decolorization of the Textile Industry Effluent: Kwang-Soo Shin; J. Microbiol. 2004;42(1):37-41.; DOI: https://doi.org/2003 [pii]

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Abstract: The textile industry wastewater has been decolorized efficiently by the white rot fungus, Irpex lacteus, without adding any chemicals. The degree of the decolorization of the dye effluent by shaking or stationary cultures is 59 and 93%, respectively, on the 8th day. The higher level of manganese-dependent peroxidase (MnP) and non-specific peroxidase (NsP) was detected in stationary cultures than in the cultures shaken. Laccase activities were equivalent in both cultures and its level was not affected significantly by the culture duration. Neither lignin peroxidase (LiP) nor Remazol Brilliant Blue R oxidase (RBBR ox) was detected in both cultures. The absorbance of the dye effluent was significantly decreased by the stationary culture filtrate of 7 days in the absence of Mn (II) and veratryl alcohol. In the stationary culture filtrate, three or more additional peroxidase bands were detected by the zymogram analysis.

Research Support, Non-U.S. Gov'ts

Chlorothalonil-Biotransformation by Glutathione S-Transferase of Escherichia coli: Young-Mog Kim , Kunbawui Park , Soon-Hyun Jung , Jun-Ho Choi , Won-Chan Kim , Gil-Jae Joo , In-Koo Rhee; J. Microbiol. 2004;42(1):42-46.; DOI: https://doi.org/2002 [pii]

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Abstract: It has recently been reported that one of the most important factors of yeast resistance to the fungicide chlorothalonil is the glutathione contents and the catalytic efficiency of glutathione S-transferase (GST) (Shin et al., 2003). GST is known to catalyze the conjugation of glutathione to a wide variety of xenobiotics, resulting in detoxification. In an attempt to elucidate the relation between chlorothalonil detoxification and GST, the GST of Escherichia coli was expressed and purified. The drug hypersensitive E. coli KAM3 cells harboring a plasmid for the overexpression of the GST gene can grow in the presence of chlorothalonil. The purified GST showed chlorothalonil-biotransformation activity in the presence of glutathione. Thus, chlorothalonil is detoxified by the mechanism of glutathione conjugation catalyzed by GST.

Effect of Titanium-Ion on the Growth of Various Bacterial Species: Tae Shick Yu; J. Microbiol. 2004;42(1):47-50.; DOI: https://doi.org/2001 [pii]

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Abstract: There are a number of studies that explain the metabolism and roles of metallic titanium and titaniumion. One of the most intriguing results from these studies is the finding of metallic titanium having no bacteriostatic effects on oral bacterial species. In this research, the effects of titanium-ion on the growth of twenty-two bacterial species, some of which are commonly found in foods such as yoghurt, kimchi, and soy fermented products, were investigated. All but two bacteria, Escherichia coli and Pseudomonas aeruginosa appeared to be sensitive to titanium-ion. These two species were grown on 360 μg/ml of titanium-ions, and they were found to be resistant to the titanium-ion. Both the wild type and plasmid cured E. coli showed good growth in a medium with 200 μg/ml of titanium-ions. These results suggest that titanium-resistance was independent from the effects of the plasmid in E. coli.

Optimal Fermentation Conditions for Enhanced Glutathione Production by Saccharomyces cerevisiae FF-8: Jae-Young Cha , Jin-Chul Park , Beong-Sam Jeon , Young-Choon Lee , Young-Su Cho; J. Microbiol. 2004;42(1):51-55.; DOI: https://doi.org/2000 [pii]

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Abstract: The influence of feedstock amino acids, salt, carbon and nitrogen sources on glutathione production by Saccharomyces cerevisiae FF-8 was investigated. Glucose, yeast extract, KH_2PO_4, and L-cysteine were found to be suitable feedstock. Highest glutathione production was obtained after cultivation with shaking for 72 h in a medium containing glucose 3.0% (w/v), yeast extract 3.0%, KH_2PO_4 0.06% and L-cysteine 0.06%. The glutathione concentration achieved using this medium increased 2.27-fold to 204 mg/l compared to YM basal medium.

Journal Article

Transposition of IntAs into the Conserved Regions of IS3 Family Elements: Chang-Gyun Han; J. Microbiol. 2004;42(1):56-59.; DOI: https://doi.org/1999 [pii]

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Abstract: Together with the previous reports, my computer survey revealed that several bacteria contain six copies of the type group II intron IntA. The sequence analysis of IntAs showed the high level of homology in the nucleotide sequence (91.9~99.8%). The consensus sequence, 2,270 base pair long, was derived from the nucleotide sequences of all IntA members. The size of the open reading frame intA was 502 amino acids long, that is homologous to reverse transcriptase-like proteins encoded within the group II introns. It was reported that EPEC.IntA and Sf.IntA were inserted into IS911 and IS629, respectively. The sequence of the flanking region IntA was analyzed here. The data show the insertion of EC.IntA into IS629, the insertion of EHEC.IntA into IS3, the insertion of Yp.IntA into IS904-like sequence, and the insertion of EK12.IntA into IS911. Interestingly, these IS elements nested by IntAs were the members of IS3 family elements. The sequences of the IS3 members correspond to the OrfB with the DDE motif conserved in retroviral integrases. Alignment of the flanking sequences of IntAs revealed that the flanking regions -25 to +10 of insertion sites, that are generally believed to be required for the retrohoming, were not strongly conserved. The data presented here suggests that the retrohoming pathway of IntA seems to differ from those of other group II introns.

Research Support, Non-U.S. Gov'ts

Use of Sucrose-Agar Globule with Root Exudates for Mass Production of Vesicular Arbuscular Mycorrhizal Fungi: Thangaswamy Selvaraj , Hoon Kim; J. Microbiol. 2004;42(1):60-63.; DOI: https://doi.org/1998 [pii]

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Abstract: A sucrose-agar globule (SAG) was newly introduced to increase production of the vesicular arbuscular mycorrhizal (VAM) fungal spores, Gigaspora gigantea and Glomus fasciculatum. An SAG inoculum and a sucrose-agar globule with root exudates (SAGE) inoculum were prepared, and their spore productions were compared with a soil inoculum. When the SAGE was used as the inoculum on sucroseagar medium plates the number of spores was increased (35% more than the soil inoculum). After the soil inoculum and SAGE were inoculated on an experimental plant, Zingiber officinale, the percentage root colonization, number of VAM spores, and dry matter content were analyzed. It was observed that the SAGE showed a higher percentage of root colonization (about 10% more), and increases in the number of spores (about 26%) and dry matter (more than 13%) for the two VAM fungal spores than the soil inoculum. The results of this study suggested that the SAGE inoculum may be useful for the mass production of VAM fungi and also for the large scale production of VAM fungal fertilizer.

Formation and Dispersion of Mycelial Pellets of Streptomyces coelicolor A3(2): Yul-Min Kim , Jae-heon Kim; J. Microbiol. 2004;42(1):64-67.; DOI: https://doi.org/1997 [pii]

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Abstract: The pellets from a culture of Streptomyces coelicolor A3(2) that were submerged shaken were disintegrated into numerous hyphal fragments by DNase treatment. The pellets were increasingly dispersed by hyaluronidase treatment, and mycelial fragments were easily detached from the pellets. The submerged mycelium grew by forming complexes with calcium phosphate precipitates or kaolin, a soil particle. Therefore, the pellet formation of Streptomyces coelicolor A3(2) can be considered a biofilm formation, including the participation of adhesive extracellular polymers and the insoluble substrates.

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