Journal of Microbiology

Volume 38(1); March 2000

Characterization of Aeromonas hydrophila Isolated from Rainbow Trouts in Korea: Soondeuk Lee , Sookyung Kim , Yoojung Oh , Yeonhee Lee; J. Microbiol. 2000;38(1):1-7.

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Abstract: Eight strains of Aeromonas hydrophila isolated from diseased trout in Korea were characterized and compared with an American type strain by various methods including biochemical and physiological tests, PCR, randomly amplified polymorphic DNA (RAPD), plasmid profiling, and gel electrophoresis of total, membrane, and extracellular proteins. Virulence factors such as surface array proteins, cytotoxin, hemolysin, haemagglutinin, and protease were also investigated. The Korean strains showed het-erogeneity in lysine decarboxylase production, utilization of various carbon sources, and production of acetoin. Five strains had the same profiles of total and membrane proteins. Six strains haemag-glutinated with trout red blood cells (RBCs) which was inhibited by fucose, galactose, and mannose, except for No. 1 where haemagglutination was inhibited by only galactose and mannose, but not by fucose. Four isolates haemagglutinated with human RBCs which was inhibited by fucose and mannose yet not by galactose. The type strain haemagglutinated only with trout RBCs which was inhibited by fucose, galactose, and mannose. Every isolate secreted protease, hemolysin, cytotoxin, and siderophore, but no enterotoxin. Results showed that the Korean isolates, except for No. 7, had very different biochemical and molecular characteristics from those of the American type strain.

Differentiation of Salmonella typhimurium from Gram-negative Intestinal Microbes by Randomly Amplified Polymorphic DNA (RAPD) Fingerprinting: Un-Ho Jin , Tae-Wook Chung , June-Ki Kim , Kyung-Soo Nam , Sang-Do Ha , Cheorl-Ho Kim; J. Microbiol. 2000;38(1):8-10.

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Abstract: In order to rapidly identify and differentiate Salmonella typhimurium from the intestinal gram-negative bacteria, randomly amplified polymorphic DNA (RAPD) fingerprinting of Salmonella typhimurium was carried out using random primers designated OPA-13 (5'-CAGCACCCAC-3'), OPB-10 (5'-CGT-CTGGGAC-3'), OPB-18 (5'-CCACAGCAGT-3'), and OPJ-10 (5'-AAGCCCGAGG-3'), and its patterns compared with 6 representive intestinal, gram-negative bacterial strains, Vibrio parahaemolyticus, V. vulnificus, Enterobacter cloacae, Escherichia coli O157:H7, Pseudomonas aeruginosa, and Proteus sp., which are often found in foods. S. typhimurium had unique and distinct fingerprinting patterns. RAPD fingerprinting is thus concluded to be a rapid and sensitive method for the identification of S. typh-imurium compared to conventional culturing procedures or immunoassays.

Development of Molecular Biological Methods to Analyze Bacterial Species Diversity in Freshwater and Soil Ecosystems: Dong-Hun Lee , Sung-Ae Noh , Chi-Kyung Kim; J. Microbiol. 2000;38(1):11-17.

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Abstract: A new method was developed for the rapid analysis of diverse bacterial species in the natural envi-ronment. Our method is based on PCR-single-strands-conformation polymorphism (PCR-SSCP) and selective isolation technique of single-stranded DNA. Variable V3 fragments of 16S rDNA were amplified by PCR with bacterial 16S rDNA primers, where one of the primers was biotinylated at the 5'-end. The biotinylated strands of the PCR products were selectively isolated by using streptavidin paramagnetic particles and a magnetic stand, to prevent SSCP analysis producing heteroduplexes from heterogeneous DNA samples. The selected strands were separated by electrophoresis on a polyacrylamide gel, and detected by silver staining. Analysis of PCR products from 8 bacterial strains demonstrated their characteristic DNA band patterns. In addition, changes in the structure of the bacterial community and species diversity in the microcosm treated with phenol could be monitored. After 3 weeks of incubation, phenol and its intermediate, 2-hydroxy-muconic-semialdehyde, were degraded by indig-enous bacteria. These dominating bacterial populations were identified as strong bands on an SSCP gel. Therefore, this study provides useful tools for microbial community analysis of natural habitats.

Isolation of the Regulator Gene Responsible for Overproduction of Catalase A in H 2 O 2 -resistant Mutant of Streptomyces coelicolor: Ji-Sook Hahn , So-Young Oh , Keith F. Chater , Jung-Hye Roe; J. Microbiol. 2000;38(1):18-23.

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Abstract: Streptomyces coelicolor produces three kinds of catalases to cope with oxidative stress and to allow nor-mal differentiation. Catalase A is the major vegetative catalase which functions in removing hydrogen peroxide generated during the process of aerobic metabolism. To understand the regulatory mechanism of response against oxidative stress, hydrogen peroxide-resistant mutant (HR40) was isolated from S. coelicolor J1501 following UV mutagenesis. The mutant overproduced catalase A more than 50-fold compared with the wild type. The mutation locus catR1 was mapped closed to the mthB2 locus by genetic crossings. An ordered cosmid library of S. coelicolor encompassing the mthB2 locus was used to isolate the regulator gene (catR) which represses catalase overproduction when introduced into HR40. A candidate catR gene was found to encode a Fur-like protein of 138 amino acids (15319 Da).

Expression of Human Cytomegalovirus Immediate Early US3 Gene in Human Fibroblast Cells: Gyu-Cheol Lee , Chong-Kyo Lee , Jin-Hyun Ahn , Chan-Hee Lee; J. Microbiol. 2000;38(1):24-30.

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Abstract: US3 gene is a member of the human cytomegalovirus (HCMV) immediate early gene. Although the precise functions of the US3 gene in HCMV replication and pathogenesis are not known, it has been reported to play a role in inhibiting major histocompatibility class I antigen presentation. For further knowledge of US3 gene expression, rabbit polyclonal antiserum of the US3 gene product was used for indirect immunofluorescence assay. In permissive human foreskin fibroblast (HFF) cells, US3 gene expression was detectable as crescent or half-moon shape in the perinuclear region at immediate early times after virus infection. HFF cells infected with mutant HCMV lacking US3 open reading frames were negative for US3 immunofluorescence assay. Double immunofluorescence assay using monoclonal antibody to gamma adaptin (specific for the Golgi complex) and rabbit anti-US3 antiserum revealed that US3 gene product could be localized to the Golgi complex. At later time after HCMV infection, US3 gene products were detected as globular aggregates in the cytosol. These aggregates were positive for gamma adaptin and stained with preimmune serum, suggesting a nonspecific reaction to the Golgi complex. Northern blot analysis revealed that transcription of US3 was observed only during immediate early times after virus infection (until 6 h postinfection). Therefore US3 gene expression appears to be confined to immediate early time and its gene products are localized to the Golgi complex as cres-cent shaped forms in the perinuclear cytoplasm.

Selection of Suitable Packing Material for Biofiltration of Toluene, m- and p-Xylene Vapors: Young-Sook Oh , Sung-Chan Choi; J. Microbiol. 2000;38(1):31-35.

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Abstract: A suitable packing material for biofiltration of monoaromatic solvent vapors was selected among various types of packing materials such as peat, bark chips, vermiculite, and Hydroballs. A previously isolated strain, Pseudomonas pseudoalcaligenes BTXO2, which could utilize toluene, m- and p-xylene as carbon and energy sources was used as a biofilter inoculum. Four glass biofilters (6 cm dia. X 60 cm) were individually packed with each of the packing materials and solvent vapors were passed through the columns. During three weeks of peat biofilter operation, average removal efficiencies of toluene, m-and p-xylene were 90.4%, 95.3%, and 82.1%, respectively. With the other packings, the efficiencies were in the range of 10.1 to 58.6% which were significantly lower than those of the peat biofilter. The peat biofilter was continually operated for approximately nine months and the biofilter sustained its degradation activity during the operation period with minimal maintenance. At steady state, average removal rates of toluene, m- and p-xylene vapors were estimated as 14.2, 5.5, and 8.1 g m^-3 packing h^-1, respectively.

Reduction of Hexavalent Chromium by Escherichia coli ATCC 33456 in Batch and Continuous Cultures: Woo Chul Bae , Tae Gu Kang , In Kyong Kang , You Jung Won , Byeong Chul Jeong; J. Microbiol. 2000;38(1):36-39.

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Abstract: Toxic hexavalent chromium, Cr(VI), was reduced to a less toxic trivalent chromium form by E. coli ATCC 33456. The suitable electron donor for Cr(VI) reduction was glucose. E. coli ATCC 33456 was more resistant to metal cations than other reported Cr(VI) reducing microorganisms. Cell growth was inhibited by the presence of Cr(VI) in a liquid medium and Cr(VI) reduction accompanied cell growth. With a hydraulic retention time of 20 h, Cr(VI) reducing efficiency was 100% to 84% when Cr(VI) concentration in the influent was in the range of 10 to 40 mg L^-1. Specific rate of Cr(VI) concentration in the influent was 2.41 mg Cr(VI) g DCW^-1 h^-1 when 40 mg :^-1 of Cr(VI) influent was used. This result suggested the potential application of E. coli ATCC 33456 for the detoxification of Cr(VI) in Cr(VI) contaminated wastewater.

Association of a Common Reductase with Multiple Aromatic Terminal Dioxygenases in Sphingomonas yanoikuyae Strain B1: Mihyun Bae , Eungbin Kim; J. Microbiol. 2000;38(1):40-43.

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Abstract: The aromatic dioxygenase system in Sphingomonas yanoikuyae strain B1 consists of three components, an oxygenase, a ferredoxin, and a reductase. The insertional knockout of the bphA4 gene encoding a reductase and subsequent complementation experiments showed that the reductase encoded by bphA4 in S. yanoikuyae strain B1 is associated with multiple dioxygenase components including that of toluate dioxygenase (XylXY).

Genetic Analysis of the VP2/NS Junction Region on Segment A of Marine Birnavirus Isolated from Cultured Flounder Paralichthys olivaceous: Seong-Joon Joh , Jeong-Ho Kim , Gang-Joon Heo; J. Microbiol. 2000;38(1):44-47.

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Abstract: The cDNA of VP2/NS junction region on segment A of the marine birnavirus (MBV) isolated from flounder (DS strain) was amplified using the reverse transcription (RT)-polymerase chain reaction (PCR). Its cDNA nucleotide and deduced amino acid sequences were analyzed, and compared with the reference strains of MBV and infectious pancreatic necrosis virus (IPNV). Analyses of the nucleotide and deduced amino acid sequences revealed that the DS strain is very similar to the reference strains of MBV, distant from the IPNV.

Monascus Red Pigment Overproduction by Coculture with Recombinant Saccharomyces cerevisiae Secreting Glucoamylase: Ho-Soo Lim , Seung-Ku Yoo , Chul-Soo Shin , Young-Min Hyun; J. Microbiol. 2000;38(1):48-51.

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Abstract: In liquid cultures using sucrose media, the coculture of Monascus with recombinant Saccharomyces cerevisiae expressing the glucoamylase gene from Aspergillus niger enhanced red pigment production by approx. 19%, compared with the coculture of wild type S. cerevisiae. Coculture with recombinant S. cerevisiae was more effective than with wild type S. cerevisiae for Monascus red pigment production. Cocultures of Monascus with commercial amylases of Aspergillus also induced high production of pigment and morphological changes in a solid culture using sucrose media.

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