• Journal of Microbiology and Biotechnology
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• v.16 no.10
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• pp.1518-1522
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• 2006

In the present study, it was observed how the phage-host system that is naturally reproduced in activated sludge is affected by the host inoculation. The system of Microlunatus phosphovorus and its phages was selected as the phage-host system native to an activated sludge system operated for 19 days under sequencing anaerobic-aerobic conditions with glutamate as the main carbon source. The phage-host system related to M. phosphovorus was monitored by plaque assay for the phages and by fluorescent in situ hybridization (FISH) for the bacterial host. In addition, the whole phage structure was also monitored by pulsed-field gel electrophoresis (PFGE). During the first 9 days, the phage-host system was more or less steady at approx. 9% (FISH/ DAPI) for M. phosphovorus and approx. 10,000 PFU/ml for its lytic phages. Microlunatus phosphovorus JCM9379 was inoculated into the activated sludge on day 10. Right after the inoculation, M. phosphovorus was approx. 24% (FISH/DAPI) whereas its lytic phages dropped down to approx. 500 PFU/ ml. After the host inoculation (within 9 days), however, the phage-host system eventually reverted to its original level in each population. On the other hand, the whole phage structure was not significantly changed by M. phosphovorus inoculation but stable throughout the process operation. Only the minor change that four phage groups gradually became abundant after the host inoculation was observed.

• Applied Microscopy
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• v.12 no.1
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• pp.33-40
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• 1982

Several phages of Bacillus thuringiensis distributed in Korea were isolated. The distribution and morphological characteristics of phages were studied. The results are as follows; 1. The isolated phages were highly specific for Bacillus thuringiensis var. thuringiensis. They were classified as YM series phages and designated as phage YM-1, YM-2 and YM-3 according to their morphological characteristics. 2. Most of these YM series phages were isolated from compost including domestic animal dung and soil under sewage. 3. The YM-1 phage was similar to Bacillus subtilis ${\phi}25$ in morphology. It has 94nm x 86nm head, contractile tail sheath and base plate with four cornered structure. 4. The YM-2 phage was similar to Bacillus subtilis GA-1 phage in morphology. It had 70nm x 56nm head and tail without contractile tail sheath. 5. The YM-3 phage was similar to Bacillus subtilis ${\phi}29$ phage. It had 56nm x 43nm head and tail with distal enlargement.

• Journal of Microbiology and Biotechnology
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• v.22 no.5
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• pp.649-653
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• 2012

A temperate phage was isolated from emetic Bacillus cereus NCTC 11143 by mitomycin C and characterized by transmission electron microscopy and DNA and protein analyses. Whole genome sequencing of Bacillus phage 11143 was performed by GS-FLX. The phage has a dsDNA genome of 39,077 bp and a 35% G+C content. Bioinformatic analysis of the phage genome revealed 49 putative ORFs involved in replication, morphogenesis, DNA packaging, lysogeny, and host lysis. Bacillus phage 11143 could be classified as a member of the Siphoviridae family by morphology and genome structure. Genomic comparisons at the DNA and protein levels revealed homologous genetic modules with patterns and morphogenesis proteins similar to those of other Bacillus phages. Thus, Bacillus phages might have a mosaic genetic relationship.

• Journal of Microbiology and Biotechnology
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• v.22 no.12
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• pp.1613-1620
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• 2012

Bacterial wilt caused by Ralstonia solanacearum is a devastating disease of many economically important crops. Since there is no promising control strategy for bacterial wilt, phage therapy could be adopted using virulent phages. We used phage PE204 as a model lytic bacteriophage to investigate its biocontrol potential for bacterial wilt on tomato plants. The phage PE204 has a short-tailed icosahedral structure and double-stranded DNA genome similar to that of the members of Podoviridae. PE204 is stable under a wide range of temperature and pH, and is also stable in the presence of the surfactant Silwet L-77. An artificial soil microcosm (ASM) to study phage stability in soil was adopted to investigate phage viability under a controlled system. Whereas phage showed less stability under elevated temperature in the ASM, the presence of host bacteria helped to maintain a stable phage population. Simultaneous treatment of phage PE204 at $10^8$ PFU/ml with R. solanacearum on tomato rhizosphere completely inhibited bacterial wilt occurrence, and amendment of Silwet L-77 at 0.1% to the phage suspension did not impair the disease control activity of PE204. The biocontrol activities of phage PE204 application onto tomato rhizosphere before or after R. solanacearum inoculation were also investigated. Whereas pretreatment with the phage was not effective in the control of bacterial wilt, post-treatment of PE204 delayed bacterial wilt development. Our results suggested that appropriate application of lytic phages to the plant root system with a surfactant such as Silwet L-77 could be used to control the bacterial wilt of crops.

• Applied Microscopy
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• v.10 no.1_2
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• pp.19-25
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• 1980

E. coli B was treated with colistin and kanamycin and the influence of these antibiotics on the absorption of $T_3$ phage was studied using the plaque counting method and the electron microscope. E. coli B treated with colistin was sharply inhibited on phage absorption and cell walls were severely damaged showing some spiny appearance around the walls. No influence of kanamycin was noted on phage absorption. Bacterial cells treated with kanamycin showed wave form in the structure of walls and a profound change was noted in the cytoplasm where it was concentrated along the periphery of the inner wall leaving the center of the cell to appear almost empty.

• Journal of Microbiology
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• v.37 no.2
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• pp.97-101
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• 1999

Random hexapeptide library on the surface of filamentous bacteriophage was constructed using the SurfZAP vector. The size of the library was approximately 105. The peptide insert was flanked by two cysteines to constrain the peptide structure with a disulfide bond. This library was screened for the topoisomerase II binding peptide. Dramatic enrichment of the fusion phage over the VCS M13 helper phage was demonstrated by bio-panning affinity selection.

• BMB Reports
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• v.40 no.5
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• pp.740-748
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• 2007

To gain insight into the structure and function of repressor proteins of bacteriophages of gram-positive bacteria, repressor of temperate Staphylococcus aureus phage ${\phi}11$ was undertaken as a model system here and purified as an N-terminal histidine-tagged variant (His-CI) by affinity chromatography. A ~19 kDa protein copurified with intact His-CI (~ 30 kDa) at low level was resulted most possibly due to partial cleavage at its Ala-Gly site. At ~10 nM and higher concentrations, His-CI forms significant amount of dimers in solution. There are two repressor binding sites in ${\phi}11$ cI-cro intergenic region and binding to two sites occurs possibly by a cooperative manner. Two sites dissected by HincII digestion were designated operators $O_L$ and $O_R$, respectively. Equilibrium binding studies indicate that His-CI binds to $O_R$ with a little more strongly than $O_L$ and binding species is probably dimeric in nature. Interestingly His-CI binding affinity reduces drastically at elevated temperatures ($32-42^{\circ}C$). Both $O_L$ and $O_R$ harbor a nearly identical inverted repeat and studies show that ${\phi}11$ repressor binds to each repeat efficiently. Additional analyses indicate that ${\phi}11$ repressor, like $\lambda$ repressor, harbors an N-terminal domain and a C-terminal domain which are separated by a hinge region. Secondary structure of ${\phi}11$ CI even nearly resembles to that of $\lambda$ phage repressor though they differ at sequence level. The putative N-terminal HTH (helix-turn-helix) motif of ${\phi}11$ repressor belongs to the HTH -XRE-family of proteins and shows significant identity to the HTH motifs of some proteins of evolutionary distant organisms but not to HTH motifs of most S. aureus phage repressors.

• Animal cells and systems
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• v.4 no.2
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• pp.141-144
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• 2000

Effects of deamido-$\textrm{NAD}^{+}$on self-splicing of primary transcripts of the phage T4 thymidylate synthase gene (td) was investigated. The self-splicing was not affected by deamido-$\textrm{NAD}^{+}$- at concentrations up to 2 mM. However, it began to decrease at 5 mM and the formation of splicing products such as the linear intron, intron-exon 2 and exon 1-exon 2, was slightly reduced. At 20 mM the self-splicing activity was almost completely abolished. This analog of the coenzyme $\textrm{NAD}^{+}$- inhibits the self-splicing of td intron RNA although it does not possess a guanidine group in its structure. The analysis of inhibitory concentrations and structural examination suggests that the key structural features of deamido-$\textrm{NAD}^{+}$ responsible for the inhibition of splicing may be the ADP-ribose moiety.

• Korean Journal of Microbiology
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• v.33 no.4
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• pp.242-246
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• 1997

The protein profile of Synechococcus sp. cyanophage was investigated employing SDS-PAGE. The phage appears to be composed of two major proteins of 97 and 52 kDa and at least seven minor proteins of 70, 65, 60, 40, 35, 28, and 6 kDa. It seems that each subunit is combined to form a multimer although any disulfide bond does not exist in the phage structure. Lytic activity of the phage particle against cell wall was detected around the 52 kDa on renaturing SDS-PAGE using heat-killed Micrococcus luteus cells as substrate. The activity has the optimal pH between 9 and 10, and slightly inhibited by EDTA.

• Journal of Microbiology and Biotechnology
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• v.21 no.5
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• pp.529-535
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• 2011

Single-chain variable fragment (scFv) is a fusion protein of the variable regions of the heavy (VH) and light (VL) chains of immunoglobulin, connected with a short linker peptide of 10 to about 20 amino acids. In this study, the scFv of a monoclonal antibody against the third domain of human CD4 was cloned from OKT4 hybridoma cells using the phage display technique and produced in E. coli. The expression, production, and purification of anti-CD4 scFv were tested using SDS-PAGE and Western blot, and the specificity of anti-CD4 scFv was examined using ELISA. A 31 kDa recombinant anti-CD4 scFv was expressed and produced in bacteria, which was confirmed by SDS-PAGE and Western blot assays. Sequence analysis proved the ScFv structure of the construct. It was able to bind to CD4 in quality ELISA assay. The canonical structure of anti-CD4 scFv antibody was obtained using the SWISS_MODEL bioinformatics tool for comparing with the scFv general structure. To the best of our knowledge, this is the first report for generating scFv against human CD4 antigen. Engineered anti-CD4 scFv could be used in immunological studies, including fluorochrome conjugation, bispecific antibody production, bifunctional protein synthesis, and other genetic engineering manipulations. Since the binding site of our product is domain 3 (D3) of the CD4 molecule and different from the CD4 immunological main domain, including D1 and D2, further studies are needed to evaluate the anti-CD4 scFv potential for diagnostic and therapeutic applications.