• Journal of Microbiology
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• v.34 no.1
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• pp.61-67
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• 1996

Double-stranded RNA (dsRNA) viruses and single-stranded RNA(ssRNA) viruses were detected in a strain of Pleurotus mushroom cultivated in a farm. Those fungal virsus were purified in the pH 6.0 or pH 7.2 using CsCI or Cs$_{2}$SO$_{4}$ buoyant density centrifugation. Each viral particles were not completely separated at any trials. However, mushroom bacili-form virus contains a single major nucleic acid with 0.7 Kb ssRNA, which might code for 20 Kd viral capsid protein. The dsRNAs are encapsidatred into spherical-form viruses, whereas ssRNA viral genomes are encapsidated into two different sizes of bacili-form particles. A healthy-looking mushroom also contained some spherical-form viruses with dsRNAs. Laboratory strains of Pleurotus ostreatus and a cultivated strain of P. sajor-caju did not show any viral particles. Mushrooms with specific disease symptoms. however, contained at least four different sizes of spherical-form viruses. Thus, we concluded that a bacilli-form virus case a severe disease symptoms of adnormal on mushroom development.

• Proceedings of the Korean Society for Food Science of Animal Resources Conference
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• 2004.05a
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• pp.353-356
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• 2004

Rotavirus and Norovirus are major causative agents of acute diarrhea and gastroenteritis. In our study, Each viral RNA was isolated from the feces of patients for viral diarrhea in Korea, respectively. And cDNA library were constructed using RT-PCR. Also, cDNAs encoding VP8 derived from Rotavirus and Capsid protein derived from norovirus were subesequently cloned and expressed in Echerichia coli as a fusion antigen. Molecular weight of fusion antigen was approximately 60kDa. Also, substantial overexpression was accomplished. We yielded egg yolk lgY which is potentially useful in controlling of Rotavirus and Norovirus which are one of the most prevalent pathogenic viruses.

• Journal of Microbiology
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• v.44 no.2
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• pp.248-253
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• 2006

Lymphocystis disease virus (LCDV) is the causative agent of lymphocystis disease. The viruses have been divided into three genotypes (genotype I for LCDV-1, II for Japanese flounder isolates, and III for rockfish isolates) on the basis of major capsid protein (MCP) gene sequences. In this study, we developed a multiplex PCR primer set in order to distinguish these genotypes. We also analyzed the MCP gene of a new LCDV isolate from the sea bass (SB98Yosu). Comparison of sequence identities between SB98Yosu and eight Japanese flounder isolates, revealed identity of more than 90.1 % at nucleotide level and 96.5% at deduced amino acid level, respectively. Phylogenetic analyses based on the MCP gene showed that SB98Yosu belongs to genotype II, along with Japanese flounder isolates. Multiplex PCR based on the MCP gene allowed us to identify these genotypes in a simple and rapid manner, even in a sample that contained two genotypes, in this case genotypes II and III.

• Journal of fish pathology
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• v.23 no.3
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• pp.273-280
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• 2010

To determine whether rock bream Oplegnathus fasciatus can be protected from rock bream iridovirus (RBIV) infection by intramuscular injection of long double-stranded RNAs (dsRNAs), we compared protective effect of virus-specific dsRNAs corresponding to major capsid protein (MCP), ORF 084, ORF 086 genes, and virus non-specific green fluorescent protein (GFP) gene. Furthermore, to determine whether the non-specific type I interferon (IFN) response was associated with protective effect, we estimated the activation of type I IFN response in fish using expression level of IFN inducible Mx gene as a marker. As a result, mortality of fish injected with dsRNAs and challenged with RBIV was delayed for a few days when comparing with PBS injected control group. However, virus-specific dsRNA injected groups exhibited no significant differences in survival period when compared to the GFP dsRNA injected group. Semi-quantitative analysis indicated that the degree of antiviral response via type I IFN response is supposedly equal among dsRNA injected fish. These results suggest that type I IFN response rather than sequence-specific RNA interference might involve in the lengthened survival period of fish injected with virus-specific dsRNAs.

• The Plant Pathology Journal
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• v.21 no.4
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• pp.334-342
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• 2005

The prototype Chlorella virus PBCV-l encodes 11 tRNA genes and over 350 protein-encoding genes in its 330 kbp genome. Initial attempts to overexpress the recombinant A189/192R protein, a putative virus attachment protein, in E. coli strain BL21(DE3) SI were unsuccessful, and multiple protein bands were detected on Western blots. However, the full-length A189/192R recombinant protein or fragments derived from it were detected when they were expressed in E. coli BL21 CodonPlus (DE3) RIL, which contains extra tRNAs. Codon usage analysis of the a189/192r gene showed highly biased usage of the AGA and AVA codons compared to genes encoded by E. coli and Chlorella. In addition, there were biases of XXA/U($56\%$) and XXG/ C($44\%$) in the codons recognized by the viral tRNAs, which correspond to the codon usage bias in the PBCV-1 genome of XXA/U ($63\%$) over those ending in XXC/G ($37\%$). Analysis of the codon usage in the major capsid protein and DNA polymerase showed preferential usage of codons that can be recognized by the viral tRNAs. The Asn (AAC) and Lys (AAG) codons whose corresponding tRNA genes are duplicated in the tRNA gene cluster were the most abundant (i.e., preferred) codons in these two proteins. The tRNA genes encoded in the PBCV-l genome seem to play a very important role during the synthesis of viral proteins through supplementing the tRNAs that are frequently used in viral proteins, but are rare in the host cells. In addition, these tRNAs would help the virus to adapt to a wide range of hosts by providing tRNAs that are rare in the host cells.

• Journal of Microbiology and Biotechnology
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• v.22 no.5
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• pp.721-728
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• 2012

The human papillomavirus (HPV) is the main cause of cervical cancer in developing countries. Rapid diagnosis and initiation of treatment of the HPV infection are critical. Various methods have been employed to reduce the immunogenicity of antibodies targeting HPV serotypes. Nanobodies are the smallest fragments of naturally occurring single-domain antibodies with their antigen-binding site compromised into a single domain. Nanobodies have remarkable properties such as high stability, solubility, and high homology to the human VH3 domain. In this study, a phagemid library was employed to enrich for nanobodies against the L1 protein of the human papilloma virus. Binding reactivity of the selected clones was evaluated using phage enzyme-linked immunosorbent assay (phage-ELISA). Finally, two nanobodies (sm5 and sm8) with the best reactivity against the Gardasil vaccine and the purified HPV-16 L1 protein were expressed and purified using a $Ni^+$-NTA column. The accuracy of expression and purification of the nanobodies was confirmed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting assays. In vitro studies demonstrated that neutralization was achieved by the selected nanobodies. The ease of generation and unique features of these molecules make nanobodies promising molecules for the new generation of HPV diagnosis and therapy.

• Journal of Microbiology and Biotechnology
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• v.30 no.1
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• pp.38-43
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• 2020

Hand, foot, and mouth disease (HFMD) is caused by enterovirus 71 (EV71) in infants and children under six years of age. HFMD is characterized by fever, mouth ulcers, and vesicular rashes on the palms and feet. EV71 also causes severe neurological manifestations, such as brainstem encephalitis and aseptic meningitis. Recently, frequent outbreaks of EV71 have occurred in the Asia-Pacific region, but currently, no effective antiviral drugs have been developed to treat the disease. In this study, we investigated the antiviral effect of salvianolic acid B (SalB) on EV71. SalB is a major component of the Salvia miltiorrhiza root and has been shown to be an effective treatment for subarachnoid hemorrhages and myocardial infarctions. HeLa cells were cultured in 12-well plates and treated with SalB (100 or 10 ㎍/ml) and 10⁶ PFU/ml of EV71. SalB treatment (100 ㎍/ml) significantly decreased the cleavage of the eukaryotic eIF4G1 protein and reduced the expression of the EV71 capsid protein VP1. In addition, SalB treatment showed a dramatic decrease in viral infection, measured by immunofluorescence staining. The Akt signaling pathway, a key component of cell survival and proliferation, was significantly increased in EV71-infected HeLa cells treated with 100 ㎍/ml SalB. RT-PCR results showed that the mRNA for anti-apoptotic protein Bcl-2 and the cell cycle regulator Cyclin-D1 were significantly increased by SalB treatment. These results indicate that SalB activates Akt/PKB signaling and inhibits apoptosis in infected HeLa cells. Taken together, these results suggest that SalB could be used to develop a new therapeutic drug for EV71-induced HFMD.

• Journal of Plant Biotechnology
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• v.31 no.4
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• pp.285-293
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• 2004

FMDV is a viral pathogen that caused foot-and-mouth disease in animals. VP1 is a major capsid protein of FMDV. It is known as one of best materials for the FMDV diagnosis and for the development of protein vaccine. In this study, 633 bp of VP1 gene was modified for the expression of VP1 in plant, based on the VP1 DNA sequence from FMDV taiwan O type and from FMDV isolated vietnam. The. deduced DNA fragment was artificially synthesized using the multiple fragment extension with long-nucleotides. A new plant transgenic vector system, pCAMBIA139011 was constructed on the basis of pBI12l and pCAMBIA1390. Using this vector system and GFP gene or modified VP1 gene, each target gene was introduced into Nicotiana tabacum. The insertion of whole target gene was successfully confirmed in each transgenic plant named GFP-A7 and VP1-4, respectively. The expression level of each gene was estimated by RT-PCR and Real-Time PCR using VP1, GFP specific primers.

• The Plant Pathology Journal
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• v.34 no.1
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• pp.59-64
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• 2018

Bacteriophages of Acidovorax citrulli, the causal agent of bacterial fruit blotch, were isolated from 39 watermelon, pumpkin, and cucumber leaf samples collected from various regions of Korea and tested against 18 A. citrulli strains. Among the six phages isolated, ACP17 forms the largest plaque, and exhibits the morphology of phages in the Myoviridae family with a head diameter of $100{\pm}5nm$ and tail length of $150{\pm}5nm$. ACP17 has eclipse and latent periods of $25{\pm}5min$ and $50{\pm}5min$, respectively, and a burst size of 120. The genome of ACP17 is 156,281 base pairs with a G + C content of 58.7%, 263 open reading frames, and 4 transfer RNA genes. Blast search and phylogenetic analysis of the major capsid protein showed that ACP17 has limited homology to two Stentrophomonas phages, suggesting that ACP17 is a new type of Myoviridae isolated from A. citrulli.

• The Plant Pathology Journal
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• v.32 no.6
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• pp.584-588
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• 2016

Several Bacillus species were isolated from rice field soils, and 16S rRNA gene sequence analysis showed that Bacillus cereus was the most abundant. A strain named BC1 showed antifungal activity against Rhizoctonia solani. Bacteriophages infecting strain BC1 were isolated from the same soil sample. The isolated phage PK16 had an icosahedral head of $100{\pm}5nm$ and tail of $200{\pm}5nm$, indicating that it belonged to the family Myoviridae. Analysis of the complete linear dsDNA genome revealed a 158,127-bp genome with G + C content of 39.9% comprising 235 open reading frames as well as 19 tRNA genes (including 1 pseudogene). Blastp analysis showed that the proteins encoded by the PK16 genome had the closest hits to proteins of seven different bacteriophages. A neighbor-joining phylogenetic tree based on the major capsid protein showed a robust clustering of phage PK16 with phage JBP901 and BCP8-2 isolated from Korean fermented food.