• Journal of fish pathology
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• v.23 no.2
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• pp.177-187
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• 2010

The Interferon stimulated gene 15 (ISG15) is strongly induced in many cell types by IFNs, viral infections, and double-stranded RNA (poly I:C). The ISG15 homologue cDNA was isolated from the rock bream LPS stimulated leukocyte cDNA library. The rock bream ISG15 homologue was found to consist of 833 bp encoding 157 amino acid residues. Compared with other known ISG15 peptide sequences, the most conserved regions of the rock bream ISG15 peptide were found to be the tandem ubiquitin-like domains and a C-terminal LRLRGG conjugating motif, characteristic of mammalian and non-mammalian ISG15 proteins. Phylogenetic analysis based on the deduced amino acid sequence revealed a homologous relationship between the ISG15 sequence of rock bream and that of Atlantic salmon, Atlantic cod, northern snake head, black rockfish and olive flounder. The expression of the rock bream ISG15 molecule was induced in the peripheral blood leukocytes (PBLs) from 1 to 24 h following poly I:C stimulation, with a peak at 3 h post-stimulation. The rock bream ISG15 gene was predominantly expressed in the PBLs, spleen and gill.

• The Plant Pathology Journal
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• v.37 no.2
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• pp.162-172
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• 2021

Soybean mosaic virus (SMV) is the predominant viral pathogen that affects the yield and quality of soybean. The natural host range for SMV is very narrow, and generally limited to Leguminosae. However, we found that SMV can naturally infect Pinellia ternata and Atractylodes macrocephala. In order to clarify the molecular mechanisms underlying the cross-family infection of SMV, we used double-stranded RNA extraction, rapid amplification of cDNA ends polymerase chain reaction and Gibson assembly techniques to carry out SMV full-length genome amplification from susceptible soybeans and constructed an infectious cDNA clone for SMV. The genome of the SMV Shanxi isolate (SMV-SX) consists of 9,587 nt and encodes a polyprotein consisting of 3,067 aa. SMV-SX and SMV-XFQ008 had the highest nucleotide and amino acid sequence identities of 97.03% and 98.50%, respectively. A phylogenetic tree indicated that SMV-SX and SMV-XFQ018 were clustered together, sharing the closest relationship. We then constructed a pSMV-SX infectious cDNA clone by Gibson assembly technology and used this clone to inoculate soybean and Ailanthus altissima; the symptoms of these hosts were similar to those caused by the virus isolated from natural infected plant tissue. This method of construction not only makes up for the time-consuming and laborious defect of traditional methods used to construct infectious cDNA clones, but also avoids the toxicity of the Potyvirus special sequence to Escherichia coli, thus providing a useful cloning strategy for the construction of infectious cDNA clones for other viruses and laying down a foundation for the further investigation of SMV cross-family infection mechanisms.

• Genomics & Informatics
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• v.20 no.1
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• pp.11.1-11.20
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• 2022

Vibrio harveyi belongs to the Vibrio genus that causes vibriosis in marine and aquatic fish species through double-stranded DNA virus replication. In humans, around 12 Vibrio species can cause gastroenteritis (gastrointestinal illness). A large amount of virus particles can be found in the cytoplasm of infected cells, which may cause death. Despite these devastating complications, there is still no cure or vaccine for the virus. As a result, we used an immunoinformatics approach to develop a multi-epitope vaccine against most pathogenic hemolysin gene of V. harveyi. The immunodominant T- and B-cell epitopes were identified using the hemolysin protein. We developed a vaccine employing three possible epitopes: cytotoxic T-lymphocytes, helper T-lymphocytes, and linear B-lymphocyte epitopes, after thorough testing. The vaccine was developed to be antigenic, immunogenic, and non-allergenic, as well as having a better solubility. Molecular dynamics simulation revealed significant structural stiffness and binding stability. In addition, the immunological simulation generated by computer revealed that the vaccination might elicit immune reactions in the actual life after injection. Finally, using Escherichia coli K12 as a model, codon optimization yielded ideal GC content and a higher codon adaptation index value, which was then included in the cloning vector pET2+ (a). Altogether, our experiment implies that the proposed peptide vaccine might be a good option for vibriosis prophylaxis.

• The Plant Pathology Journal
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• v.38 no.6
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• pp.646-655
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• 2022

Pepper mild mottle virus (PMMoV), one of the most prevalent viruses in chili pepper (Capsicum annuum L.) is a non-enveloped, rod-shaped, single-stranded positive-sense RNA virus classified in the genus Tobamovirus. The supernatants of five bacterial cultures (Pseudomonas putida [PP], Bacillus licheniformis [BLI], P. fluorescens [PF], Serratia marcescens [SER], and B. amyloliquifaciens [BA]) were analyzed to find novel antiviral agents to PMMoV in chili pepper. Foliar spraying with supernatants (1:1, v/v) obtained from Luria-Bertani broth cultures of PP, BLI, PF, SER, and BA inhibited PMMoV infection of chili pepper if applied before the PMMoV inoculation. Double-antibody sandwich enzyme-linked immunosorbent assay showed that treatments of five supernatants resulted in 51-66% reductions in PMMoV accumulation in the treated chili pepper. To identify key compounds in supernatants of PP, BLI, PF, SER, and BA, the supernatants were subjected to gas chromatography-mass spectrometry. The 24 different types of compounds were identified from the supernatants of PP, BLI, PF, SER, and BA. The compounds vary from supernatants of one bacterial culture to another which includes simple compounds-alkanes, ketones, alcohols, and an aromatic ring containing compounds. The compounds triggered the inhibitory effect on PMMoV propagation in chili pepper plants. In conclusion, the cultures could be used to further conduct tissue culture and field trial experiments as potential bio-control agents.

• Biomolecules & Therapeutics
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• v.30 no.2
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• pp.170-178
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• 2022

The airway epithelium is equipped with the ability to resist respiratory disease development and airway damage, including the migration of airway epithelial cells and the activation of TLR3, which recognizes double-stranded (ds) RNA. Primary cilia on airway epithelial cells are involved in the cell cycle and cell differentiation and repair. In this study, we used Beas-2B human bronchial epithelial cells to investigate the effects of the TLR3 agonist polyinosinic:polycytidylic acid [Poly(I:C)] on airway cell migration and primary cilia (PC) formation. PC formation increased in cells incubated under serum deprivation. Migration was faster in Beas-2B cells pretreated with Poly(I:C) than in control cells, as judged by a wound healing assay, single-cell path tracking, and a Transwell migration assay. No changes in cell migration were observed when the cells were incubated in conditioned medium from Poly(I:C)-treated cells. PC formation was enhanced by Poly(I:C) treatment, but was reduced when the cells were exposed to the ciliogenesis inhibitor ciliobrevin A (CilioA). The inhibition of Beas-2B cell migration by CilioA was also assessed and a slight decrease in ciliogenesis was detected in SARS-CoV-2 spike protein (SP)-treated Beas-2B cells overexpressing ACE2 compared to control cells. Cell migration was decreased by SP but restored by Poly(I:C) treatment. Taken together, our results demonstrate that impaired migration by SP-treated cells can be attenuated by Poly(I:C) treatment, thus increasing airway cell migration through the regulation of ciliogenesis.

• Molecules and Cells
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• v.41 no.4
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• pp.271-281
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• 2018

IFIT1 (also known as ISG56) is a member of the interferon-inducible protein with tetratricopeptide repeats (IFITs) family. IFITs are strongly induced by type I interferon (IFN), double-stranded RNA and virus infection. Here, we investigated IFIT1 expression in human umbilical vein endothelial cells (HUVECs) and in human bronchus epithelial cells (BEAS-2Bs) induced by the H9N2 virus and inactivated viral particle at different time points. We also investigated the effect of H9N2 virus and viral particle infection on $IFN-{\alpha}/{\beta}$ production, and assessed whether hemagglutinin or neuraminidase protein induced IFIT1 expression. Results showed that both H9N2 virus infection and viral particle inoculation induced the expression of IFIT1 at mRNA and protein levels in the two cell lines. Hemagglutinin or neuraminidase protein binding alone is not sufficient to induce IFIT1 expression. Surprisingly, the expression patterns of IFIT1 in response to H9N2 virus and viral particles in the two cell lines were opposite, and production kinetics of $IFN-{\alpha}/{\beta}$ also differed. An additional finding was that induction of IFIT1 in response to H9N2 virus infection or viral particle inoculation was more sensitive in HUVECs than in BEAS-2Bs. Our data offers new insight into the innate immune response of endothelial cells to H9N2 virus infection.

• Journal of Microbiology and Biotechnology
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• v.25 no.1
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• pp.98-108
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• 2015

Staphylococcus aureus is an important foodborne pathogen that causes diverse diseases ranging from minor infections to life-threatening conditions in humans and animals. To further understand its pathogenesis, the genome of the strain S. aureus FORC_001 was isolated from a contaminated food. Its genome consists of 2,886,017 bp double-stranded DNA with a GC content of 32.8%. It is predicted to contain 2,728 open reading frames, 57 tRNAs, and 6 rRNA operons, including 1 additional 5S rRNA gene. Comparative phylogenetic tree analysis of 40 complete S. aureus genome sequences using average nucleotide identity (ANI) revealed that strain FORC_001 belonged to Group I. The closest phylogenetic match was S. aureus MRSA252, according to a whole-genome ANI (99.87%), suggesting that they might share a common ancestor. Comparative genome analysis of FORC_001 and MRSA252 revealed two non-homologous regions: Regions I and II. The presence of various antibiotic resistance genes, including the SCCmec cluster in Region I of MRSA252, suggests that this strain might have acquired the SCCmec cluster to adapt to specific environments containing methicillin. Region II of both genomes contains prophage regions but their DNA sequence identity is very low, suggesting that the prophages might differ. This is the first report of the complete genome sequence of S. aureus isolated from a real foodborne outbreak in South Korea. This report would be helpful to extend our understanding about the genome, general characteristics, and virulence factors of S. aureus for further studies of pathogenesis, rapid detection, and epidemiological investigation in foodborne outbreak.

• Applied Biological Chemistry
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• v.37 no.3
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• pp.148-153
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• 1994

Rice black-streaked dwarf virus (RBSDV), a member of the plant reoviridae fijivirus group, causes a serious damage for rice production in Korea. To characterize the RBSDV genome, virus particles were produced by feeding of planthopper (Laodelphax striatellus F.) carring RBSDV to maize plants for 2 days. In $30{\sim}40$ days after feeding, the viral particles were purified from the infected maize roots by using $10{\sim}40%$ sucrose gradient centrifugation. After treatment of 10% SDS to remove the viral coat proteins, ten viral double-stranded RNAs were resolved in agrose gel electrophoresis. Total dsRNA was then used to synthesize cDNA by reverse transcriptase and a cDNA library was constructed in the ${\lambda}gt11$ vector. The phages that contain RBSDV cDNA fragments were selected by hybridizing with the random-primed probe prepared from RBSDV dsRNAs. After subcloning of several cDNA fragments into the pUC19 plasmid vector, one clone (pRV3) was chosen for sequencing. The pRV3 clone was shown to be located on the RBSDV genome fragment No.3 by RNA gel-blot analysis. Sequence analysis of the clone revealed that the pRV3 contains two partial open reading frames.

• Proceedings of the Korean Society of Plant Pathology Conference
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• 1995.06b
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• pp.77-96
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• 1995

To obtain basic informations for the improvement of seed potato production in Korea, some etiological properties of potato virus Y(PVY) distributed in the major seed potato production area(Daekwanryeong) were characterized, and the nucleotide and amino acid sequences of the coat protein gene of the PVY strains isolated were analyzed. PVY strains in Daekwonryeong, an alpine area, were identified to be two strains, PVYo and PVYN by symptoms of indicator plants, and their distribution in potato fields was similar. Major symptom on potato varieties by PVY was grouped as either mosaic alone or mosaic accompanied with veinal necrosis in the lower leaves. The symptom occurrence of the two symptoms was similar with Irish Cobbler, but Superior showed a higher rate of mosaic symptom than the other. The PVY strain which was isolated from potato cv. Superior showing typical mosaic symptoms produced symptoms of PVY-O on the indicator plants of Chenopodium amaranticolor, Nicotiana tabacum cv. Xanthi nc and Physalis floridana, but no symptom o Capsicum annum cv. Ace. Moreover, results from the enzyme-linked immunosorbent assay with monoclonal and polyclonal antibodies showed that the isolated PVY reacts strongly with PYV-O antibodies but does not react specifically with PVY-T antibodies. The purified virus particles were flexious with a size of 730$\times$11nm. On the basis of the above characteristics, the strain was identified to be a PVY-O and named as of PVY-K strain. The flight of vector aphids was observed in late May, however, the first occurrence of infected plants was in mid June with the bait plants surrounded with PVY-infected potato plants and early July with the bait plants surrounded with PVY-free potato plants. PVY infection rates by counting symptoms on bait plants (White Burley) were 1.1% with the field surrounded with PVY-free potato plants and 13.7% the fields surrounded with PVY-infected potato plants, showing the effect of infection pressure. The propagated PVY-K strain on tobacco(N. sylvestris) was purified, and the RNA of the virus was extracted by the method of phenol extraction. The size of PVY-K RNA was measured to be 9, 500 nucleotides on agarose gel electrophoresis. The double-stranded cDNAs of PVY-K coat protein(CP) gene derived by the method of polymerase chain reaction were transformed into the competent cells of E. coli JM 109, and 2 clones(pYK6 and pYK17) among 11 clones were confirmed to contain the full-length cDNA. Purified plasmids from pYK17 were cut with Sph I and Xba I were deleted with exonuclease III and were used for sequencing analysis. The PVY-K CP gene was comprised of 801 nucleotides when counted from the clevage site of CAG(Gln)-GCA(Ala) to the stop codon of TGA and encoded 267 amino acids. The molecular weight of the encoded polypeptides was calculated to be 34, 630 daltons. The base composition of the CP gene was 33.3% of adenine, 25.2% of guanine, 20.1% of cytosine and 21.4% of uracil. The polypeptide encoded by PVY-K CP gene was comprised of 22 alanines, 20 threonines, 19 glutamic acids and 18 glycines in order. The homology of nucleotide sequence of PVY-K CP gene with those of PVY-O(Japan), PVY-T(Japan), PVY-TH(Japan), PVYN(the Netherlands), and PVYN(France) was represented as 97.3%, 88.9%, 89.3%, 89.6% and 98.5%, respectively. The amino acid sequence homology of the polypeptide encoded by PVY-K CP gene with those encoded by viruses was represented as 97.4%, 92.5%, 92.9%, 92.9%, and 98.5%, respectively.