• The Plant Pathology Journal
• /
• v.35 no.2
• /
• pp.172-177
• /
• 2019

A duplex PCR method was developed for simultaneous detection and identification of tobacco root rot pathogens Phytophthora nicotianae and Thielaviopsis basicola. The specific primers for P. nicotianae were developed based on its internal transcribed spacer (ITS) regions of ribosomal gene, ras gene and hgd gene, while the specific primers for T. basicola were designed based on its ITS regions and ${\beta}$-tubulin gene. The specificity of the primers was determined using isolates of P. nicotianae, T. basicola and control samples. The results showed that the target pathogens could be detected from diseased tobacco plants by a combination of the specific primers. The sensitivity limitation was $100fg/{\mu}l$ of pure genomic DNA of the pathogens. This new assay can be applied to screen out target pathogens rapidly and reliably in one PCR and will be an important tool for the identification and precise early prediction of these two destructive diseases of tobacco.

• BMB Reports
• /
• v.40 no.5
• /
• pp.670-677
• /
• 2007

Although polysaccharide is believed to play an important role in the medicinal effect of Dendrobium chrysotoxum Lindl (DCL), its role as an antioxidant and in anti-hyperglycemic induction was not reported. In this study, polysaccharide with molecular weight of approximately 150 kDa, herein named DCLP, was isolated from the stem of DCL, and its antioxidative, hypoglycemic and immune stimulating effects were evaluated using various in vitro and in vivo assay systems. DCLP inhibited hydroxyl radicals ($^{\cdot}$OH)-mediated deoxyribose degradation by scavenging hydroxyl radicals directly as well as by chelating iron ions. DCLP also showed dose-dependent scavenging activity on superoxide anions ($O_2^{{\cdot}-}$) and offered significant protection (p < 0.001) against glucose oxidase-mediated cytotoxicity in Jurkat cells. DCLP had immune stimulating effects, as evidenced by the DCLP-mediated increases in the level of DNA synthesis, viability, and cytokine secretion in mouse lymphocytes. Moreover, oral administration of DCLP produced a significant reduction in blood glucose level in alloxan-induced diabetic mice. These findings suggest that DCLP has a potential utility in treating patients who require enhanced antioxidation, immune function and/or hypoglycemic activity.

• Journal of the Society of Cosmetic Scientists of Korea
• /
• v.27 no.1
• /
• pp.73-81
• /
• 2001

For an attempt to develop safe materials protecting UV-induced skin damage, plant extracts were evaluated for their antioxidative and free radical scavenging activities. From the results of these screening procedures, the ethanol extract of the flowers of Prunus persica was selected for further study. It was found that Prunus persica (50-200 $\mu\textrm{g}$/㎖) inhibited UVB-induced DNA damage measured by tail moment in the Single Cell Gel Electrophoresis(COMET assay) and inhibited UV-induced lipid peroxidation, expecially against UVB-induced peroxidation at higher than 10 $\mu\textrm{g}$/㎖. Also P.persica(100∼1,000 $\mu\textrm{g}$/㎖) inhibited the amount of $\^$14/C-arachidonic acid metabolites release from UVB-irradiated keratinocytes and it possessed the protective activity against UV-induced cytotoxicity of keratinocytes. All these results indicate that the flowers of P. persica extract may be beneficial for protection UV-induced skin damage when topically applied.

• The Plant Pathology Journal
• /
• v.31 no.1
• /
• pp.41-49
• /
• 2015

Sweet cherry is an important fruit crop with increasing economical value in Turkey and the world. A number of viruses cause diseases and economical losses in sweet cherry. Prune dwarf virus (PDV), is one of the most common viruses of stone fruits including sweet cherry in the world. In this study, PDV was detected from 316 of 521 sweet cherry samples collected from 142 orchards in 10 districts of Isparta province of Turkey by double antibody sandwich-enzyme linked immunosorbent assay (DAS-ELISA). The presence of PDV in ELISA positive samples was confirmed in 37 isolates by reverse transcription- polymerase chain reaction (RT-PCR) method. A genomic region of 862 bp containing the coat protein (CP) gene of PDV was re-amplified from 21 selected isolates by RT-PCR. Amplified DNA fragments of these isolates were purified and sequenced for molecular characterization and determining genetic diversity of PDV. Sequence comparisons showed 84-99% to 81-100% sequence identity at nucleotide and amino acid level, respectively, of the CP genes of PDV isolates from Isparta and other parts of the world. Phylogenetic analyses of the CP genes of PDV isolates from different geographical origins and diverse hosts revealed that PDV isolates formed different phylogenetic groups. While isolates were not grouped solely based on their geographical origins or hosts, some association between phylogenetic groups and geographical origins or hosts were observed.

• Journal of Microbiology and Biotechnology
• /
• v.24 no.12
• /
• pp.1728-1735
• /
• 2014

Scrub typhus, caused by infection with Orientia tsutsugamushi, is a mite-borne zoonotic disease endemic to the Asian-Pacific region. In Korea, the incidence of this disease has increased with climate changes, and over 10,000 cases of infection were reported in 2013. Although this infection is treatable with antibiotics such as doxycycline and azithromycin, an effective prophylactic vaccine against O. tsutsugamushi would be more desirable for preventing scrub typhus in endemic areas. In this study, we investigated the 56-kDa type-specific antigen (TSA56), which is a major outer membrane protein of O. tsutsugamushi, as a vaccine candidate. Intranasal immunization of recombinant TSA56 (rec56) induced a higher level of TSA56-specific IgG than that induced by intramuscular immunization of tsa56-expressing DNA (p56). Both types of immunization induced a cell-mediated immune response to TSA56, as demonstrated by the splenic cell proliferation assay. Mice immunized with p56, followed by rec56 plus heat-labile enterotoxin B subunit from E. coli, had a stronger protection from a homologous challenge with the O. tsutsugamushi Boryong strain than with other combinations. Our preliminary results suggest that an effective human vaccine for scrub typhus can include either recombinant TSA56 protein or tsa56-expressing DNA, and provide the basis for further studies to optimize vaccine performance using additional antigens or different adjuvants.

• Journal of Radiation Protection and Research
• /
• v.44 no.1
• /
• pp.15-25
• /
• 2019

Background: Psammaplin A (PsA) is a radiosensitizer whereas its clinical application is hampered by poor bioavailability. This study aimed to synthesize novel radiosensitizers using PsA as the lead compound. Materials and Methods: Eight homodimeric disulfides were synthesized from corresponding acid and cystamine dihydrochloride in N-hydroxysuccinimide and dicyclohexylcarbodiimide coupling conditions. One monomeric thiol analog was obtained by reduction of homodimeric disulfide with dithiothreitol. Clonogenic assay was used to measure cell survival after irradiation and drug treatment in human lung cancer (A549) and glioblastoma (U373MG) cells. Results and Discussion: Using the PsA backbone, nine compounds were synthesized. Eight compounds showed variable cytotoxicity with 50% inhibitory concentrations ranging $16.14{\mu}M$ to $150.10{\mu}M$ (A549), and $13.25{\mu}M$ to $50.15{\mu}M$ (U373MG). Four and six compounds radiosensitized A549 and U373MG cells, respectively. Two compounds that radiosensitized both cell lines were tested for its inhibitory effects on DNMT1. One of them was shown to significantly inhibit DNMT1 activity. Conclusion: Novel compounds with radiosensitizing activity were synthesized. These compounds have a great potential to serve as a basis for the development of future radiosensitizers. Further investigation is warranted for their clinical application.

• The Plant Pathology Journal
• /
• v.38 no.4
• /
• pp.313-322
• /
• 2022

Seed-borne pathogens in crops reduce the seed germination rate and hamper seedling growth, leading to significant yield loss. Due to the growing concerns about environmental damage and the development of resistance to agrochemicals among pathogen populations, there is a strong demand for eco-friendly alternatives to synthetic chemicals in agriculture. It has been well established during the last few decades that plant seeds harbor diverse microbes, some of which are vertically transmitted and important for plant health and productivity. In this study, we isolated culturable endophytic bacteria and fungi from soybean seeds and evaluated their antagonistic activities against common bacterial and fungal seed-borne pathogens of soybean. A total of 87 bacterial isolates and 66 fungal isolates were obtained. Sequencing of 16S rDNA and internal transcribed spacer amplicon showed that these isolates correspond to 30 and 15 different species of bacteria and fungi, respectively. Our antibacterial and antifungal activity assay showed that four fungal species and nine bacterial species have the potential to suppress the growth of at least one seed-borne pathogen tested in the study. Among them, Pseudomonas koreensis appears to have strong antagonistic activities across all the pathogens. Our collection of soybean seed endophytes would be a valuable resource not only for studying biology and ecology of seed endophytes but also for practical deployment of seed endophytes toward crop protection.

• Journal of Microbiology and Biotechnology
• /
• v.26 no.11
• /
• pp.1881-1890
• /
• 2016

Manganese superoxide dismutase (MnSOD) is a vital enzyme that protects cells from free radicals through eliminating superoxide radicals ($O^{2-}$). Hirudin, a kind of small active peptide molecule, is one of the strongest anticoagulants that can effectively cure thrombus diseases. In this study, we fused Hirudin to the C terminus of human MnSOD with the GGGGS linker to generate a novel dual-feature fusion protein, denoted as hMnSOD-Hirudin. The hMnSOD-Hirudin gene fragment was cloned into the pET15b (SmaI, CIAP) vector, forming a recombinant pET15b-hMnSOD-Hirudin plasmid, and then was transferred into Escherichia coli strain Rosetta-gami for expression. SDS-PAGE was used to detect the fusion protein, which was expected to be about 30 kDa upon IPTG induction. Furthermore, the hMnSOD-Hirudin protein was heavily detected as a soluble form in the supernatant. The purification rate observed after Ni NTA affinity chromatography was above 95%. The hMnSOD-Hirudin protein yield reached 67.25 mg per liter of bacterial culture. The identity of the purified protein was confirmed by western blotting. The hMnSOD-Hirudin protein activity assay evinced that the antioxidation activity of the hMnSOD-Hirudin protein obtained was $2,444.0{\pm}96.0U/mg$, and the anticoagulant activity of the hMnSOD-Hirudin protein was $599.0{\pm}35.0ATU/mg$. In addition, in vitro bioactivity assay showed that the hMnSOD-Hirudin protein had no or little cytotoxicity in H9c2, HK-2, and H9 (human $CD_4{^+}$, T cell) cell lines. Transwell migration assay and invasion assay showed that the hMnSOD-Hirudin protein could suppress human lung cancer 95-D cell metastasis and invasion in vitro.

• Journal of The Korean Society of Clinical Toxicology
• /
• v.17 no.2
• /
• pp.58-65
• /
• 2019

Purpose: Alpha-amanitin induces potent oxidative stress and apoptosis, and may play a significant role in the pathogenesis of hepatotoxicity. This study examined the mechanisms of α-amanitin-induced apoptosis in vitro, and whether green tea extract (GTE) offers protection against hepatic damage caused by α-amanitin (AMA) induced apoptosis in vivo. Methods: The effects of GTE and SIL on the cell viability of cultured murine hepatocytes induced by AMA were evaluated using an MTT assay. Apoptosis was assessed by an analysis of DNA fragmentation and caspase-3. In the in vivo protocol, mice were divided into the following four groups: control group (0.9% saline injection), AMA group (α-amanitin 0.6 mg/kg), AMA+SIL group (α-amanitin and silibinin 50 mg/kg), and AMA+GTE group (α-amanitin and green tea extract 25 mg/kg). After 48 hours of treatment, the hepatic aminotransferase and the extent of hepatonecrosis of each subject was evaluated. Results: In the hepatocytes exposed to AMA and the tested antidotes, the cell viability was significantly lower than the AMA only group. An analysis of DNA fragmentation showed distinctive cleavage of hepatocyte nuclear DNA in the cells exposed to AMA. In addition, the AMA and GTE or SIL groups showed more relief of the cleavage of the nuclear DNA ladder. Similarly, values of caspase-3 in the AMA+GTE and AMA+SIL groups were significantly lower than in the AMA group. The serum AST and ALT levels were significantly higher in the AMA group than in the control and significantly lower in the AMA+GTE group. In addition, AMA+GTE induced a significant decrease in hepatonecrosis compared to the controls when a histologic grading scale was used. Conclusion: GTE is effective against AMA-induced hepatotoxicity with its apoptosis regulatory properties under in vitro and in vivo conditions.

• Journal of the Korean Society of Food Science and Nutrition
• /
• v.40 no.1
• /
• pp.78-83
• /
• 2011

The flower of Chrysanthemum morifolium Ramat. with antioxidant, anticancer, and anti-inflammatory functions has been a widely used traditional herb as a healthy beverage and medicine. The aim of the present study was to investigate a herb tea consisting of C. morifolium Ramat., Corni fructus and Schizandra chinensis Baillon for its hepatoprotective activity against $CCl_4$-induced toxicity in freshly isolated rat hepatocytes and antigenotoxic effect against oxidative stress induced DNA damage in human leukocytes. Three different compositions of the herb tea (Mix I, II, and III) were prepared by extracting with water at $90^{\circ}C$. Freshly isolated rat hepatocytes were exposed to $CCl_4$ along with/without various concentrations of each tea. Protection of rat primary cells against $CCl_4$-induced damage was determined by the MTT assay. The significant antihepatotoxic effect of the tea was shown in Mix I and II. The increased transaminase (AST and/or ALT) release in media of $CCl_4$ treated hepatocytes was significantly lowered by all the teas tested. The effect of the tea on DNA damage in human leukocytes was evaluated by Comet assay. All teas showed a protective effect against $H_2O_2$-induced DNA damage. From these results, it is assumed that herb tea based on C. morifolium Ramat., Corni fructus and Schizandra chinensis Baillon exerted antihepatotoxic and antigenotoxic effects.