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Inactivation of Mycobacteria by Radicals from Non-Thermal Plasma Jet

  • Lee, Chaebok;Subhadra, Bindu;Choi, Hei-Gwon;Suh, Hyun-Woo;Uhm, Han. S;Kim, Hwa-Jung
    • Journal of Microbiology and Biotechnology
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    • v.29 no.9
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    • pp.1401-1411
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    • 2019
  • Mycobacterial cell walls comprise thick and diverse lipids and glycolipids that act as a permeability barrier to antibiotics or other chemical agents. The use of OH radicals from a non-thermal plasma jet (NTPJ) for the inactivation of mycobacteria in aqueous solution was adopted as a novel approach. Addition of water vapor in a nitrogen plasma jet generated OH radicals, which converted to hydrogen peroxide ($H_2O_2$) that inactivated non-pathogenic Mycobacterium smegmatis and pathogenic Mycobacterium tuberculosis H37Rv. A stable plasma plume was obtained from a nitrogen plasma jet with 1.91 W of power, killing Escherichia coli and mycobacteria effectively, whereas addition of catalase decreased the effects of the former. Mycobacteria were more resistant than E. coli to NTPJ treatment. Plasma treatment enhanced intracellular ROS production and upregulation of genes related to ROS stress responses (thiolrelated oxidoreductases, such as SseA and DoxX, and ferric uptake regulator furA). Morphological changes of M. smegmatis and M. tuberculosis H37Rv were observed after 5 min treatment with $N_2+H_2O$ plasma, but not of pre-incubated sample with catalase. This finding indicates that the bactericidal efficacy of NTPJ is related to the toxicity of OH and $H_2O_2$ radicals in cells. Therefore, our study suggests that NTPJ treatment may effectively control pulmonary infections caused by M. tuberculosis and nontuberculous mycobacteria (NTM) such as M. avium or M. abscessus in water.

Fusaric Acid Production in Fusarium oxysporum Transformants Generated by Restriction Enzyme-Mediated Integration Procedure (Restriction Enzyme-Mediated Integration 방법으로 확보한 Fusarium oxysporum 형질전환체의 후자리산 생성능 분석)

  • Lee, Theresa;Shin, Jean Young;Son, Seung Wan;Lee, Soohyung;Ryu, Jae-Gee
    • Research in Plant Disease
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    • v.19 no.4
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    • pp.254-258
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    • 2013
  • Fusaric acid (FA) is a mycotoxin produced by Fusarium species. Its toxicity is relatively low but often associated with other mycotoxins, thus enhancing total toxicity. To date, biosynthetic genes or enzymes for FA have not been identified in F. oxysporum. In order to explore the genetic element(s) for FA biosynthesis, restriction enzyme mediated integration (REMI) procedure as an insertional mutagenesis was employed using FA producing-F. oxysporum strains. Genetic transformation of two F. oxysporum strains by REMI yielded more than 7,100 transformants with efficiency of average 3.2 transformants/${\mu}g$ DNA. To develop a screening system using phytotoxicity of FA, eleven various grains and vegetable seeds were tested for germination in cultures containing FA: Kimchi cabbage seed was selected as the most sensitive host. Screening for FA non-producer of F. oxysporum was done by growing each fungal REMI transformant in Czapek-Dox broth for 3 weeks at $25^{\circ}C$ then observing if the Kimchi cabbage seeds germinated in the culture filtrate. Of more than 5,000 REMI transformants screened, fifty-three made the seeds germinated, indicating that they produced little or fewer FA. Among them, twenty-six were analyzed for FA production by HPLC and two turned out to produce less than 1% of FA produced by a wild type strain. Sequencing of genomic DNA regions (252 bp) flanking the vector insertion site revealed an uncharacterized genomic region homologous (93%) to the F. fujikuroi genome. Further study is necessary to determine if the vector insertion sites in FA-deficient mutants are associated with FA production.

Studies on the Host Range of Colletotrichum dematium Isolated from Anthracnose of Pepper and Toxic Metabolites Produced by the Pathogen (고추 탄저병균(炭疽病菌) Colletotrichum dematium의 기주범위(寄主範圍) 및 대사독소(代謝毒素)에 관(關)한 연구(硏究))

  • Kang, Hi Wang;Yu, Seung Hun;Park, Jong Seong
    • Korean Journal of Agricultural Science
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    • v.14 no.1
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    • pp.26-37
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    • 1987
  • This studies were conducted to investigate pathgenicity and host range of Colletotrichum dematium isolated from anthracnose of pepper, and phytoxicity of its culture filtrate and the partially purified toxin. The results obtained were as follows. 1. Investigation on the host range of C. dematium has revealed that pepper as well as soybean, tomato, spinach, and beet were highly susceptible, egg plant and water melon were moderately susceptible and stone leek was slightly suceptible, but no symptoms were produced on carrot, tabacco, cucumber and melon. 2. The culture filtrates of C. dematium in Czapeck dox liquid media were toxic to leaves of pepper and caused necrosis and wilting of the plant. The toxicity of culture filtrates was most active at 15 days after fungal growth in Czapeck dox liquid media and the toxin productivity in still culture was higher than that in shaking culture. 3. The partially purified toxic substance was isolated from the culture filtrates by the acetone precipitation method. When cuttings of various pepper cultivars were placed in the toxin solutions, suceptible cultivars and resistant cultivars were equally toxic and showed necrosis and wilting of the leaves. 4. Several other plants such as soybean, tomato and carrot were also affected with the toxin solution by shoot cutting bioassay and showed veinal necrosis, leaf spots and wilting of the shoots. 5. The acetone precipitation toxin affected seed germination of pepper, cucumber, sesame and egg plant and inhibited the growth of root and hypocotyl of the seedlings.

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The physiological and degradational characteristics of Fomers fomentarius (말굽버섯균의 생리적 특성과 부후특성)

  • Kim, Yu-I;Chai, Jyung-Ki
    • Journal of Mushroom
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    • v.2 no.4
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    • pp.200-206
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    • 2004
  • The optimal temperature for mycelial growth the F. fomentarius was $30^{\circ}C$ and the range of the temperature for mycelial growth wsa about 10~30. The optimal pH for the growth was 4.0. The percentage of weight loss percentage wsa 17.4%. The percentage of WEC extractives wsa increased to 2.24%. The observation of micromorphological showed that the detected cell wall were erosive and thinning as typical degradation pattern of white-rot fungi.

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Studies on the Varietal Resistance and Effects of Nutrients for Fungal Growth of Pepper Anthracnose Disease Caused by Colletotrichum dematium f. sp. capsicum (고추 탄달병(炭疸病)에 대한 품종(品種) 저항성(抵抗性) 및 병원균(病原菌) 생장(生長)에 미치는 영양원(營養源)의 효과(效果))

  • Chang, Sun-Hwa;Chung, Bong-Koo
    • The Korean Journal of Mycology
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    • v.13 no.4
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    • pp.227-233
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    • 1985
  • Studies on the varietal resistance and effects of nutrients for fungal growth were carried out in order to obtain basic materials for breeding resistant variety and control measures of the red pepper anthracnose disease caused by Colletotrichum dematium f. sp. capsicum. Four cultivars such as Kumchang No. 2, Bulamhouse, Pakistan, Hongilpum were resistant among twenty-one pepper cultivars, and five cultivars including Taiwan pepper were moderate. The remaining twelve cultivars including H-038 and Saegochu were susceptible. These susceptible cultivars were mostly belonged to sweet taste cultivars. Glucose was known the best source for fungal growth, and near 3 percent of carbon concentration was the best for mycelial growth of the fungus. Conidial sporulation was rather decreased by adding high concentrations of C-source, whereas fungal dry weight was a positive tendency in proportion to increasing carbon concentrations. N-sources and vitamins were not remarkable as that for carbon, and rather a decreasing trend for mycelial growth by adding N-source. Especially, the lowest of mycelial growth was in the case of Czapek-Dox plus ammonium sulfate. The medium plus vitamins either niacin or thiamine was slightly increased to mycelial growth.

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Cultural Characteristics of Cauliflower Mushroom, Sparassis crispa (꽃송이버섯(Sparassis crispa)의 배양적 특성)

  • Cheong, Jong-Chun;Park, Jeong-Sik;Hong, In-Pyo;Seok, Soon-Ja;Jhune, Chang-Sung;Lee, Chan-Jung
    • The Korean Journal of Mycology
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    • v.36 no.1
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    • pp.16-21
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    • 2008
  • This study was carried out to obtain the basic data for the mycelial growth conditions of cauliflower mushroom, Sparassis crispa. Twenty-one isolates were collected from domestic and abroad. The optimal temperature and pH for the mycelial growth of these isolates were $23^{\circ}C$ and pH 6.0, respectively. The mycelial growth was the best in the HBA medium, but very poor in the Lilly medium. However no mycelial growth in the CzapekDox medium. The utilization of carbon source was the best with fructose, and that of nitrogen source was the best with glutamine when compared to other tested sources. The selected isolate ASI150010 produced the highest mycelial weight in liquid culture containing soybean mill ($15\;g/{\ell}$) and potato ($200\;g/{\ell}$) extract. And uncleaned rice, wheat and barley were found to be good substrates for the mycelial growth S. crispa.

Antitumor Activity of Methylene Chloride Fraction from Angelica Keiskei Through Induction of Apoptosis in Human Prostate Carcinoma DU145 Cells (전립선 암세포주 DU145의 세포고사 유도를 통한 신선초 (Angelica keiskei) 메틸렌 클로라이드 추출물의 항암효과)

  • Kang, Yoon-Mook;Kim, Sung-Moo;Kim, Hyun-Jung;Park, Kyung-Ran;Shim, Bum-Sang;Kim, Sung-Hoon;Choi, Seung-Hoon;Ahn, Kyoo-Seok;Ahn, Kwang-Seok
    • Journal of Korean Traditional Oncology
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    • v.15 no.1
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    • pp.19-27
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    • 2010
  • The roots and leaves of Angelica keiskei (AK) have been used for the treatment of various diseases including coronary heartdisease, hypertension, and cancer in the Korean folk medicine. However, the mechanism by which methylenechloride fraction (MF) from AK exerts anti-tumorigenic activity in human prostate cancer cells has not been fully understood. In the present study, we report the MF exerted the highest cytotoxicity against prostate cancer DU145 cells compared with other fractions. Especially, MF caused the accumulation of sub-G1 DNA contents of cell cycle and increased annexin V-positive apoptotic bodies and DNA fragmentation. MF down-regulated several proliferative (Cyclin D1) and anti-apoptotic (Bcl-xl, Bcl-2, IAP-1/2, and survivin)gene products in these cells. Hence, MF induced apoptosis through the caspase-3 activation in DU145 cells. We further confirmed that caspase-3 plays an importance role in MF-induced apoptosis in DU145 cells by using caspase-3 inhibitor. Additionally, we observed that MF potentiated Dox-induced apoptosis in DU145 cells. Taken together, our data demonstrate the evidence that MF induces apoptosis depend on caspase-3 activation of and overcomes resistance to chemotherapy in human prostate cancer cells.

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Comparative Outcome of Thai Pediatric Osteosarcoma Treated with Two Protocols: the Role of High-Dose Methotrexate (HDMTX) in a Single Institute Experience

  • Choeyprasert, Worawut;Pakakasama, Samart;Sirachainan, Nongnuch;Songdej, Duantida;Chuansumrit, Ampaiwan;Anurathapan, Usanarat;Hongeng, Suradej;Nartthanarung, Adisak
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.22
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    • pp.9823-9829
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    • 2014
  • Background: High-dose methotrexate (HD-MTX) is recognized as an efficient component of therapy against pediatric osteosarcoma in combination with other drugs such as cisplatin (CDP), carboplatin (CBDCA), doxorubicin (ADM), etoposide (VP-16) and ifosfamide (IFO). Objectives: To demonstrate the feasibility and effectiveness of the HD-MTX/CDP/DOX/VP-16/IFO [MTX(+)] protocol comparable to CDP/ADM/CBDCA/IFO [MTX(-)] for treating childhood osteosarcoma at Ramathibodi Hospital (1999-2014). Materials and Methods: A retrospective analysis was conducted of osteosarcoma patients aged less than 18 years treated with two chemotherapeutic regimens between 1999 and 2014. A total of 45 patients received the MTX(-) and 21 the MTX(+) protocol. Results: Overall limb-salvage and amputation rate were 12.9% and 77.7%, respectively. Kaplan-Meier analysis results for 3-year disease free survival (DFS) and overall survival (OS) regardless of treatment regimens were $43.4{\pm}6.0%$ and $53.2{\pm}6.1%$ respectively. The 3-year DFS and OS were improved significantly with the MTX(+) protocol compared to MTX(-) protocol (p=0.010 and p=0.009, log rank test) [$69.8{\pm}10.5%$, $79.8{\pm}9.1%$ for MTX(+) and $31.1{\pm}6.9%$, $42.2{\pm}7.4%$ for MTX(-) protocol, respectively]. Patients with metastatic osteosarcoma treated with the MTX(+) protocol had statistically significant higher 3-year DFS and OS than those treated with the MTX(-) protocol ($66.7{\pm}13.6%$ and $15.0{\pm}8.0%$ for 3-year DFS, p=0.010, $73.3{\pm}13.2%$ and $20{\pm}8.9%$ for 3-year OS, p=0.006, respectively). The independent risk factors for having inferior 3-year DFS and OS were poor histological response (tumor necrosis <90%) and treatment with the MTX(-) protocol. The multivariate analysis identified only the treatment with the MTX(-) protocol as an independent predictor of inferior OS with a hazard ratio (HR) of 3.53 (95% confidence interval of 1.2-10.41, p=0.022). Conclusions: Our study demonstrated the tolerability, feasibility and efficacy of the HDMTX-based regimen improving the survival rate in pediatric osteosarcoma cases, in line with reports from developed countries.

Composition of a New Medium for Mycelial Growth of Hericium erinaceus (노루궁뎅이버섯(Hericium erinaceus)의 새로운 균사배양기의 조성)

  • Ko, Han-Gyu;Kim, Dong-Myong;Park, Won-Mok
    • The Korean Journal of Mycology
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    • v.25 no.4 s.83
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    • pp.369-376
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    • 1997
  • These researches were carried out for improvement of medium for mycelial growth of Hericium erinaceus isolate KU-1. It grew well at pH 4 and $25^{\circ}C$. Glucose and sucrose were favorable carbon sources for mycelial growth. As nitrogen sources, ammonium acetate and arginine enhanced mycelial growth. Optimum C/N ratio was 200. Based On the results, the following recipe is suggested for synthetic medium for the mycelial growth: glucose 18.02 g, arginine 2.613 g, ammonium acetate 2.313 g, $CaCl_2\;0.33\;g$, $KH_2PO_4\;8.5\;g$, $MgSO_4{\cdot}7H_2O\;2.0\;g$, $FeSO_4{\cdot}7H_2O\;0.02\;g$, $ZnSO_4{cdot}7H_2O\;0.02\;g$, $MnSO_4{\cdot}7H_2O\;0.02\;g$, water 1 liter. This medium was superior for the mycelial growth to other conventional media such as Yeast malt extract agar (YMA), Park medium, Potato dextrose agar (PDA), Malt extract agar, Czapek-dox agar, Macaya-lizano medium and Yeast extract agar. This new synthetic medium is designated as Ko medium.

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Salvage Chemotherapy in Recurrent Platinum-Resistant or Refractory Epithelial Ovarian Cancer with Carboplatin and Distearoylphosphatidylcholine Pegylated Liposomal Doxorubicin (Lipo-Dox®)

  • Khemapech, Nipon;Oranratanaphan, S.;Termrungruanglert, W.;Lertkhachonsuk, R.;Vasurattana, A.
    • Asian Pacific Journal of Cancer Prevention
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    • v.14 no.3
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    • pp.2131-2135
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    • 2013
  • Background: To evaluate the efficacy and safety of distearoylphosphatidylcholine pegylated liposomal doxorubicin (DPLD) combined with carboplatin for the treatment of platinum resistant or refractory epithelial ovarian cancer (EOC) or fallopian tube cancer. Materials and Methods: A retrospective analysis of women who received DPLD with carboplatin for recurrent EOC or fallopian tube cancer in King Chulalongkorn Memorial Hospital Thailand from January 2006 to August 2011 was conducted. Patients were identified from the medical records and data on demographic factors, stage, histology, surgical findings, cytoreduction status, and prior chemotherapies were abstracted. The efficacy and toxicity of DPLD/carboplatin were evaluated. Progression-free (PFS) and overall survival (OS) were estimated by the Kaplan-Meier method. Results: A total of 65 patients, 64 with platinum resistant or refractory epithelial ovarian cancer and 1 with fallopian tube cancer, were enrolled. DPLD and carboplatin were given for an average of 4.46 cycles per patient with a total of 273 cycles. Among the 65 evaluable patients, 0% achieved CR, 7.69% PR, 15.4% SD and 76.% PD. The overall response rate was 23.1%. With a median follow-up of 27.4 months, the median progression-free and median overall survival in the 36 patients was 4.46 months and 8.76 months respectively. In the aspect of side effects, palmar-plantar erythrodysesthesia (PPE) occurred in 33.3% (Grade I 22.2%, Grade II 11.1%) and mucositis in 41.7% (Grade I 27.8%, Grade II 13.9%) of all treatment cycles, all Grade 1 or 2. Anemia, leukopenia and thrombocytopenia occurred in 58.3% (Grade I 41.7%, Grade II 16.7%), 66.7% (Grade I 47.2%, Grade II 19.4%), and 22.2% (Grade I 16.6%, Grade II 5.56%) of cycle respectively, and were mostly Grade 1 or 2. Conclusions: DPLD, the second-generation PLD drug combined with carboplatin every 4 weeks, is effective and has low toxicity for treatment of patients with recurrent platinum-resistant or refractory epithelial ovarian cancer.