Effects of Rhizobacteria for Growth Promotion and Induction of Fungal Disease Resistance in Vegetable Crops

Gurung, Sun Kumar Kangwon National University 2019 국내석사

Increasing public concerns about adverse effect of pesticides and fertilizer resulted in renewed interest in beneficial soil microorganisms that can improve soil quality and enhance plant health. Plant growth promoting rhizobacteria (PGPR) are beneficial microorganisms that have potentiality for both plant growth promotion and plant diseases control. Most studies of PGPR as antagonists begin by assessing the efficacy of single PGPR strains against plant pathogens. However, in the field several diseases often occur on individual crops. This study was conducted to select PGPR strains for broad-spectrum pathogen suppression and growth promotion in the presence of plant pathogens. First, preliminary screening was carried out for antibiosis capacity of PGPR strains against nine different fungal pathogens in vitro. Nine fungal pathogens include Pythium ultimum, Phytopthora capsici, Cercospora capsici, Fusarium oxysporium f. sp. melonis, Sclerotium rolfsi, Stemphylium lycopersicum, Stemphylium botryosum, Collectotrichum dematium, and Monosporascus cannonbullus. All the tested pathogens were obtained from Korean Agriculture Culture Collection (KACC). The bacterial isolates were collected from soils of Gangwon-do, Korea. 16S rRNA gene sequence analysis was performed for the identification of the isolates, RR8 (Panibacillus polymyxa), EN23 (Bacillus siamensis), AB05 (Bacillus valenzensis), AB12 (Bacillus subtilis) and PA5 (Bacillus subtilis). The rhizobacteria Paenibacillus polymyxa (RR8) showed the highest antagonistic activity against all tested fungal pathogens except Monosporascus cannonbullus. According to the results, RR8 exhibited mycelial suppression at 47.8 %, 46.5 %, 44.1 % 42.3 %, 38.2 %, 18.7 %, 17.8 % and 5.0 % against Fusarium oxysporium f. sp. melonis, Cercospora capsici, Stemphylium lycopersici, Phytopthora capsici, Stemphylium botryosum, Pythium ultimum, Colletotrichum dematium, and Sclerotiorum rolfsii respectively. Moreover, EN23 exhibited the highest efficacy against Sclerotinia rolfsii in I- plate assay. In vitro biochemical test revealed that EN23 exhibited the highest protein hydrolysis, RR8 was positive for zinc solubilization and phosphate solubilization and EN23, AB05 and PA05 were positive for ammonia production. In vitro plant assay was conducted to evaluate the growth promotion and disease inhibition in studied crops (melon, tomato and lettuce) by using the effective single and combined bacterial isolates (RR8, EN23 and PA5) from in vitro tests. According to the result, the highest shoot dry weight (89.40 mg) was observed in combination treatments of RR8 and EN23 in melon seedling as compared to other treatments and control. In tomato, combination treatments of RR8 and EN23 showed the highest shoot dry weight (113.0 mg) as compared to other treatments and control. In lettuce, combination treatments of RR8 and AB12 exhibited the highest shoot dry weight (31.80 mg) as compared to other treatments and control. Greenhouse evaluation of effective bacterial isolates (RR8, EN23 and PA5) and their combinations were carried out to assess their efficacy in promoting growth and reducing disease in melon, tomato and lettuce. Results showed that in melon, the highest shoot dry weight (10.56 g) was observed in combination treatments of RR8 and EN23. In tomato, RR8 and EN23 combination treatments showed the highest shoot dry weight (26.2 g). In lettuce, leaf area was found the highest (174.7 cm2) in combination treatments of RR8 and AB12. In addition, reduction of disease rate of Fusarium oxysporium f. sp. melonis on melon was observed at 46.2 % by the combination treatments of EN23 and AB 12. In tomato, combination treatments of EN23 and AB12 showed 37.04 % reduction of disease rate of Stemphylium lycopersicum. In lettuce, Pythium ultimum was reduced by 44.78 % with the combination treatments of RR8 and AB12. In addition, field evaluations with effective bacterial isolates (RR8, EN23 and PA5) and their combinations results exhibited that combination of RR8 and EN23 showed the highest plant height 198.2 cm compared to other treatments and control. In case of tomato crop highest number of fruit per plant (158) was recorded in combination treatments of RR8 and AB12. In lettuce crop, leaf area (192.6 cm2) was recorded at the highest in combination treatments of RR8 and EN23. Reduction of disease rate of Fusarium oxysporium f. sp. melonis in melon was recorded the highest (46.02 %) in combination treatments of EN23 and AB12. In lettuce, disease rate of Pythium ultimum was reduced by 42.2 % in combination treatments of RR8 and AB12. In tomato, disease reduction rate of Stemphylium lycopersicum was recorded the highest (37.80 %) with the combination treatments of AB12 and EN23 as compared to control. In summary, both the tested individual and combination treatments of PGPR exhibited the high levels of disease control and plant growth promotion compared to control but the magnitude of the efficacy was better in combination treatments than that of individual PGPR treatments.

Fundamental Understanding and Fabrication of Microbial Fuel Cells for Generation of Sustainable Renewable Energy

Gurung, Anup Kangwon National Univ 2013 국내박사

Energy is considered as one of the most important component for social and economic development. It is indispensable for securing a reasonable standard of living, as it fulfills basis human requirements. Therefore, per capita consumption is considered as a fundamental driver as well as indicator of wealth and quality of a nation. The current global energy demand is primarily dominated by fossil fuels in which more than 80% of this demand is met by fossil fuels sector. The heavy reliance on fossil fuels, especially oil and gas has triggered the global energy crisis as well as become one of the major drives for degrading the environment including air, water and soil. In recent years, the global energy demand has increased dramatically due to economically booming developing countries like Brazil, China and India. To become less dependent on fossil fuels, current world is shifting paradigm in energy by developing alternative energy sources mainly through the utilization of renewable energy sources. In particular, bioenergy recovery from wastes with the help of microorganism is viewed as one of the promising ways to mitigate the current global warming crisis as well as to supply global energy. It has been proved that microorganism can generate power by converting organic matter into electricity using microbial fuel cells (MFCs). MFC is a bioelectrochemical device that employs microbes to generate electricity from bio-convertible substrate such as wastewaters including municipal, solid waste, industrial and agricultural wastes, and sewage. Sustainability, carbon neutral and generation of renewable energy are some of the major features of MFCs. The possibility of bioelectricity generation in two chambered mediator-less MFCs was investigated using fermentable and non-fermentable substrates in batch mode. Two different electron acceptors namely dissolved oxygen and potassium ferricyanide-[K3Fe(CN)6] have been tested in the cathode chambered for the effects of reducing agent on the power generation. The average voltages generated with both substrates were more or less similar. However, with [K3Fe(CN)6] as the reducing agent, the power output increased by approximately 40%. In order to increase voltage generation, three individual H-type two chambered MFCs were connected together in series and parallel connections. The stackable MFCs in series produce an open circuit voltage (OCV) of 2.033 V which is equal to the mathematical sum of the individual MFCs. Similarly, the connection of three MFC units in parallel produced the maximum power of 0.4 mW, corresponding to the maximum current of 1.25 mA. For the better understanding of the voltage reversal phenomenon, a dead cell (without substrate) was connected with the two other good working cells. It was found that voltage reversal occurred when the dead cell connected in the circuit. Reactor configuration can influence the voltage generation significantly in MFCs. In order to investigate this, two chambered rectangular MFC was designed and operated in semi-batch mode. The maximum power density of 503±55 mW/m2 was obtained corresponding to the current density of 1107±326 mA/m2. In order to obtain high voltage, two MFCs were connected together in series or parallel. The stacked voltages of 1.3 V (OCV) and 1.22 V (1000 Ω) were produced, respectively, when two MFCs were connected together in series. When the MFCs were connected in parallel mode, the maximum power of 2.9 mW was obtained corresponding to the maximum current of 6.26 mA. Voltage reversal is a critical problem in a serially-connected MFC unit. The voltage reversal often occurs when substrate concentration is relatively low in the anodic compartment. In this study two rectangular individual cells were stacked together in a series in continuous mode. In order to investigate the effect of substrate on power generation, one MFC (MFC1) was constantly fed with 1 g/L glucose while another MFC (MFC2) was fed with varied concentrations of glucose (0.1, 0.2, 0.3, 0.5 and 0.8 g/L). Voltage reversal occurred when the MFC2 was fed with 0.1 g/L glucose, whereas in the remaining substrate concentrations voltage reversal did not occur. In an effort to reduce dependency on non-renewable energy sources, lignocellulosic biomass is considered as the potential sources of renewable energy in recent years. We investigated the electricity generation from rice straw in two chambered MFCs. The MFC produced the average working potential of 0.601±0.02 V (1000Ω). The MFC produced the maximum power of 0.6 mW corresponding to the maximum current of 1.41 mA. The present study demonstrates that stable power could be generated using rice straw as the electron donor source in MFCs. 미생물연료전지는 생활하수, 산업폐수, 농업부산물을 포함하는 폐수와 같은 물질로부터 미생물을 이용하여 전기를 생산하는 전기화학적 장치이다. 메디에이터가 없는 미생물연료전지의 전기 생산 가능성은 글루코오스, 아세테이트를 가지고 회분식 운전으로 연구하였다. 환원반응조에서 파워발생에 미치는 환원제의 효과는 두 개의 다른 전자 수용체 즉, 용존산소와 Potassium ferricyanide를 이용하여 실험하였다. 두 물질(글루코오스, 아세테이트)에 의해 발생된 평균전압은 유사하였다. 환원제로 Potassium ferricyanide를 사용하였을 때 파워는 약 40% 증가하였다. 파워 발생을 높이기 위해 세 개의 H-타입의 미생물연료전지를 직렬과 병렬로 각각 연결하였다. 직렬로 연결된 미생물연료전지의 OCV는 각각의 미생물연료전지의 총 전압(OCV)의 합과 비슷한 2.033V가 측정되었다. 병렬로 연결된 세 개의 미생물연료전지는 최대전류인 1.25mA에 상응하는 0.4mW의 최고 파워가 발생되었다. 전압 변환 (voltage reversal) 현상을 더 잘 이해하기 위해서 기질이 없는 죽은 반응조에 두 개의 정상적인 반응조를 연결했다. 회로에 죽은 반응조가 연결되었을 때 전압 변환이 발생했다. 반응조 배치는 미생물연료전지에서 전압 생산에 상당한 영향을 줄 수 있다. 이것을 연구하기 위해 두 개의 사각형 모양의 미생물연료전지가 제작되었고 이를 이용해 반회분식으로 실험하였다. 최고 전류밀도인 1107±326mA/cm2에 상응하는 503±55 mW/m2의 최고 파워밀도가 측정되었다. 높은 전압을 얻기 위해 두 개의 미생물연료전지가 직렬, 병렬로 연결되었다. 미생물연료전지가 직렬로 연결되었을 때 발생한 전압은 OCV의 경우 1.3 V, 1000Ω의 저항을 연결했을 경우 1.22 V가 각각 측정되었다. 미생물연료전지가 병렬로 연결되는 경우 최고전류 6.26 mA에 상응하는 최고 파워 2.9mW가 측정되었다. 전압 전환은 미생물연료전지가 직렬로 연결되었을 때 발생하는 심각한 문제이다. 전압 전환은 음극전지에 기질의 농도가 비교적 낮을 때 발생한다. 이번 연구에서 두 개의 사각형 모양의 미생물연료전지가 연속운전방식으로 직렬연결되었다. 기질의 파워 발생에 미치는 효과를 연구하기 위해 한 개의 미생물연료전지(MFC1)에 1 g/L의 글루코스를 연속적으로 주입하였고 이 때 다른 미생물연료전지(MFC2)에는 다양한 농도 (0.1, 0.2, 0.3, 0.5, 0.8 g/L)의 글루코스를 주입하였다. MFC2에 0.1 g/L 글루코스를 주입했을 때만 전압 변환이 발생했고 다른 농도의 글루코스 주입에서는 발생하지 않았다. 재이용불가능한 자원의 의존을 줄이기 위한 노력으로 리그노셀룰로직 바이오매스가 최근 재이용에너지원으로 고려되었다. 우리는 미생물연료전지를 이용, 볏짚으로 부터의 전기생산을 연구했다. 미생물연료전지는 1000Ω의 저항을 연결했을 때 0.601±0.02V의 포텐셜을 발생했다. 본 연구를 통해 미생물연료전지에서 전자공여체로 볏짚을 이용하여 안정적인 파워 발생이 가능하다는 것을 입증하였다

Study on the PROTAC Platform by Harnessing Targeted Protein Degradation Part I. Discovery of Novel KEAP1 Degrader Part II. Applicability of KEAP1 E3 Ligase to the PROTAC Platform : Part I. Discovery of Novel KEAP1 Degrader Part II. Applicability of KEAP1 E3 Ligase to the PROTAC Platform Part III.

RAJU GURUNG 가천대학교 메디컬캠퍼스 일반대학원 2024 국내박사

Synthesis and Anti-inflammatory Activity of Structurally Relevant Flavonoids, Bichalcones and Unnatural Biflavonoids

Gurung, Santosh Kumar Kangwon National University 2010 국내박사

Synthesis and Anti-inflammatory activity of bichalcone derivatives Bichalcones are molecules comprised of two identical or non-identical chalcone units linked in a symmetrical or unsymmetrical manner through an alkyl or an alkoxy based linker of varying length. A few numbers of natural or synthetic bichalcones have been reported and their biological data are limited to date. Here we report the synthesis of bichalcone derivatives having C-C and C-O-C at 4-4’’’ linkage and their anti-inflammatory activity. The synthetic bichalcones exhibited somewhat different inhibitory activities against production of PGE2 and NO. Though the degree of suppression against production of PGE2 and NO were not stronger, even some of the derivatives slightly promote production of PGE2 and NO. However, the methodology employed for the synthesis of bichalcone should be amenable to the preparation of additional derivatives for SAR studies to enhance the bioactivity. Bichalcones은 두 개의 동일하거나 서로 다른 chalcone이 대칭 혹은 비대칭 되게 alkyl 혹은 alkoxy구조로 연결되어 있다. 천연 bichalcones 화합물과 합성하여 얻은 일부 bichalcones 화합물은 그 생리활성에 대하여 많은 연구가 이미 진행되었다. 본 연구에서 4-4’’’ C-C와 C-O-C 연결구조를 가진 bichalcone 유도체를 합성하고 그들의 항 염증 생리활성을 검증하였다. 합성한 bichalcones은 PGE2와 NO에 대해 어느 정도의 생성 억제력을 보였다. . 그 중 대부분 화합물은 PGE2와 NO에 대한 생성 억제력이 그리 높지 않았으며 극 소수의 화합물이 PGE2와 NO에 대하여 조금 상승된 생성 억제력을 보였다. 하지만 본 연구에서 얻은 bichalcones의 합성방법은 다른 bichalcone 유도체의 합성에 유용하게 응용할 수 있다. 핵심어: Bichalcones 합성, Claisen-Schmidt condensation, Suzuki C-C 와 Ulmann C-O-C 결합 반응, PGE2, COX-2, NO, NO 생성, 항 염증 . Wogonin 유도체의 항염증 생리활성에 대한 SAR 연구 Wogonins A-ring의 작용기전 구조 변경을 통하여 얻은 물질을 LPS-induced RAW 264.7세포를 사용하여 COX-2 조건하에서 PGE2 생성 억제력을 검증하였다. Wogonin의 A-ring의 작용기전 그룹을 삭제한 유도체는 wogonin보다 낮은 생리활성을 나타냈다. 본 연구를 통하여 wogonin의 A-ring의 작용기전 모두가 생리활성에 있어서 각자의 중요한 역할을 한다는 것을 입증하였다. 핵심어: 구조 변경, wogonin, PGE2 생성, COX-2, 항 염증 Wogonin B-ring 유도체의 합성 및 항 염증 생리활성 Wogonin은 5,7-dihydroxy-8-methoxyflavone구조이다. Wogonin과 그 유도체 화합물은 COX-2와 NO 생성 억제력을 나타낸다는 연구보가가 있었다. 그 중에는 wogonin의 B-ring을 다른 구조로 대체하여도 유효한 생리활성이 나타났다는 연구결과가 있었다. 기존의 연구를 바탕으로 우리는 wogonin B-ring 구조를 bioequivalent 구조인 phenyl 그룹으로 변경한 유도체를 설계하고 합성하였다. 핵심어: Wogonin, bioequivalent 그룹, COX-2, NO 생성, 항 염증 생리활성. Oroxylin A의 6-Alkyl (Aryl) Amino 유도체 합성 Oroxylin A는 5, 7-dihydroxy-6-methoxyflavone 구조이다. Oroxylin A와 그 유도체는 COX-2 억제, 항균, 독성 등 생리활성을 나타낸다는 연구결과가 있었다. Oroxylin A는 넓은 범위에 적용될 수 있는 치료제로 인식되고 있다. 다양한 형태의 oroxylin A 유도체는 이미 합성되었고 그 생리활성도 검증되었다. 본 연구에서는 기존의 oroxylin A의 생리활성 연구결과를 참고로 우리는 새로운 시리즈의 유도체를 합성하였다. Oroxylin A 구조 중 6-metoxyl 그룹을 amine 그룹으로 변경하고 다양한 형태의 alky (aryl) chloride 또는 sulphonyl chloride로 6-alky (aryl) amino oroxylin A 유도체 시리즈를 합성하였다. 핵심어: Oroxylin A, Wessley-Moser rearrangement, alky (aryl) chloride와 sulphonyl chloride Unnatural Biflavonoids 설계 및 합성 Biflavonoids는 자연계에 존재하는 화합물로서 C-C 혹은 C-O-C 연결 형태로 이루어진 flavonoid dimmers 구조이며 광범하게 유효한 생리활성을 지니고 있다. 최근 합성한 C-C와 C-O-C결합의 biflavonoid 시리즈 화합물에서는 항염증 생리활성에 효능이 있는 것으로 나타났다. 높은 효능의 신규 항 염증 치료제의 발견, 합성을 목표로 C-NH-C, C-S-C와 C-CH2-C 결합으로 이루어진 다양한 유형의 신규unnatural biflavonoid를 설계하였다. 신규 unnatural biflavonoid 화합물은 C-O-C biflavonoid의 bioisosters 화합물로서 이들에 대해서는 아직 연구보고가 없다. 본 연구의 목표는 신규 unnatural biflavonoid을 합성하고 그 생리활성을 검증하는 것이다. 우리는 Buchwald-Hartwig과 Suzuki 결합반응을 이용한 신규 unnatural biflavonoid 합성경로와 반응에 필요한 주요 중간체 물질들을 얻었다. 이 합성방법을 사용하여 각 유형별 unnatural biflavonoid유도체를 합성할 수 있다. 핵심어: unnatural biflavonoid, bioisoster, Buchwald-Hartwig 반응과 Suzuki 반응.

(THE) IMPACT OF FOREIGN DIRECT INVESTMENT ON NEPAL'S ECONOMIC GROWTH

Gurung, Kapil 아주대학교 국제대학원 2011 국내석사

The federal democratic republic of Nepal is among one of the land locked least developed country. Even though Nepal opened the market to foreign economy long time ago but still the foreign investment and economic transaction has been quiet less. This paper has conducted a descriptive statistical analysis and an empirical analysis to establish the relationship between real gross domestic product and foreign direct investment stock. Using time series econometrics tests for the period 1980-2008, the empirical results indicate the existence of a long-run relationship between the determinants. Even though the relationships are positive the foreign investment has not been consistent through the years. That means definitely Nepal has lot of weakness to attract foreign investment. Some of the main reasons could be political instability, outdated foreign investment law, rigid labor regulation and poor physical infrastructure.

Determinants of Stock Prices : A case study of NEPSE

Gurung, Arun 아주대학교 국제대학원 2010 국내석사

The purpose of the study is to examine the relationship between the macroeconomic variables and the Nepalese stock market. The analysis is based on Ordinary least square method find the co?integration between the set of variables and the stock market. Granger?s causality test was applied to find the direction of causal relationship between the variables and the stock market. The set of variables include boarder money supply, Consumer Price Index, Risk free return ,Exchange rate and the dependent variable is NEPSE(Nepal Stock exchange).The results showed that the variables did have influence but were weak when differentiated at order one. Tags: NEPSE, macroeconomic variables, Cointregration

Electricity Trade between Nepal and India : Problems and Prospects

Gurung, Ritu 아주대학교 국제대학원 2009 국내석사

Abstract Nepal is one of the most trade dependent countries in South Asia. India is the major trading partner of Nepal. Because of geographical proximity, trade with India is more convenient than with any other neighboring countries. Trade relation between India and Nepal has a long history. However, despite of all the trade treaties between two countries, India rejoices the economies of scale where as Nepal is facing a severe trade deficit. According to the “Nepal Quarterly Economic Update” published by Asian Development Bank, total export to India decreased by 5.4% and to total imports from India rose 2.7% this year. Through this thesis, I have tried to analyze the problems of bilateral trade in electricity between two countries and what are the significances of the trade. If Nepal continues trading with India then what would be the future prospects of Nepalese economy. The main objectives of the study are to a) Review of Nepal and India electricity trade relation b) Analysis of the electricity market growth of both nations c) Study of Nepal’s water resources as a comparative advantage d) Analysis of the impact of electricity trade, in particular, in improving trade relation in terms of friendly and historical relationship between the two countries. e). Identify the problems and scope and areas of improvement in Indo-Nepal electricity trade relations. Economic growth of Nepal on the basis of hydroelectricity trade is the primary focus of the study. The electricity deficit has the major role in economy growth of the country. At the present context, Nepal and India electricity trade is based on the intra industry trade theory but it will no longer exists in the future when Nepal will only export the hydroelectricity to India. With available data I have tried to analyze numerically that Nepal and enjoy the electricity export led growth. The projected ten years revenue growth is positive and supports the theory of comparative advantage. The cost analysis of producing electricity by using coal (69%) in India since it is the highly dependent factor for power generation for India and by water resources (63%) in Nepal has been shown. Nepal also has the advantage on cost of production because of run of river type which naturally discourages the construction cost of big dams’ reservoirs. Finally, on the basis of my study, it is illustrated that Nepal and India electricity trade relationship is going to grow stronger with the market demand growth. Nepal can rejoice the comparative advantage in producing hydroelectricity and sell it to the highly potential market of India and gain the economic growth for both the countries. Acknowledgements This dissertation would not have been completed without the guidance and support from my advisor Professor Young-Kyun Song. I am very thankful for his encouragements and suggestions during the preparation of thesis. Similarly, I am also grateful to my committee members Professor Soobok Lee and Professor Min-je Sung for their valuable inputs during the defence of my thesis. It would be unthinkable of me not to thank my father and Dhiraj for always encouraging me to pursue my beliefs and dreams. My brothers and aunty with their faithful and loyal support have been instrumental in my endeavours. All of my loved ones’ absolute support gave me the courage to follow this complicated journey and focus on my goals. My sincere thank goes to Mr. R.P Devkota for his immense support on my thesis completion without which my thesis would have remained incomplete. Likewise, my extended thank goes to Mr. Sachidananda Joshi for his priceless suggestions and feedback on my draft thesis. I would also like to thank Mr. Arun Ranjitkar, Mr. Deepesh Man Shrestha, Mr. Jeevan Baniya, Munna, Rita and to all of those who helped me on the entire process of writing my thesis. I am especially grateful to GSIS, all the professors of GSIS and the entire family of Ajou University for giving me the opportunity to learn in a multicultural environment and giving me an edge to explore the world in a different way. Dedication This dissertation is solely dedicated to my dad and my mom whose love and support made me reach so far. Mom, you are always missed and your memories can never fed away. It is your love and blessing that holds me high when I am low and tormented. Dad, thank you for your faith in me and keeping my moral always up. I am so proud to be your daughter.

Effects of COVID-19 on customer visiting restaurants behavior: A case study of Sokcho, Korea.

GURUNG YUJAL 세종대학교 관광대학원 2024 국내석사

This study was focused on identifying consumers' perceptions of risk and various forms of trust that influence their intention to frequent restaurants during the pandemic. As COVID-19 spread all over the world in 2020, which has primarily impacted the hospitality industry. The COVID-19 pandemic has caused a significant decline in the number of customers at most South Korean eateries, while some have managed to remain open. It all depends on consumer’s mindset and the trust they have in certain business brands. To prevent corona as much as possible, several laws and guidelines were created, including mask-wearing, social separation, groups, and others. The study's findings demonstrate how consumers had faith in restaurant brands, were supportive of the restaurant industry also known as Solidarity with the restaurant sector, found restaurants to be reasonably priced, and the age factor led to disease denial to visit restaurants during those pandemic situations. 164 questionnaires in all were gathered and reviewed. SPSS software was used for data analysis to provide descriptive statistics and gather respondent demographic data. The study's hypothesis was tested using multiple regression analysis and Principle Component Analysis (PCA). The findings of multiple regression revealed that there isn't a statistically significant positive correlation between any of the factors and the behavior of customers visiting restaurants. Hence, the hypothesis of this study was not accepted. Keywords: COVID-19 pandemic, food quality, perceived risk and safety, solidarity with food services

Theoretical Seismograms for 3-D Heterogeneous Models with Variable Surface Curvatures : Torsional Modes

Gyanendra, Gurung 전북대학교 일반대학원 2012 국내박사

Secretion of Membrane Vesicles from Staphylococcus aureus and Their Cytotoxicity to Host Cells : 황색포도알균이 분비한 막소포체에 의한 숙주세포사

Gurung, Mamata 경북대학교 대학원 2012 국내박사

Gram-negative bacteria produce outer membrane vesicles that play a role in the delivery of virulence factors to host cells. OMVs are released by the budding of the Gram-negative bacterial envelope. Secreted OMVs can disseminate far from the cell and impart biological functions on the environment and on other cells, including playing a role in pathogenesis, quorum signaling, nutrient acquisition, biofilm formation and horizontal gene transfer. OMVs are spherical and bilayered nanovesicles with an average diameter of 20-300 nm. They are composed of LPS, phospholipids, outer membrane proteins, periplasmic proteins, cytosolic proteins, PAMPs, and even nucleic acids. OMVs allow enzymes to reach distant targets in a concentrated, protected, and targeted form. An important mechanism of OMV delivery to consider is endocytosis into eukaryotic cells which bypasses the need for heterotypic fusion of a bacterial outer membrane bilayer into a eukaryotic plasma membrane and results in the entry of the entire OMV into the interior of the cell. Several virulence factors which are packaged in OMVs have been well characterised in Gram-negative bacteria. However, little is known about the membrane-derived vesicles produced by Gram-positive bacteria. The present study examined the production of MVs from S. aureus and investigated the delivery of MVs to host cells and subsequent cytotoxicity. Four S. aureus strains tested, two type strains and two clinical isolates, produced spherical nanovesicles during in vitro culture. MVs were also produced during in vivo infection of a clinical S. aureus isolate in a mouse pneumonia model. Proteomic analysis showed that 143 different proteins were identified in the S. aureus-derived MVs The cytoplasmic proteins were the most abundant (67.1%), followed by membrane proteins (19.6%). Pathogenesis-associated proteins, including IgG-binding protein, ferritin, putative ferrichrome-binding lipoprotein precursor, ABC transporter extracellular binding protein, and lipoprotein for high-affinity iron ion transport, and proteins associated with antibiotic resistance, including β-lactamase, penicillin-binding protein 2, and membrane protein oxaA, were found in the S. aureus MVs, which suggest that bacterial effectors packaged in the MVs are directly associated with bacterial pathogenesis. The delivery of bacterial components to host cells via the MVs and delivery mechanism of MVs into host cells were determined and showed that MVs were interacted with the plasma membrane of host cells via a cholesterol-rich membrane microdomain and then delivered their component protein A to host cells within 30 min. These suggest that MVs are a vehicle for the delivery of bacterial components to host cells. Intact S. aureus MVs induced apoptosis of HEp-2 cells in a dose-dependent manner, whereas lysed MVs neither delivered their component into the cytosol of host cells nor induced cytotoxicity. In conclusion, this study is the first report that S. aureus MVs are an important vehicle for delivery of bacterial effector molecules to host cells. 그람 음성균의 외막소포체(outer membrane vesicle)는 숙주 감염시 각종 병원성인자를 숙주세포에 전달하는 중요한 역할을 하는 것으로 알려져 있지만, 그람 양성균에서의 막소포체(membrane vesicle) 분비와 이에 의한 숙주세포 상해에 대해서는 거의 알려져 있지 않다. 본 연구는 다양한 인체 감염증을 일으키는 중요한 병원체인 황색포도알균을 대상으로 막소포체의 분비. 분비된 막소포체의 숙주세포 전달 및 막소포체에 의한 숙주세포 상해 등을 조사하였다. 2주의 황색포도알균 표준균주(ATCC 25923과 ATCC 700699)와 2주의 임상분리균주 (toxic shock syndrome toxin-1 생산 Staphylococcus aureus 103과 methicillin-resistant S. aureus 06ST1048)는 액체배양에서 모두 막소포체를 생산하였다. 황색포도알균의 생체감염에서의 막소포체 분비를 확인하기 위하여 임상분리균주인 S. aureus 06ST1048 균주를 마우스의 기도에 주입한 후 폐렴을 유발하여 폐 조직을 관찰한 결과 황색포도알균의 표면과 균 주위에서 조직에서 균이 분비한 100 nm 정도되는 nanovesicle을 투과전자현미경으로 관찰 할 수 있었다. 황색포도알균이 분비하는 막소포체의 단백을 규명하기 위하여 S. aureus 06ST1048균주가 분비한 막소포체의 단백체를 분석한 결과 총 143 종류의 단백을 동정할 수 있었으며, immunoglobulin G와 결합 할 수 있는 protein A, ß-lactamase 등의 병원성 관련 단백도 함께 동정되었다. 황색포도알균의 막소포체는 숙주세포의 cholesterol 이 풍부한 membrane microdomain인 lipid raft를 통하여 숙주세포 내로 이동하는 것을 protein A 항체를 이용하여 공촛점 현미경으로 확인하였다. 또한 완전한 구조를 가진 막소포체는 숙주세포의 세포질로 이동이 잘 되는 반면, 용해된 막소포체는 숙주세포 내로 이동이 거의 되지 않았다. 막소포체를 HEp-2 세포에 처리한 결과 숙주세포사가 나타났으며, 처리한 막소포체의 농도가 증가함에 따라 숙주세포사도 함께 증가하였다. 그러나 용해된 막소포체는 숙주세포 내로의 이동과 세포사가 모두 일어 나지 않았다. 이러한 결과를 종합하면 황색포도알균은 시험관 배양 및 생체감염에서 막소포체를 분비하며, 다양한 병원성 관련 단백이 함유된 막소포체는 숙주세포막의 lipid raft를 통하여 숙주세포 내로 전달되어 숙주세포사를 유도하였다. 따라서 그람양성균이 분비하는 막소포체는 그람음성균이 분비하는 외막소포체처럼 병원성인자 등 다양한 세균분자를 숙주세포로 전달하는 중요한 매개체임을 본 연구를 통하여 규명하였다.