• Proceedings of the Korean Vacuum Society Conference
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• 2013.08a
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• pp.88-89
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• 2013

A variety of influenza A viruses from animal hosts are continuously prevalent throughout the world which cause human epidemics resulting millions of human infections and enormous industrial and economic damages. Thus, early diagnosis of such pathogen is of paramount importance for biomedical examination and public healthcare screening. To approach this issue, here we propose a fully integrated Rotary genetic analysis system, called Rotary Genetic Analyzer, for on-site detection of influenza A viruses with high speed. The Rotary Genetic Analyzer is made up of four parts including a disposable microchip, a servo motor for precise and high rate spinning of the chip, thermal blocks for temperature control, and a miniaturized optical fluorescence detector as shown Fig. 1. A thermal block made from duralumin is integrated with a film heater at the bottom and a resistance temperature detector (RTD) in the middle. For the efficient performance of RT-PCR, three thermal blocks are placed on the Rotary stage and the temperature of each block is corresponded to the thermal cycling, namely $95^{\circ}C$ (denature), $58^{\circ}C$ (annealing), and $72^{\circ}C$ (extension). Rotary RT-PCR was performed to amplify the target gene which was monitored by an optical fluorescent detector above the extension block. A disposable microdevice (10 cm diameter) consists of a solid-phase extraction based sample pretreatment unit, bead chamber, and 4 ${\mu}L$ of the PCR chamber as shown Fig. 2. The microchip is fabricated using a patterned polycarbonate (PC) sheet with 1 mm thickness and a PC film with 130 ${\mu}m$ thickness, which layers are thermally bonded at $138^{\circ}C$ using acetone vapour. Silicatreated microglass beads with 150~212 ${\mu}L$ diameter are introduced into the sample pretreatment chambers and held in place by weir structure for construction of solid-phase extraction system. Fig. 3 shows strobed images of sequential loading of three samples. Three samples were loaded into the reservoir simultaneously (Fig. 3A), then the influenza A H3N2 viral RNA sample was loaded at 5000 RPM for 10 sec (Fig. 3B). Washing buffer was followed at 5000 RPM for 5 min (Fig. 3C), and angular frequency was decreased to 100 RPM for siphon priming of PCR cocktail to the channel as shown in Figure 3D. Finally the PCR cocktail was loaded to the bead chamber at 2000 RPM for 10 sec, and then RPM was increased up to 5000 RPM for 1 min to obtain the as much as PCR cocktail containing the RNA template (Fig. 3E). In this system, the wastes from RNA samples and washing buffer were transported to the waste chamber, which is fully filled to the chamber with precise optimization. Then, the PCR cocktail was able to transport to the PCR chamber. Fig. 3F shows the final image of the sample pretreatment. PCR cocktail containing RNA template is successfully isolated from waste. To detect the influenza A H3N2 virus, the purified RNA with PCR cocktail in the PCR chamber was amplified by using performed the RNA capture on the proposed microdevice. The fluorescence images were described in Figure 4A at the 0, 40 cycles. The fluorescence signal (40 cycle) was drastically increased confirming the influenza A H3N2 virus. The real-time profiles were successfully obtained using the optical fluorescence detector as shown in Figure 4B. The Rotary PCR and off-chip PCR were compared with same amount of influenza A H3N2 virus. The Ct value of Rotary PCR was smaller than the off-chip PCR without contamination. The whole process of the sample pretreatment and RT-PCR could be accomplished in 30 min on the fully integrated Rotary Genetic Analyzer system. We have demonstrated a fully integrated and portable Rotary Genetic Analyzer for detection of the gene expression of influenza A virus, which has 'Sample-in-answer-out' capability including sample pretreatment, rotary amplification, and optical detection. Target gene amplification was real-time monitored using the integrated Rotary Genetic Analyzer system.

• Pediatric Infection and Vaccine
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• v.25 no.3
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• pp.156-164
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• 2018

Purpose: We aimed to identify the epidemiology and the clinical characteristics of human parainfluenza virus type 4 (HPIV-4) infection compared to HPIVs 1-3 infections in Korean children. Methods: We reviewed medical records of children with HPIV infection who visited Seoul National University Children's Hospital from 2015 to 2017. Detection of respiratory viruses was performed using real time-polymerase chain reaction (rt-PCR), which could differentiate HPIVs 1-4. Diagnosis was classified as a febrile illness, upper respiratory tract infection (URI), croup, bronchiolitis, or pneumonia. The epidemiology, demographic features, and clinical characteristics among HPIV types were compared. The clinical data were analyzed only for the previously healthy children. Results: Of the 472 children diagnosed with HPIV infection, 108 (22.9%) were previously healthy: 24 (22.2%), 19 (17.6%), 39 (36.1%), and 26 (24.1%) in HPIV types 1, 2, 3, and 4, respectively. The median age of children with HPIV-4 infection was 11 (0-195) months: the proportion of children aged < 2 years and 2 to < 5 years were 65.4% and 19.2%, respectively. Clinical diagnoses of HPIV-4 infection were bronchiolitis (38.5%), pneumonia (30.8%), and URI (30.8%). Croup was the most prevalent in HPIV-2 (21.1%) and none in HPIV-4 infection (P=0.026). Hospital admission rates among HPIV types were not significantly different (P>0.05). Conclusions: We observed seasonal peak of HPIV-4 infection in 2015 and 2017. HPIV-4 was a common respiratory pathogen causing lower respiratory tract infection in hospitalized children.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• v.38 no.1
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• pp.14-19
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• 2012

Introduction: This study was conducted to evaluate ssrA expression resulting from adaptation of Escherichia coli (E. coli) to oral pathogens through signal exchange. Materials and Methods: Human cell lines Hep2 and HT29, wild-type E. coli (WT K-12), ssrA knock-out E. coli (${\Delta}K$-12), and Scleropages aureus (S. aureus) were used. A single culture consisting of Hep2, HT29, WT K-12, and ${\Delta}K$-12, and mixed cultures consisting of Hep2 and WT K-12, Hep2 and ${\Delta}K$-12, WT K-12 and S. aureus, ${\Delta}K$-12 and S. aureus, and Hep2, WT K-12, and S. aureus were prepared. For HT29, a mixed culture was prepared with WT K-12 and with WT K-12 and S. aureus. Total RNA was extracted from each culture with the resulting expression of ssrA, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-${\kappa}B$), and p53 was evaluated by Reverse transcription polymerase chain reaction (RT-PCR). Results: The expression of ssrA in a single culture of WT K-12 was lower than that observed in the mixed culture of WT K-12 with S. aureus. Greater ssrA expression was observed in the mixed culture of WT K-12 with Hep2 than in the single culture of WT K-12. The expression of NF-${\kappa}B$ was higher in the mixed culture of Hep2 with ${\Delta}K$-12 than that in the mixed culture of Hep2 with WT K-12, and was lowest in the single culture of Hep2. The expression of ssrA was higher in the mixed culture of WT K-12 with Hep2 and S. aureus than in the mixed culture of WT K-12 with Hep2. Conclusion: These results suggest that ssrA plays an important role in the mechanism of E. coli adaptation to a new environment.

• Clinical and Experimental Pediatrics
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• v.46 no.2
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• pp.143-153
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• 2003

Purpose : Infection with Shiga-like toxin (SLT)-producing Escherichia coli, an emerging human pathogen found particularly in young children under 5 years of age, causes a spectrum of illnesses with high morbidity and mortality, ranging from diarrhea to hemorrhagic colitis and hemolytic uremic syndrome. Host mediators play an important role in the pathogenesis of SLT-I toxicity. The experiments described here were designed to investigate the effect of SLT-I on TNF-${\alpha}$ production and to understand the effect of TNF-${\alpha}$ on GB3 expression. We also further examine the relationship between the Gb3 level and the differential susceptibility of cells to the cytotoxic action of SLT-I. Methods : The effect of purified SLT-1 from E. coli O157 : H7 (ATCC 43890) on tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$) production in Raw264.7 cells was investigated. Many mediators regulate endothelial cell membrane expression of the glycolipid globotriaosyleramide (Gb3), which serves as the toxin receptor, suggesting that the host response to the toxin or other bacterial products may contribute to pathogenesis by regulating target cell sensitivity to the toxins. Therefore, the relationships between Gb3 expression and cytotoxicity against SLT-I on three types of cells were evaluated. Results : Detectable levels of TNF-${\alpha}$ were produced as early as six hours after induction and continued to increase during 48 hours by SLT-I. It was also found that Vero cells and dendritic cells (DC2.4 cells) expressed high levels of Gb3, 83% and 68%, respectively, and that Raw264.7 cells had a low level of Gb3 (29%) and appeared refractory to cytotoxicity against SLT-I. Vero cells and DC2.4 cells expressing high levels of Gb3 were highly susceptible to SLT-I. Furthermore, macrophages showed a resistance to SLT-I cytotoxicity, despite the fact that Gb3 expression was enhanced. Conclusion : These results strongly suggest that the expression of Gb3 is necessary but not sufficient to confer sensitivity of macrophages to SLT-I and further underpin the important role of SLT-I and its Gb3 receptors in the pathogenesis of E. coli O157 infection.

• Korean Journal of Soil Science and Fertilizer
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• v.45 no.4
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• pp.555-559
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• 2012

Manures contain a variety of pathogenic microorganisms that pose a risk to human or animal. On-farm contaminations through contaminated manure were considered likely sources of the pathogen for several outbreak. Pathogenic microorganisms may survive in low numbers during the composting process and subsequently regrow to high levels under favorable conditions. The objective of this study was to investigate effect of temperature on survival of Salmonella enterica and Staphylococcus aureus in livestock manure compost. Commercial livestock manure compost (manure 60%, sawdust 40%) was inoculated with S. enterica and S. aureus. Compost was incubated at four different temperatures (10, 25, 35, and $55^{\circ}C$) for 20 weeks. Samples were taken every week during incubation depending on the given conditions. S. enterica persisted for up to 1 day in livestock manure compost at $55^{\circ}C$, over 140 days at $10^{\circ}C$, 140 days at $25^{\circ}C$, and 70 days at $35^{\circ}C$, respectively. S. aureus persisted for up to 1 day in livestock manure compost at $55^{\circ}C$ and 90 days at $10^{\circ}C$, 70 days at $25^{\circ}C$, and 40 days at $35^{\circ}C$, respectively. The results indicate that S. enterica and S. aureus persisted longer under low temperature condition. S. enterica survived longer than S. aureus at three different temperatures (10, 25, and $35^{\circ}C$). This study will provide useful and practical guidelines to applicators of soil in deciding appropriate handling and time frames for land application of livestock manure compost for sustainable agriculture. Results from these studies provide useful information in identifying manure handling practices to reduce the risk of S. enterica and S. aureus transmission to fresh produce.

• The Korean Journal of Pesticide Science
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• v.18 no.2
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• pp.88-94
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• 2014

Among pre-harvest environmental factors, increasing attention has been paid to the effects of chemical and microbiological factors on fresh produce. The occurrence and prevalence of these factors have been usually studied with regard to the final products at the post-harvesting stage and/or when they are sold in the market. However, the origin and routes of transmission of both factors remain to be clarified. In the present study, we examined the contamination levels of food-borne pathogens and chemical factors such as pesticide residues and heavy metals in 83 and 43 samples, respectively, including various soil, water, and fertilizer samples, as well as post-harvested and processed samples. Among the organic farming samples, only one pesticide, dimethomorph, was detected in the soil sample, however no pesticides were observed from any other samples in organic farming system. Thus, it was thought that might be contaminated from conventional farm land in the vicinity. Whereas many pesticide residues were detected in conventional farming systems such as soil, fertilizer, water, and fresh produce as expected. Furthermore, heavy metals detected from all tested samples did not shown contamination levels higher than the standard limit. We comparatively assessed the levels of contamination by food-borne pathogens on the samples from organic and conventional farming systems, and found aerobic bacteria at approximately 7 log CFU/g, with no significant differences observed between the two systems. Coliforms were present at lower levels than aerobic bacteria. No human pathogens were present among the coliforms detected, indicating that these bacteria are saprophytes without the ability to cause food-borne illnesses. In contrast, among the high-risk food-borne pathogens, only sporadic cells of Bacillus cereus were found on samples of organic farming system. These data extend previous findings that the most prevalent food-borne pathogen is B. cereus and demonstrate that it spreads to whole living plants via soil.

• Journal of Oral Medicine and Pain
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• v.33 no.2
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• pp.159-166
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• 2008

An ideal anti-bacterial medication for oral infection requires selective effect on pathogens causing dental caries and periodontal disease but not on normal flora. In addition, it should be less toxic for human and even for environment. This study was to seek such a natural anti-bacterial medication and thus anti-bacterial effect of Hamamelis virginiana was evaluated. Many recent researches on the anti-bacterial effect of natural plant extract and essential oil have reported that natural products can be used as medication for prevention and restrainment of dental caries, halitosis and periodontitis. It has been also reported that Hamamelis virginiana has anti-bacterial effect on Porphyromonas gingivalis, Fusobacterium nucleatum, Capnocytophaga gingivalis, Veilonella parvula, Eikenella corrodens, Peprostreptococcus micros, and Actinomyces odontolyticus. This study evaluated anti-bacterial effect of Hamamelis virginiana on Streptoccoccus mutans, Haemophylus actinomycetemcomitans, and Klebsiella pneumoniae to expand its anti-bacterial effect on other important oral pathogens and eventually to develop its oral care products or apply to clinical purpose. In this study, anti-bacterial tests for antibiotic disk susceptibility, minimal inhibitory concentration and minimal bactericidal concentration were performed to evaluate anti-bacterial effect of Hamamelis virginiana against Streptoccoccus mutans, Haemophylus actinomycetemcomitans, and Klebsiella pneumoniae. The results showed that Hamamelis virginiana has anti-bacterial effect on all pathogen strains tested in this study and furthermore Hamamelis virginiana possesses bactericidal effect other than bacteriostatic effect on Streptoccoccus mutans, Haemophylus actinomycetemcomitans, Klebsiella pneumoniae. This study indicates that a natural anti-bacterial medication for oral diseases can be developed using Hamamelis virginiana.

• Journal of Plant Biotechnology
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• v.45 no.3
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• pp.171-182
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• 2018

Papaya (Carica papaya L.) is one of the crops widely planted in tropical and subtropical areas. The papaya fruit has low calories and are plentiful in vitamins A and C and in minerals. A major problem in papaya production is a plant disease caused by the papaya ringspot virus (PRSV). The first PRSV-resistant GM papaya expressing a PRSV coat protein gene was developed by USA scientists in 1992. The first commercial GM papaya cultivars derived from the event was approved by the US government in 1997. Development of transgenic papayas has been focused on vaccine production and limited agricultural traits, including insect and pathogen resistance, long shelf life, and aluminum and herbicide tolerance. Approximately 17 countries, including the USA and China, produced transgenic papayas and/or commercialized them, which provoked studies on biosafety assessment and development of GM-detection technologies. For the biosafety assessment of potential effects on human health, effects of long-term feeding to model animals have been studied in terms of toxicity and allergenicity. Studies on environmental safety assessment include influence on soil-microbial biodiversity and transfer to soil bacteria of GM selection markers. Many countries, such as Korea, the European Union, and Japan, that have strict regulations for GM crops have serious concerns about unintended introduction of GM cultivars and food commodities using unauthorized GM crops. Transgene- and/or GM event-specific molecular markers and technologies for genomics-based detection of unauthorized GM papaya have been developed and have resulted in the robust detection of GM papayas.

• Korean Journal of Ecology and Environment
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• v.38 no.2 s.112
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• pp.225-236
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• 2005

The protection of public health In wastewater reclamation and reuse is one of the most important issues. Monitoring data of Escherichia coli were collected from paddy rice plots in 2003 and 2004 experiments. Five treatments were used and each one was triplicated to evaluate the changes of E. coli: surface water, biofilter effluent (secondary level), UV-disinfected water and pond treatment. Microbial risk was quantified to assess human health risk by exposure to E. coli in paddy rice plots, which were irrigated with reclaimed wastewater. Beta-Poisson model was used to estimate the microbial risk of pathogen ingestion that may occur to farmer and neighbor children. Monte-Carlo analysis (10,000 trials) was used to estimate the risk characterization of uncertainty. In the following analysis, two scenarios were related to the reduction of risk against direct ingestion and exposure times. Scenarios A and B were assumed that the risk was 1,000 and 10,000 times lower than direct ingestion.'Golfers were assumed to be 0.001 L of reclaimed water by contact with balls and their cloths. Opportunity of contact in paddy rice field with pathogens was more frequent than handing golf balls, because of agricultural activity was practiced in ponded water in paddy rice culture. As a result of microbial risk assessment using total data of experimental period, risk value of E. coli in 2003 and 2004 experiment ranged from $10^{-5}$ to $10^{-8}$ and $10^{-4}$ to $10^{-8}$, respectively. The risk values in biofilter effluent irrigation was the highest, which is $10^{-4}$ in 2003 and $10^{-5}$ in 2004 experiments with scenario A. Ranges of $10^{-6}$ to $10^{-8}$ were considered at reasonable levels of risk for communicable disease transmission from environmental exposure and the risk value above $10^{-4}$ was considered to be attributable to the risk of infection. Irrigation with UV-disinfected water in the paddy field during the agricultural Period showed significantly lower microbial risk than others, and their levels of risk value were within the range of actual paddy rice field with surface water.

• Korean Journal of Microbiology
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• v.43 no.3
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• pp.186-192
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• 2007

This study was conducted to survey the hygienic status of chicken meats on the microbial levels, which were collected from poultry processing plants located in the local provinces in nationwide including the JeJu island (n=15) in 1997. In particular, Salmonella spp., Campylobacter jejuni, and Listeria monocytogenes, which were retarded as one of the most important entero-pathogens relating to food home illness from poultry, were investigated on their isolation frequency including the other pathogens related on the food-borne illness. A total of 115 processed chickens were submitted on the present study. In general, the bacterial contamination frequency showed more or less lower $(10{\sim}100 cells)$ than those of sold on the retail and super markets and department stores because of lacking of cross-contamination incidences, depending on the total cells, Coliforms and Staphylococcal cells count. While, Salmonella species, Campylobacter jejuni, Listeria monocytogenes, and coagulase positive Staphylococcus aureus isolation frequency of chicken meats from slaughter houses were 58.3%, 37.4%, 43.5%, and 30.4%, in order. But the present microbial isolation data were a little lower levels than those of sold on the retail and super markets and famous department stores in Seoul and GyeongGi province at the same period. It seemed that the cross-contamination problems (including the human, environmental and instrumental factors) during the marketing stage (after the last processing procedure; rinsing step) had the major roles on the increasing of the microbial contamination frequency on the chicken meats after the slaughter houses.