• Title/Summary/Keyword: Limit of Detection

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Specific and Sensitive Primers Developed by Comparative Genomics to Detect Bacterial Pathogens in Grains

  • Baek, Kwang Yeol;Lee, Hyun-Hee;Son, Geun Ju;Lee, Pyeong An;Roy, Nazish;Seo, Young-Su;Lee, Seon-Woo
    • The Plant Pathology Journal
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    • v.34 no.2
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    • pp.104-112
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    • 2018
  • Accurate and rapid detection of bacterial plant pathogen is the first step toward disease management and prevention of pathogen spread. Bacterial plant pathogens Clavibacter michiganensis subsp. nebraskensis (Cmn), Pantoea stewartii subsp. stewartii (Pss), and Rathayibacter tritici (Rt) cause Goss's bacterial wilt and blight of maize, Stewart's wilt of maize and spike blight of wheat and barley, respectively. The bacterial diseases are not globally distributed and not present in Korea. This study adopted comparative genomics approach and aimed to develop specific primer pairs to detect these three bacterial pathogens. Genome comparison among target pathogens and their closely related bacterial species generated 15-20 candidate primer pairs per bacterial pathogen. The primer pairs were assessed by a conventional PCR for specificity against 33 species of Clavibacter, Pantoea, Rathayibacter, Pectobacterium, Curtobacterium. The investigation for specificity and sensitivity of the primer pairs allowed final selection of one or two primer pairs per bacterial pathogens. In our assay condition, a detection limit of Pss and Cmn was $2pg/{\mu}l$ of genomic DNA per PCR reaction, while the detection limit for Rt primers was higher. The selected primers could also detect bacterial cells up to $8.8{\times}10^3cfu$ to $7.84{\times}10^4cfu$ per gram of grain seeds artificially infected with corresponding bacterial pathogens. The primer pairs and PCR assay developed in this study provide an accurate and rapid detection method for three bacterial pathogens of grains, which can be used to investigate bacteria contamination in grain seeds and to ultimately prevent pathogen dissemination over countries.

Improvement of the detection limit of rapid detection kit for Salmonella Typhimurium using image analysis system (이미지 분석을 이용한 살모넬라 신속 진단키트의 측정감도 향상)

  • Lee, Sangdae;Kim, Giyoung;Park, Saet-Byeol;Moon, Ji-Hea
    • Korean Journal of Agricultural Science
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    • v.39 no.3
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    • pp.421-425
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    • 2012
  • The objective of this study was to improve the detection limit of rapid detection kit for Salmonella Typhimurium by image analysis system. The rapid detection kit was comprised of four elements: sample pad, conjugate pad, nitrocellulose pad and absorbent pad. Gold nanoparticle and Salmonella antibody were used as a tag and a receptor. Salmonella antibody and goat rabbit IgG antibody were used as test and control lines on nitrocellulose membrane. The color intensity of test line began to increase from $10^5CFU/mL$ of Salmonella sample. A multiple linear regression analysis was employed to explain the relationship between predicted and measured number of Salmonella cells. The developed model could successfully predict the cell number of Salmonella with validation against extra-experimental result.

Rapid and Visual Detection of Vibrio parahaemolyticus in Aquatic Foods Using blaCARB-17 Gene-Based Loop-Mediated Isothermal Amplification with Lateral Flow Dipstick (LAMP-LFD)

  • Hu, Yuan-qing;Huang, Xian-hui;Guo, Li-qing;Shen, Zi-chen;LV, Lin-xue;Li, Feng-xia;Zhou, Zan-hu;Zhang, Dan-feng
    • Journal of Microbiology and Biotechnology
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    • v.31 no.12
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    • pp.1672-1683
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    • 2021
  • Vibrio parahaemolyticus is recognized as one of the most important foodborne pathogens responsible for gastroenteritis in humans. The blaCARB-17 gene is an intrinsic β-lactamase gene and a novel species-specific genetic marker of V. parahaemolyticus. In this study, a loop-mediated isothermal amplification (LAMP) assay combined with a lateral flow dipstick (LFD) was developed targeting this blaCARB-17 gene. The specificity of LAMP-LFD was ascertained by detecting V. parahaemolyticus ATCC 17802 and seven other non-V. parahaemolyticus strains. Finally, the practicability of LAMP-LFD was confirmed by detection with V. parahaemolyticus-contaminated samples and natural food samples. The results showed that the optimized reaction parameters of LAMP are as follows: 2.4 mmol/l Mg2+, 0.96 mmol/l dNTPs, 4.8 U Bst DNA polymerase, and an 8:1 ratio of inner primer to outer primer, at 63℃ for 40 min. The optimized reaction time of the LFD assay is 60 min. Cross-reactivity analysis with the seven non-V. parahaemolyticus strains showed that LAMP-LFD was exclusively specific for V. parahaemolyticus. The detection limit of LAMP-LFD for V. parahaemolyticus genomic DNA was 2.1 × 10-4 ng/μl, corresponding to 630 fg/reaction and displaying a sensitivity that is 100-fold higher than that of conventional PCR. LAMP-LFD in a spiking study revealed a detection limit of approximately 6 CFU/ml, which was similar with conventional PCR. The developed LAMP-LFD specifically identified the 10 V. parahaemolyticus isolates from 30 seafood samples, suggesting that this LAMP-LFD may be a suitable diagnostic method for detecting V. parahaemolyticus in aquatic foods.

MOTION VECTOR DETECTION ALGORITHM USING THE STEEPEST DESCENT METHOD EFFECTIVE FOR AVOIDING LOCAL SOLUTIONS

  • Konno, Yoshinori;Kasezawa, Tadashi
    • Proceedings of the Korean Society of Broadcast Engineers Conference
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    • 2009.01a
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    • pp.460-465
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    • 2009
  • This paper presents a new algorithm that includes a mechanism to avoid local solutions in a motion vector detection method that uses the steepest descent method. Two different implementations of the algorithm are demonstrated using two major search methods for tree structures, depth first search and breadth first search. Furthermore, it is shown that by avoiding local solutions, both of these implementations are able to obtain smaller prediction errors compared to conventional motion vector detection methods using the steepest descent method, and are able to perform motion vector detection within an arbitrary upper limit on the number of computations. The effects that differences in the search order have on the effectiveness of avoiding local solutions are also presented.

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Selective Detection of Campylobacter sp. and Campylobacter jejuni in Meat Food by Polymerase Chain Reaction (PCR을 이용한 육류 내 Campylobacter sp. 및 Campylobacter jejuni의 분리 검출)

  • Joo, Jong-Won;Hong, Kyung-Pyo;Kim, Yong-Hui;Cho, Sang-Buem
    • Journal of the East Asian Society of Dietary Life
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    • v.18 no.5
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    • pp.753-759
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    • 2008
  • The principal objective of this study was to develop the optimum oligonucleotide primers for the simple detection of Campylobacter in food samples. In order to achieve this goal, a variety of oligonucleotide primers were designed via the modification of 16S rDNA, ceuE and mapA sequences of Campylobacter. Through the subsequent analysis of the specificity and sensitivity of primers, two types of oligonucleotide primers, CB4 and CJ1, were selected for Campylobacter genus-specific and C. jejuni species-specific primers, respectively. The detection limit was found to be $10^0{\sim}10^1$ cells per reaction with the prepared cell suspension, however, the sensitivity in the meat samples was less, at $10^1{\sim}10^2$. We suggested that PCR inhibitors such as hemoglobin or immunoglobulin in pork or beef influenced.

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A Study on the Development of Early Acetone Gas Detection to Prevent the Acetone Leakage Accident (아세톤 누출사고 예방을 위한 아세톤 가스 조기감지 기술개발에 관한 연구)

  • Seung Jin Jeon;Youngbo Choi
    • Journal of the Korean Society of Safety
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    • v.38 no.2
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    • pp.30-35
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    • 2023
  • Acetone is a widely used Volatile Organic Compound (VOC) in industries and laboratories. However, acetone affects human health adversely and causes fires and explosions. Early acetone detection and improved personnel training in safety and emergency management are necessary to prevent acetone-related accidents. The multi-VOC acetone detectors used currently have a sensitivity and selectivity limit. In this study, we discovered that Pt-loaded iron oxide (a metal oxide semiconductor) conversely, has high detection and selectivity for very low-levels of acetone gas. The loaded Pt catalyzes the reaction between the sensing materials' surface and the oxygen molecules in the air; this optimizes acetone detection and can decrease acetone-related illnesses, fires and explosions.

A Study on Occupational Environment Assessment Strategies for Respirable Particulate Matter at Coal-Fired Power Plants (석탄화력발전소 호흡성분진 작업환경 평가 전략 사례에 관한 연구)

  • Eun-Seung Lee;Yun-Keun Lee;Dong-Il Shin
    • Journal of Korean Society of Occupational and Environmental Hygiene
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    • v.33 no.3
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    • pp.375-383
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    • 2023
  • Objectives: Coal-fired power plants feature diverse working conditions, including multi-layered employment structures and irregular work cycles due to outsourcing and non-standardized tasks. The current uniform occupational environment measurement systems have limitations in accurately assessing and evaluating these varied conditions. This study aims to propose alternative measurement and assessment strategies to supplement existing methods. Methods: Major domestic coal-fired power plants were selected as the study targets. To prepare for the study and establish strategies, work processes were identified and existing occupational environment measurement results were compared and analyzed. The study proceeded by employing three strategies: specific exposure groups (SEGs) measurement, continuous monitoring, and supplementary measurements, which were then compared and discussed. Results: Previous exposure index evaluations (5,268 cases) indicated that crystalline silica, a type of respirable particulate matter, had detection limits below the threshold (non-detectable) in 82.6% (4,349 cases) of instances. Exposures below 10% of the exposure limit were observed at a very low concentration of 96.1%. Similar exposure group measurements yielded results where detection limits were below the threshold in 38.2% of cases, and exposures below 10% of the limit were observed in 70.6%. Continuous monitoring indicated detection limits below the threshold in 12.6% of cases, and exposures below 10% of the limit were observed in 75.6%. Instances requiring active workplace management accounted for more than 30% of cases, with SEGs at 11.8% (four cases), showing a higher proportion compared to 3.0% (four cases) in continuous monitoring. For coal dust, exposures below 10% of the limit were highest in legal measurements at 90.2% (113 cases), followed by 74.0% (91 cases) in continuous monitoring, and 47.0% (16 cases) in SEGs. Instances exceeding 30% were most prevalent in SEGs at 14.7% (five cases), followed by legal measurements at 5.0% (eight cases), and continuous monitoring at 2.4% (three cases). When examining exposure levels through arithmetic means, crystalline silica was found to be 104.7% higher in SEGs at 0.0088 mg/m3 compared to 0.0043 mg/m3 in continuous monitoring. Coal dust measurements were highest in SEGs at 0.1247 mg/m3, followed by 0.1224 mg/m3 in legal measurements, and 0.0935 mg/m3 in continuous monitoring. Conclusions: Strategies involving SEGs measurement and continuous monitoring can enhance measurement reliability in environments with irregular work processes and frequent fluctuations in working conditions, as observed in coal-fired power plants. These strategies reduce the likelihood of omitting or underestimating processes and enhance measurement accuracy. In particular, a significant reduction in detection limits below the threshold for crystalline silica was observed. Supplementary measurements can identify worker exposure characteristics, uncover potential risks in blind spots of management, and provide a complementary method for legal measurements.

Detection of Cymbidium Mosaic Virus and Odontoglosum Ringspot Virus by ELISA and RT-PCR from Cultivated Orchids in Korea (ELISA와 RT-PCR에 의한 국내재배난에서 심비디움 모자이크 바이러스와 오돈토글로섬 윤문 바이러스이 검정)

  • 박원목;심걸보;김수중;류기현
    • Korean Journal Plant Pathology
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    • v.14 no.2
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    • pp.130-135
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    • 1998
  • This study was carried out to detect cymbidium mosaic potexvirus (CymMV) and odontoglossum ringspot tobamovirus (ORSV) in cultivated orchid plants in Korea. The standard double antibody sandwich enzyme-linked immunosorbent assay (ELISA) and reverse transcription polymerase chain reaction (RT-PCR) were carried out for detection of the viruses in the collected orchid samples. ELISA was suitable for massive-scale diagnostic method for virus detection in orchids. RT-PCR was rapid, time-saving and reliable detective method, and detection limit data showed that RT-PCR was 103 times more sensitive than ELISA. Of the 321 individual orchids representing 5 orchids genera tested by the ELISA, CymMV and ORSV were detected in 15.6% and 22.4%, and mixed infection of the both viruses with 4.9%, respectively. Of the Cymbidium plants tested, cultivated plants showed 52.5% virus infection rate with either CymMV or ORSV and both viruses.

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Survey on Annual Excess Trend for Permissible Exposure Limit of Trichloroethylene (트리클로로에틸렌의 허용기준 적용에 따른 연도별 초과 경향 연구)

  • Kim, Ki-Youn
    • Journal of Korean Society of Occupational and Environmental Hygiene
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    • v.29 no.1
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    • pp.21-26
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    • 2019
  • Objective: The aim of this study is to analyze an excess trend for domestic permissible exposure limit of trichloroethylene based on previous literature review. Materials and Methods: The research object is a trichloroethylene among 13 chemical substances regulated with PEL(Permissible Exposure Limit) in Occupational Safety and Health Act. The information utilized from this study is the work environment measurement data from 2004 to 2013. The highest level among concentration data measured at various workplaces was selected as a representative value through data process. N.D. (Not Detected) data was considered as 1/2 of LOD(Limit Of Detection). Results: Among work environment measurement data between 2004 and 2013, the highest number of excess workplace and excess rate(24 sites & 1.15%) was observed in 2008's data when applying the PEL(50 ppm) of trichloroethylene. When they are compared with the ACGIH's TLV-TWA(10 ppm), 2008's data showed the highest number of excess workplace and excess rate(175 sites & 8.37%). The number of excess workplace and excess rate related to PEL of trichloroethylene showed increase trend in 2005 but tended to decrease after 2008. Conclusions: Based on the results obtained from this study, the exposure level of trichloroethylene in case of domestic workers is not considered as a safe phase regardless of the year of work environment measurement year. Thus, a strictly preventive management in workplace should be provided for reducing exposure level of trichloroethylene.

Simultaneous Analysis of 17 Organophosphorous Pesticides in Blood by Automated Head Space-SPME GC/MS (HS-SPME-GC/MS에 의한 혈액중 17종 유기인계 농약의 동시분석법)

  • Rhee, Jong-Sook;Jung, Jin-Mi;Lee, Han-Sun;Yeom, Hye-Sun;Lee, Sang-Ki;Park, Yoo-Sin;Chung, Hee-Sun
    • YAKHAK HOEJI
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    • v.54 no.6
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    • pp.429-440
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    • 2010
  • HS-SPME-GC/MS was studied and optimized for the determination of 17 orgarnophosphorous pesiticides (OPPs: chlorpyrifos, chlorpyrifos-methyl, demeton-s-methyl, diazinon, dimethoate, EPN, fenitrothion, fenthion, malathion, methidathion, monocrotophos, parathion, phenthoate, phosphamidon, sulfotep, terbufos, triazophos) in blood. Optimum SPME parameters were selected: choice of SPME fiber (85 ${\mu}m$ polyacrylate), pH effect (0.5 N HCl), salt effect ($Na_2SO_4$, 0.2 g; 20%), headspace incubation temperature ($80^{\circ}C$), headspace incubation time (1 min), headspace adsorption time (30 min) and GC desorption time (2 min). These parameters were optimized using HS-SPME autosampler coupled with gas chromatography-mass spectrometry (GC-MS). Method validation was carried out in terms of linearity, limit of detection (LOD), limit of quantitation (LOQ) and recovery in blood. The assay was linear over 0.5~5.0 mg/l ($r^2$=0.955~1.000). Limit of detection (LOD) and limit of quantitation (LOQ) in blood were determined 0.03~0.3 mg/l (S/N=3) and 0.1~1.1 mg/l (S/N=10), respectively. Relative recovery with 0.5, 1 and 5 mg/l (in blood) were 90.8%, 98.5% and 94.1%, respectively. This method will be applied to the determination of the orgarnophosphorous pesticides in postmortem blood. The proposed protocol can be an attractive alternative to be used in routine toxicological analysis.