• 한국환경농학회:학술대회논문집
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• 한국환경농학회 2009년도 정기총회 및 국제심포지엄
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• pp.157-168
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• 2009

Frequent outbreaks of foodborne illness have been increasing the awareness of agro-food safety. Conventional methods for pathogen detection and identification are labor.intensive and take days to complete. The increasing use of rapid food safety testing is receiving more and more attention. The major reason for this trend is that the food industry requires quick and accurate results. The rapid detection of contaminants in food is critical for ensuring the safety of consumers. Recent advances in technology make detection and identification faster, more sensitive and more specific than traditional method. In this paper, technology trends and recent developments in rapid methods for agro-food safety are discussed.

• 대한수의학회지
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• 제47권4호
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• pp.399-407
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• 2007

Cause of high lethality and dissemination to human being, new development of rapid method for the detection of highly pathogenic Avian Influenza Virus (AIV) is still necessary. For the detection of AIV subtype H5N1, typical pathogenic AIV, new method to confirm sub-typing of this virus is also needed. For the purpose of ultra-rapid detection and sub-typing of hemagglutinin and neuraminidase of AIV, this study was planned. As the results we could demonstrate an ultra-rapid multiplex real-time PCR (URMRT PCR) for the detection of AIV In this study, the URMRT PCR were optimized with synthesized AIV H5- and AIV Nl-specific DNA templates and GenSpector TMC, which is a semiconductor process technology based real-time PCR system with high frequencies of temperature monitoring. Under eight minutes, the amplifications of two AIV subtype-specific PCR products were successfully and independently detected by 30 cycled ultra-rapid PCR, including melting point analysis, from $1{\times}10^3$ copies of mixed template DNA. The URMRT PCR for the detection of AIV H5N 1 developed in this study could be expected to apply not only detections of different AIVs, but also various pathogens. It was also discussed that this kind of the fastest PCR based detection method could be improved by advance of related technology in near future.

• 대한임상검사과학회지
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• 제46권2호
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• pp.64-67
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• 2014

Vancomycin-resistant Enterococci (VRE) are a leading cause of a nosocomial infection. While seven glycopeptide resistance genotypes have been found in Enterococci, vanA and vanB are the most common resistance genotypes. Aims of this study were to detect antibiotic susceptibilities of 23 Enterococcus spp, which broke out in a university hospital by the disk diffusion test, to investigate specific genes of vanA and vanB by conventional and real-time PCR. PCR for vanA and vanB was performed on 23 Enterococci, all 23 were positive for vanA type. This study reports the validation of a simple and rapid VRE detection method that can be easily incorporated into the daily routine of a clinical laboratory. Early detection of VRE strains, including those with susceptibility to Vancomycin, is of paramount clinical importance, as it allows a rapid initiation of strict infection control practices as well as a therapeutic guidance for a confirmed infection. The real-time PCR method is a rapid technique to detect vanA in Enterococci. It is simple and reliable for the rapid characterization of VRE.

• Journal of Veterinary Science
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• 제22권3호
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• pp.40.1-40.12
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• 2021

Background: The microsporidian parasite Nosema ceranae is a global problem in honeybee populations and is known to cause winter mortality. A sensitive and rapid tool for stable quantitative detection is necessary to establish further research related to the diagnosis, prevention, and treatment of this pathogen. Objectives: The present study aimed to develop a quantitative method that incorporates ultra-rapid real-time quantitative polymerase chain reaction (UR-qPCR) for the rapid enumeration of N. ceranae in infected bees. Methods: A procedure for UR-qPCR detection of N. ceranae was developed, and the advantages of molecular detection were evaluated in comparison with microscopic enumeration. Results: UR-qPCR was more sensitive than microscopic enumeration for detecting two copies of N. ceranae DNA and 24 spores per bee. Meanwhile, the limit of detection by microscopy was 2.40 × 10⁴ spores/bee, and the stable detection level was ≥ 2.40 × 10⁵ spores/bee. The results of N. ceranae calculations from the infected honeybees and purified spores by UR-qPCR showed that the DNA copy number was approximately 8-fold higher than the spore count. Additionally, honeybees infected with N. ceranae with 2.74 × 10⁴ copies of N. ceranae DNA were incapable of detection by microscopy. The results of quantitative analysis using UR-qPCR were accomplished within 20 min. Conclusions: UR-qPCR is expected to be the most rapid molecular method for Nosema detection and has been developed for diagnosing nosemosis at low levels of infection.

• Journal of Microbiology and Biotechnology
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• 제19권11호
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• pp.1464-1469
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• 2009

To achieve more accurate and rapid detection of macrolide-lincosamide-streptogramin B resistance genes, erm(A), erm(B), and erm(C), we developed a TaqMan probe-based real-time PCR (Q-PCR) method and compared it with conventional PCR (C-PCR), which is the most widely using erm gene identification method. The detection limit of Q-PCR was 5 fg of genomic DNA or 5-8 CFU of bacterial cells of Staphylococcus aureus. The utilization of Q-PCR might shorten the time to erm detection from 3-4 h to about 50 min. These data indicated that Q-PCR assay appears to be not only highly sensitive and specific, but also the most rapid diagnostic method. Therefore, the appropriate application of the Q-PCR assay will permit rapid and accurate identification of erm genes from clinical and other samples.

• The Plant Pathology Journal
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• 제34권6호
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• pp.499-505
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• 2018

Huanglongbing (HLB, Citrus greening disease) is one of the most devastating diseases that threaten citrus production worldwide. Although HLB presents systemically, low titer and uneven distribution of these bacteria within infected plants can make reliable detection difficult. It was known loop-mediated isothermal amplification (LAMP) method has the advantages of being highly specific, rapid, efficient, and laborsaving for detection of plant pathogens. We developed a new LAMP method targeting gene contained tandem repeat for more rapid and sensitive detection of Candidatus Liberibacter asiaticus (CLas), putative causal agent of the citrus huanglongbing. This new LAMP method was 10 folds more sensitive than conventional PCR in detecting the HLB pathogen and similar to that of real-time PCR in visual detection assay by adding SYBR Green I to mixture and 1% agarose gel electrophoresis. Positive reactions were achieved in reaction temperature 57, 60 and $62^{\circ}C$ but not $65^{\circ}C$. Although this LAMP method was not more sensitive than real-time PCR, it does not require a thermocycler for amplification or agarose gel electrophoresis for resolution. Thus, we expect that this LAMP method shows strong promise as a reliable, rapid, and cost-effective method of detecting the CLas in citrus and can be applied for rapid diagnosis is needed.

• Journal of Biosystems Engineering
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• 제34권4호
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• pp.254-259
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• 2009

Conventional methods for pathogen detection and identification are labor-intensive and take several days to complete. Recently developed biosensors have shown potential for the rapid detection of foodborne pathogens. In this study, an impedimetric biosensor was developed for rapid detection of Salmonella typhimurium. To develop the biosensor, an interdigitated microelectrode (IME) was fabricated by using semiconductor fabrication process. Anti-Salmonella antibodies were immobilized based on either avidin-biotin binding or self assembled monolayer (SAM) on the surface of the IME to form an active sensing layer. To evaluate effect of antibody immobilization methods on sensitivity of the sensor, detection limit of the biosensor was analyzed with Salmonella samples innoculated in phosphate buffered saline (PBS) or food extract. The impedimetric biosensor based on SAM immobilization method produced better detection limit. The biosensor could detect 107 CFU/mL of Salmonella in pork meat extract. This method may provide a simple, rapid, and sensitive method to detect foodborne pathogens.

• Journal of Microbiology and Biotechnology
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• 제17권3호
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• pp.516-519
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• 2007

Enterobacter sakazakii is an emerging food pathogen, which induces severe meningitis and sepsis in neonates and infants, with a high fatality rate. The disease is generally associated with the ingestion of contaminated infant formula. In this study, we describe the development of a real-time PCR protocol to identify E. sakazakii using a TaqMan probe, predicated on the nucleotide sequence data of the 168 rRNA gene obtained from a variety of pathogens. To detect E. sakazakii, four primer sets and one probe were designed. Five strains of E. sakazakii and 28 non-E. sakazakii bacterial strains were used in order to ensure the accuracy of detection. The PCR protocol successfully identified all of the E. sakazakii strains, whereas the 28 non-E. sakazakii strains were not detected by this method. The detection limits of this method for E. sakazakii cells and purified genomic DNA were 2.3 CFU/ assay and 100 fg/assay, respectively. These findings suggest that our newly developed TaqMan real-time PCR method should prove to be a rapid, sensitive, and quantitative method for the detection of E. sakazakii.

• Food Science and Biotechnology
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• 제18권1호
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• pp.89-94
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• 2009

Frequent outbreaks of foodborne illness have been increasing the awareness of food safety. Conventional methods for pathogen detection and identification are labor-intensive and take days to complete. Some immunological, rapid assays are developed, but these assays still require prolonged enrichment steps. Recently developed biosensors have shown potential for the rapid detection of foodborne pathogens. In this study, an impedimetric biosensor was developed for rapid detection of Salmonella entritidis in food sample. To develop the biosensor, an interdigitated microelectrode (IME) was fabricated by using a semiconductor fabrication process. Anti-Salmonella antibodies were immobilized based on neutravidin-biotin binding on the surface of the IME to form an active sensing layer. To evaluate the effect of electrode gap on sensitivity of the sensor, 3 types of sensors with different electrode gap sizes (2, 5, and $10{\mu}m$) were fabricated and tested. The impedimetric biosensor could detect $10^3\;CFU/mL$ of Salmonella in pork meat extract with an incubation time of 5 min. This method may provide a simple, rapid, and sensitive method to detect foodborne pathogens.

• Journal of Dairy Science and Biotechnology
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• 제38권4호
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• pp.169-188
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• 2020

Salmonella spp. is the most common cause of gastrointestinal food poisoning worldwide, and human salmonellosis is mostly caused by the consumption of contaminated food. Therefore, the development of rapid detection methods for Salmoenlla spp. and rapid identification of the source of infection by subtyping are important for the surveillance and monitoring of food-borne salmonellosis. Therefore, this review introduces (1) History and nomenclature of Salmoenlla spp., (2) Epidemiology of Salmoenlla spp., (3) Detection methods for Salmoenlla spp. - conventional culture method, genetic detection method, molecular detection methods, and aptamer, and (4) Subtyping methods for Salmoenlla spp. - pulsed-field gel electrophoresis and repetitive sequence-based polymerase chain reaction (PCR).