• Journal of Microbiology and Biotechnology
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• 제32권1호
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• pp.91-98
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• 2022

Tetanus is a potentially fatal public health illness resulted from the neurotoxins generated by Clostridium tetani. C. tetani is not easily culturable and culturing the relevant bacteria from infected wounds has rarely been useful in diagnosis; PCR-based assays can only be conducted at highly sophisticated laboratories. Therefore, a real-time recombinase polymerase amplification assay (Exo-RPA) was constructed to identify the fragments of the neurotoxin gene of C. tetani. Primers and the exo probe targeting the conserved region were designed, and the resulting amplicons could be detected in less than 20 min, with a detection limit of 20 copies/reaction. The RPA assay displayed good selectivity, and there were no cross-reactions with other infectious bacteria common in penetrating wounds. Tests of target-spiked serum and pus extract revealed that RPA is robust to interfering factors and has great potential for further development for biological sample analysis. This method has been confirmed to be reliable for discriminating between toxic and nontoxic C. tetani strains. The RPA assay dramatically improves the diagnostic efficacy with simplified device architecture and is a promising alternative to real-time PCR for tetanus detection.

• 한국양봉학회지
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• 제32권2호
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• pp.119-131
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• 2017

벌집꼬마밑빠진벌레 (Small hive beetle; SHB; Aethina tumida)의 신속검출과 대량 조사를 위하여 SHB 특이 초고속 유전자 증폭법을 개발하였다. 3쌍의 Aethina tumida-특이 유전자 증폭용 프라이머들은 벌집꼬마밑빠진벌레의 미토콘드리아 유전체 중 cytochrome oxidase subunit I (COI) 유전자에 근거하여 선발하였다. 최적화된 초고속 PCR은 $2.1{\times}10^1$ 분자의 작은벌집딱정벌레 COI 유전자를 18분 40초만에 특이적으로 그리고 정량적으로 검출할 수 있었다. 양봉현장의 적용을 위하여, 봉변으로부터 쉽게 DNA를 추출하는 방법을 고안하였으며, 봉변 1g 중 $10^5$ 분자의 벌집꼬마밑빠진벌레 COI 유전자가 존재할 경우(1/1000의 SHB 유충 사체), 10분 이내에 벌집꼬마밑빠진벌레의 존재와 분자적 정량을 마칠 수 있었다. 제안하는 이 실험법이 양봉현장에 널리 적용되어, 벌통 내 벌집꼬마밑빠진벌레의 침입여부 판단, 증식의 수준, 그리고 침입지역의 파악 및 제어에 활용되기를 기대한다.

• 한국습지학회지
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• 제15권2호
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• pp.265-269
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• 2013

본 연구는 아질산성 질소의 간이분석을 위한 기초 연구로 수행되었다. 수중 질소는 부영양화를 유발하는 암모니아, 아질산 및 질산으로 주로 구성되어 있다. 질소화합물을 제거하기 위한 많은 연구들이 진행되었고, 이에 따른 분석기술도 발달했다. 현장 모니터링을 위한 간이 분석방법의 개발은 수질관리를 위해 필요하므로, 본 연구는 아질산성 질소의 현장 분석법 개발을 위해 진행되었다. BCDMA와 바이페닐로 코팅된 PVC 흡착 칼럼을 이용한 아질산성 질소 분석의 기초 연구를 수행하였다. 흡착칼럼내의 흡착제는 4~20 $mgNO_2-N/L$ 범위내에서 발색길이를 나타내었고, 흡광도는 pH의 영향을 받는 것으로 나타났다.

• 대한의생명과학회지
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• 제26권3호
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• pp.149-156
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• 2020

We developed a simple and rapid method for detecting vancomycin resistance genes, such as vanA and vanB, using loop-mediated isothermal amplification (LAMP). To identify not only vancomycin resistance genes, but also the genus Enterococcus, primers were designed for vanA, vanB, and 16S rRNA. Screening for vancomycin susceptibility in Enterococcus was performed using Etest (bioMérieux Inc). The results of the LAMP assay were compared to those of real-time RT-PCR. The optimal conditions for the LAMP assay were 65℃ for 60 min. The detection limits of the LAMP assay for vanA, and vanB were 2 × 10² copies/reaction. Compared to RT-PCR, the sensitivities and specificities of LAMP for 16S rRNA, vanA, and vanB were 100/100%, 100/100%, and 100/100%, respectively. The vanA genotype-vanB phenotype accounted for 57.5% (46/80) of the vancomycin-resistant Enterococci samples collected from 2016 to 2019. In conclusion, the LAMP assay developed in this study showed high sensitivity and specificity for vancomycin-resistant genes. Moreover, due to the simplicity and rapidity of the LAMP assay, its use can be very useful in clinical microbiology laboratories.

• Journal of Microbiology and Biotechnology
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• 제21권3호
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• pp.274-276
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• 2011

Distinction of Bacillus cereus from other closely related bacilli is challenging and new efficient methods are continually demanded. From our previous work on RAPD profiles of bacilli, we found a possibility that B. cereus strains could be distinguished from other bacilli. In this work, RAPD-PCR profiles of B. cereus strains were obtained using a 10-mer (S30) as a primer, and a B. cereus specific 0.91-kb band was produced from all tested strains. The RAPD-PCR procedure also successfully detected B. cereus from spiked cheonggukjang when B. cereus cells were present at more than $10^2$/g sample.

• Tuberculosis and Respiratory Diseases
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• 제85권3호
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• pp.264-272
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• 2022

Background: The current conventional drug susceptibility test (DST) for Mycobacterium tuberculosis (Mtb) takes several weeks of incubation to obtain results. As a rapid method, molecular DST requires only a few days to get the results but does not fully cover the phenotypic resistance. A new rapid method based on the ability of viable Mtb bacilli to hydrolyze fluorescein diacetate to free fluorescein with detection of fluorescent mycobacteria by flow cytometric analysis, was recently developed. Methods: To evaluate this cytometric method, we tested 39 clinical isolates which were susceptible or resistant to isoniazid (INH) or rifampin (RIF), or ethambutol (EMB) by phenotypic or molecular DST methods and compared the results. Results: The susceptibility was determined by measuring the viability rate of Mtb and all the isolates which were tested with INH, RIF, and EMB showed susceptibility results concordant with those by the phenotypic solid and liquid media methods. The isolates having no mutations in the molecular DST but resistance in the conventional phenotypic DST were also resistant in this cytometric method. These results suggest that the flow cytometric DST method is faster than conventional agar phenotypic DST and may complement the results of molecular DST. Conclusion: In conclusion, the cytometric method could provide quick and more accurate information that would help clinicians to choose more effective drugs.

• 식물병연구
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• 제21권3호
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• pp.180-185
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• 2015

In the present work, a pair of primers specific to Tomato yellow leaf curl virus (TYLCV) was designed to allow specific amplification of DNA fragments from any TYLCV isolates using an extensive alignment of the complete genome sequences of TYLCV isolates deposited in the GenBank database. A pair of primers which allows the specific amplification of tomato ${\beta}$-tubulin gene was also analyzed as an internal PCR control. A duplex PCR method with the developed primer sets showed that TYLCV could be directly detected from the leaf crude sap of infected tomato plants. In addition, our developed duplex PCR method could determine PCR errors for TYLCV diagnosis, suggesting that this duplex PCR method with the primer sets is a good tool for specific and sensitive TYLCV diagnosis. The developed duplex PCR method was further verified from tomato samples collected from some farms in Korea, suggesting that this developed PCR method is a simple and reliable tool for rapid and large-scale TYLCV detections in tomato plants.

• Journal of Dairy Science and Biotechnology
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• 제31권1호
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• pp.59-65
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• 2013

Antibiotic residues are undesirable in milk and milk products for a number of reasons. In particular, they can have harmful effects on public health and harm to the manufacturer of the cultured milk products, e.g. MRSA etc. Although government regulatory agencies and the dairy industry have been successful in decreasing the presence of high concentrations of antibiotic residues, violations still occur and lead to contaminated products. As a result, several rapid and reliable methods for the detection of antibiotic residues have been developed, including microbiological and instrumental analysis methods. The conventional methods are time consuming, but recent improvements have allowed for better detection time, sensitivity, and accuracy. An example of an advanced detection instrument is the biosensor, which has several applications in food and environmental science, e.g. food-born pathogen detection, antimicrobial residues etc. In the present review, the recent trends in the methods used to test for antibiotic residues in milk and dairy products, as well as their specific applications, have been discussed.

• 한국수의병리학회지
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• 제6권1호
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• pp.1-5
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• 2002

A1celaphine herpesvirus 1 (AHV-1) is a causative agent of malignant catarrhal fever which is a fatal and a lymphoproliferative syndrome. AHV-1 is a gamma herpesvirus, which induces frequent latent infection and often difficult to detect its antigens or specific nucleic acids because of its low viral copies in the infected tissues. A new method, in situ PCR, is developed for the detection of AHV-1 nucleic acid in this study. Target sequences of AHV-1 open reading frame 50 gene were detected within AHV-1 infected MDBK cells. As compare with other molecular biological methods for the detection of AHV-1, in situ PCR was found to be more sensitive than in situ hybridization and to be less sensitive than nested PCR. However, nested PCR cannot afford to observe and differentiate AHV-1 infected cells. In situ PCR amplifies a target sequence within cells that can be visualized microscopically with increased sensitivity compared to detection by in situ hybridization. In situ PCR has wide applications for sensitive localization of low copy AHV-1 viral sequences within cells to investigate the role of viruses in a variety of clinical conditions and also provide the rapid, sensitive, and specific detection of AHV-1 infection.

• 정보보호학회논문지
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• 제14권4호
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• pp.111-121
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• 2004

인터넷의 급속한 발달과 더불어 네트워크 환경에서의 침입은 빠르게 증가하고 있으며, 그 피해 또한 급격히 증가하고 있다. 최근의 인터넷 공격은 특정 호스트나 네트워크에 대한 피해를 초래할 뿐만 아니라, 네트워크 전반의 성능저하를 유발한다. 기존의 침입 탐지 시스템은 각 지역망 및 특정한 대상 시스템을 보호하기 위한 솔루션들로, 기간망 수준의 실시간 공격 탐지에 적용하기 힘든 문제점을 가지고 있다. 본 논문에서는 네트워크 수준의 실시간 공격탐지를 위하여 각 포트별 트래픽을 대상으로 지수평활법을 적용하는 광대역 네트워크 침입 탐지 기법 제안하였다. 8일간의 기간망의 트래픽 데이터를 대상으로 한 실험에서, 제안한 기법은 공격으로 추정되는 급격한 트래픽의 증가를 적절히 탐지함을 보여주었다.