• 식물병연구
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• 제24권4호
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• pp.348-352
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• 2018

A multiplex reverse transcription-polymerase chain reaction (mRT-PCR) assay was developed for the detection of Clover yellow vein virus (ClYVV), Peanut mottle virus (PeMoV), and Tomato spotted wilt virus (TSWV), which were recently reported to infect soybean and azuki bean in Korea. Species-specific primer sets were designed for the detection of each virus, and their specificity and sensitivity were tested using mixed primer sets. From among the designed primer sets, two combinations were selected and further evaluated to estimate the detection limits of uniplex, duplex, and multiplex RT-PCR. The multiplex RT-PCR assay could be a useful tool for the field survey of plant viruses and the rapid detection of ClYVV, PeMoV, and TSWV in leguminous plants.

• 한국식물병리학회:학술대회논문집
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• 한국식물병리학회 2003년도 정기총회 및 추계학술발표회
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• pp.150.1-150
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• 2003

A single-step multiplex reverse transcription polymerase chain reaction (RT-PCR) assay was developed for the simultaneous detection of five cucurbit-infecting viruses： cucumber mosaic virus (CMV), watermelon mosaic virus 2 (WMV2), zucchini yellow mosaic virus (ZYMV), cucumber green mottle mosaic virus (CGMMV), and kyuri green mottle mosaic virus (KGMMV). The multiplex RT-PCR provides a simple and rapid method for detecting various viruses in cucurbit plants, which will help diagnose many cucurbit plants at a time.

• The Plant Pathology Journal
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• 제29권3호
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• pp.338-343
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• 2013

A detection system based on a multiplex reverse transcription (RT) polymerase chain reaction (PCR) was developed to simultaneously identify multiple viruses in the lily plant. The most common viruses infecting lily plants are the cucumber mosaic virus (CMV), lily mottle virus (LMoV), lily symptomless virus (LSV). Leaf samples were collected at lily-cultivation facilities located in the Kangwon province of Korea and used to evaluate the detection system. Simplex and multiplex RT-PCR were performed using virus-specific primers to detect single- or mixed viral infections in lily plants. Our results demonstrate the selective detection of 3 different viruses (CMV, LMoV and LSV) by using specific primers as well as the potential of simultaneously detecting 2 or 3 different viruses in lily plants with mixed infections. Three sets of primers for each target virus, and one set of internal control primers were used to evaluate the detection system for efficiency, reliability, and reproducibility.

• Journal of Microbiology and Biotechnology
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• 제19권11호
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• pp.1470-1474
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• 2009

This paper describes the development a of direct multiplex reverse transcription-nested polymerase chain reaction (PCR) method, devised for simultaneous detection and typing of influenza viruses. This method combines the direct reverse transcription reaction without RNA purification with the enhancement of sensitivity and specificity of nested PCR. The method successfully detected three major human influenza viruses: influenza virus A subtype 1 (H1N1) and subtype 3 (H3N2), and influenza B virus (B). The minimum number of virus particles (pfu/ml) necessary for detection in spiked saliva samples was 200 (H1N1), 140 (H3N2), and 4.5 (B). The method's sensitivity and simplicity will be convenient for use in clinical laboratories for the detection and subtyping of influenza and possibly other RNA viruses.

• Clinical and Experimental Pediatrics
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• 제52권12호
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• pp.1358-1363
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• 2009

목 적:호흡기 감염의 증상이 없는 소아들을 대상으로 다중 역전사중합효소연쇄반응법(multiplex reverse transcription-polymerase chain reaction; mRT-PCR)을 이용하여 비인두 상주균의 이환율을 알아보고자 하였다. 방 법:2008년 7월 25일부터 28일까지 33명의 소아들을 대상으로 비강 면봉채취법으로 검체를 채취하였으며, 이들은 검체 채취 당시 심각한 호흡기 감염의 증상이 없었다. 모아진 검체에서 DNA를 추출한 후 multiplex primer set ($Seeplex^{(R)}$ PneumoBacter ACE Detection, Seegene, Seoul, Korea)로 PCR을 진행하였다. 증폭된 반응산물은 2% agarose gel과 전기 영동 자동화 시스템인 screen tape system (Lab901, Scottland, UK)에 각각 전기 영동하여 확인하였다. 결 과:전체 33명의 소아 중 남아는 15명 여아는 18명이었으며, 나이는 3.2세에서 16.3세로 중앙값은 8.2세였다. mRT-PCR 결과 30명(90.9%)의 소아들에서 양성을 보였으며(S. pneumoniae, H. influenzae, C. pneumoniae, B. pertussis), 이들 중 13명(39.4%)에서 2가지 이상의 균이 검출되었다. 균의 종류로는 12명(36.4%)에서는 S. pneumoniae와 H. influenzae, 1명(3.0%)에서는 S. pneumoniae, H. influenzae와 C. pneumoniae이 검출되었다.. 결 론:mRT-PCR은 비인두 상주균의 동정에 있어서 민감도가 높은 방법으로 생각된다. 하지만 비인두 상주균에 대한 PCR 결과가 소아들의 임상 양상과 어느 정도 일치할지에 대해서는 더 많은 연구가 필요할 것으로 생각된다.

• Journal of Plant Biotechnology
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• 제49권3호
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• pp.193-206
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• 2022

Astringent persimmon (Diospyros kaki Thunb.) is an important fruit crop in Korea; it possesses significant medicinal potential. However, knowledge regarding the pathogens affecting this crop, particularly, viruses and viroids, is limited. In the present study, reverse transcription-polymerase chain reaction (RT-PCR) and high-throughput transcriptome sequencing (HTS) were used to investigate the viruses and viroids infecting astringent persimmons cultivated in Korea. A one-step multiplex RT-PCR (mRT-PCR) method for the simultaneous detection of the pathogens was developed by designing species-specific primers and selecting the primer pairs via combination and detection limit testing. Seven of the sixteen cultivars tested were found to be infection-free. The RT-PCR and HTS analyses identified two viruses and one viroid in the infected samples (n = 51/100 samples collected from 16 cultivars). The incidence of single infections (n = 39/51) was higher than that of mixed infections (n = 12/51); the infection rate of the Persimmon cryptic virus was the highest (n = 31/39). Comparison of the monoplex and mRT-PCR results using randomly selected samples confirmed the efficiency of mRT-PCR for the identification of pathogens. Collectively, the present study provides useful resources for developing disease-free seedlings; further, the developed mRT-PCR method can be extended to investigate pathogens in other woody plants.

• Journal of Microbiology and Biotechnology
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• 제22권8호
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• pp.1165-1169
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• 2012

In April 2009, the H1N1 pandemic influenza virus emerged as a novel influenza virus. The aim of this study was to compare the performances of several molecular assays, including conventional reverse transcription polymerase chain reaction (RT-PCR), two real-time reverse transcription (rRT)-PCRs, and two multiplex RTPCRs. A total of 381 clinical specimens were collected from patients (223 men and 158 women), and both the Seeplex RV7 assay and rRT-PCR were ordered on different specimens within one week after collection. The concordance rate for the two methods was 87% (332/381), and the discrepancy rate was 13% (49/381). The positive rates for the molecular assays studied included 93.1% for the multiplex Seeplex RV7 assay, 93.1% for conventional reverse transcription (cRT)-PCR, 89.7% for the multiplex Seeplex Flu ACE Subtyping assay, 82.8% for protocol B rRT-PCR, and 58.6% for protocol A rRT-PCR. Our results showed that the multiplex Seeplex assays and the cRT-PCR yielded higher detection rates than rRT-PCRs for detecting the influenza A (H1N1) virus. Although the multiplex Seeplex assays had the advantage of simultaneous detection of several viruses, they were time-consuming and troublesome. Our results show that, although rRT-PCR had the advantage, the detection rates of the molecular assays varied depending upon the source of the influenza A (H1N1)v virus. Our findings also suggest that rRT-PCR sometimes detected virus in extremely low abundance and thus required validation of analytical performance and clinical correlation.

• Journal of Veterinary Science
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• 제21권2호
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• pp.22.1-22.9
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• 2020

Rabid raccoon dogs (Nyctereutes procyonoides koreensis) have been responsible for animal rabies in South Korea since the 1990s. A recombinant rabies vaccine strain, designated as ERAGS, was constructed for use as a bait vaccine. Therefore, new means of differentiating ERAGS from other rabies virus (RABV) strains will be required in biological manufacturing and diagnostic service centers. In this study, we designed two specific primer sets for differentiation between ERAGS and other RABVs based on mutation in the RABV glycoprotein gene. Polymerase chain reaction analysis of the glycoprotein gene revealed two DNA bands of 383 bp and 583 bp in the ERAGS strain but a single DNA band of 383 bp in the field strains. The detection limits of multiplex reverse transcription polymerase chain reaction (RT-PCR) were 80 and 8 FAID₅₀/reaction for the ERAGS and Evelyn-Rokitnicki-Abelseth strains, respectively. No cross-reactions were detected in the non-RABV reference viruses, including canine distemper virus, parvovirus, canine adenovirus type 1 and 2, and parainfluenza virus. The results of multiplex RT-PCR were 100% consistent with those of the fluorescent antibody test. Therefore, one-step multiplex RT-PCR is likely useful for differentiation between RABVs with and those without mutation at position 333 of the RABV glycoprotein gene.

• 한국수의병리학회:학술대회논문집
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• 한국수의병리학회 2003년도 추계학술대회초록집
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• pp.27-27
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• 2003

Porcine epidemic diarrhea virus (PEDV) and transmissible gastroenteritis virus (TGEV) infections are considered difficult to distinguish clinically and histopathologically. Prompt differentiation between PEDV- and TGEV-associated enteritis would greatly facilitate the management of disease in countries where PEDV and TGEV are epizootic. Rapid differential diagnosis and treatment are crucial to reducing mortality and morbidity from PEDV- and TGEV-induced enteritis in piglets. The objective for this study was to develop a protocol to differentiate between PEDV and TGEV directly from formalin-fixed, paraffin-embedded tissue, using a multiplex reverse transcription-nested polymerase chain reaction (RT-nPCR) assay. (omitted)

• Clinical and Experimental Pediatrics
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• 제59권3호
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• pp.120-125
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• 2016

Purpose: Viral gastroenteritis among children is mainly caused by rotavirus, norovirus, astrovirus, or adenovirus strains. However, changing socioeconomic conditions and a rotavirus vaccination program may be affecting the prevalence of these viral infections. Therefore, we aimed to elucidate the season-specific trends in viral infections for facilitating prophylaxis and surveillance in our region. Methods: We evaluated 345 pediatric patients (203 males, 142 females; age, 1 month to 16 years) who visited the CHA Bundang Medical Center because of gastroenteric symptoms between June 2014 and May 2015. The specimens were simultaneously tested for norovirus, rotavirus, astrovirus, and adenovirus via multiplex reverse transcription polymerase chain reaction. Clinical characteristics of patients were analyzed retrospectively. Results: The most common virus was norovirus, followed by rotavirus, adenovirus, and astrovirus. Of all viral infections, 45.2% occurred mainly between 6 and 24 months of age; in particular, norovirus infection mostly occurred in all age groups except those below 6 months of age, when rotavirus was most prevalent. In addition, seasonal variation was observed, such as norovirus infection from December to February, rotavirus infection from February to April, and adenovirus infection from July to October. Conclusion: Our results showed that the most common cause of acute pediatric viral gastroenteritis had changed from rotavirus to norovirus in our patients, because of effective rotaviral vaccination. We recommend the management of food and personal hygiene in accordance with age or seasons as well as active vaccination for preventing viral gastroenteritis.