• Mycobiology
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• v.38 no.1
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• pp.33-38
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• 2010

We have successfully applied the nested PCR to detect Cylindrocarpon destructans, a major pathogen causing root rot disease from ginseng seedlings in our former study. The PCR assay, in this study, was used to detect the pathogen from soils. The nested PCR using internal transcribed spacer (ITS) 1, 4 primer set and Dest 1, 4 primer set maintained the specificity in soils containing various microorganisms. For a soil DNA extraction method targeting chlamydospores, when several cell wall disrupting methods were tested, the combination of lyophilization and grinding with glass beads, which broke almost all the chlamydospores, was the strongest. The DNA extraction method which was completed based on the above was simple and time-saving because of exclusion of unnecessary stages, and efficient to apply in soils. As three ginseng fields whose histories were known were analyzed, the PCR assay resulted as our expectation derived from the field information. The direct PCR method will be utilized as a reliable and rapid tool for detecting and monitoring C. destructans in ginseng fields.

• The Plant Pathology Journal
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• v.35 no.5
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• pp.530-537
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• 2019

Fuji, a major apple cultivar in Korea, is susceptible to white rot. Apple white rot disease appears on the stem and fruit; the development of which deteriorates fruit quality, resulting in decreases in farmers' income. Thus, it is necessary to characterize molecular markers related to apple white rot resistance. In this study, we screened for differentially expressed genes between uninfected apple fruits and those infected with Botryosphaeria dothidea, the fungal pathogen that causes white rot. Antimicrobial tests suggest that a gene expression involved in the synthesis of the substance inhibiting the growth of B. dothidea in apples was induced by pathogen infection. We identified seven transcripts induced by the infection. The seven transcripts were homologous to genes encoding a flavonoid glucosyltransferase, a metallothionein-like protein, a senescence-induced protein, a chitinase, a wound-induced protein, and proteins of unknown function. These genes have functions related to responses to environmental stresses, including pathogen infections. Our results can be useful for the development of molecular markers for early detection of the disease or for use in breeding white rotresistant cultivars.

• Toxicological Research
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• v.30 no.2
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• pp.77-81
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• 2014

In contrast to animals, plants do not have a circulatory system as well as mobile immune cells that allow them to protect themselves against pathogens. Instead, plants exclusively depend on the innate immune system to defend against pathogens. As typically observed in the animal innate immunity, plant immune responses are composed of pathogen detection, defense signaling which includes transcriptional reprogramming, and secretion of antimicrobial compounds. Although knowledge on recognition and subsequent signaling of pathogen-derived molecules called elicitors is now expanding, the mechanisms of how these immune molecules are excreted are yet poorly understood. Therefore, current understandings of how plants secrete defense products especially via exocytosis will be discussed in this review.

• Journal of Microbiology and Biotechnology
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• v.23 no.6
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• pp.878-884
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• 2013

Salmonella is a major foodborne pathogen that causes a variety of human diseases. Development of ligands directly and specifically binding to the Salmonella will be crucial for the rapid detection of, and thus for efficient protection from, the virulent bacteria. In this study, we identified a RNA aptamer-based ligand that can specifically recognize Salmonella Typhimurium through SELEX technology. To this end, we isolated and characterized an RNase-resistant RNA aptamer that bound to the OmpC protein of Salmonella Typhimurium with high specificity and affinity ($K_d$ ~ 20 nM). Of note, the selected aptamer was found to specifically bind to Salmonella Typhimurium, but neither to Gram-positive bacteria (Staphylococcus aureus) nor to other Gram-negative bacteria (Escherichia coli O157:H7). This was evinced by aptamer-immobilized ELISA and aptamer-linked precipitation experiments. This Salmonella species-specific aptamer could be useful as a diagnostic ligand against pathogen-caused foodborne sickness.

• Korean Journal Plant Pathology
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• v.14 no.6
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• pp.589-593
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• 1998

DNA amplification by polymerase chain reaction (PCR) was used to specifically detect Plasmodiophora brassicae, causing clubroot of crucifers. On the basis of DNA sequence informations, an oligonucleotide primer set specific for the pathogen was designed form small subunit gene (18S-like) and internal transcribed spacer (ITS) region of ribosomal DNA. Primer ITS 5/PB-C produced an amplification product of approximately 520 bp in length with DNA from P. brassicae. However, no amplification product was produced with DNAs from several soil-borne fungi, Didymella bryoniae and Rhizopus stolonifer. Using these primers, the clubroot pathogen was readily detected from infected roots of crucifers, but not from healthy roots. Southern hybridization analysis further confirmed that the amplification product was originated from P. brassicae.

• Research in Plant Disease
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• v.20 no.1
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• pp.21-24
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• 2014

Clubroot caused by the obligate biotrophic protist Plasmodiophora brassicae Woronin is one of the most damaging diseases of Brassicaceae family. In this study, we developed species-specific primer sets for rapid and accurate detection of P. brassicae. The primer sets developed amplified a specific fragment only from P. brassicae DNA while they did not amplify a band from 10 other soilborne pathogens or from Kimchi cabbage. In sensitivity test, the species-specific primer set ITS1-1/ITS1-2 could work for approximately 10 spores/ml of genomic DNA showing more sensitivity and accuracy than previous methods. With quantitative real-time PCR test, the primer set detected less spores of P. brassicae than before, confirming that the species-specific primer set could be useful for rapid and accurate detection of P. brassicae.

• The Korean Journal of Mycology
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• v.44 no.2
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• pp.103-107
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• 2016

To establish a rapid and accurate detection of Phytophthora pinifolia, which is a quarantine pathogenic fungus in Korea, a species-specific primer was developed based on the ras-related protein (Ypt1) gene. Species-specific primer based on the DNA sequences of Ypt1 gene amplified 193 bp polymerase chain reaction (PCR) product for P. pinifolia. The primer pair yielded the predicted PCR product size exactly in testing with target pathogen DNAs, but not from the other 10 species of Phytophthora and 14 species of other phytopathogenic fungi. The primer pair also showed only the species-specific amplification curve on realtime PCR on target pathogen DNA. The detection sensitivity of real time PCR using species-specific primer pair was 10 to 100 times higher than conventional PCR, with 1 to $10pg/{\mu}L$.

• The Korean Journal of Mycology
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• v.46 no.4
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• pp.467-477
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• 2018

Colletotrichum circinans, an anthracnose pathogen, causes serious damage to onions worldwide. In this study, specific molecular markers were developed to detect C. circinans accurately and quickly with both conventional and real-time PCR methods. The cirTef-F/cirTef-R and cirTu-F/cirTu-R primer sets, which are specific for C. circinans, were constructed by analyzing $tef-1{\alpha}$ and ${\beta}-tubulin$ genes in the fungus. Using the conventional PCR method, 100 pg and 1 ng of fungal DNA could be detected using the cirTef-F/cirTef-R and cirTu-F/cirTu-R sets, respectively. Using the real-time PCR method, 10 pg and 100 pg of fungal DNA could be detected more sensitively with the cirTef-F/cirTef-R and cirTu-F/cirTu-R sets, respectively. Detection of C. circinans from the artificially infected onion seeds was possible by using both conventional and real-time PCR methods and the developed cirTef-F/cirTef-R primer set. The PCR markers specific for C. circinans developed in this study may enhance the efficiency of fungal pathogen detection in imported vegetables and seeds.

• Korean Journal of Veterinary Research
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• v.42 no.3
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• pp.327-333
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• 2002

The study was carried out to examine the distribution and survival rate of Listeria monocytogenes (L monocytogenes) from various source of waters using improved isolation method. In comparision of enrichment media for isolation of L monocytogenes from water, the isolation rate and 50% detection limit of the pathogen were higher in UVM modified Listeria enrichment broth (UVM) than Listeria enrichment broth (LEB). On the other hand, when compared the selective media for isolation of the pathogen from water, the isolation rate was highest in culture at Oxford agar followed by Fraser agar, and LEB agar. In order to improve enrichment method, 100 ml of water samples with 0.1 CFU/ml of L monocytogenes was inoculated into 10 ml of UVM concentrated at 10-fold, and incubated for 24 h at $36^{\circ}C$. Isolated frequency of the pathogens in improved enrichment method completely corresponded with common (filter) method. Of a total mumber of 147 water samples from river, lake and sea, the pathogen was isolated from 1 of 39 (2.6%) river water samples and 1 of 75 (1.3%) sea water samples, but no pathogen was isolated from 33 lake water samples. Serotypes of 2 isolates were identified as type 1. L monocytogenes decreased in number from 7.2-7.4 to 4.2-4.7 log CFU/ml for 1 week poststorage (5 and $20^{\circ}C$), but the pathogens were able to be detected in river and sea water until 8 weeks after storage. However, in tap water, L monocytogenes were decreased to undetectable level after 2 weeks of storage.

• Journal of Animal Science and Technology
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• v.57 no.3
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• pp.7.1-7.7
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• 2015

To meet the increasing demands of specific pathogen free (SPF) minipigs in biomedical researches, 8 pregnant Chinese Wuzhishan minipigs (WZSP) sows with clear background were chosen to obtain SPF WZSP by hysterectomy. At $111{\pm}days2$ of the pregnancy, piglets were aseptically taken out from the sows and artificially suckled for 40 to 45 days in the positive isolators. Then, the piglets defined as F0 were transferred to barrier environment and fed with standard feeds. The original SPF colony was formed for breeding by selected piglets from F0 group of 6-8 months old. Biological characteristics of SPF WZSP were collected and further compared to those of conventional (CV) WZSP, including growth performance, reproductive performance, hematology and blood biochemistry, and major pathogens detection. As a result, 61 F0 piglets were obtained from 8 candidate sows, and 55 out of them survived. After strictly selection, 35 F0 piglets were used to form the original SPF colony, which produced 14 litters of SPF piglets defined as F1. Piglet survival rates, growth performance, and reproductive performance of SPF WZSP were similar to CV WZSP. Some hematology and blood biochemistry parameters showed significant differences between SPF and CV WZSP. Eighteen kinds of pathogens were identified to be free in F0 and F1 SPF colony by repeated pathogen detections. In conclusion, we established a satisfied SPF WZSP colony maintaining original characteristics, free of controlled diseases, and being proved to be a suitable laboratory animal.