• Pediatric Infection and Vaccine
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• 제14권1호
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• pp.69-74
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• 2007

Purpose : Human coronanviruses (hCovs) including hCoV-229E and hCoV-OC43 have been known as etiologic agents of the common colds and were regarded as clinically insignificant agents. However, recent identification of hCoV-NL63 and hCoV-HKU1 in children with lower respiratory tract infections has evoked the clinical concerns about their prevalence and the clinical significance of these hCoVs in children. This study was performed to investigate the prevalence of hCoVs in children with community-acquired pneumonia. Methods : From March 2006 to January 2007, nasopharyngeal specimens collected from children hospitalized with pneumonia, were tested for the presence of common respiratory viruses (respiratory syncytial virus, influenza A, influenza B, parainfluenza viruses, and adenovirus) using multiplex reverse transcriptase polymerase chain reaction (RT-PCR). Human metapneumovirus (hMPV) infection was excluded by nested RT-PCR using primers for the F-gene. To detect the different strains of hCoVs, nested RT-PCR assays specific for hCoVNL63, hCoV-OC43, hCoV-229E, and hCoV-HKU1 were performed. Results : Out of the 217 nasopharyngeal aspirate from children aged under 15 years, respiratory syncytial virus (RSV) was detected in 32 patients, hMPV in 18, human parainfluenza virus in 10, influenza virus A in 2, and adenovirus in 6. HCoVs were detected by RT-PCR in 8 (3.7%) of the 217 patients, hCoV-229E in 1, hCoV-NL63 in 3, and hCoVOC43 in 4 patients. HCoV-HKU1 was not detected in this study population. Conclusion : Recently identified hCoV-NL63 and hCoV-HKU1 seemed to have a little clinical significance in Korean children with severe or hospitalized community-acquired pneumonia.

• The Journal of the Korean Society for Microbiology
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• 제35권2호
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• pp.191-201
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• 2000

The viruses of Herpes Simplex Virus 1 (HSV-1), Herpes Simplex Virus 2 (HSV-2) and Varicella-Zoster virus (VZV) which belong to the alpha herpes subfamily are important human pathogens. When eruptions were not fully developed from these viral infections, clinical diagnosis was not always easy and required virological confirmation test. The above viruses were reactivated in individuals who were compromised in immune competence for one reason or another. Polymerase chain reaction (PCR) enables rapid and sensitive detection of HSV and VZV DNAs. Its sensitivity was largely influenced by choice of primers. Authors conducted a study to detect of those three viruses in human specimens including vesicle fluid and joint fluid and serum using PCR methods. Primers used for this study were the general primer pair GPHV-RU which was known to amplify within the genes enjoying the highest degree of homology between UL15 of HSV and UL42 of VZV. PCR with primers hybridized pair GPHV-RU amplifies a 396 bp with THP-1 and HSV-2 standard strain DNA and 405 bp with VZV standard strain DNA. Restriction enzyme cleavage with HpaII and DdeI were used to detect and distinguish DNAs of THP-1 and HSV-2 and VZV. The purpose of this study was a rapid and easy detection of VZV and THP-1 or HSV-2 from various clinical specimens (vesicle fluid, serum and joint fluid) by PCR method. Used methods were: HSV PCR with primer 1, 2 and HpaII RE digestion; VZV nested PCR; HSV PCR with primer A, Band BssHII RE digestion. 1) In 33 cases (33/42, 78.6%) VZV was detected single or mixed infection from 42 clinical specimens which included vesicle fluid (5), serum form respiratory infected children (10), serum from immune suppressed adult cancer patients (7) and joint fluid from arthritis patients (20). 2) In 20 cases (20/42, 47.6%) HSV was detected singly or mixed infection and 19 of those cases were HSV-2 and 1 case was THP-1. 3) In 19 cases (19/42, 45.2%) VZV was singly detected which included serum from respiratory infected children (6 cases), joint fluid from arthritis patients (9 cases), vesicle fluid (2 cases) and serum form immunosuppressed cancer patients (2 cases). 4) HSV was singly detected in 6 cases (6/42, 14.3%) which included joint fluid from arthritis patients (5 cases) and serum form respiratory infected children (1 cases). 5) 14 cases of VZV and HSV mixed infection (14/42, 33.3%) were detected. They included vesicle fluid (3 cases), serum form immunosuppressed cancer patients (4 cases), serum from respiratory infected children (2 cases) and joint fluid from arthritis patients (5 cases). 6) HSV-1 and HSV-2 detection and typing by HSV PCR with primer A, Band BssHII RE digestion method was more sensitive and the results were easier to detect than on other method.

• Research in Plant Disease
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• 제14권1호
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• pp.15-20
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• 2008

Three isolates of Cucumber mosaic virus (CMV) were isolated from weed hosts showing typical mosaic symptoms, and some properties of the viruses were investigated. CMV isolates, designated as Is-CMV, Jd-CMV and Pla-CMV from Isodon inflexus, Jeffersonia dubia and Phryma leptostachya var. asiatica, respectively, were identified and characterized by biological reaction in several host plants, serological property, dsRNA analysis, reverse transcription-polymerase chain reaction (RT-PCR), restriction fragment-length polymorphism (RFLP). All isolates systemically infected in Nicotiana benthamiana, Cucurbita pepo cv. Black beauty and Cucumis sativus, and did not reveal any differences in these host plants between the isolates. However, remarkable difference in the symptoms was found between the CMVs in Capsicum annuum. Is-CMV induced an asymptomatic symptoms, while Jd-CMV and Pla-CMV produced severe mosaic symptoms in C. annuum plants. In dsRNA analysis, all isolates revealed four major bands with estimated molecular size of 3.4, 3.2, 2.1 and 1.0 kbp. The cDNAs of coat protein gene of the isolates were amplified by RT-PCR using a genus-specific single pair primers that designed to amplify a DNA fragment of approximately ranging from 938 to 966 bp. By restriction mapping analysis using RFLP of the RT-PCR products as well as by serological properties of gel diffusion test, the CMV isolates belong to a typical members of CMV subgroup IA. This is the first report on the occurrence of CMV in the three weed hosts.

• Tuberculosis and Respiratory Diseases
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• 제63권4호
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• pp.331-336
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• 2007

Background: Although pulmonary tuberculosis (TB) is a respiratory disease, the presence of Mycobacterium tuberculosis (Mtb) DNA or Mtb itself has been reported in the peripheral blood (PB) of several patients with pulmonary TB. Additionally, it was recently announced that active pulmonary TB patients donated PB, and that this blood was then transfused to other individuals in Korea. Methods: Sixty-nine patients with bacteriologically-confirmed pulmonary TB (35), non-tuberculous mycobacterial (NTM) lung disease (6), and other lung diseases (28) were enrolled in this study, which was conducted to determine if Mtb DNA could be detected in the PB by PCR. In addition, 10 pulmonary TB patients with high-burden bacilli were also enrolled in this study for the culture of Mtb in PB. Results: PCR detected the presence of Mtb in 22.8% (8/35) of the pulmonary TB patients, in 16.7% (1/6) of the patients with NTM lung disease, and in none of the patients with other diseases (0%). In addition, no Mtb was cultured from the PB of the 10 pulmonary TB patients. Conclusion: Although Mtb DNA was detected in the PB of some patients with pulmonary TB, viable Mtb was not isolated from the PB of those patients, which indicates that patients that viable Mth may not be transmitted via trasfusion of blood of pulmonary TB patients.

• Journal of Food Hygiene and Safety
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• 제30권3호
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• pp.281-288
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• 2015

Vibrio is a genus of Gram-negative, curved, halophilic, and non-spore-forming bacteria. Some of the Vibrio species, such as V. cholerae and V. parahaemolyticus, often contaminate seafood products and occasionally cause human diseases when the seafood products are ingested. A total of 24 Vibrio strains were isolated from shrimp samples on Thiosulphate citrate bile salt sucrose (TCBS) media in this study. All of the 24 isolates were confirmed to belong to the genus Vibrio by using 16S rRNA gene sequence analyses. Vitek 2 system and species-specific polymerase chain reaction (PCR) methods were used to further identify a total of 29 Vibrio strains at the species level, including the 24 shrimp Vibrio isolates and five Vibrio reference strains. The specificities of the two methods to identify Vibrio strains at the species level were compared in this study. The species-specific PCR method was designed to detect five different Vibrio species, such as Vibrio cholerae, Vibrio parahaemolyticus, Vibrio vulnificus, Vibrio alginolyticus, and Vibrio mimicus. From the 24 Vibrio shrimp isolates, the Vitek 2 system method could identify 15 (62.5%) strains as Vibrio species and 7 (29.2%) strains as non-Vibrio species, but could not identify the rest 2 (8.3%) strains. But species-specific PCR method could identify 16 (66.7%) strains as Vibrio species and could not identify the rest 8 (33.3%) strains. Among the 24 Vibrio shrimp strains, these two methods could unanimously identify 7 (7/24, 29.2%) strains (2 V. parahaemolyticus, 4 V. alginolyticus, and 1 V. mimicus). Considering that such different identification results were obtained using the two different methods in this study, identification method for Vibrio species must be carefully chosen.

• Clinical and Experimental Pediatrics
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• 제49권9호
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• pp.977-982
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• 2006

Purpose : There has been an increasing amount of literature concerning the association between Mycoplasma pneumoniae and asthma pathogenesis. Interleukin(IL)-6 stimulates the differentiation of monocytes, and can promote Th2 differentiation and simultaneously inhibit Th1 polarization. IL-8 is a potent chemoattractant and, it has been suggested, has a role in asthma pathogenesis. Nitric oxide (NO) synthesized by airway epithelium may be important in the regulation of airway inflammation and reactivity. Vascular endothelial growth factor(VEGF) has been reported to be a mediator of airway remodeling in asthma. We investigated the effects of M. pneumoniae on IL-6, IL-8, NO and VEGF production in human respiratory epithelial cells. Methods : A549 cells were cultured and inoculated with M. pneumoniae at a dose of 20 cfu/cell. After infection, the presence of M. pneumoniae in epithelial cell cultures was monitored by immunofluorescence and confirmed by polymerase chain reaction(PCR) detection. IL-6, IL-8 and VEGF were determined by an enzyme-linked immunosorbent assay and reverse transcriptase-polymerase chain reaction. NO was measured using the standard Griess reaction. Results : In A549 cells, M. pneumoniaeinduced IL-6, IL-8, NO and VEGF release in time-dependent manners. It also induced mRNA expression of IL-6, IL-8 and VEGF in similar manners. Conclusion : These observations suggest that M. pneumoniae might have a role in the pathogenesis of the allergic inflammation of bronchial asthma.

• Asian Pacific Journal of Cancer Prevention
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• 제16권10호
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• pp.4369-4376
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• 2015

Background: To investigate in-vitro antagonistic effect of low-dose liquiritigenin on gemcitabine-induced capillary leak syndrome (CLS) in pancreatic adenocarcinoma via inhibiting reactive oxygen species (ROS)-mediated signalling pathways. Materials and Methods: Human pancreatic adenocarcinoma Panc-1 cells and human umbilical vein endothelial cells (HUVECs) were pre-treated using low-dose liquiritigenin for 24 h, then added into gemcitabine and incubated for 48 h. Cell viability, apoptosis rate and ROS levels of Panc-1 cells and HUVECs were respectively detected through methylthiazolyldiphenyl-tetrazoliumbromide (MTT) and flow cytometry. For HUVECs, transendothelial electrical resistance (TEER) and transcellular and paracellular leak were measured using transwell assays, then poly (ADP-ribose) polymerase 1 (PARP-1) and metal matrix proteinase-9 (MMP9) activity were assayed via kits, mRNA expressions of p53 and Rac-1 were determined through quantitative polymerase chain reaction (qPCR); The expressions of intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) and PARP-1 were measured via western blotting. Results: Low-dose liquiritigenin exerted no effect on gemcitabine-induced changes of cell viability, apoptosis rate and ROS levels in Panc-1 cells, but for HUVECs, liquiritigenin ($3{\mu}M$) could remarkably elevate gemcitabine-induced decrease of cell viability, transepithelial electrical resistance (TEER), pro-MMP9 level and expression of ICAM-1 and VCAM-1 (p<0.01). Meanwhile, it could also significantly decrease gemcitabine-induced increase of transcellular and paracellular leak, ROS level, PARP-1 activity, Act-MMP9 level, mRNA expressions of p53 and Rac-1, expression of PARP-1 and apoptosis rate (p<0.01). Conclusions: Low-dose liquiritigenin exerts an antagonistic effect on gemcitabine-induced leak across HUVECs via inhibiting ROS-mediated signalling pathways, but without affecting gemcitabine-induced Panc-1 cell apoptosis. Therefore, low-dose liquiritigenin might be beneficial to prevent the occurrence of gemcitabine-induced CLS in pancreatic adenocarcinoma.

• Journal of Food Hygiene and Safety
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• 제27권4호
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• pp.466-472
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• 2012

In this study, ribulose bisphosphate carboxylase (rbcL), RNApolymeraseC (rpoC1), intergenic spacer (psbA-trnH), and second internal transcribed spacer (ITS2) as identification markers for discrimination of P. mirifica in foods were selected. To be primer design, we obtained 719 bp, 520 bp, 348 bp, and 507 bp amplicon using universal primers from selected regions of P. mirifica. The regions of rbcL, rpoC1, and psbA-trnH were not proper for design primers because of high homology about P. mirifica, P. lobata, and B. superba. But, we had designed 4 pairs of oligonucleotide primers from ITS2 gene. Predicted amplicon from P. mirifica were obtained 137 bp and 216 bp using finally designed primers SFI12-miri-6F/SFI12-miri-7R and SFI12-miri-6F/SFI12-miri-8R, respectively. The species-specific primers distinguished P. mirifica from related species were able to apply food materials and processed foods. The developed PCR method would be applicable to food safety management for illegally distributed products in markets and internet shopping malls.

• Korean Journal of Poultry Science
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• 제44권1호
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• pp.1-9
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• 2017

The objective of the present study was to determine the expression of genes associated with lipopolysaccharide (LPS)-induced stressor in two breeds of chickens: the Korean native chicken (KNC) and the White Leghorn chicken (WLH). Forty chickens per breed, aged 40 weeks, were randomly allotted to the control (CON, administered the saline vehicle) and LPS-injected stress groups. Samples were collected at 0 and 48 h post-LPS injection, and total RNA was extracted from the chicken livers for RNA microarray and quantitative real-time polymerase chain reaction (qRT-PCR) analyses. In response to LPS, 1,044 and 1,193 genes were upregulated, and 1,000 and 1,072 genes were downregulated in the KNC and WLH, respectively, using a ${\geq}2$-fold cutoff change. A functional network analysis revealed that stress-related genes were downregulated in both KNC and WLH after LPS infection. The results obtained from the qRT-PCR analysis of mRNA expression of heat shock 90 (HSP90), 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR), activating transcription factor 4 (ATF4), sterol regulatory element-binding protein 1 (SREBP1), and X-box binding protein 1 (XBP1) were confirmed by the results of the microarray analysis. There was a significant difference in the expression of stress-associated genes between the control and LPS-injected KNC and WLH groups. The qRT-PCR analysis revealed that the stress-related $HSP90{\alpha}$ and HMGCR genes were downregulated in both LPS-injected KNC and WLH groups. However, the HSP70 and $HSP90{\beta}$ genes were upregulated only in the LPS-injected KNC group. The results suggest that the mRNA expression of stress-related genes is differentially affected by LPS stimulation, and some of the responses varied with the chicken breed. A better understanding of the LPS-induced infective stressors in chicken using the qRT-PCR and RNA microarray analyses may contribute to improving animal welfare and husbandry practices.

• Journal of Food Hygiene and Safety
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• 제28권2호
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• pp.124-129
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• 2013

The method for detection foods containing allergenic materials by PCR was developed in this study. To detect allergenic raw material from processed food, species specific primer which up to 200bp for PCR product were designed or selected from advanced research. As target materials, 14 items were selected (12 target materials for allergen in Korea, 2 target materials for allergen in foreign countries). The amplicon size for eggs, milk, buckwheat, peanuts, beans, wheat, mackerel, crab, shrimp, pork, peach, tomato, almond, and sesame were confirmed 281, 131, 138, 120, 118, 127, 211, 174, 231, 138, 174, 132, 103, and 220bp, respectively. And any non-specific bands were not detected among each others. Detection method for allergenic material developed in this study could be used to investigate inaccurate goods for allergen labeling or non-intentional contaminant during processed foods manufacturing. In addition, the system will be usefully to detection accurate allergenic raw materials of export for other countries.