• Journal of Animal Science and Technology
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• 제64권6호
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• pp.1105-1116
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• 2022

Recently, we reported the robust in vitro three-dimensional (3D) expansion of intestinal organoids derived from adult bovine (> 24 months) samples. The present study aimed to establish an in vitro 3D system for the cultivation of intestinal organoids derived from growing cattle (12 months old) for practical use as a potential alternative to in vivo systems for various purposes. However, very few studies on the functional characterization and 3D expansion of adult stem cells from livestock species compared to those from other species are available. In this study, intestinal crypts, including intestinal stem cells, from the small intestines (ileum and jejunum) of growing cattle were isolated and long-term 3D cultures were successfully established using a scaffold-based method. Furthermore, we generated an apical-out intestinal organoid derived from growing cattle. Interestingly, intestinal organoids derived from the ileum, but not the jejunum, could be expanded without losing the ability to recapitulate crypts, and these organoids specifically expressed several specific markers of intestinal stem cells and the intestinal epithelium. Furthermore, these organoids exhibited key functionality with regard to high permeability for compounds up to 4 kDa in size (e.g., fluorescein isothiocyanate [FITC]-dextran), indicating that apical-out intestinal organoids are better than other models. Collectively, these results indicate the establishment of growing cattle-derived intestinal organoids and subsequent generation of apical-out intestinal organoids. These organoids may be valuable tools and potential alternatives to in vivo systems for examining host-pathogen interactions involving epithelial cells, such as enteric virus infection and nutrient absorption, and may be used for various purposes.

• Journal of Veterinary Science
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• 제21권3호
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• pp.46.1-46.18
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• 2020

Background: High concentrations of particulate matter less than 2.5 ㎛ in diameter (PM2.5) in poultry houses is an important cause of respiratory disease in animals and humans. Pseudomonas aeruginosa is an opportunistic pathogen that can induce severe respiratory disease in animals under stress or with abnormal immune functions. When excessively high concentrations of PM2.5 in poultry houses damage the respiratory system and impair host immunity, secondary infections with P. aeruginosa can occur and produce a more intense inflammatory response, resulting in more severe lung injury. Objectives: In this study, we focused on the synergistic induction of inflammatory injury in the respiratory system and the related molecular mechanisms induced by PM2.5 and P. aeruginosa in poultry houses. Methods: High-throughput 16S rDNA sequence analysis was used for characterizing the bacterial diversity and relative abundance of the PM2.5 samples, and the effects of PM2.5 and P. aeruginosa stimulation on inflammation were detected by in vitro and in vivo. Results: Sequencing results indicated that the PM2.5 in poultry houses contained a high abundance of potentially pathogenic genera, such as Pseudomonas (2.94%). The lung tissues of mice had more significant pathological damage when co-stimulated by PM2.5 and P. aeruginosa, and it can increase the expression levels of interleukin (IL)-6, IL-8, and tumor necrosis factor-α through nuclear factor (NF)-κB pathway in vivo and in vitro. Conclusions: The results confirmed that poultry house PM2.5 in combination with P. aeruginosa could aggravate the inflammatory response and cause more severe respiratory system injuries through a process closely related to the activation of the NF-κB pathway.

• The Korean Journal of Mycology
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• 제25권1호통권80호
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• pp.10-19
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• 1997

Fusarium wilt of radish (Raphanus sativus L.) is caused by the Fusarium oxysporum f. sp. raphani (FOR) which mainly attacks Raphanus spp. The pathogen is a soil-borne and forms chlamydospores in infected plant residues in soil. Infected pathogen colonizes the vascular tissue, leading to necrosis of the vascular tissue. Growth promoting beneficial organisms such as Pseudomonas fluorescens WCS374 (strain WCS374), P. putida RE10 (strain RE10) and Pseudomonas sp. EN415 (strain EN415) were used for microorganisms-mediated induction of systemic resistance in radish against Fusarium wilt. In this bioassy, the pathogens and bacteria were treated into soil separately or concurrently, and mixed the bacteria with the different level of combination. Significant suppression of the disease by bacterial treatments was generally observed in pot bioassy. The disease incidence of the control recorded 46.5% in the internal observation and 21.1% in the external observation, respectively. The disease incidence of P. putida RE10 recorded 12.2% in the internal observation and 7.8% in the external observation, respectively. However, the disease incidence of P. fluorescens WCS374 which was proved to be highly suppressive to Fusarium wilt indicated 45.6% in the internal observation and 27.8% in the external observation, respectively. The disease incidence of P. putida RE10 mixed with P. fluorescens WCS374 or Pseudomonas sp. EN415 was in the range of 10.0-22.1%. On the other hand, the disease incidence of P. putida RE10 mixed with Pseudomonas sp. EN415 was in the range of 7.8-20.2%. The colonization by FOR was observed in the range of $2.4-5.1{\times}10^3/g$ on the root surface and $0.7-1.3{\times}10^3/g$ in the soil, but the numbers were not statistically different. As compared with $3.8{\times}10^3/g$ root of the control, the colonization of infested ROR indicated $2.9{\times}10^3/g$ root in separate treatments of P. putida RE10, and less than $3.8{\times}10^3/g$ root of the control. Also, the colonization of FOR recorded $5.1{\times}10^3/g$ root in mixed treatments of 3 bacterial strains such as P. putida RE10, P. fluorescens WCS374 and Pseudomonas sp. EN415. The colonization of FOR in soil was less than that of FOR in root part. Based on soil or root part, the colonization of ROR didn't indicate a significant difference. The colonization of introduced 3 fluorescent pseudomonads was observed in the range of $2.3-4.0{\times}10^7/g$ in the root surface and $0.9-1.8{\times}10^7/g$ in soil, but the bacterial densities were significantly different. When growth promoting organisms were introduced into the soil, the population of Pseudomonas sp. in the root part treated with P. putida RE10 was similar in number to the control and recorded the low numerical value as compared with any other treatments. The population density of Pseudomonas sp. in the treatment of P. putida RE10 indicated significant differences in the root part, but didn't show significant differences in soil. The population densities of infested FOR and introduced bacteria on the root were high in contrast to those of soil. P. putida RE10 and Pseudomonas sp. EN415 used in this experiment appeared to induce the resistance of the host against Fusarium wilt.

• Tuberculosis and Respiratory Diseases
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• 제41권1호
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• pp.11-19
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• 1994

Background: Mycobacterium tuberculosis is a facultative intracellular pathogen which persists and multiplies within macrophage. Competent cell mediated immunity by cooperation of both T lymphocyte and macrophage of the host is required to kill the Mycobacterium tuberculosis. But a precise understanding of the pathogenesis of tuberculosis infection in pulmonary alveolar macrophage has not been achived. Research on the macrophage's basic microbicidal mechanism has elucidated the importance of oxygen-dependent or oxygen-independent components. Oxygen dependent processing begins with the reduction of oxygen by NADPH oxidase and generation of superoxide. In this study, the oxidative metabolic status of blood monocyte and pulmonary alveolar macrophage in patients with active pulmonary tuberculosis was accessed and compared with that of healthy control subjects to know whether there was a basic difference in superoxide generation by mononuclear cells between two groups. Methods: Pulmonary alveolar macrophage was purified after performing BAL(bronchoalveolar lavage) through the bronchi of infected lesion by plastic adhesion method. Blood monocyte was purified by Ficoll-Hypaque method. Superoxide generation by blood monocyte and pulmonary alveolar macrophage was measured by ferricytochrome-C reduction method after either stimulated with PMA(phorbol myristate acerate) or non-stimulated states. We also measured the effect of pulmonary tuberculosis patient's serum on superoxide generation by monocyte. Results: 1) Generation of superoxide by alveolar macrophage obtained from patients with pulmonary tuberculosis was little higher than those of controls, and PMA enhanced the generation of 2) Generation of superoxide by blood monocyte obtained from patients with pulmonary tuberculosis was little higher than those of control(p>0.05), and PMA more enhanced the generation of superoxide in patientswith pulmonary tuberculosis than those in controls(p<0.02). 3) Patient's serum enhanced the generation of superoxide by blood monocyte obtained from patients with pulmonary tuberculosis and controls, but not in the case of PMA stimulated blood monocyte. Conclusion: The present study suggest that the phenomenon of M.tuberculosis escape the microbicidal action of macrophage was not result of suppressed superoxide generation by blood monocyte and pulmonary alveolar macrophage, rather there might be a factor to stimulate the generation of superoxide by blood monocyte in pulmonary tuberculosis patient serum, but the comparision with effect of control's serum on superoxide generation needs further elucidation.

• Microbiology and Biotechnology Letters
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• 제26권2호
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• pp.122-129
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• 1998

It were reported that antifungal mechanism of Enterobacter cloacae is a volatile ammonia that produced by the strain in soil, and the production of ammonia is related to the bacterial urease activity. A powerful bacterium SH14 against soil-borne pathogen Fusarium solani, which cause root rot of many important crops, was selected from a ginseng pathogen suppressive soil. The strain SH14 was identified as Bacillus subtilis by cultural, biochemical, morphological method, and $API^{circledR}$ test. From several in vitro tests, the antifungal substance that is produced from B. subtilis SH14 was revealed as heat-stable and low-molecular weight antibiotic substance. In order to construct the multifunctional biocontrol agent, the urease gene of Bacillus pasteurii which can produce pathogenes-suppressive ammonia transferred into antifungal bacterium. First, a partial BamH I digestion fragment of plasmid pBU11 containing the alkalophilic B. pasteurii l1859 urease gene was inserted into the BamH I site of pEB203 and expressed in Escherichia coli JM109. The recombinant plasmid was designated as pGU366. The plasmid pGU366 containing urease gene was introduced into the B. subtilis SH14 with PEG-induced protoplast transformation (PIP) method. The urease gene was very stably expressed in the transformant of B. subtilis SH14. Also, the optimal conditions for transformation were established and the highest transformation frequency was obtained by treatment of lysozyme for 90 min, and then addition of 1.5 ${mu}g$/ml DNA and 40% PEG4000. From the in vitro antifungal test against F. solani, antifungal activity of B. subtilis SH14(pGu366) containing urease gene was much higher than that of the host strain. Genetical development of B. subtilis SH14 by transfer of urease gene can be responsible for enhanced biocontrol efficacy with its antibiotic action.

• Clinical and Experimental Pediatrics
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• 제46권2호
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• pp.143-153
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• 2003

Purpose : Infection with Shiga-like toxin (SLT)-producing Escherichia coli, an emerging human pathogen found particularly in young children under 5 years of age, causes a spectrum of illnesses with high morbidity and mortality, ranging from diarrhea to hemorrhagic colitis and hemolytic uremic syndrome. Host mediators play an important role in the pathogenesis of SLT-I toxicity. The experiments described here were designed to investigate the effect of SLT-I on TNF-${\alpha}$ production and to understand the effect of TNF-${\alpha}$ on GB3 expression. We also further examine the relationship between the Gb3 level and the differential susceptibility of cells to the cytotoxic action of SLT-I. Methods : The effect of purified SLT-1 from E. coli O157 : H7 (ATCC 43890) on tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$) production in Raw264.7 cells was investigated. Many mediators regulate endothelial cell membrane expression of the glycolipid globotriaosyleramide (Gb3), which serves as the toxin receptor, suggesting that the host response to the toxin or other bacterial products may contribute to pathogenesis by regulating target cell sensitivity to the toxins. Therefore, the relationships between Gb3 expression and cytotoxicity against SLT-I on three types of cells were evaluated. Results : Detectable levels of TNF-${\alpha}$ were produced as early as six hours after induction and continued to increase during 48 hours by SLT-I. It was also found that Vero cells and dendritic cells (DC2.4 cells) expressed high levels of Gb3, 83% and 68%, respectively, and that Raw264.7 cells had a low level of Gb3 (29%) and appeared refractory to cytotoxicity against SLT-I. Vero cells and DC2.4 cells expressing high levels of Gb3 were highly susceptible to SLT-I. Furthermore, macrophages showed a resistance to SLT-I cytotoxicity, despite the fact that Gb3 expression was enhanced. Conclusion : These results strongly suggest that the expression of Gb3 is necessary but not sufficient to confer sensitivity of macrophages to SLT-I and further underpin the important role of SLT-I and its Gb3 receptors in the pathogenesis of E. coli O157 infection.

• Asian Journal of Turfgrass Science
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• 제20권1호
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• pp.65-76
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• 2006

Scz1, an isolate of Sclerotinia homoeocarpa, was recently reported as a novel pathogen responsible for dollar spot disease in Zoysiagrass, a warm season turfgrass. Scz1 possessed different characteristics on mycelial pigment, mycelial affinity and host pathogenecity compared to those of Scb1, a typical isolate, obtained from creeping bentgrass, a cool season turfgrass. In this study, only three isolates, Scz1, Scz2(another analogous isolate of Sclerotinia homoeocarpa from zoysiagrass), and Scb1, were examined at the molecular level using the internal transcribed spacer(ITS) and random amplified polymorphic DNA(RAPD) assays to verify their identification and genetic variation. As a result of ITS assay, partial ITS sequences of three isolates showed 94-97% similarity with a standardized ITS sequence of S. homoeocarpa registered on BLAST. In the analysis of RAPD, range value through similarity matrix was 0.167 between Scz1 and Scb1, 0.139 between Scz2 and Scb1, and 0.713 between Scz1 and Scz2, respectively. Furthermore, tendegram analysis indicated that Scz1 and Scz2, unlike Scb1, were clustered together as accompanying a high genetic similarity. In in vitro fungicide bioassay, $EC_{50}$ value representing the sensitivity degree to propiconazole, a well-known fungicide for dollar spot disease, was 0.012 ${\mu}g/ml$ for Sczl, 0.003 ${\mu}g/ml$ for Scz2, and 0.030 ${\mu}g/ml$ for Scb1. From all data taken, we concluded that both Scz1 and Scz2 belonged to one group of S. homoeocarpa, since they exhibit the same host range and high level of genetic similarity, whereas their chemical competences to a fungicide were different. This study would provide further approach for assessing genetic diversity of S. homoeocarpa isolates as well as characterizing individual isolate against chemical exposure.

• Research in Plant Disease
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• 제29권4호
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• pp.331-344
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• 2023

More than a year has passed after the 6th edition of 'List of Plant Diseases in Korea (LPDK)' was published in April 2022. The 6.1st edition (2023) of List of Plant Diseases in Korea was made by correcting errors found in the 6th edition of list and adding new diseases reported after the 6th edition. There were 397 corrections from the 6th edition, most of which were simple spelling errors or minor issues. However, 12 diseases were deleted due to duplication or unclear literature proof, and 2 diseases had their diseases' common names changed. We added 158 diseases that were reported before 2021 but not included in the 6th edition, or reported after the 6th edition. After all, 146 diseases were added to the 6,534 diseases in the 6th edition, resulting in a total of 6,680 diseases in the 6.1st edition. Thirty host taxa were also added, increasing the number from 1,390 in the 6th edition to 1,420 in the 6.1st edition. Pathogens were also added to 62 taxa, from 2,400 in the 6th edition, bringing the total to 2,462 taxa in the 6.1st edition. Ultimately, the 6.1st edition (2023) of 'The List of Plant Diseases in Korea' contains 6,680 diseases caused by pathogens of 2,462 taxa on 1,420 hosts. The 6.1st edition is not printed as a book, but is provided through the online 'List of Plant Diseases in Korea' (https://genebank. rda.go.kr/kplantdisease.do).

• 한국균학회소식:학술대회논문집
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• 한국균학회 2015년도 춘계학술대회 및 임시총회
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• pp.59-59
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• 2015

Cryptococcus neoformans causes life-threatening meningoencephalitis in humans, but the treatment of cryptococcosis remains challenging. To develop novel therapeutic targets and approaches, signaling cascades controlling pathogenicity of C. neoformans have been extensively studied but the underlying biological regulatory circuits remain elusive, particularly due to the presence of an evolutionarily divergent set of transcription factors (TFs) in this basidiomycetous fungus. In this study, we constructed a high-quality of 322 signature-tagged gene deletion strains for 155 putative TF genes, which were previously predicted using the DNA-binding domain TF database (http://www.transcriptionfactor.org/). We tested in vivo and in vitro phenotypic traits under 32 distinct growth conditions using 322 TF gene deletion strains. At least one phenotypic trait was exhibited by 145 out of 155 TF mutants (93%) and approximately 85% of the TFs (132/155) have been functionally characterized for the first time in this study. Through high-coverage phenome analysis, we discovered myriad novel TFs that play critical roles in growth, differentiation, virulence-factor (melanin, capsule, and urease) formation, stress responses, antifungal drug resistance, and virulence. Large-scale virulence and infectivity assays in insect (Galleria mellonella) and mouse host models identified 34 novel TFs that are critical for pathogenicity. The genotypic and phenotypic data for each TF are available in the C. neoformans TF phenome database (http://tf.cryptococcus.org). In conclusion, our phenome-based functional analysis of the C. neoformans TF mutant library provides key insights into transcriptional networks of basidiomycetous fungi and ubiquitous human fungal pathogens.

• Clinical and Experimental Pediatrics
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• 제45권6호
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• pp.719-726
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• 2002

Purpose : Neonates in neonatal intensive care units(NICU) have a high risk of acquiring nosocomial infection because of their impaired host defence mechanism and invasive procedures. Nosocomial infection result in considerable morbidity and mortality among neonates. This study was carried out to survey both the epidemiology of nosocomial infection in our NICU and the annual trends of pathogens. Methods : We retrospectively reviewed culture proven nosocomial infection which occurred in our NICU from January 1995 to December 1999. The data included clinical characteristics, site of infection, pathogens, and mortality. Results : Nosocomial infection rates was 9.0 per 100 NICU admissions during the five-year period. Major sites of infection were bloodstream(32.3%), skin(18.4%), endotracheal tube(17.2%), and catheter(10.6%). The most common pathogen was S. aureus(29.9%). and the others were coagulase- negative staphylococci(CONS)(14.8%), Enterobacter(12.4%), and Candida(9.0%). During the five-year period, nosocomial infection rates increased from 9.5 to 11.6 per 100 admissions with the increase of CONS, Candida, Klebsiella, and Acinetobacter baumannii. The infection rate of S. aureus decreased. Multiple episodes of nosocomial infection occurred in 26.1% of all nosocomial infections. Overall bloodstream infection rates were 3.6 per 100 NICU admissions during five years. CONS(29.1%) and S. aureus(27.1%) were the two most common pathogens. Increasing rates of bloodstream infection by CONS, Candida, Klebsiella, and Acinetobacter baumannii were observed. Bloodstream infection related mortality was 11.9%. Conclusion : The predominant pathogens of nosocomial infection in NICU were S. aureus and CONS. Bloodstream infection, the most frequent nosocomial infection, should be a major focus of surveillance and prevention efforts in NICU.