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vfr, A Global Regulatory Gene, is Required for Pyrrolnitrin but not for Phenazine-1-carboxylic Acid Biosynthesis in Pseudomonas chlororaphis G05

  • Wu, Xia;Chi, Xiaoyan;Wang, Yanhua;Zhang, Kailu;Kai, Le;He, Qiuning;Tang, Jinxiu;Wang, Kewen;Sun, Longshuo;Hao, Xiuying;Xie, Weihai;Ge, Yihe
    • The Plant Pathology Journal
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    • v.35 no.4
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    • pp.351-361
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    • 2019
  • In our previous study, pyrrolnitrin produced in Pseudomonas chlororaphis G05 plays more critical role in suppression of mycelial growth of some fungal pathogens that cause plant diseases in agriculture. Although some regulators for pyrrolnitrin biosynthesis were identified, the pyrrolnitrin regulation pathway was not fully constructed. During our screening novel regulator candidates, we obtained a white conjugant G05W02 while transposon mutagenesis was carried out between a fusion mutant $G05{\Delta}phz{\Delta}prn::lacZ$ and E. coli S17-1 (pUT/mini-Tn5Kan). By cloning and sequencing of the transposon-flanking DNA fragment, we found that a vfr gene in the conjugant G05W02 was disrupted with mini-Tn5Kan. In one other previous study on P. fluorescens, however, it was reported that the deletion of the vfr caused increased production of pyrrolnitrin and other antifungal metabolites. To confirm its regulatory function, we constructed the vfr-knockout mutant $G05{\Delta}vfr$ and $G05{\Delta}phz{\Delta}prn::lacZ{\Delta}vfr$. By quantifying ${\beta}-galactosidase$ activities, we found that deletion of the vfr decreased the prn operon expression dramatically. Meanwhile, by quantifying pyrrolnitrin production in the mutant $G05{\Delta}vfr$, we found that deficiency of the Vfr caused decreased pyrrolnitrin production. However, production of phenazine-1-carboxylic acid was same to that in the wild-type strain G05. Taken together, Vfr is required for pyrrolnitrin but not for phenazine-1-carboxylic acid biosynthesis in P. chlororaphis G05.

Bleomycin Inhibits Proliferation via Schlafen-Mediated Cell Cycle Arrest in Mouse Alveolar Epithelial Cells

  • Jang, Soojin;Ryu, Se Min;Lee, Jooyeon;Lee, Hanbyeol;Hong, Seok-Ho;Ha, Kwon-Soo;Park, Won Sun;Han, Eun-Taek;Yang, Se-Ran
    • Tuberculosis and Respiratory Diseases
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    • v.82 no.2
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    • pp.133-142
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    • 2019
  • Background: Idiopathic pulmonary fibrosis involves irreversible alveolar destruction. Although alveolar epithelial type II cells are key functional participants within the lung parenchyma, how epithelial cells are affected upon bleomycin (BLM) exposure remains unknown. In this study, we determined whether BLM could induce cell cycle arrest via regulation of Schlafen (SLFN) family genes, a group of cell cycle regulators known to mediate growth-inhibitory responses and apoptosis in alveolar epithelial type II cells. Methods: Mouse AE II cell line MLE-12 were exposed to $1-10{\mu}g/mL$ BLM and $0.01-100{\mu}M$ baicalein (Bai), a G1/G2 cell cycle inhibitor, for 24 hours. Cell viability and levels of pro-inflammatory cytokines were analyzed by MTT and enzyme-linked immunosorbent assay, respectively. Apoptosis-related gene expression was evaluated by quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR). Cellular morphology was determined after DAPI and Hoechst 33258 staining. To verify cell cycle arrest, propidium iodide (PI) staining was performed for MLE-12 after exposure to BLM. Results: BLM decreased the proliferation of MLE-12 cells. However, it significantly increased expression levels of interleukin 6, tumor necrosis factor ${\alpha}$, and transforming growth factor ${\beta}1$. Based on Hoechst 33258 staining, BLM induced condensation of nuclear and fragmentation. Based on DAPI and PI staining, BLM significantly increased the size of nuclei and induced G2/M phase cell cycle arrest. Results of qRT-PCR analysis revealed that BLM increased mRNA levels of BAX but decreased those of Bcl2. In addition, BLM/Bai increased mRNA levels of p53, p21, SLFN1, 2, 4 of Schlafen family. Conclusion: BLM exposure affects pulmonary epithelial type II cells, resulting in decreased proliferation possibly through apoptotic and cell cycle arrest associated signaling.

Analysis of the Genome Sequence of Strain GiC-126 of Gloeostereum incarnatum with Genetic Linkage Map

  • Jiang, Wan-Zhu;Yao, Fang-Jie;Fang, Ming;Lu, Li-Xin;Zhang, You-Min;Wang, Peng;Meng, Jing-Jing;Lu, Jia;Ma, Xiao-Xu;He, Qi;Shao, Kai-Sheng;Khan, Asif Ali;Wei, Yun-Hui
    • Mycobiology
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    • v.49 no.4
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    • pp.406-420
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    • 2021
  • Gloeostereum incarnatum has edible and medicinal value and was first cultivated and domesticated in China. We sequenced the G. incarnatum monokaryotic strain GiC-126 on an Illumina HiSeq X Ten system and obtained a 34.52-Mb genome assembly sequence that encoded 16,895 predicted genes. We combined the GiC-126 genome with the published genome of G. incarnatum strain CCMJ2665 to construct a genetic linkage map (GiC-126 genome) that had 10 linkage groups (LGs), and the 15 assembly sequences of CCMJ2665 were integrated into 8 LGs. We identified 1912 simple sequence repeat (SSR) loci and detected 700 genes containing 768 SSRs in the genome; 65 and 100 of them were annotated with gene ontology (GO) terms and KEGG pathways, respectively. Carbohydrate-active enzymes (CAZymes) were identified in 20 fungal genomes and annotated; among them, 144 CAZymes were annotated in the GiC-126 genome. The A mating-type locus (MAT-A) of G. incarnatum was located on scaffold885 at 38.9 cM of LG1 and was flanked by two homeodomain (HD1) genes, mip and beta-fg. Fourteen segregation distortion markers were detected in the genetic linkage map, all of which were skewed toward the parent GiC-126. They formed three segregation distortion regions (SDR1-SDR3), and 22 predictive genes were found in scaffold1920 where three segregation distortion markers were located in SDR1. In this study, we corrected and updated the genomic information of G. incarnatum. Our results will provide a theoretical basis for fine gene mapping, functional gene cloning, and genetic breeding the follow-up of G. incarnatum.

A research on the emotion classification and precision improvement of EEG(Electroencephalogram) data using machine learning algorithm (기계학습 알고리즘에 기반한 뇌파 데이터의 감정분류 및 정확도 향상에 관한 연구)

  • Lee, Hyunju;Shin, Dongil;Shin, Dongkyoo
    • Journal of Internet Computing and Services
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    • v.20 no.5
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    • pp.27-36
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    • 2019
  • In this study, experiments on the improvement of the emotion classification, analysis and accuracy of EEG data were proceeded, which applied DEAP (a Database for Emotion Analysis using Physiological signals) dataset. In the experiment, total 32 of EEG channel data measured from 32 of subjects were applied. In pre-processing step, 256Hz sampling tasks of the EEG data were conducted, each wave range of the frequency (Hz); Theta, Slow-alpha, Alpha, Beta and Gamma were then extracted by using Finite Impulse Response Filter. After the extracted data were classified through Time-frequency transform, the data were purified through Independent Component Analysis to delete artifacts. The purified data were converted into CSV file format in order to conduct experiments of Machine learning algorithm and Arousal-Valence plane was used in the criteria of the emotion classification. The emotions were categorized into three-sections; 'Positive', 'Negative' and 'Neutral' meaning the tranquil (neutral) emotional condition. Data of 'Neutral' condition were classified by using Cz(Central zero) channel configured as Reference channel. To enhance the accuracy ratio, the experiment was performed by applying the attributes selected by ASC(Attribute Selected Classifier). In "Arousal" sector, the accuracy of this study's experiments was higher at "32.48%" than Koelstra's results. And the result of ASC showed higher accuracy at "8.13%" compare to the Liu's results in "Valence". In the experiment of Random Forest Classifier adapting ASC to improve accuracy, the higher accuracy rate at "2.68%" was confirmed than Total mean as the criterion compare to the existing researches.

Molecular detection of blaVIM, blaBIC, blaKPC, and blaSIM genes from isolated bacteria in retail meats (육류용 고기로부터 분자진단을 이용한 항생제내성 유전자 양상)

  • Hwang, You Jin
    • Journal of the Korea Academia-Industrial cooperation Society
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    • v.22 no.6
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    • pp.413-419
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    • 2021
  • The purpose of this study was to investigate the ability to treat and prevent infection by multiple Gram-negative bacterial pathogens as a last choice option in the treatment of serious infections in clinical settings. The global spread of extended-spectrum 𝛽-lactamases (ESBLs) and/or carbapenemases in microorganisms are of enormous concern to health services because they are often associated with multi-drug resistance which significantly restricts the antibiotic treatment options. In this study, the antimicrobial resistance profiles of bacteria isolated from South Korean market-derived meat samples were determined by the disc diffusion method. PCR was used to detect the presence of antibiotic resistance genes and ESBL producing genes. In total, we tested 181 isolated colonies from 36 market-derived meat samples. Single PCR and DNA sequencing results revealed that genes blaVIM, blaBIC, blaKPC, and blaSIM were present in the bacteria isolated from retail meat. The bacteria in the meat were separately sequenced and based on alignment, four different bacteria were identified. These findings suggest that bacteria found in retail meats are a reservoir for the spreading of ESBL blaVIM, blaBIC, blaKPC, and blaSIM resistance genes and bacteria strains.

Risk assessment for norovirus foodborne illness by raw oyster (Ostreidae) consumption and economic burden in Korea

  • Yoo, Yoonjeong;Oh, Hyemin;Lee, Yewon;Sung, Miseon;Hwang, Jeongeun;Zhao, Ziwei;Park, Sunho;Choi, Changsun;Yoon, Yohan
    • Fisheries and Aquatic Sciences
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    • v.25 no.5
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    • pp.287-297
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    • 2022
  • The objective of this study was to evaluate the probability of norovirus foodborne illness by raw oyster consumption. One hundred fifty-six oyster samples were collected to examine the norovirus prevalence. The oyster samples were inoculated with murine norovirus and stored at 4℃-25℃. A plaque assay determined norovirus titers. The norovirus titers were fitted with the Baranyi model to calculate shoulder period (h) and death rate (Log PFU/g/h). These kinetic parameters were fitted to a polynomial model as a function of temperature. Distribution temperature and time were surveyed, and consumption data were surveyed. A dose-response model was also searched through literature. The simulation model was prepared with these data in @RISK to estimate the probability of norovirus foodborne. One sample of 156 samples was norovirus positive. Thus, the initial contamination level was estimated by the Beta distribution (2, 156), and the level was -5.3 Log PFU/g. The developed predictive models showed that the norovirus titers decreased in oysters under the storage conditions simulated with the Uniform distribution (0.325, 1.643) for time and the Pert distribution (10, 18, 25) for temperature. Consumption ratio of raw oyster was 0.98%, and average consumption amount was 1.82 g, calculated by the Pert distribution [Pert {1.8200, 1.8200, 335.30, Truncate (0, 236.8)}]. 1F1 hypergeometric dose-response model [1 - (1 + 2.55 × 10-3 × dose)-0.086] was appropriate to evaluate dose-response. The simulation showed that the probability of norovirus foodborne illness by raw oyster consumption was 5.90 × 10-10 per person per day. The annual socioeconomic cost of consuming raw oysters contaminated with norovirus was not very high.

Analysis of 16S rRNA gene sequencing data for the taxonomic characterization of the vaginal and the fecal microbial communities in Hanwoo

  • Choi, Soyoung;Cha, Jihye;Song, Minji;Son, JuHwan;Park, Mi-Rim;Lim, Yeong-jo;Kim, Tae-Hun;Lee, Kyung-Tai;Park, Woncheoul
    • Animal Bioscience
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    • v.35 no.11
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    • pp.1808-1816
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    • 2022
  • Objective: The study of Hanwoo (Korean native cattle) has mainly been focused on meat quality and productivity. Recently the field of microbiome research has increased dramatically. However, the information on the microbiome in Hanwoo is still insufficient, especially relationship between vagina and feces. Therefore, the purpose of this study is to examine the microbial community characteristics by analyzing the 16S rRNA sequencing data of Hanwoo vagina and feces, as well as to confirm the difference and correlation between vaginal and fecal microorganisms. As a result, the goal is to investigate if fecal microbiome can be used to predict vaginal microbiome. Methods: A total of 31 clinically healthy Hanwoo that delivered healthy calves more than once in Cheongju, South Korea were enrolled in this study. During the breeding season, we collected vaginal and fecal samples and sequenced the microbial 16S rRNA genes V3-V4 hypervariable regions from microbial DNA of samples. Results: The results revealed that the phylum-level microorganisms with the largest relative distribution were Firmicutes, Actinobacteria, Bacteroidetes, and Proteobacteria in the vagina, and Firmicutes, Bacteroidetes, and Spirochaetes in the feces, respectively. In the analysis of alpha, beta diversity, and effect size measurements (LefSe), the results showed significant differences between the vaginal and fecal samples. We also identified the function of these differentially abundant microorganisms by functional annotation analyses. But there is no significant correlation between vaginal and fecal microbiome. Conclusion: There is a significant difference between vaginal and fecal microbiome, but no significant correlation. Therefore, it is difficult to interrelate vaginal microbiome as fecal microbiome in Hanwoo. In a further study, it will be necessary to identify the genetic relationship of the entire microorganism between vagina and feces through the whole metagenome sequencing analysis and meta-transcriptome analysis to figure out their relationship.

A Comparison of Analysis Methods for Work Environment Measurement Databases Including Left-censored Data (불검출 자료를 포함한 작업환경측정 자료의 분석 방법 비교)

  • Park, Ju-Hyun;Choi, Sangjun;Koh, Dong-Hee;Park, Donguk;Sung, Yeji
    • Journal of Korean Society of Occupational and Environmental Hygiene
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    • v.32 no.1
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    • pp.21-30
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    • 2022
  • Objectives: The purpose of this study is to suggest an optimal method by comparing the analysis methods of work environment measurement datasets including left-censored data where one or more measurements are below the limit of detection (LOD). Methods: A computer program was used to generate left-censored datasets for various combinations of censoring rate (1% to 90%) and sample size (30 to 300). For the analysis of the censored data, the simple substitution method (LOD/2), β-substitution method, maximum likelihood estimation (MLE) method, Bayesian method, and regression on order statistics (ROS)were all compared. Each method was used to estimate four parameters of the log-normal distribution: (1) geometric mean (GM), (2) geometric standard deviation (GSD), (3) 95th percentile (X95), and (4) arithmetic mean (AM) for the censored dataset. The performance of each method was evaluated using relative bias and relative root mean squared error (rMSE). Results: In the case of the largest sample size (n=300), when the censoring rate was less than 40%, the relative bias and rMSE were small for all five methods. When the censoring rate was large (70%, 90%), the simple substitution method was inappropriate because the relative bias was the largest, regardless of the sample size. When the sample size was small and the censoring rate was large, the Bayesian method, the β-substitution method, and the MLE method showed the smallest relative bias. Conclusions: The accuracy and precision of all methods tended to increase as the sample size was larger and the censoring rate was smaller. The simple substitution method was inappropriate when the censoring rate was high, and the β-substitution method, MLE method, and Bayesian method can be widely applied.

Development and Validation of an Analytical Method for β-Agonists in Livestock and Fishery Products Using LC-MS/MS (LC-MS/MS를 이용한 축·수산물 중 β-agonist계 시험법 개발 및 검증)

  • Lee, Tae Ho;Kim, Yu Ra;Park, Su Jeong;Kim, Ji Young;Choi, Jang Duck;Moon, Gui Im
    • Korean Journal of Environmental Agriculture
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    • v.41 no.2
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    • pp.135-151
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    • 2022
  • BACKGROUND: The β-agonists known as phenyl ethanolamine derivatives have a conjugated aromatic ring with amino group. They are used as tocolytic agents and bronchodilator to human and animal generally, and some of them are used as growth promoters to livestock. METHODS AND RESULTS: β-agonists in samples were extracted by 0.4 N perchloric acid and ethyl acetate. The target compounds were analyzed by liquid chromatography-electrospray tandem mass spectrometry (LC-ESI-MS/MS). Validation of method was performed according to CODEX guidelines (CAC/GL-71). The matrix matched calibration gave correlation coefficients>0.98, and the obtained recoveries were in the range of 62.0-109.8%, with relative standard deviation ≤ 20.1%. In addition, a survey was performed to inspect any residual β-agonist from 100 samples of livestock and fishery products and ractopamine was detected in one of the 100 samples. CONCLUSION(S): In this study, we established the analytical method for β-agonists through using the expanded target compounds and samples. And we anticipate that the established method would be used for analysis to determine veterinary drug residues in livestock and fishery products.

Changes in Electrophysiological Activation Due to Different Levels of Cognitive Load (인지부하의 정도에 따른 뇌신경생리학적 변화)

  • Kwon, Joo-Hee;Kim, Euijin;Kim, Jeonghui;Im, Chang-Hwan;Kim, Do-Won
    • Journal of Biomedical Engineering Research
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    • v.43 no.1
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    • pp.52-60
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    • 2022
  • Purpose: For now, cognitive load is assessed based on survey-based methods, which can be difficult to track the amount of cognitive load in real-time. In this study, we investigated the difference in electrophysiological activation due to different levels of cognitive load not only at sensor-level but also at source-level using electroencephalogram that might be potentially used for quantitative cognitive load evaluation. Materials and Methods: In this study, ten healthy subjects (mean age 24.3 ± 2.1, three female) participated the experiment. All participants performed 4 sessions of n-back task in different difficulties: 0-, 1-, 2-, and 3-back during electroencephalogram recording. For sensor-level analysis, we calculated the event-related potential and event-related spectral perturbation while low resolution brain electromagnetic tomography (LORETA) to estimate the source activation. Each result was compared between different workload conditions using statistical analysis. Results: Statistical results revealed that the accuracy of the task performance was significantly different between different cognitive loads (p = 0.018). The post-hoc analysis confirmed that the accuracy of the 3-back task was significantly decreased compared to 1-back condition (p = 0.018), but not with 2-back condition (p = 0.180). ERP results showed that P300 target amplitude between 1-back and 3-back had a marginal difference in Cz (p = 0.059) and Pz(p = 0.093). A significant inhibition in Cz high-beta activation (p = 0.017) and decrease in source activation of right parahippocampal gyrus was found in 3-back condition compared to 1-back condition (p < 0.05). Conclusion: In this study, we compared the sensor- and source-level differences in electroencephalogram between different levels of cognitive load, that were found to be in line with the previous reports related to cognitive load evaluation. We expect that the outcome of the current study can be used as a feature to establish a quantitative cognitive load assessment system.