2.High-throughput screening of Saccharomyces cerevisiae efficiently producing tyrosine.
Tanghao LIU ; Youran LI ; Liang ZHANG ; Zhongyang DING ; Zhenghua GU ; Guiyang SHI ; Sha XU
Chinese Journal of Biotechnology 2021;37(9):3348-3360
Tyrosine is an important aromatic amino acid. Besides its nutritional value, tyrosine is also an important precursor for the synthesis of coumarins and flavonoids. Previously, our laboratory constructed a Saccharomyces cerevisiae strain LTH0 (ARO4K229L, ARO7G141S, Δaro10, Δzwf1, Δura3) where tyrosine feedback inhibition was released. In the present study, heterologous expression of betaxanthins synthesis genes DOD (from Mirabilis jalapa) and CYP76AD1 (from sugar beet B. vulgaris) in strain LTH0 enabled production of yellow fluorescence. The engineered strain LTH0-DOD-CYP76AD1 was subjected to UV combined with ARTP mutagenesis, followed by flow cytometry screening. Among the mutants screened, the fluorescence intensity of the mutant strain LTH2-5-DOD-CYP76AD1 at the excitation wavelength of 485 nm and emission wavelength of 505 nm was (5 941±435) AU/OD, which was 8.37 times higher than that of strain LTH0-DOD-CYP76AD1. Fourteen mutant strains were subjected to fermentation to evaluate their tyrosine producing ability. The highest extracellular tyrosine titer reached 26.8 mg/L, which was 3.96 times higher than that of strain LTH0-DOD-CYP76AD1. Heterologous expression of the tyrosine ammonia lyase FjTAL derived from Flavobacterium johnsoniae further increased the titer of coumaric acid to 119.8 mg/L, which was 1.02 times higher than that of the original strain LTH0-FjTAL.
Flavobacterium
;
High-Throughput Screening Assays
;
Mirabilis
;
Saccharomyces cerevisiae/genetics*
;
Tyrosine
3.Cloning and expression of duck C4BPα and verification of its interaction with Riemerella anatipestifer.
Delong LI ; Lijuan TAN ; Jiulong GU ; Siyuan WANG ; Ting LIU ; Sihuai CHEN ; Jiye GAO ; Fashu TANG ; Jixiang LI
Chinese Journal of Biotechnology 2020;36(4):693-699
To study the interaction between C4b-binding protein (C4BP) and Riemerella anatipestifer (RA), we cloned duck C4BPα, conducted prokaryotic expression and prepared the polyclonal antibody by immunizing mice. Then indirect immunofluorescence assay and dot blotting hybridization assay were used to verify the interaction between C4BP and RA. The full length of duck C4BPα nucleotide sequence was 1 230 bp, with the highest similarity to chicken C4BPα (82.1%). Phylogenetic tree analysis showed that duck C4BPα and chicken C4BPα were on the same phylogenetic tree branch and the genetic evolution relationship between them was the closest. C4BPα was efficiently expressed in Escherichia coli BL21 (DE3). The recombinant proteins existed in intracellular soluble form. The titer of polyclonal antibody was more than 1:10 000 and polyclonal antibodies could specifically recognize the recombinant proteins. The results of indirect immunofluorescence assay and dot blot hybridization assay showed that RA could interact with duck C4BP. The results provide a basis to further reveal the pathogenesis of RA.
Animals
;
Cloning, Molecular
;
Complement C4b-Binding Protein
;
chemistry
;
genetics
;
metabolism
;
Ducks
;
classification
;
genetics
;
microbiology
;
Gene Expression Regulation
;
Mice
;
Phylogeny
;
Riemerella
;
metabolism
4.A Case of Chryseobacterium hominis Isolated from Human Blood Drawn Through Peripherally Inserted Central Catheter
Dongju WON ; Jung Hyun BYUN ; Myungsook KIM ; Dongeun YONG
Laboratory Medicine Online 2019;9(4):246-248
Chryseobacterium hominis is non-fermenting Gram-negative rod that was first identified as a novel species in 2007. Here, we report the first clinical case of C. hominis bacteremia, which was confirmed by MALDI-TOF MS and 16S rRNA gene sequencing. A 16-year-old boy diagnosed with acute lymphoblastic leukemia was hospitalized for three months. Two sets of blood culture test through a peripherally inserted central catheter (PICC), which was inserted a month ago, was performed when his white blood cell count declined and he had a high fever. Colonies of medium sizes that looked round, mucoid, sticky, and grayish on blood and chocolate agar plates were observed. Identification of bacteria using the VITEK MALDI-TOF MS system (BioMérieux, France) was not successful and the VITEK 2 system (BioMérieux, USA) indicated Sphingomonas paucimobilis, with a questionable level of confidence (92%). However, Microflex LT Biotyper (Bruker Daltonics, Germany) showed C. homins (log score: 1.81) and sequence of 16S rRNA showed a 100% identity with C. hominis. Piperacillin-tazobactam was administered since the isolate was susceptible to piperacillin-tazobactam but C. hominis showed growth in the next four follow-up culture of blood drawn through PICC. The fever subsided only after PICC was changed. The clinical prognosis and antimicrobial susceptibility test of C. hominis should be further studied.
Adolescent
;
Agar
;
Bacteremia
;
Bacteria
;
Cacao
;
Catheters
;
Chryseobacterium
;
Fever
;
Follow-Up Studies
;
Genes, rRNA
;
Humans
;
Leukocyte Count
;
Male
;
Precursor Cell Lymphoblastic Leukemia-Lymphoma
;
Prognosis
;
Sphingomonas
5.Serosurvey of Avian metapneumovirus, Orithobacterium rhinotracheale, and Chlamydia psittaci and Their Potential Association with Avian Airsacculitis.
Zong Hui ZUO ; Tian Yuan ZHANG ; Yong Xia GUO ; Jun CHU ; Guang Gang QU ; Li Zhong MIAO ; Zhi Qiang SHEN ; Cheng HE
Biomedical and Environmental Sciences 2018;31(5):403-406
Seasonal outbreaks of airsacculitis in China's poultry cause great economic losses annually. This study tried to unveil the potential role of Avian metapneumovirus (AMPV), Ornithobacterium rhinotracheale (ORT) and Chlamydia psittaci (CPS) in avian airsacculitis. A serological investigation of 673 breeder chickens and a case-controlled study of 430 birds were undertaken. Results showed that infection with AMPV, ORT, and CPS was highly associated with the disease. The correlation between AMPV and CPS were positively robust in both layers and broilers. Finally, we determined the co-infection with AMPV, ORT, and CPS was prevalent in the sampled poultry farms suffering from respiratory diseases and the outbreak of airsacculitis was closely related to simultaneous exposure to all three agents.
Air Sacs
;
microbiology
;
pathology
;
Animals
;
Antibodies, Bacterial
;
blood
;
Antibodies, Viral
;
blood
;
Case-Control Studies
;
Chickens
;
Chlamydia
;
Chlamydia Infections
;
microbiology
;
pathology
;
veterinary
;
Coinfection
;
Flavobacteriaceae Infections
;
microbiology
;
pathology
;
veterinary
;
Humans
;
Metapneumovirus
;
Ornithobacterium
;
Paramyxoviridae Infections
;
pathology
;
veterinary
;
virology
;
Poultry Diseases
;
microbiology
;
pathology
;
virology
;
Respiratory Tract Diseases
;
microbiology
;
veterinary
;
virology
;
Seroepidemiologic Studies
6.A two-component signal transduction system contributes to the virulence of Riemerella anatipestifer
Qing WANG ; Mianmian CHEN ; Wei ZHANG
Journal of Veterinary Science 2018;19(2):260-270
Similar to other studies of bacterial pathogens, current studies of the pathogenesis of Riemerella anatipestifer (RA) are focused mainly on in vitro culture conditions. To elucidate further the pathogenesis of RA in vivo, bacterial RNA was extracted from overnight tryptic soy broth cultures (in vitro) and from the blood of infected ducks (in vivo) for comparative RNA sequencing analysis. In total, 682 upregulated genes were identified in vivo. Among the upregulated genes, a signal transduction response regulator (ArsR) and a signal transduction histidine kinase (SthK) were predicted to be located on the same operon. A mutant was constructed by deletion of both of these genes. Duck infection tests showed that genes ArsR and SthK were related to the virulence of the pathogen in vivo. Differentially expressed genes identified by comparison of in vitro and in vivo conditions provided an insight into the physiological process of RA infection and provided an opportunity to identify additional virulence factors.
Ducks
;
Genes, vif
;
Histidine
;
In Vitro Techniques
;
Operon
;
Phosphotransferases
;
Physiological Processes
;
Riemerella
;
RNA, Bacterial
;
Sequence Analysis, RNA
;
Signal Transduction
;
Virulence Factors
;
Virulence
7.Peritoneal Dialysis-associated Peritonitis Caused by Chryseobacterium indologenes
Myung Woo CHOI ; Sung Sik OH ; Mi Rim CHOI ; Jong Hwa LEE ; Hyun Ju YANG ; In O SUN ; Kwang Young LEE
Korean Journal of Medicine 2018;93(1):65-67
Chryseobacterium indologenes (C. indologenes) is a nonmotile, gram-negative bacillus that is widely distributed in nature. Generally considered nonpathogenic, C. indologenes rarely infects humans and is not normally present in the human microflora. C. indologenes infections have been observed in cases of peritoneal dialysis (PD)-associated peritonitis, although the incidence of these infections is low. Although C. indologenes is generally susceptible to trimethoprim-sulfamethoxazole, levofloxacin, ciprofloxacin, piperacillin-tazobactam, and cefepime, no guidelines have been established for the treatment of PD-associated peritonitis. Here we report the first case of PD-associated peritonitis in Korea with C. indologenes identified as the sole etiologic agent. The patient recovered after intraperitoneal antibiotic treatment without the need for Tenckhoff catheter removal.
Bacillus
;
Catheters
;
Chryseobacterium
;
Ciprofloxacin
;
Humans
;
Incidence
;
Korea
;
Levofloxacin
;
Peritoneal Dialysis
;
Peritonitis
;
Trimethoprim, Sulfamethoxazole Drug Combination
8.Biological control of Flavobacterium psychrophilum infection in ayu (Plecoglossus altivelis altivelis) using a bacteriophage PFpW-3
Korean Journal of Veterinary Research 2018;58(1):39-43
The efficacy of using a bacteriophage (phage) to control Flavobacterium psychrophilum (F. psychrophilum) infection of ayu (Plecoglossus altivelis altivelis) was evaluated in this study. Intramuscular challenge failed to induce sufficient infection levels; therefore, a newly designed net-scratch challenge method was also used to induce bacterial infection. Administration of phage PFpW-3 in F. psychrophilum-infected ayu showed notable protective effects, increased survival rates and mean times to death. Additionally, the fate of inoculated bacteria and phage in ayu were investigated. Our results suggest that the phage PFpW-3 could be considered an alternative biocontrol agent against F. psychrophilum infections in ayu culture.
Bacteria
;
Bacterial Infections
;
Bacteriophages
;
Flavobacterium
;
Methods
;
Osmeriformes
;
Survival Rate
9.Effects of supplementation with antifreeze proteins on the follicular integrity of vitrified-warmed mouse ovaries: Comparison of two types of antifreeze proteins alone and in combination.
Min Kyung KIM ; Hyun Sun KONG ; Hye Won YOUM ; Byung Chul JEE
Clinical and Experimental Reproductive Medicine 2017;44(1):8-14
OBJECTIVE: The aim of this study was to analyze the effect of supplementing vitrification and warming solutions with two types of antifreeze proteins (AFPs) and the combination thereof on the follicular integrity of vitrified-warmed mouse ovaries. METHODS: Ovaries (n=154) were obtained from 5-week-old BDF1 female mice (n=77) and vitrified using ethylene glycol and dimethyl sulfoxide with the supplementation of 10 mg/mL of Flavobacterium frigoris ice-binding protein (FfIBP), 10 mg/mL of type III AFP, or the combination thereof. Ovarian sections were examined by light microscopy after hematoxylin and eosin staining, and follicular intactness was assessed as a whole and according to the type of follicle. Apoptosis within the follicles as a whole was detected by a terminal deoxynucleotidyl transferase deoxyuridine triphosphate nick-end labeling assay. RESULTS: The proportion of overall intact follicles was significantly higher in the type III AFP-supplemented group (60.5%) and the combination group (62.9%) than in the non-supplemented controls (43.8%, p<0.05 for each). The proportion of intact primordial follicles was significantly higher in the FfIBP-supplemented (90.0%), type III AFP-supplemented (92.3%), and combination (89.7%) groups than in the non-supplemented control group (46.2%, p<0.05 for each). The proportions of non-apoptotic follicles were similar across the four groups. CONCLUSION: Supplementation of the vitrification and warming solutions with FfIBP, type III AFP, or the combination thereof was equally beneficial for the preservation of primordial follicles in vitrified mouse ovaries.
Animals
;
Antifreeze Proteins*
;
Apoptosis
;
Deoxyuridine
;
Dimethyl Sulfoxide
;
DNA Nucleotidylexotransferase
;
Eosine Yellowish-(YS)
;
Ethylene Glycol
;
Female
;
Fertility Preservation
;
Flavobacterium
;
Hematoxylin
;
Humans
;
Mice*
;
Microscopy
;
Ovary*
;
Vitrification
10.A Case of Flavobacterium ceti Meningitis.
Annals of Laboratory Medicine 2016;36(6):614-616
No abstract available.
Aged
;
Aneurysm/surgery
;
Anti-Bacterial Agents/pharmacology
;
Brain Diseases/surgery
;
Craniotomy/adverse effects
;
DNA, Bacterial/chemistry/genetics/metabolism
;
Female
;
Flavobacteriaceae Infections/etiology/microbiology
;
Flavobacterium/classification/drug effects/*isolation & purification
;
Humans
;
Meningitis/*diagnosis/microbiology
;
Microbial Sensitivity Tests
;
Phylogeny
;
Postoperative Complications
;
Sequence Analysis, DNA

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