1.Patterns of Iron Utilization According to the Growth of Staphylococcus aureus.
Yong LIM ; Sung Heui SIN ; Nam Woong YANG ; Seung Il LEE ; Jae Su KIM ; Sae Heuk JOO ; Jong Hoon PARK
Journal of the Korean Society for Microbiology 1998;33(5):435-439
To elucidate iron utilization patterns of Staphylococcus aureus according to the growth, we checked the residual iron concentration and the production of siderophores at the indicated times while culturing S. aureus ATCC 6538 and 25923 strains in brain heart infusion broth. By using streptonigrin susceptibility test and investigating growth curves in three culture media of which iron concentration is 0.2, 20, 45 uM, respectively, we found out that iron metabolism of 6538 strain was more active than that of 25923 strain. In point of tendency of iron consumption, 6538 strain steeply consumed iron just before the onset of stationary phase, but 25923 strain did gradually iron throughout the growth phase. Nevertheless, total amount of iron consumed by each strain during the growth was almost no difference between the strains. CAS diffusion assay in detecting siderophores showed that siderophore production followed iron consumption. These results suggest that the siderophores play significant role in iron utilization in vitro.
Brain
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Culture Media
;
Diffusion
;
Heart
;
Iron*
;
Metabolism
;
Siderophores
;
Staphylococcus aureus*
;
Staphylococcus*
;
Streptonigrin
2.Cell Wall Proteins of Staphylococcus aureus Responsive to Oxygen Tension and Iron Concentration.
Seung Il LEE ; Yong LIM ; Sung Heui SHIN
Korean Journal of Infectious Diseases 2001;33(4):292-297
BACKGROUND: We previously reported that activity of iron-uptake systems (IUS) influenced on the growth of Staphylococcus aureus in laboratory medium and body fluids according to the iron and oxygen concentrations, which they are closely related each other in several microbial metabolism. In the present study, we tried to investigate the profiles of cell wall proteins of S. aureus according to the change of iron and oxygen concentrations. METHODS: SR-1 strain, whose IUS are defective, was isolated from the standard strain ATCC 6538 by repeated exposure against streptonigrin. These two strains were cultured under the aerobic, microaerobic and anaerobic conditions in the iron-sufficient BHI and iron-depleted BHI, respectively. Cell wall proteins were visualized by Coomassie staining after SDS-PAGE. RESULTS: Cell wall proteins of the both strains were expressed more than under the aerobic condition than under the anaerobic condition in the iron-sufficient medium as well as in the iron-deficient medium. However, expression of cell wall proteins of SR-1 strain was markedly inhibited compared to that of parental ATCC 6538 strain, especially in the iron-deficient medium. Among the proteins more expressed under the aerobic culture condition in the iron-deficient medium, about 88, 55, 39, 36, 35 and 33 kDa of proteins were iron-repressible and oxygen-inducible, and corresponded to the iron-repressible proteins which other researchers reported. CONCLUSION: Expression of cell wall proteins of S. aureus was affected by simultaneous and respective change of iron and oxygen concentrations. Activity of IUS influenced more on the expression of cell wall proteins of S. aureus in the iron-deficient environment than in the iron-sufficient environment. These results suggest the possibility that the iron-repressible and oxygen-inducible proteins mimic those (antigens) found commonly in clinical infections.
Body Fluids
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Cell Wall*
;
Electrophoresis, Polyacrylamide Gel
;
Humans
;
Iron*
;
Metabolism
;
Oxygen*
;
Parents
;
Staphylococcus aureus*
;
Staphylococcus*
;
Streptonigrin
3.Effect of Iron-Uptake System on the Growth of Staphylococcus aureus according to the Iron Concentration and Oxygen Tension.
Korean Journal of Infectious Diseases 2001;33(3):214-218
BACKGROUND: We previously reported that activity of iron-uptake systems (IUS) influenced on the growth of Staphylococcus aureus in body fluids which are relatively iron-restrictive conditions. Iron and oxygen are closely related each other in several microbial metabolism. In the present study, we tried to investigate the effect of IUS on the growth of S. aureus according to the iron concentration and oxygen tension. METHODS: SR-1 strain, whose IUS are defective, was isolated from the standard strain, ATCC 6538. These two strains were cultured under the aerobic, microaerobic and anaerobic condition in the iron-sufficient BHI and iron-depleted BHI, respectively. Bacterial growth was measured by optical density. RESULTS: Growth of both strains was inhibited in the iron-depleted BHI. Growth of parental strain was more active in the iron-sufficient BHI as well as in the ion-depleted BHI than that of SR-1 strain. Growth of both strains was more active under the aerobic condition than under the microaerobic or anaerobic condition. In the iron-depleted BHI, parental strain showed a striking difference of growth according to the oxygen tension. In the iron-depleted BHI, growth of SR-1 strain was markedly inhibited regardless of oxygen tension. CONCLUSION: IUS influenced more on the growth of S. aureus in the iron-depleted environments than in the iron-sufficient environments, and under the aerobic condition than under the microaerobic or anaerobic condition in the iron-depleted environments. These results indicated the possibility that oxygen as well as iron regulate activity and expression of IUS.
Body Fluids
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Humans
;
Iron*
;
Metabolism
;
Oxygen*
;
Parents
;
Staphylococcus aureus*
;
Staphylococcus*
;
Strikes, Employee
4.Characterization of plantaricin IIA-1A5 biosynthesized by Lactobacillus plantarum IIA-1A5 in corn steep liquor based medium
Tuti Rostianti Maulani ; Betty Sri Laksmi Jenie ; Irma Isnafia Arief ; Sukarno Sukarno
Malaysian Journal of Microbiology 2021;17(3):313-320
Aims:
To characterize the plantaricin IIA-1A5 crude extract that biosynthesized by Lactobacillus plantarum IIA-1A5
using corn steep liquor (CSL) based medium.
Methodology and results:
Lactobacillus plantarum IIA-1A5 was grown in several media containing different components including corn steep liquor (CSL), molasses and MRS (de Man Rogosa Sharpe) as control medium for 24 h at 37 °C. Antibacterial activities of the cell-free supernatant were expressed as diameter of inhibition zones observed by paper disc method. The results showed that CSL medium produced cell-free supernatant of L. plantarum IIA-1A5 with significantly higher antibacterial activity againts Staphylococcus aureus ATCC 25923 (9.81 mm), Lactobacillus monocytogenes ATCC 7644 (9.61 mm), Bacillus cereus (8.97 mm) and Escherichia coli ATCC 25922 (9.23 mm) were not significantly different compared to control MRS broth media (9.59 mm). CSL medium added only with 3% yeast extract and Tween 80 produced supernatant which showed similar antibacterial activity either to 10% molasses or control medium (Medium K and B). The CSL medium was considered more efficient and low cost, therefore this medium was selected for production and characterization of plantaricin IIA-1A5 crude extract. Further characterization performed by SDS PAGE analysis showed that crude plantaricin had molecular weight of approximately 9.9 kDa, higher than that produced in control medium (8.0 kDa). However, both plantaricins were categorized under the same class for small bacteriocin (class II). This study also revealed the plantaricin IIA-1A5 produced in CSL medium was stable to heat and pH and not significantly different compared to control MRS broth media. The antibacterial activity of plantaricin IIA-1A5 crude extract against S. aureus ATCC 25923 (10.09 mm) was not significantly different with 1000 ppm sodium benzoate (9.70 mm) and 300 ppm sodium nitrite (9.82 mm).
Conclusion, significance and impact of study
The CSL medium produced cell-free supernatant of L. plantarum IIA 1A5 had significant antibacterial activity characterization againts S. aureus ATCC 25923, L. monocytogenes ATCC 7644, B. cereus and E. coli ATCC 25922. Comparison of the inhibition activity of plantaricin IIA-1A5 crude extract against pathogen with synthetic preservatives indicated that plantaricin IIA-1A5 crude extract have the potency to replace synthetic preservatives. CSL based medium is potential to be used for low-cost plantaricin IIA-1A5 production.
Anti-Bacterial Agents--metabolism
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Bacteriocins--metabolism
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Lactobacillus plantarum
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Microbial Viability--drug effects
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Staphylococcus aureus
5.Immunogenicity of Staphylococcus aureus recombinant clumping factor A.
Hao FENG ; Lefeng LIU ; Jiaqi CHI ; Ning WANG ; Runting LI ; Chunyu TONG ; Jinzhu MA ; Zhanbo ZHU ; Yudong CUI
Chinese Journal of Biotechnology 2009;25(8):1180-1186
In order to characterize the immunogenicity and immunoprotection of the Staphylococcus aureus (S. aureus) surface protein Clumping factor A (ClfA), we amplified clfa genes from S. aureus Newman strain, Wood46 strain and HLJ23-1. The clfa gene from Newman strain was subsequently inserted into pQE-30 vector and the recombinant plasmid was transformed into Escherichia coli strain M15 (pREP4). The recombinant ClfA protein was expressed and purified. Then, we immunized mice with the purified recombinant protein. The antibody level and the concentration of cytokines were measured by enzyme-linked immunosorbent assay. Finally, immunized mice were challenged with S. aureus Newman, Wood46 and HLJ23-1. These results suggested that clfa gene sequences were highly conserved, and the recombinant ClfA was expressed correctly with good antigenicity. The antibody titer and the concentration of cytokines in the immunized groups increased significantly (P < 0.05) compared with control, and the mice in the immunized groups were protected against the challenge strains to some extent. These results showed that the ClfA had high immunogenicity and immunoprotective potential.
Animals
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Coagulase
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genetics
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immunology
;
metabolism
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Escherichia coli
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genetics
;
metabolism
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Immunization
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Mice
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Recombinant Proteins
;
genetics
;
immunology
;
metabolism
;
Staphylococcus aureus
;
metabolism
;
pathogenicity
6.Expression and lytic efficacy assessment of the Staphylococcus aureus phage SA4 lysin gene.
Anil Kumar MISHRA ; Mayank RAWAT ; Konasagara Nagaleekar VISWAS ; ABHISHEK ; Sujeet KUMAR ; Manjunatha REDDY
Journal of Veterinary Science 2013;14(1):37-43
Treatment of bovine mastitis caused by Staphylococcus (S.) aureus is becoming very difficult due to the emergence of multidrug-resistant strains. Hence, the search for novel therapeutic alternatives has become of great importance. Consequently, bacteriophages and their endolysins have been identified as potential therapeutic alternatives to antibiotic therapy against S. aureus. In the present study, the gene encoding lysin (LysSA4) in S. aureus phage SA4 was cloned and the nucleotide sequence was determined. Sequence analysis of the recombinant clone revealed a single 802-bp open reading frame encoding a partial protein with a calculated mass of 30 kDa. Results of this analysis also indicated that the LysSA4 sequence shared a high homology with endolysin of the GH15 phage and other reported phages. The LysSA4 gene of the SA4 phage was subsequently expressed in Escherichia coli. Recombinant LysSA4 induced the lysis of host bacteria in a spot inoculation test, indicating that the protein was expressed and functionally active. Furthermore, recombinant lysin was found to have lytic activity, albeit a low level, against mastitogenic Staphylococcus isolates of bovine origin. Data from the current study can be used to develop therapeutic tools for treating diseases caused by drug-resistant S. aureus strains.
Animals
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Base Sequence
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Cloning, Molecular
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Gene Expression Regulation, Viral/physiology
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Mucoproteins/genetics/*metabolism
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Phylogeny
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Polymerase Chain Reaction/methods
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Recombinant Proteins
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Staphylococcus Phages/genetics/*metabolism/physiology
;
Staphylococcus aureus/*virology
7.Biochemical characters and antibiotic susceptibility of Staphylococcus aureus isolates.
Subhankari Prasad CHAKRABORTY ; Santanu Kar MAHAPATRA ; Somenath ROY
Asian Pacific Journal of Tropical Biomedicine 2011;1(3):212-216
OBJECTIVETo observe the biochemical characters and antibiotic susceptibility of isolated Staphylococcus aureus (S. auerus) strains against some conventional and traditional antibiotics.
METHODSThirty post operative pathogenic isolated S. aureus strains were used in this study. Bacterial culture was done in Mueller-Hinton broth at 37 °C. Characters of these strains were determined by traditional biochemical tests such as hydrolysis test of gelatin, urea, galactose, starch and protein, and fermentation of lactose and sucrose. Antibiotic susceptibility were carried out by minimum inhibitory concentration test, minium bactericidal concentration test, disc agar diffusion test and brain heart infusion oxacillin screening agar.
RESULTSFrom this study, it was observed that 100% S. aureus isolates showed positive results in gelatin, urea and galactose hydrolysis test, 50% isolates were positive in starch hydrolysis test, 35% in protein hydrolysis test, 100% isolates in lactose fermenting test, but no isolate was positive in sucrose fermenting test. Antibiotic susceptibility testing suggested that 20% of isolates were resistant to kanamycin and 46.67% were resistant to oxacillin.
CONCLUSIONSThese findings show that all these isolates have gelatin, urea, galactose hydrolysis and lactose fermenting activity. 20% of these isolates were resistant to kanamycin and 46.67% were resistant to oxacillin.
Anti-Bacterial Agents ; pharmacology ; Disk Diffusion Antimicrobial Tests ; Galactose ; metabolism ; Gelatin ; metabolism ; Hydrolysis ; Microbial Sensitivity Tests ; Staphylococcus aureus ; drug effects ; isolation & purification ; metabolism ; Starch ; metabolism ; Urea ; metabolism
8.Enzymatic cyclization of peptides using immobilized sortase A.
Shu-xiang ZHANG ; Min-zhi LIU ; Yan YANG ; Ke-di CHENG ; Jian-qiang KONG ; Wei WANG
Acta Pharmaceutica Sinica 2015;50(5):627-632
Peptide cyclization, a pivotal approach to modifying linear precursors of proteins and pepticles, has been used to enhance their biological activities and serum stabilities. Recently, sortase A (SrtA) from Staphyloccus aureus becomes a promising new technology for efficiently incorporating site specific modifications into proteins, conjugating the cell surface and cyclizing the linear peptides. In this study, we constructed two recombinant expression systems, one with chitin binding domain and the other with six-histidine tag and chitin binding domain on the N-terminal of SrtA, separately. The results of enzymatic kinetics indicate that the two recombinant tags do not impair the transpeptidase activity of SrtA compared with the standard reaction reported under the same reaction condition. The two synthesized peptides with N-ternimal three glycines and C-terminal penta-amino acid motif, LPETG, were cyclized using immobilized and recycled SrtA. The SrtA-based cyclization promises to represent a simple method for easy and efficient enzymatic synthesis of large cyclic peptides.
Aminoacyltransferases
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metabolism
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Bacterial Proteins
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metabolism
;
Cyclization
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Cysteine Endopeptidases
;
metabolism
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Enzymes, Immobilized
;
metabolism
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Kinetics
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Peptides
;
metabolism
;
Peptides, Cyclic
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biosynthesis
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Staphylococcus aureus
;
enzymology
9.Non-myeloperoxidase-mediated system activity of neutrophil in newborn infants.
Xiao-dong ZHU ; Tong-xin CHEN ; Ruo-xu JI ; Xiao-ling ZHOU ; Lian-wen WANG ; Jian-xing ZHU
Chinese Journal of Pediatrics 2003;41(4):286-289
OBJECTIVETo evaluate the variety of non-myeloperoxidase-mediated system activity of neutrophils in newborns during bacterial infection and the effect of cord plasma on the activation of non-myeloperoxidase-mediated system.
METHODSAn infection model with Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) and a non-infection model with phorbol-12-myristate-13-acetate (PMA) were established to investigate the activation of non-myeloperoxidase-mediated system in neutrophils. According to the intensity of fluorescence, the activation of non-myeloperoxidase-mediated system of neutrophils was detected by flow cytometry (FCM). The blood cells and plasma were separated from cord blood and adult blood and cross-mixed in order to investigate the opsonic activity.
RESULTSIn the non-infection model, the activation of non-myeloperoxidase-mediated system with PMA stimulation in cord blood was lower compared with that in adult blood, the statistical difference was significant (t = 3.378, P < 0.01). In the infection model, the activations of non-myeloperoxidase-mediated system in cord blood were also lower compared with those in adult blood, while the statistical difference could only be found in the model with E. coli stimulation (t = 12.150, P < 0.001). Furthermore the experiments demonstrated that cord plasma could deeply depress the non-myeloperoxidase-mediated system activity with E. coli stimulation. On the contrary, adult plasma could successfully recruit the potential of non-myeloperoxidase-mediated system activity of neutrophils in newborns.
CONCLUSIONThe function of neonatal neutrophils might not developed very well. As a stimulant, E. coli failed to induce the non-myeloperoxidase-mediated system activity in neonates, which might be related to the lower level of immunoglobulins in cord blood. This result indicated that immunoglobulins played a more important modulating role in bacterial killing during gram-negative bacterial infections.
Escherichia coli ; immunology ; Fetal Blood ; immunology ; Flow Cytometry ; Humans ; Infant, Newborn ; Neutrophils ; enzymology ; immunology ; Peroxidase ; metabolism ; Staphylococcus aureus ; immunology
10.Human soluble dendritic cell-specific intercellular adhesion molecule 3-grabbing nonintegrin inhibits phagocytosis of Staphylococcus aureus by immature dendritic cells.
Hui-Jie LI ; Tian-Yu XU ; Jia ZHOU ; Ling-Yan ZHU ; Li-Yun ZHANG ; Xiao LU ; Zheng-Liang CHEN
Journal of Southern Medical University 2015;35(4):544-548
OBJECTIVETo study the effect and mechanism of soluble dendritic cell-specific intercellular adhesion molecule 3-grabbing nonintegrin (sDC-SIGN) on the phagocytosis of Staphylococcus aureus (S. aureus) by immature dendritic cells (imDCs).
METHODSFlow cytometry was employed to examine the effect of sDC-SIGN on the phagocytosis of S. aureus by imDCs. Enzyme-linked immunosorbent assay (ELISA) was used to analyze the binging of sDC-SIGN to S. aureus, lipoteichoic acid (LTA) and lipopolysaccharides (LPS) and investigate the effect of the ligands mannan and LTA and anti-DC-SIGN antibodies 1C6 and 4H3 on the binging of sDC-SIGN to S. aureus.
RESULTSsDC-SIGN inhibited the phagocytosis of S. aureus by imDCs. sDC-SIGN bound to S. aureus in a Ca(2+)-dependent manner. sDC-SIGN concentration-dependently bound to LTA, but not to LTA, and the binging of sDC-SIGN to S. aureus was blocked by mannan, LTA, 1C6 and 4H3.
CONCLUSIONsDC-SIGN preferentially binds to the carbohydrate constituents on S. aureus to affect the binding between membrane-bound DC-SIGN and S. aureus, thus suppressing the phagocytosis of S. aureus by imDCs.
Cell Adhesion Molecules ; metabolism ; Dendritic Cells ; cytology ; metabolism ; Humans ; Lectins, C-Type ; metabolism ; Lipopolysaccharides ; Phagocytosis ; Receptors, Cell Surface ; metabolism ; Staphylococcus aureus ; Teichoic Acids