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REFERENCE LINKING PLATFORM OF KOREA S&T JOURNALS
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Journal of Microbiology and Biotechnology
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The Korean Society for Applied Microbiology and Biotechnology
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Volume & Issues
Volume 20, Issue 12 - Dec 2010
Volume 20, Issue 11 - Nov 2010
Volume 20, Issue 10 - Oct 2010
Volume 20, Issue 9 - Sep 2010
Volume 20, Issue 8 - Aug 2010
Volume 20, Issue 7 - Jul 2010
Volume 20, Issue 6 - Jun 2010
Volume 20, Issue 5 - May 2010
Volume 20, Issue 4 - Apr 2010
Volume 20, Issue 3 - Mar 2010
Volume 20, Issue 2 - Feb 2010
Volume 20, Issue 1 - Jan 2010
Selecting the target year
A Brief Overview of Escherichia coli O157:H7 and Its Plasmid O157
Lim, Ji-Youn ; Yoon, Jang-W. ; Hovde, Carolyn J. ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 5~14
DOI : 10.4014/jmb.0908.08007
Enterohemorrhagic Escherichia coli O157:H7 is a major foodborne pathogen causing severe disease in humans worldwide. Healthy cattle are a reservoir of E. coli O157:H7, and bovine food products and fresh produce contaminated with bovine waste are the most common sources for disease outbreaks in the United States. E. coli O157:H7 also survives well in the environment. The abilities to cause human disease, colonize the bovine gastrointestinal tract, and survive in the environment require that E. coli O157:H7 adapt to a wide variety of conditions. Three major virulence factors of E. coli O157:H7 have been identified including Shiga toxins, products of the pathogenicity island called the locus of enterocyte effacement, and products of the F-like plasmid pO157. Among these virulence factors, the role of pO157 is least understood. This review provides a board overview of E. coli O157:H7 with an emphasis on pO157.
Caulobacter ginsengisoli sp. nov., a Novel Stalked Bacterium Isolated from Ginseng Cultivating Soil
Liu, Qing-Mei ; Ten, Leonid N. ; Im, Wan-Taek ; Lee, Sung-Taik ; Yoon, Min-Ho ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 15~20
DOI : 10.4014/jmb.0903.03026
A Gram negative, aerobic, nonspore-forming, straight or curved rod-shaped bacterium, designated Gsoil
, was isolated from soil of a ginseng field in Pocheon Province (South Korea) and was characterized using a polyphasic approach. Cells were dimorphic, with stalk (or prostheca) and nonmotile or nonstalked and motile, by means of a single polar flagellum. Comparative analysis of 16S rRNA gene sequences revealed that strain Gsoil
was most closely related to Caulobacter mirabilis LMG
(97.2%), Caulobacter fusiformis ATCC
(97.1 %), Caulobacter segnis LMG
(97.0%), Caulobacter vibrioides DSM
(96.8%), and Caulobacter henricii ATCC
(96.7%). The sequence similarities to any other recognized species within Alphaproteobacteria were less than 96.0%. The detection of Q-10 as the major respiratory quinone and a fatty acid profile with summed feature 7 (
(15.9%) as the major fatty acids supported the affiliation of strain Gsoil
to the genus Caulobacter. The G+C content of the genomic DNA was 65.5 mol%. DNA-DNA hybridization experiments showed that the DNA-DNA relatedness values between strain Gsoil
and its closest phylogenetic neighbors were below 11%. On the basis of its phenotypic properties and phylogenetic distinctiveness, strain Gsoil
should be classified as representing a novel species in the genus Caulobacter, for which the name Caulobacter ginsengisoli sp. novo is proposed. The type strain is Gsoil
Monitoring the Bacterial Community Dynamics in a Petroleum Refinery Wastewater Membrane Bioreactor Fed with a High Phenolic Load
Silva, Cynthia C. ; Viero, Aline F. ; Dias, Ana Carolina F. ; Andreote, Fernando D. ; Jesus, Ederson C. ; De Paula, Sergio O. ; Torres, Ana Paula R. ; Santiago, Vania M.J. ; Oliveira, Valeria M. ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 21~29
DOI : 10.4014/jmb.0906.06001
The phenolic compounds are a major contaminant class often found in industrial wastewaters and the biological treatment is an alternative tool commonly employed for their removal. In this sense, monitoring microbial community dynamics is crucial for a successful wastewater treatment. This work aimed to monitor the structure and activity of the bacterial community during the operation of a laboratory-scale continuous submerged membrane bioreactor (SMBR), using PCR and RT-PCR followed by denaturing gradient gel electrophoresis (DGGE) and 16S rRNA libraries. Multivariate analyses carried out using DGGE profiles showed significant changes in the total and metabolically active dominant community members during the 4-week treatment period, explained mainly by phenol and ammonium input. Gene libraries were assembled using 16S rDNA and 16S rRNA PCR products from the fourth week of treatment. Sequencing and phylogenetic analyses of clones from the 16S rDNA library revealed a high diversity of taxa for the total bacterial community, with predominance of Thauera genus (ca. 50%). On the other hand, a lower diversity was found for metabolically active bacteria, which were mostly represented by members of Betaproteobacteria (Thauera and Comamonas), suggesting that these groups have a relevant role in the phenol degradation during the final phase of the SMBR operation.
Unbalanced Restriction Impairs SOS-induced DNA Repair Effects
Katna, Anna ; Boratynski, Robert ; Furmanek-Blaszk, Beata ; Zolcinska, Natalia ; Sektas, Marian ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 30~38
DOI : 10.4014/jmb.0907.07005
The contribution of a type II restriction-modification system (R-M system) to genome integrity and cell viability was investigated. We established experimental conditions that enabled the achievement of hemimethylated and unmethylated states for the specific bases of the recognition sequences of the host's DNA. To achieve this, we constructed the MboII R-M system containing only one (i.e., M2.MboII) out of two functional MboII methyltransferases found in Moraxella bovis. Using the incomplete R-M system, we were able to perturb the balance between methylation and restriction in an inducible manner. We demonstrate that upon the SOS-induced DNA repair in mitomycin C treated cells, restriction significantly reduces cell viability. Similar results for the well-studied wild-type EcoRI R-M system, expressed constitutively in Escherichia coli, were obtained. Our data provide further insights into the benefits and disadvantages of maintaining of a type II R-M system, highlighting its impact on host cell fitness.
Method Development for Electrotransformation of Acidithiobacillus caldus
Chen, Linxu ; Lin, Jianqun ; Li, Bing ; Lin, Jianqiang ; Liu, Xiangmei ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 39~44
DOI : 10.4014/jmb.0905.05023
Acidithiobacillus caldus is an acidophilic, chemolithotrophic bacterium that plays an important role in bioleaching. Gene transformation into A. caldus is difficult, and only the conjugation method was reported successful, which was a relatively sophisticated method. In this research, electrotransformation of A. caldus species was achieved for the first time using A. caldus Y-3 and plasmid pJRD215. Transformants were confirmed by colony PCR specific to the str gene on pJRD215, and the recovery of the plasmid from the presumptive transformants. Optimizations were made and the transformation efficiency was increased from 0.8 to
plasmid DNA. The developed electrotransformation method was convenient in introducing foreign genes into A. caldus.
Efficient and Precise Construction of Markerless Manipulations in the Bacillus subtilis Genome
Yu, Haojie ; Yan, Xin ; Shen, Weiliang ; Shen, Yujia ; Zhang, Ji ; Li, Shunpeng ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 45~53
DOI : 10.4014/jmb.0904.04051
We have developed an efficient and precise method for genome manipulations in Bacillus subtilis that allows rapid alteration of a gene sequence or multiple gene sequences without altering the chromosome in any other way. In our approach, the Escherichia coli toxin gene mazF, which was used as a counter-selectable marker, was placed under the control of a xylose-inducible expression system and associated with an antibiotic resistance gene to create a "mazF-cassette". A polymerase chain reaction (PCR)-generated fragment, consisting of two homology regions joined to the mazF-cassette, was integrated into the chromosome at the target locus by homologous recombination, using positive selection for antibiotic resistance. Then, the excision of the mazF-cassette from the chromosome by a single-crossover event between two short directly repeated (DR) sequences, included in the design of the PCR products, was achieved by counter-selection of mazF. We used this method efficiently and precisely to deliver a point mutation, to inactivate a specific gene, to delete a large genomic region, and to generate the in-frame deletion with minimal polar effects in the same background.
HpaXm from Xanthomonas citri subsp. malvacearum is a Novel Harpin with Two Heptads for Hypersensitive Response
Miao, Wei-Guo ; Song, Cong-Feng ; Wang, Yu ; Wang, Jin-Sheng ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 54~62
DOI : 10.4014/jmb.0905.05027
A novel harpin-like protein, HpaXm, was described from cotton leaf blight bacteria, Xanthomonas citri subsp. malvacearum. The hpaXm was found to be localized between hrp2 and hrcC. A phylogenetic analysis of the complete amino acid sequence or solely the 13 highly conserved residues
-SEKQLDQLLTQLI-COOH in the N-terminal
-helix indicates that HpaXm is evolutionarily closer to HpaGXag and HpaXac than to Hpa1Xoo and Hpa1Xoc. A synthesized peptide containing two heptads, 39-LDQLLTQLIMALLQ-52, from the N-terminal a-helical region of HpaXm displayed comparable activity in inducing a hypersensitive response, but two other synthesized derivatives,
, showed reduced HR-triggering activity. The data from a GST trap test revealed that HpaXm was released into the extracellular medium, hpaXm mutant deficient for the leader peptide (1-MNSLNTQIGANSSFL-15) was unable to be secreted outside cells but still induced HR in tobacco leaves.
Comparative Study of the Nucleotide Bias Between the Novel H1N1 and H5N1 Subtypes of Influenza A Viruses Using Bioinformatics Techniques
Ahn, In-Sung ; Son, Hyeon-Seok ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 63~70
DOI : 10.4014/jmb.0908.08004
Novel influenza A (H1N1) is a newly emerged flu virus that was first detected in April 2009. Unlike the avian influenza (H5N1), this virus has been known to be able to spread from human to human directly. Although it is uncertain how severe this novel H1N1 virus will be in terms of human illness, the illness may be more widespread because most people will not have immunity to it. In this study, we compared the codon usage bias between the novel H1N1 influenza A viruses and other viruses such as H1N1 and H5N1 subtypes to investigate the genomic patterns of novel influenza A (H1N1). Totally, 1,675 nucleotide sequences of the hemagglutinin (HA) and neuraminidase (NA) genes of influenza A virus, including H1N1 and H5N1 subtypes occurring from 2004 to 2009, were used. As a result, we found that the novel H1N1 influenza A viruses showed the most close correlations with the swine-origin H1N1 subtypes than other H1N1 viruses, in the result from not only the analysis of nucleotide compositions, but also the phylogenetic analysis. Although the genetic sequences of novel H1N1 subtypes were not exactly the same as the other H1N1 subtypes, the HA and NA genes of novel H1N1s showed very similar codon usage patterns with other H1N1 subtypes, especially with the swine-origin H1N1 influenza A viruses. Our findings strongly suggested that those novel H1N1 viruses seemed to be originated from the swine-host H1N1 viruses in terms of the codon usage patterns.
Real-Time PCR Analysis of Metabolic Pathway of PHB in Acidiphilium cryptum DX1-1
Xu, Ai-Ling ; Xia, Jin-Lan ; Liu, Ke-Ke ; Li, Li ; Yang, Yu ; Nie, Zhen-Yuan ; Qiu, Guan-Zhou ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 71~77
DOI : 10.4014/jmb.0906.06054
The time, yield, and related genes expression of PHB accumulation of Acidiphilium cryptum DX1-1 were investigated under four different initial C/N ratios, 1.2, 2.4, 7.5, and 24. The results of time and yield of poly-
-hydroxybutyrate (PHB) accumulation show that the initial C/N ratio of 2.4 was optimum for strain DX1-1 to accumulate PHB, but both higher and lower initial C/N ratios did not favor that process. Based on the genome of Acidiphilium cryptum JF-5, 13 PHB accumulation related genes in strain JF-5 were chosen and successfully cloned from strain DX1-1. The differential expressions of the 13 functional genes, in different C/N ratios as cited above, were then studied by real-time PCR. The results show that all the 13 genes were most upregulated when the initial C/N ratio was 2.4, and among which the gene Acry_3030 encoding poly-
-hydroxybutyrate polymerase and Aery_0626 encoding acetyl-CoA synthetase were much more upregulated than the other genes, which proved that they play the most important role for PHB accumulation, and acetate is the main initial substance for PHB accumulation for strain DX1-1. Potential regulatory motifs analysis showed that the genes related to PHB accumulation are regulated by different promoters and that the motif had weak similarity to the model promoters, suggesting that PHB metabolism in Acidiphilium cryptum may be mediated by a different mechanism.
Secondary Metabolites of Volvariella bombycina and Their Inhibitory Effects on Melanogenesis
Xu, Guang-Hua ; Choo, Soo-Jin ; Kim, Young-Hee ; Ryoo, In-Ja ; Seok, Soon-Ja ; Ahn, Jong-Seog ; Yoo, Ick-Dong ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 78~81
DOI : 10.4014/jmb.0906.06017
Four compounds were isolated from the culture broth of Volvariella bombycina and they were identified as ergosta-4,6,8(14),22-tetraene-3-one (1), ergosterol peroxide (2), indole-3-carboxaldehyde (3), and indazole (4) by interpretation of spectroscopic data. Among them, compound 2 exhibited melanogenesis inhibitory effect in cultured B16 mouse melanoma cells.
Evaluation of the Antibacterial Activity of Rhapontigenin Produced from Rhapontin by Biotransformation Against Propionibacterium acnes
Kim, Jeong-Keun ; Kim, Na-Rae ; Lim, Young-Hee ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 82~87
DOI : 10.4014/jmb.0907.07022
Biotransformation is often used to improve chemical activity. We evaluated the antimicrobial activity of rhapontigeuin, converted from rhapontin after treatment with Pectinex. Rhapontigenin showed 4-16 times higher antimicrobial activity than rhapontin. The activity was higher against Gram-positive strains than Gram-negative strains. Minimum inhibitory concentrations (MICs) of rhapontigenin, retinol, and five antibiotics were determined by the microbroth dilution method for antibiotic-sensitive and -resistant Propionibacterium acnes. We also investigated the in vitro antibacterial activity of rhapontigenin in combination with antibiotic against antibiotic-resistant P. acnes. The antibiotic combination effect against resistant P. acnes was studied by the checkerboard method. The combination formulations (rhapontigenin and clindamycin, retinol and clindamycin) showed synergistic effects on the inhibition of the growth of clindamycin-resistant P. acnes. It is predictable that the combination of antibiotics with rhapontigenin is helpful to treat acne caused by antibiotic-resistant P. acnes. The antibacterial activity of rhapontigenin was enhanced by biotransformation.
A Novel Selenium- and Copper-Containing Peptide with Both Superoxide Dismutase and Glutathione Peroxidase Activities
Zou, Xian-Feng ; Ji, Yue-Tong ; Gao, Gui ; Zhu, Xue-Jun ; Lv, Shao-Wu ; Yan, Fei ; Han, Si-Ping ; Chen, Xing ; Gao, Chang-Cheng ; Liu, Jun-Qiu ; Luo, Gui-Min ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 88~93
DOI : 10.4014/jmb.0907.07014
Superoxide dismutase (SOD), glutathione peroxidase (GPX), and catalase (CAT) play crucial roles in balancing the production and decomposition of reactive oxygen species (ROS) in living organisms. These enzymes act cooperatively and synergistically to scavenge ROS. In order to imitate the synergism of these enzymes, we designed and synthesized a novel 32-mer peptide (32P) on the basis of the previous 15-mer peptide with GPX activity and a 17-mer peptide with SOD activity. Upon the selenation and chelation of copper, the 32-mer peptide was converted to a new Se- and Cu-containing 32-mer peptide (Se-Cu-32P) that displayed both SOD and GPX activities, and its kinetics was studied. Moreover, the novel peptide was demonstrated to be able to better protect vero cells from the injury induced by the xanthine oxidase (XOD)/xanthine/
damage system than its parents. Thus, this bifunctional enzyme imitated the synergism of SOD and GPX and could be a better candidate of therapeutic medicine.
Improvement of Ethanol Production by Electrochemical Redox Combination of Zymomonas mobilis and Saccharomyces cerevisiae
Jeon, Bo-Young ; Park, Doo-Hyun ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 94~100
DOI : 10.4014/jmb.0904.04029
Zymomonas mobilis was immobilized in a modified graphite felt cathode with neutral red (NR-cathode) and Saccharomyces cerevisiae was cultivated on a platinum plate anode. An electrochemical redox reaction was induced by 3 volts of electric potential charged to the cathode and anode. The Z. mobilis produced 1.3-1.5 M of ethanol in the cathode compartment, whereas the S. cerevisiae produced 1.7-1.9 M in the anode compartment after 96 h. The ethanol produced by the Z. mobilis immobilized in the NR-cathode and S. cerevisiae cultivated on the platinum plate was 1.5-1.6 times higher than that produced under conventional conditions. The electrochemical oxidation potential inhibited Z. mobilis, but activated S. cerevisiae. The SDS-PAGE pattern of the total soluble proteins extracted from the Z. mobilis cultivated under the electrochemical oxidation conditions was gradually simplified in proportion to the potential intensity. Z. mobilis and S. cerevisiae were cultivated in the cathode and anode compartments, respectively, of an electrochemical redox combination system. The Z. mobilis culture cultivated in the cathode compartment for 24 h was continuously transferred to the S. cerevisiae culture in the anode compartment at a rate of 300 ml/day. Approx. 1.0-1.2 M of ethanol was produced by the Z. mobilis in the cathode compartment within 24 h, and an additional 0.8-0.9 M produced by the S. cerevisiae in the anode compartment within another 24 h. Thus, a total of 2.0-2.1 M of ethanol was produced by the electrochemical redox combination of Z. mobilis and S. cerevisiae within 48 h.
Enhancement of L-Lactic Acid Production in Lactobacillus casei from Jerusalem Artichoke Tubers by Kinetic Optimization and Citrate Metabolism
Ge, Xiang-Yang ; Qian, He ; Zhang, Wei-Guo ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 101~109
DOI : 10.4014/jmb.0905.05032
Efficient L-lactic acid production from Jerusalem artichoke tubers, by Lactobacillus casei G-02, using simultaneous saccharification and fermentation (SSF) in a fed-batch culture, is demonstrated. A kinetic analysis of the SSF revealed that the inulinase activity was subjected to product inhibition, whereas the fermentation activity of G-02 was subjected to substrate inhibition. It was also found that the intracellular NADH oxidase (NOX) activity was enhanced by the citrate metabolism, which dramatically increased the carbon flux of the Embden-Meyerhof-Parnas (EMP) pathway, along with the production of ATP. As a result, when the SSF was carried out at
after an initial hydrolysis of 1 h and included a sodium citrate supplement of 10 g/l, an L-lactic acid concentration of 141.5 g/l was obtained after 30 h, with a volumetric productivity of 4.7 g/l/h. The conversion efficiency and product yield were 93.6% of the theoretical lactic acid yield and 52.4 g lactic acid/l00 g Jerusalem artichoke flour, respectively. Such a high concentration of lactic acid with a high productivity from Jerusalem artichokes has not been reported previously, making G-02 a potential candidate for the economic production of L-lactic acid from Jerusalem artichokes on a commercial scale.
Immobilization of Lactobacillus salivarius ATCC 11741 on Loofa Sponge Coated with Chitosan for Lactic Acid Fermentation
Chantawongvuti, R. ; Veerajetbodithat, J. ; Jaturapiree, P. ; Muangnapoh, C. ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 110~116
DOI : 10.4014/jmb.0905.05039
Lactic acid (LA) fermentation by Lactobacillus salivarius ATCC 11741 immobilized on loofa sponge (LS) was evaluated. To increase the surface area of LS for cell immobilization,
and chitosan were introduced as surface modifying reagents. Four chitosans of different molecular weights were separately coated on LS. All experiments were conducted in shaking flask mode at 100 rpm rotating speed and
as a pH regulating agent. The effects of initial glucose concentration were investigated in the range of 20-100 g/l on LA fermentation by free cells. The results indicate that the maximum concentration of LA was produced with 50 g/l glucose concentration. The immobilized cell system produced 1.5 times higher concentration than free cells for 24 h of fermentation. Moreover, immobilized cells can shorten the fermentation time by 2-fold compared with free cells at the same level of LA concentration. At 1% (w/v) chitosan in 2% (v/v) acetic acid, the Yp/s and productivities of various molecular weights of chitosans were insignificantly different. Repeated batch fermentations showed 5 effective recycles with Yp/s and productivity in the range of 0.55-0.85 and 0.90-1.20 g/l.h, respectively. It is evident that immobilization of L. salivarius onto LS permits reuse of the system under these fermentation conditions. Scanning electron micrographs indicated that there were more intact cells on the chitosan-treated LS than on the untreated LS, thus confirming the effectiveness of the LS-chitosan combination when being utilized as a promising immobilization carrier for LA fermentation.
Conversion of Shrimp Shell by Using Serratia sp. TKU017 Fermentation for the Production of Enzymes and Antioxidants
Wang, San-Lang ; Li, Jeng-Yu ; Liang, Tzu-Wen ; Hsieh, Jia-Lin ; Tseng, Wan-Nine ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 117~126
DOI : 10.4014/jmb.0905.05045
A chitinase (CHT) and a protease (PRO) were purified from the culture supernatant of Serratia sp. TKU017, with shrimp shell as the sole carbon/nitrogen source. The molecular masses of CHT and PRO determined by SDS-PAGE were approximately 65 kDa and 53 kDa, respectively. CHT was inhibited by
, and PRO was inhibited by most tested divalent metals and EDTA. The optimum pH, optimum temperature, pH stability, and thermal stability of CHT and PRO were pH 5,
, pH 5-7, and <
, and pH 9,
, pH 5-11, and <
, respectively. PRO retained 95% of its protease activity in the presence of 0.5 mM SDS. The result demonstrates that PRO is an SDS-resistant protease and probably has a rigid structure. The
-day supernatant showed the strongest antioxidant activity (70%, DPPH scavenging ability) and the highest total phenolic content (
of gallic acid equiv./ml). Significant associations between the antioxidant potency and the total phenolic content, as well as between the antioxidant potency and free amino groups, were found for the supernatant. With this method, we have shown that shrimp shell wastes can be utilized and it is effective in the production of enzymes and antioxidants, facilitating its potential use in industrial applications and functional foods.
Enhancement of Ornithine Production in Proline-Supplemented Corynebacterium glutamicum by Ornithine Cyclodeaminase
Lee, Soo-Youn ; Cho, Jae-Yong ; Lee, Hyun-Jeong ; Kim, Yang-Hoon ; Min, Ji-Ho ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 127~131
DOI : 10.4014/jmb.0907.07034
In this study, Corynebacterium glutamicum and its derived mutants were used to demonstrate the relationship between proline, glutamate, and ornithine. The maximum ornithine production was shown in the culture medium (3,295.0 mg/l) when the cells were cultured with 20 mM proline, and was 15.5 times higher than in the presence of 1 mM proline. However, glutamate, which is known as an intermediate in the process of converting proline to ornithine, did not have any positive effect on ornithine production. This suggests that the conversion of proline to ornithine through glutamate, is not possible in C. glutamicum. Comparative analysis between the wild-type strain, SJC 8043 (
), and SJC 8064 (
), showed that C glutamicum could regulate ornithine production by ornithine cyclodeaminase (Ocd) under proline-supplemented conditions. Therefore, proline directly caused an increase in the endogenous level of ornithine by Ocd, which would be a primary metabolite in the ornithine biosynthesis pathway.
Heterologous Production of Streptokinase in Secretory Form in Streptomyces lividans and in Nonsecretory Form in Escherichia coli
Kim,, Mi-Ran ; Choeng, Yong-Hoon ; Chi, Won-Jae ; Kang, Dae-Kyung ; Hong, Soon-Kwang ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 132~137
DOI : 10.4014/jmb.0906.06005
The skc gene encoding streptokinase (SK) with a molecular mass of approximately 47.4 kDa was cloned from Streptococcus equisimilis ATCC 9542 and heterologously overexpressed in Streptomyces lividans TK24 and E. coli using various strong promoters. When the promoter for sprT [Streptomyces griseus trypsin (SGT)] was used in the host S. lividans TK24, a 47.4-kDa protein was detected along with a smaller hydrolyzed protein (44 kDa), suggesting that posttranslational hydrolysis had occurred as has been reported in other expression systems. The casein/plasminogen plate assay revealed that the plasmid construct containing the SGT signal peptide was superior to that containing the SK signal peptide in terms of SK production. Maximal production of SK was calculated to be about 0.25 unit/ml of culture broth, a value that was five times higher than that obtained with other expression systems using ermE and tipA promoters in the same host. When the skc gene was expressed in E. coli BL21(
)pLys under the control of the T7 promoter, a relatively large amount of SK was expressed in soluble form without hydrolysis. SK activity in E. coli/pET28a-
was more than 2 units/ml of culture broth, even though about half of the expressed protein formed an inactive inclusion body.
Production of Biosurfactant Lipopeptides Iturin A, Fengycin, and Surfactin A from Bacillus subtilis CMB32 for Control of Colletotrichum gloeosporioides
Kim, Pyoung-Il ; Ryu, Jae-Won ; Kim, Young-Hwan ; Chi, Youn-Tae ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 138~145
DOI : 10.4014/jmb.0905.05007
A bacterial strain isolated from soil for its potential to control the anthracnose disease caused by Colletotrichum gloeosporioides was identified as a Bacillus subtilis. Bacillus subtilis CMB32 produced antifungal agents on M9 broth at
. Biosurfactant lipopeptides produced by Bacillus subtilis CMB32 were precipitated by adjusting to pH 2 and extracting using chloroform/methanol, and then were purified using column chromatography and reverse-phase HPLC. The molecular masses of the lipopeptides were estimated by MALDI-TOF mass spectrometry as (a) 1,080, (b) 1,486, and (c) 1,044 Da, respectively. They had cyclic structures and amino acid compositions of (a) Pro, Asx, Ser, Tyr, Glx, (b) Glx, Tyr, Thr, Ala, Pro, lie, and (c) Glx, Leu, Val, Asx, respectively. Further analysis revealed that Bacillus subtilis CMB32 produced three antifungal lipopeptides: (a) iturin A, (b) fengycin, and (c) surfactin A.
Enhancement of Clavulanic Acid Production by Expressing Regulatory Genes in gap Gene Deletion Mutant of Streptomyces clavuligerus NRRL3585
Jnawali, Hum Nath ; Lee, Hei-Chan ; Sohng, Jae-Kyung ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 146~152
DOI : 10.4014/jmb.0907.07020
Streptomyces clavuligerus NRRL3585 produces a clinically important
-lactamase inhibitor, clavulanic acid (CA). In order to increase the production of CA, the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene (gap) was deleted in S. clavuligerus NRRL3585 to overcome the limited glyceraldehyde-3-phosphate pool; the replicative and integrative expressions of ccaR (specific regulator of the CA biosynthetic operon) and claR (Lys-type transcriptional activator) genes were transformed together into a deletion mutant to improve clavulanic acid production. We constructed two recombinant plasmids to enhance the production of CA in the gap1 deletion mutant of S. clavuligerus NRRL3585: pHN11 was constructed for overexpression of ccaR-claR, whereas pHN12 was constructed for their chromosomal integration. Both pHN11 and pHN12 transformants enhanced the production of CA by 2.59-fold and 5.85-fold, respectively, compared with the gap1 deletion mutant. For further enhancement of CA, we fed the pHN11 and pHN12 transformants ornithine and glycerol. Compared with the gap1 deletion mutant, ornithine increased CA production by 3.24- and 6.51-fold in the pHN11 and pHN12 transformants, respectively, glycerol increased CA by 2.96- and 6.21-fold, respectively, and ornithine and glycerol together increased CA by 3.72- and 7.02-fold, respectively.
Bacteriocinogenic Potential of Newly Isolated Strains of Enterococcus faecium and Enterococcus faecalis from Dairy Products of Pakistan
Javed, Imran ; Ahmed, Safia ; Ali, Muhammad Ishtiaq ; Ahmad, Bashir ; Ghumro, Pir Bux ; Hameed, Abdul ; Chaudry, Ghulam Jilani ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 153~160
DOI : 10.4014/jmb.0904.04024
The present study was carried out for the isolation of bacteriocin-producing enterococci from indigenous sources. Gram-positive enterococci are known for having the ability to produce enterocins with good antimicrobial potential. A total of 34 strains were isolated from processed dairy products of Pakistan and seven out of them were found to be member of genus Enterococcus on selective enumeration. Biochemical and molecular characterization revealed that four of these isolates (IJ-03, IJ-07, IJ-11, and IJ-12) were Enterococcus faecalis and three (IJ-06, IJ-21, and IJ-31) were Enterococcus faecium. Local processed cheese was the source of all enterococcal isolates, except E. faecium IJ-21 and IJ-31, which were isolated from indigenous yoghurt and butter samples, respectively. Bacterial isolates were sensitive to commonly used antibiotics except methicillin and kanamycin. They also lacked critical virulence determinants, mainly cytolysin (cyl), gelatinase (gel), enterococcal surface protein (esp), and vancomycin resistance (vanA and vanB). Polymerase chain reaction amplification identified that enterocin A and P genes were present in the genome of E. faecium IJ-06 and IJ-21, whereas the E. faecium IJ-31 genome showed only enterocin P genes. No amplification was observed for genes that corresponded with the enterocins 31, AS-48, L50A, and L50B, and ent 1071A and 1071B. There were no signals of amplification found for E. faecalis IJ-11, indicating that the antimicrobial activity was because of an enterocin different from those checked by PCR. Hence, the indigenous bacterial isolates have great potential for bacteriocin production and they had antibacterial activity against a variety of closely related species.
Assessment of Potential Probiotic and Starter Properties of Pediococcus spp. Isolated from Turkish-Type Fermented Sausages (Sucuk)
Yuksekdag, Z. Nur ; Aslim, Belma ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 161~168
DOI : 10.4014/jmb.0904.04019
In this study, the metabolic activities of five strains of Pediococcus spp., in terms of the quantities they produced of lactic acid, hydrogen peroxide, exopolysaccharides, and proteolytic activity, were determined. Lactic acid levels produced by these strains were found to be in the range of 2.5-5.6 mg/ml. All strains produced hydrogen peroxide. The P. pentosaceus Z13P strain produced the maximum amount (0.25 mg/ml) of proteolytic activity. Exopolysaccharide (EPS) production by the Pediococcus strains during growth in MRS (de Man, Rogosa, and Sharpe) medium was in the range 25-64 mg/l. The susceptibility of 10 different antibiotics against these strains was also tested. All strains were found to be resistant to amoxicillin, gentamicin, and vancomycin. Antimicrobial effects of the Pediococcus spp. on pathogens were also determined by an agar diffusion method. All of the strains were able to inhibit L. monocytogenes. The tolerance of the strains to low pH, their resistance to bile salts of strains, and their abilities to autoaggregate and coaggregate with L. monocytogenes were also evaluated.
Structure and Diversity of Arsenic-Resistant Bacteria in an Old Tin Mine Area of Thailand
Jareonmit, Pechrada ; Sajjaphan, Kannika ; Sadowsky, Michael J. ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 169~178
DOI : 10.4014/jmb.0906.06026
The microbial community structure in Thailand soils contaminated with low and high levels of arsenic was determined by denaturing gradient gel electrophoresis. Band pattern analysis indicated that the bacterial community was not significantly different in the two soils. Phylogenetic analysis obtained by excising and sequencing six bands indicated that the soils were dominated by Arthrobacter koreensis and
-Proteobacteria. Two hundred and sixty-two bacterial isolates were obtained from arsenic-contaminated soils. The majority of the As-resistant isolates were Gramnegative bacteria. MIC studies indicated that all of the tested bacteria had greater resistance to arsenate than arsenite. Some strains were capable of growing in medium containing up to 1,500 mg/l arsenite and arsenate. Correlations analysis of resistance patterns of arsenite resistance indicated that the isolated bacteria could be categorized into 13 groups, with a maximum similarity value of 100%. All strains were also evaluated for resistance to eight antibiotics. The antibiotic resistance patterns divided the strains into 100 unique groups, indicating that the strains were very diverse. Isolates from each antibiotic resistance group were characterized in more detail by using the repetitive extragenic palindromic-PCR (rep-PCR) DNA fingerprinting technique with ERIC primers. The PCR products were analyzed by agarose gel electrophoresis. The genetic relatedness of 100 bacterial fingerprints, determined by using the Pearson product-moment similarity coefficient, showed that the isolates could be divided into four clusters, with similarity values ranging from 5-99%. Although many isolates were genetically diverse, others were clonal in nature. Additionally, the arsenic-resistant isolates were examined for the presence of arsenic resistance (ars) genes by using PCR, and 30% of the isolates were found to carry an arsenate reductase encoded by the arsC gene.
Anaerobic Lipid Degradation Through Acidification and Methanization
Kim,, I-Jung ; Kim, Sang-Hyoun ; Shin, Hang-Sik ; Jung, Jin-Young ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 179~186
DOI : 10.4014/jmb.0903.03020
In biological wastewater treatment, high lipid concentrations can inhibit the activity of microorganisms critical to the treatment process and cause undesirable biomass flotation. To reduce the inhibitory effects of high lipid concentrations, a two-phase anaerobic system, consisting of an anaerobic sequencing batch reactor (ASBR) and an upflow anaerobic sludge blanket (UASB) reactor in series, was applied to synthetic dairy wastewater treatment. During 153 days of operation, the two-phase system showed stable performance in lipid degradation. In the ASBR, a 13% lipid removal efficiency and 10% double-bond removal efficiency were maintained. In the UASB, the chemical oxygen demand (COD), lipid, and volatile fatty acid (VFA) removal efficiencies were greater than 80%, 70%, and 95%, respectively, up to an organic loading rate of 6.5 g COD/l/day. No serious operational problems, such as significant scum formation or sludge washout, were observed. Protein degradation was found to occur prior to degradation during acidogenesis.
Investigation of Possible Horizontal Gene Transfer from Transgenic Rice to Soil Microorganisms in Paddy Rice Field
Kim, Sung-Eun ; Moon, Jae-Sun ; Kim, Jung-Kyu ; Choi, Won-Sik ; Lee, Sang-Han ; Kim, Sung-Uk ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 187~192
DOI : 10.4014/jmb.0905.05050
In order to monitor the possibility of horizontal gene transfer between transgenic rice and microorganisms in a paddy rice field, the gene flow from a bifunctional fusion (TPSP) rice containing trehalose-6-phosphate synthase and phosphatase to microorganisms in soils was investigated. The soil samples collected from the paddy rice field during June 2004 to March 2006 were investigated by multiplex PCR, Southern hybridization, and amplified fragment length polymorphism (AFLP). The TPSP gene from soil genomic DNAs was not detected by PCR. Soil genomic DNAs did not show homologies on the Southern blotting data, indicating that gene transfer did not occur during the last two years in the paddy rice field. In addition, the AFLP band patterns produced by soil genomic DNAs from both transgenic and non-transgenic rice fields appeared similar to each other when analyzed by the NTSYSpc program. Thus, these data suggest that transgenic rice does not give a significant impact on the communities of soil microorganisms, although long-term observation may be needed.
A Real-Time PCR Assay for the Quantitative Detection of Ralstonia solanacearum in Horticultural Soil and Plant Tissues
Chen, Yun ; Zhang, Wen-Zhi ; Liu, Xin ; Ma, Zhong-Hua ; Li, Bo ; Allen, Caitilyn ; Guo, Jian-Hua ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 193~201
DOI : 10.4014/jmb.0906.06019
A specific and rapid real-time PCR assay for detecting Ralstonia solanacearum in horticultural soil and plant tissues was developed in this study. The specific primers RSF/RSR were designed based on the upstream region of the UDP-3-O-acyl-GlcNAc deacetylase gene from R. solanacearum, and a PCR product of 159 bp was amplified specifically from 28 strains of R. solanacearum, which represent all genetically diverse AluI types and all 6 biovars, but not from any other nontarget species. The detection limit of
tomato stem and horticultural soil was achieved in this real-time PCR assay. The high sensitivity and specificity observed with field samples as well as with artificially infected samples suggested that this method might be a useful tool for detection and quantification of R. solanacearum in precise forecast and diagnosis.
Isolation of a Gibberellin-producing fungus (Penicillium sp. MH7) and Growth Promotion of Crown Daisy (Chrysanthemum coronarium)
Hamayun, Muhammad ; Khan, Sumera Afzal ; Iqbal, Ilyas ; Ahmad, Bashir ; Lee, In-Jung ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 202~207
DOI : 10.4014/jmb.0905.05040
Plant growth promoting fungi (PGPF) are well known for the production of useful secondary metabolites. However, limited information is available on the gibberellin (GA) production capacity of PGPF of endophytic origin. In the current study, 15 fungal endophytes were isolated from the roots of Crown daisy, and then screened on Waito-c rice, in order to identify plant growth promoting fungi. The fungal isolate MH7 significantly increased the shoot length (12.1 cm) of Waito-c in comparison with control treatment (7.9 cm). In a separate experiment, the culture filtrate (CF) of MH7 significantly promoted the growth attributes of Crown daisy. The MH7 CF was analyzed for gibberellins and it contained all physiologically active gibberellins (
, 1.37 ng/ml;
, 5.88 ng/ml;
, 8.62 ng/ml; and
, 2.05 ng/ml) in conjunction with physiologically inactive
(1.16 ng/ml), and
(0.98 ng/ml). The CF of MH7 produced higher amounts of
than wild-type Fusarium fujikuroi, which was used as a control for GA production. The fungal isolate MH7 was later identified as a new strain of Penicillium on the basis of its morphological characteristics and phylogenetic analysis of the 188 rDNA sequence.
The Gene fpk1, Encoding a cAMP-dependent Protein Kinase Catalytic Subunit Homolog, is Required for Hyphal Growth, Spore Germination, and Plant Infection in Fusarium verticillioides
Pei-Bao, Zhao ; Ren, Ai-Zhi ; Xu, Hou-Juan ; Li, Duo-Chuan ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 208~216
DOI : 10.4014/jmb.0905.05044
Fusarium verticillioides is an important pathogen of maize, being responsible for ear rots, stalk rots, and seedling blight worldwide. During the past decade, F. verticillioides has caused several severe epidemics of maize seedling blight in many areas of China, which lead to significant losses. In order to understand the molecular mechanisms regulating fungal development and pathogenicity in this pathogen, we isolated and characterized the gene fpk1 (GenBank Accession No. EF405959) encoding a homolog of the cAMP-dependent protein kinase catalytic subunit, which included a 1,854-bp DNA sequence from ATG to TAA, with a 1,680-bp coding region, and three introns (lengths: 66 bp, 54 bp, and 54 bp), and the predicated protein precursor had 559 aa. The mutant
, which was disrupted of the fpkl gene, showed reduced vegetative growth, fewer and shorter aerial mycelia, strongly impaired conidiation, and reduced spore germination rate. After germinating, the fresh hypha was stubby and lacking of branch. When inoculated in susceptible maize varieties, the infection of the mutant
was delayed and the infection efficiency was reduced compared with that of the wild-type strain. AU this indicated that gene fpk1 participated in hyphal growth, conidiophore production, spore germination, and virulence in F. verticillioides.
Antiallergic Effects of Fermented Ixeris sonchifolia and Its Constituents in Mice
Trinh, Hien-Trung ; Bae, Eun-Ah ; Hyun, Yang-Jin ; Jang, Yoon-Ah ; Yun, Hyung-Kwon ; Hong, Seong-Sig ; Kim, Dong-Hyun ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 217~223
DOI : 10.4014/jmb.0904.04015
To evaluate the antiallergic effect of fermented Ixeris sonchifolia (IS, family Compositae), we prepared IS kimchi, isolated lactic acid bacteria (LAB) from it, fermented IS with these LAB, and investigated their antiallergic effects. IS kimchi inhibited the passive cutaneous anaphylaxis (PCA) reaction induced by an IgE-antigen complex as well as the scratching behavior induced by compound 48/80 or histamine more potently than IS. When IS was fermented with LAB isolated from IS kimchi, its antiallergic effects was also increased. Of LAB used for fermentation, Lactobacillus brevis more potently increased the antiallergic effects. Its main constituents, chlorogenic acid and luteolin, potently inhibited the PCA reaction induced by the IgE-antigen complex as well as the pruritis induced by compound 48/80 or histamine. These constituents inhibited the expression of pro inflammatory and allergic cytokines, TNF-
. and IL-4, and transcription factor NF-
activation induced by the IgE-antigen complex in RBL-2H3 cells, as well as the degranulation of RBL-2H3 cells induced by the IgE-antigen complex. Luteolin more potently inhibited these allergic reactions than chlorogenic acid. These findings suggest that the antiallergic effect of IS can be increased by LAB fermentation, and the fermented IS might improve allergic reactions such as pruritus, anaphylaxis, and inflammation.
Induced Death of Escherichia coli Encapsulated in a Hollow Fiber Membrane as Observed In Vitro or After Subcutaneous Implantation
Granicka, L. H. ; Zolnierowicz, J. ; Wasilewska, D. ; Werynski, A. ; Kawiak, J. ;
Journal of Microbiology and Biotechnology, volume 20, issue 1, 2010, Pages 224~228
DOI : 10.4014/jmb.0906.06056
The encapsulation of bacteria may be used to harness them for longer periods of time in order to make them viable, whereas antibiotic treatment would result in controlled release of therapeutic molecules. Encapsulated Escherichia coli GFP (green fluorescent protein) (E. coli GFP) was used here as a model for therapeutic substance - GFP fragments release (model of bioactive substances). Our aim was to evaluate the performance of bacteria encapsulated in hollow fibers (HFs) treated with antibiotic for induction of cell death. The polypropylene-surface-modified HFs were applied for E. coli encapsulation. The encapsulated bacteria were treated with tetracycline in vitro or in vivo during subcutaneous implantation into mice. The HF content was evaluated in a flow cytometer, to assess the bacteria cell membrane permeability changes induced by tetracycline treatment. It was observed that the applied membranes prevented release of bacteria through the HF wall. The E. coli GFP culture encapsulated in HF in vitro proved the tetracycline impact on bacteria viability and allows the recognition of the sequence of events within the process of bacteria death. Treatment of the SCID mice with tetracycline for 8 h proved the tetracycline impact on bacteria viability in vivo, raising the necrotic bacteria-releasing GFP fragments. It was concluded that the bacteria may be safely enclosed within the HF at the site of implantation, and when the animal is treated with antibiotic, bacteria may act as a local source of fragments of proteins expressed in the bacteria, a hypothetical bioactive factor for the host eukaryotic organism.