Journal of Microbiology

Volume 48(2); April 2010

Research Support, Non-U.S. Gov'ts

Microbial Community on Healthy and Diseased Leaves of an Invasive Plant Eupatorium adenophorum in Southwest China: Zhen-Xin Zhou , Huan Jiang , Chen Yang , Ming-Zhi Yang , Han-Bo Zhang; J. Microbiol. 2010;48(2):139-145. Published online May 1, 2010; DOI: https://doi.org/10.1007/s12275-010-9185-y

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27 Citations

Abstract: Invasive plants have caused great economic losses and environmental problems worldwide. Eupatorium adenophorum is one of the most invasive weeds in China. To better understand its invasive mechanisms, in the present paper, the microbial communities of healthy and diseased leaves of E. adenophorum were obtained using both culture-independent and -dependent methods and their diversities were compared. The bacteria obtained from culture-independent method belong to Proteobacteria (95.8%), Actinobacteria (2.1%), and Firmicutes (2.1%) and fungi belong to Ascomycota (65.2%) and Basidiomycota (34.8%). Very few overlapped microbial species were found by culture-dependent and -independent methods. Healthy leaves display higher bacterial diversity than diseased leaves. Phylogenetic structures are very different between healthy and diseased phyllosphere microbial communities. Bacteria close to Acinetobacter and Pseudomonas were dominant on healthy leaves, whereas those close to Shigella were dominant on diseased leaves. 52.9% of fungal clones from healthy leaves were Ustilaginomycetes, close to Rhodotorula phylloplana and uncultured basidomycete; by contrast, 60% of clones from diseased leaves were Lecanoromycetes, close to Umbilicaria muehlenbergii. No bacteria but four fungal strains phylogenetically close to Myrothecium sp. and Alternaria alternate were pathogenic to seedlings and detached leaves of the invasive plant. Therefore, this plant may be resistant to pathogens from bacteria but not fungi in its introduced range.

Diversity of Thermophilic Fungi in Tengchong Rehai National Park Revealed by ITS Nucleotide Sequence Analyses: Wen-Zheng Pan , Xiao-Wei Huang , Kang-Bi Wei , Chun-Mei Zhang , Dong-Mei Yang , Jun-Mei Ding , Ke-Qin Zhang; J. Microbiol. 2010;48(2):146-152. Published online May 1, 2010; DOI: https://doi.org/10.1007/s12275-010-9157-2

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36 Citations

Abstract: The geothermal sites near neutral and alkalescent thermal springs in Tengchong Rehai National Park were examined through cultivation-dependent approach to determine the diversity of thermophilic fungi in these environments. Here, we collected soils samples in this area, plated on agar media conducive for fungal growth, obtained pure cultures, and then employed the method of internal transcribed spacer (ITS) sequencing combined with morphological analysis for identification of thermophilic fungi to the species level. In total, 102 strains were isolated and identified as Rhizomucor miehei, Chaetomium sp., Talaromyces thermophilus, Talaromyces byssochlamydoides, Thermoascus aurantiacus Miehe var. levisporus, Thermomyces lanuginosus, Scytalidium thermophilum, Malbranchea flava, Myceliophthora sp. 1, Myceliophthora sp. 2, Myceliophthora sp. 3, and Coprinopsis sp. Two species, T. lanuginosus and S. thermophilum were the dominant species, representing 34.78% and 28.26% of the sample, respectively. Our results indicated a greater diversity of thermophilic fungi in neutral and alkaline geothermal sites than acidic sites around hot springs reported in previous studies. Most of our strains thrived at alkaline growth conditions.

NtrC-Sensed Nitrogen Availability Is Important for Oxidative Stress Defense in Pseudomonas putida KT2440: Sujin Yeom , Jinki Yeom , Woojun Park; J. Microbiol. 2010;48(2):153-159. Published online May 1, 2010; DOI: https://doi.org/10.1007/s12275-010-0075-0

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15 Citations

Abstract: The zwf, which encodes glucose-6-phosphate dehydrogenase, is repressed by NtrC under nitrogen-limited condition. Previously, we demonstrated that induction of zwf-1 is required for protecting Pseudomonas putida cells under oxidative stress, which could be possible probably because of derepression of HexR on the zwf-1 gene under oxidative stress. These findings led us investigate that NtrC still represses the zwf-1 under nitrogen-limited oxidative stress condition, which makes cells more sensitive under such condition. Interestingly, deletion of the ntrC gene significantly reduces growth rate, but renders cells more resistant to oxidative stress, under nitrogen limited condition in P. putida. More vitality of the ntrC mutant under oxidative stress condition was also confirmed by the fluorogenic redox dye using flow cytometry. The results of transcriptome analysis demonstrated that the derepression of several oxidative stress genes along with the zwf-1 gene might confer high resistance to oxidative stress in the ntrC mutant. Here, we presented the data for the first time, showing that different sets of genes are involved in nitrogen-rich and nitrogen-limited oxidative stress conditions and NtrC-sensed nitrogen availability is one of the most important prerequisite for full cellular defense against oxidative stress in P. putida.

Journal Article

Psychroflexus lacisalsi sp. nov., a Moderate Halophilic Bacterium Isolated from a Hypersaline Lake (Hunazoko-Ike) in Antarctica: Hongyan Zhang , Shoko Hosoi-Tanabe , Syuhei Ban , Satoshi Imura; J. Microbiol. 2010;48(2):160-164. Published online May 1, 2010; DOI: https://doi.org/10.1007/s12275-010-0018-9

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11 Citations

Abstract: A novel Gram-negative, aerobic, moderate halophilic, and psychrotolerant bacterium, designated as strain H7T, was isolated from a hypersaline lake located in Skarvsnes, Antarctica. Cells were filaments with varying lengths. Coccoid bodies developed in old cultures. Growth occurred with 0.5-15% (w/v) NaCl (optimum, 5.8-7.0%), at pH 6.0-10.0 (optimum, pH 7.0-8.0), and at 10-28°C (optimum, 25°C). The strain had a G+C content of 34.9 mol%, which is within the range of 32-36 mol% reported for the genus Psychroflexus. Chemotaxonomic data (major respiratory quinone: MK-6; major fatty acids: aC15:0, iC16:0 3-OH, and aC15: 1 A) supported the classification of strain H7T within the genus Psychroflexus. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain H7T should be assigned to the genus Psychroflexus and has a homology with Psychroflexus salinarum (98.2%), P. sediminis (96.1%), P. torquis (95.2%), P. tropicus (95.8%), and P. gondwanense (92.2%). Strain H7 is not identified as P. salinarum because that DNA-DNA hybridization data were 8.5% between strain H7T and P. salinarum. The combination of phylogenetic analysis, DNA-DNA hybridization data, phenotypic characteristics, and chemotaxonomic differences supported the view that strain H7T represents a novel species of the genus Psychroflexus. The name Psychroflexus lacisalsi is proposed, and the type strain is H7T (=JCM 16231T =KACC 14089T).

Research Support, Non-U.S. Gov'ts

Sphingomonas humi sp. nov., Isolated from Soil: Tae Hoo Yi , Chang-Kyun Han , Sathiyaraj Srinivasan , Kang Jin Lee , Myung Kyum Kim; J. Microbiol. 2010;48(2):165-169. Published online May 1, 2010; DOI: https://doi.org/10.1007/s12275-010-0011-3

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12 Citations

Abstract: A Gram-negative, non-motile, non-spore-forming, small, orange, rod-shaped bacterium was isolated from soil in South Korea and characterized to determine its taxonomic position. Phylogenetic analysis based on 16S rRNA gene sequence examination revealed that strain PB323T belongs to the family Sphingomonadaceae. The highest degree of sequence similarity was found with Sphingomonas kaistensis PB56T (98.9%), followed by Sphingomonas astaxanthinifaciens TDMA-17T (98.3%). Chemotaxonomic characteristics (the G+C content of the genomic DNA 69.0 mol%, Q-10 quinone system, C18:1ω7c/ω9t/ω12t, C16:1ω7c/C15:0 iso 2OH, C17:1ω6c, and C16:0 as the major fatty acids) corroborated assignment of strain PB323T to the genus Sphingomonas. Results of physiological and biochemical tests clearly demonstrate that strain PB323T represents a distinct species and support its affiliation with the genus Sphingomonas. Based on these data, PB323T (=KCTC 12341T =JCM 16603T =KEMB 9004-003T) should be classified as a type strain of a novel species, for which the name Sphingomonas humi sp. nov. is proposed.

In Vivo Studies with a Candida tropicalis Isolate Exhibiting Paradoxical Growth In Vitro in the Presence of High Concentration of Caspofungin: Sedigh Bayegan , Laszlo Majoros , Gabor Kardos , Adam Kemény-Beke , Cecilia Miszti , Renato Kovacs , Rudolf Gesztelyi; J. Microbiol. 2010;48(2):170-173. Published online May 1, 2010; DOI: https://doi.org/10.1007/s12275-010-9221-y

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22 Citations

Abstract: We investigated the activity of caspofungin against a Candida tropicalis clinical isolate showing paradoxical growth in vitro. BALB/c mice immunosuppressed by cyclophosphamide were infected intraperitoneally using 107 CFU/mouse. Caspofungin was administered intraperitoneally once daily for 5 days or as a single dose using the following doses: 0.12, 0.25, 1, 2, 3, 5, and 15 mg/kg. The single dose of caspofungin was effective only at 5 and 15 mg/kg concentrations (100% survival). Five-day caspofungin treatment led to 100% survival at doses of 1 mg/kg or higher. Caspofungin treatment significantly decreased the number of viable yeasts in the peritoneal lavage samples as well as in the infected abscesses at doses 1, 3, 5, and 15 mg/kg caspofungin as compared to the untreated control (P<0.001 in all cases), and even to the group treated with 0.12 mg/kg caspofungin (P<0.05 in all cases). At 2 mg/kg caspofungin dose, sterilization of the internal organs was reproducibly incomplete, suggesting that the role of paradoxical growth in the late clinical failure cannot be excluded.

Experimental and Computational Characterization of the Ferric Uptake Regulator from Aliivibrio salmonicida (Vibrio salmonicida): Hege Lynum Pedersen , Rafi Ahmad , Ellen Kristin Riise , Hanna-Kirsti Schrøder Leiros , Stefan Hauglid , Sigrun Espelid , Bjørn Olav Brandsdal , Ingar Leiros , Nils-Peder Willassen , Peik Haugen; J. Microbiol. 2010;48(2):174-183. Published online May 1, 2010; DOI: https://doi.org/10.1007/s12275-010-9199-5

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5 Citations

Abstract: The Ferric uptake regulator (Fur) is a global transcription factor that affects expression of bacterial genes in an iron-dependent fashion. Although the Fur protein and its iron-responsive regulon are well studied, there are still important questions that remain to be answered. For example, the consensus Fur binding site also known as the “Fur box” is under debate, and it is still unclear which Fur residues directly interact with the DNA. Our long-term goal is to dissect the biological roles of Fur in the development of the disease cold-water vibriosis, which is caused by the psychrophilic bacteria Aliivibrio salmonicida (also known as Vibrio salmonicida). Here, we have used experimental and computational methods to characterise the Fur protein from A. salmonicida (AS-Fur). Electrophoretic mobility shift assays show that AS-Fur binds to the recently proposed vibrio Fur box consensus in addition to nine promoter regions that contain Fur boxes. Binding appears to be dependent on the number of Fur boxes, and the predicted “strength” of Fur boxes. Finally, structure modeling and molecular dynamics simulations provide new insights into potential AS-Fur–DNA interactions.

Journal Article

Sequence Analysis of the Gene Encoding H Antigen in Escherichia coli Isolated from Food in Morocco: Samira Badri , Aziz Fassouane , Ingrid Filliol , Mohammed Hassar , Nozha Cohen; J. Microbiol. 2010;48(2):184-187. Published online May 1, 2010; DOI: https://doi.org/10.1007/s12275-010-9182-1

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2 Citations

Abstract: In order to develop other molecular method useful for typing of motile and non motile Escherichia coli strains, a total of 207 strains of E. coli (133 reference strains, 74 food strains) were characterized by analysis of sequences of their amplified flagellin-encoding (fliC) gene products. The collection of reference strains was used for database building of fliC gene sequences. Application of this identification system to 74 E. coli food isolates revealed a reproducible and clear cut classification with very good correlation to results obtained by HhaI restriction of the amplified flagellin gene. The proposed determination of fliC sequences variations should be helpful for epidemiological studies.

Research Support, Non-U.S. Gov'ts

Differential Expression of citA Gene Encoding the Mitochondrial Citrate Synthase of Aspergillus nidulans in Response to Developmental Status and Carbon Sources: In Sook Min , Ji Young Bang , Soon Won Seo , Cheong Ho Lee , Pil Jae Maeng; J. Microbiol. 2010;48(2):188-198. Published online May 1, 2010; DOI: https://doi.org/10.1007/s12275-010-0096-8

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6 Citations

Abstract: As an extension of our previous studies on the mitochondrial citrate synthase of Aspergillus nidulans and cloning of its coding gene (citA), we analyzed differential expression of citA in response to the progress of development and change of carbon source. The cDNA consisted of 1,700 nucleotides and was predicted to encode a 474-amino acid protein. By comparing the cDNA sequence with the corresponding genomic sequence, we confirmed that citA gene contains 7 introns and that its transcription starts at position -26 (26-nucleotide upstream from the initiation codon). Four putative CreA binding motifs and three putative stress-response elements (STREs) were found within the 1.45-kb citA promoter region. The mode of citA expression was examined by both Northern blot and confocal microscopy using green fluorescent protein (sGFP) as a vital reporter. During vegetative growth and asexual development, the expression of citA was ubiqiutous throughout the whole fungal body including mycelia and conidiophores. During sexual development, the expression of citA was quite strong in cleistothecial shells, but significantly weak in the content of cleistothecia including ascospores. Acetate showed a strong inductive effect on citA expression, which is subjected to carbon catabolite repression (CCR) caused by glucose. The recombinant fusion protein CitA40::sGFP (sGFP containing the 40-amino acid N-terminal segment of CitA) was localized into mitochondria, which supports that a mitochondrial targeting signal is included within the 40-amino acid N-terminal segment of CitA.

Functional Analysis of the Inhibitor of Apoptosis Genes in Antheraea pernyi Nucleopolyhedrovirus: Feng Yan , Xiaobei Deng , Junpeng Yan , Jiancheng Wang , Lunguang Yao , Songya lv , Yipeng Qi , Hua Xu; J. Microbiol. 2010;48(2):199-205. Published online May 1, 2010; DOI: https://doi.org/10.1007/s12275-010-9108-y

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14 Citations

Abstract: The inhibitor of apoptosis proteins (IAP) plays an important role in cell apoptosis. We cloned two novel IAP family members, Ap-iap1 and Ap-iap2, from Antheraea pernyi nucleopolyhedrovirus (ApNPV) genome. Ap-IAP1 contains two baculoviral IAP repeat (BIR) domains followed by a RING domain, but Ap-IAP2 has only one BIR domain and RING. The result of transient expression in Spodoptera frugiperda (Sf21) showed that Ap-iap1 blocked cell apoptosis induced by actinomycin D treatment and also rescued the p35 deficient Autographa californica nucleopolyhedrovirus (AcNPV) to replicate in Sf9 cells, while Ap-iap2 does not have this function. Several Ap-IAP1 truncations were constructed to test the activity of BIRs or RING motif to inhibit cell apoptosis. The results indicated that BIRs or RING of Ap-IAP1 had equally function to inhibit cell apoptosis. Therefore deletion of above both of the above domains could not block apoptosis induced by actinomycin D or rescue the replication of AcMNPV△p35. We also screened two phage-display peptides that might interact with Ap-IAP1.

Kaposi’s Sarcoma-Associated Herpesvirus Viral Protein Kinase Interacts with RNA Helicase A and Regulates Host Gene Expression: Jae Eun Jong , Junsoo Park , Sunmi Kim , Taegun Seo; J. Microbiol. 2010;48(2):206-212. Published online May 1, 2010; DOI: https://doi.org/10.1007/s12275-010-0021-1

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13 Citations

Abstract: RNA helicase A (RHA) containing the DExH motif is a human homolog of maleless protein that regulates expression of genes located in the Drosophila X chromosome during dosage compensation. RHA exerts helicase activity that unwinds double-stranded RNA and DNA to a single-strand form. The protein acts as a bridging factor mediating interactions of CBP/p300 and RNA pol II, and consequently affects gene expression. Kaposi’s sarcoma-associated herpesvirus (KSHV) is a member of the γ-herpesvirus subfamily that causes several disorders. The majority of herpesviruses commonly encode predicted viral protein kinases. KSHV open reading frame 36 (ORF36) codes for protein kinase domains, and functions as a serine/threonine protein kinase. KSHV ORF36 is classified as a late gene, as it is expressed during lytic replication and localized in the nuclei of KSHV-infected cells. Recent studies show that viral protein kinase (vPK) interacts with cellular proteins. In this study, we determined the cellular localization of vPK in KSHVinfected BCBL-1 cells using confocal microscopy. Proteomic analysis indicates that cellular proteins interacted with vPK, and co-immunoprecipitation reactions further reveal interactions between vPK and RHA. Moreover, KSHV vPK appeared to regulate the transcriptional activation of Cre promoter, and plays an important role in cellular transcription of RHA.

Phenotypic Characterization and Genomic Analysis of the Shigella sonnei Bacteriophage SP18: Kyoung-Ho Kim , Ho-Won Chang , Young-Do Nam , Seong Woon Roh , Jin-Woo Bae; J. Microbiol. 2010;48(2):213-222. Published online May 1, 2010; DOI: https://doi.org/10.1007/s12275-010-0055-4

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12 Citations

Abstract: A novel bacteriophage that infects Shigella sonnei was isolated from the Gap River in Korea, and its phenotypic and genomic characteristics were investigated. The virus, called SP18, showed morphology characteristic of the family Myoviridae, and phylogenetic analysis of major capsid gene (gp23) sequences classified it as a T4-like phage. Based on host spectrum analysis, it is lytic to S. sonnei, but not to Shigella flexneri, Shigella boydii or members of the genera Escherichia and Salmonella. Pyrosequencing of the SP18 bacteriophage genome revealed a 170-kb length sequence. In total, 286 ORFs and 3 tRNA genes were identified, and 259 ORFs showed similarity (BLASTP e-value<0.001) to genes of other bacteriophages. The results from comparative genomic analysis indicated that the enterophage JS98, isolated from human stool, is the closest relative of SP18. Based on phylogenetic analysis of gp23 protein-coding sequences, dot plot comparison and BLASTP analysis of genomes, SP18 and JS98 appear to be closely related to T4-even phages. However, several insertions, deletions, and duplications indicate differences between SP18 and JS98. Comparison of duplicated gp24 genes and the soc gene showed that duplication events are responsible for the differentiation and evolution of T4-like bacteriophages.

Protection Against Helicobacter pylori Infection by a Trivalent Fusion Vaccine Based on a Fragment of Urease B-UreB414: Li Wang Wang , Xiao-Fei Liu , Shi Yun , Xiao-Peng Yuan , Xu-Hu Mao , Chao Wu , Wei-Jun Zhang , Kai-Yun Liu , Gang Guo , Dong-Shui Lu , Wen-De Tong , Ai-Dong Wen , Quan-Ming Zou; J. Microbiol. 2010;48(2):223-228. Published online May 1, 2010; DOI: https://doi.org/10.1007/s12275-009-0233-4

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14 Citations

Abstract: A multivalent fusion vaccine is a promising option for protection against Helicobacter pylori infection. In this study, UreB414 was identified as an antigenic fragment of urease B subunit (UreB) and it induced an antibody inhibiting urease activity. Immunization with UreB414 partially protected mice from H. pylori infection. Furthermore, a trivalent fusion vaccine was constructed by genetically linking heat shock protein A (HspA), H. pylori adhesin A (HpaA), and UreB414, resulting in recombinant HspA-HpaA-UreB414 (rHHU). Its protective effect against H. pylori infection was tested in BALB/c mice. Oral administration of rHHU significantly protected mice from H. pylori infection, which was associated with H. pylori-specific antibody production and Th1/Th2-type immune responses. The results show that a trivalent fusion vaccine efficiently combats H. pylori infection, and that an antigenic fragment of the protein can be used instead of the whole protein to construct a multivalent vaccine.

Virulence Attenuation of Streptococcus pneumoniae clpP Mutant by Sensitivity to Oxidative Stress in Macrophages via an NO-Mediated Pathway: Chul-Yong Park , Eun-Hye Kim , Sang-Yoon Choi , Thao Dang-Hien Tran , In-Hye Kim , Su-Nam Kim , Suhkneung Pyo , Dong-Kwon Rhee; J. Microbiol. 2010;48(2):229-235. Published online May 1, 2010; DOI: https://doi.org/10.1007/s12275-010-9300-0

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23 Citations

Abstract: ClpP protease is essential for virulence and survival under stress conditions in several pathogenic bacteria. The clpP mutation in a murine infection model has demonstrated both attenuation of virulence and a sensitivity to hydrogen peroxide. However, the underlying mechanisms for these changes have not been resolved. Because macrophages play a major role in immune response and activated macrophages can kill microbes via oxygen-dependant mechanisms, we investigated the effect of the clpP mutation on its sensitivity to macrophage-mediated oxygen-dependant mechanisms. The clpP mutant derived from D39 (serotype 2) exhibited a higher sensitivity to oxidative stresses such as reactive oxygen intermediates, reactive nitrogen intermediates, and H2O2, but no sensitivity to osmotic stress (NaCl) and pH. Moreover, viability of the clpP mutant was significantly increased in murine macrophage cells by treatment with S-methylisothiourea sulfate, which inhibits inducible nitric oxide synthase (iNOS) activity and subsequently elicits lower level secretions of nitric oxide (NO). However, viability of wild type was unchanged. Taken together, these results indicate that ClpP is involved in the resistance to oxidative stresses after entrapment by macrophages and subsequently contributes to virulence via NO mediated pathway.

Journal Article

Antimicrobial Resistance Patterns and Characterization of Integrons of Shigella sonnei Isolates in Seoul, 1999-2008: Young-hee Jin , Young-hee Oh , Ji-hun Jung , Soo-jin Kim , Jin-ah Kim , Ki-young Han , Min-young Kim , Seog-gee Park , Young-ki Lee; J. Microbiol. 2010;48(2):236-242. Published online May 1, 2010; DOI: https://doi.org/10.1007/s12275-010-9220-z

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15 Citations

Abstract: A total of 66 Shigella sonnei isolates from 1999 to 2008 in Seoul was analyzed for their antimicrobial resistance, carriage of integron, and the patterns of Pulsed-field gel electrophoresis (PFGE). A high level of antimicrobial resistance to streptomycin (100%), trimethoprim/sulfamethoxazole (95%), tetracycline (94%), nalidixic acid (65%), and ampicillin (41%) was observed among S. sonnei isolates. Fourteen profiles of antimicrobial resistance were identified with the most common resistance profile being nalidixic acid, streptomycin, tetracycline, and trimethoprim/sulfamethoxazole (35%). PCR and DNA sequencing analysis revealed the presence of class 2 integron in all isolates, and class 1 and 2 integrons in 7 isolates. The class 2 integron carried two types of gene cassettes. One cassette array was dfrI, sat2, and aadA1 (91%), and the other was dfr1 and sat1 (8%). dfrA12 and aadA2 gene cassette was found in one isolate containing class 1 integron. PFGE was carried out to examine the genetic relatedness among isolates. All isolates except for one showed similar PFGE patterns (similarity of 80.1%). These results suggest that the S. sonnei isolated during 1999-2008 in Seoul have similar lineages that have not undergone evolutionary changes with time.

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