bluuebunny/crossref_metadata_2025 huggingface dataset:
Role of the outer membrane in the accumulation of quinolones bySerratia marcescens
Authors: Berlanga Mercedes , Ruiz Neus , Hernandez-Borrell Jordi , Montero Teresa , Viñas Miguel
Published: April 2011

Accumulation of four quinolones by Serratia marcescens was measured fluorometrically. The passage of quinolones through the outer membrane was studied in both lipopolysaccharide-deficient and porin-deficient mutants. The lipopolysaccharide (LPS) layer formed a partially effective barrier for highly hydrophobic quinolones such as nalidixic acid. Quinolones with a low relative hydrophobicity coefficient seemed to pass preferentially through the water-filled Omp3 porin channels. Results were confirmed when Omp3 was cloned in a porin-defective Escherichia coli.
Localization of fungal fimbriae by immunocytochemistry in pathogenic and nonpathogenic isolates of Venturia inaequalis
Authors: Svircev Antonet M , Smith Ronald J , Zhou Ting , Day Alan W
Published: April 2011

Pathogenic and nonpathogenic isolates of Venturia inaequalis were grown in liquid culture. Hyphae were treated with two types of fimbrial antiserum (AU- and AV-1) and examined by immunofluorescent microscopy, in order to establish the distribution of fimbrial epitopes in whole cell mounts. The AV-1 antiserum was specific for the glycoprotein subunits while the AU- antiserum was specific for the protein moieties present on the fimbriae of Mycobotryum violaceum. The use of fimbrial antiserum with immunocytochemistry and transmission electron microscopy demonstrated a clear distinction between pathogenic and nonpathogenic isolates of V. inaequalis, based on the appearance of the fungal cell wall and the distribution of fimbrial epitopes labeled with AV-1 antiserum and immunogold complex. In actively growing hyphae of the pathogenic isolate, characterized by distinct cellular organelles, small vacuoles, and lipid bodies, fimbrial epitopes were concentrated in the fungal cell wall and were present minimally on the outer surface. In contrast, actively growing hyphae of the nonpathogenic isolate of V. inaequalis had extensive fine hair-like protrusions in the fungal cell wall which labeled with the AV-1 antiserum and immunogold. The distribution of fimbrial epitopes in V. inaequalis was highly dependent on the developmental growth stage of the fungal mycelium. Aging mycelia in both the pathogenic and nonpathogenic isolates of V. inaequalis were characterized by a large central vacuole and no label. In the pathogenic and nonpathogenic isolates of V. inaequalis grown in vitro, the distribution of fimbrial glycoprotein epitopes provided a more complex profile than that seen in M. violaceum.Key words: fimbriae, immunocytochemistry, Venturia inaequalis, Mycobotryum violaceum.
Cloning of the histidine biosynthetic genes fromCorynebacterium glutamicum: Organization and analysis of thehisGandhisEgenes
Authors: Kwon Joon-Hye , Chun Jae-Yeon , Lee Heung-Shick , Cheon Choong-Ill , Song Eun-Sook , Min Kyung-Hee , Lee Myeong-Sok
Published: April 2011

The physically linked hisG and hisE genes, encoding for ATP-phosphoribosyltransferase and phosphoribosyl-ATP-pyrophosphohydrolase were isolated from the Corynebacterium glutamicum gene library by complementation of Escherichia coli histidine auxotrophs. They are two of the nine genes that participate in the histidine biosynthetic pathway. Molecular genetics and sequencing analysis of the cloned 9-kb insert DNA showed that it carries the hisG and hisE genes. In combining this result with our previous report, we propose that all histidine biosynthetic genes are separated on the genome by three unlinked loci. The coding regions of the hisG and hisE genes are 279 and 87 amino acids in length with a predicted size of about 30 and 10 kDa, respectively. Computer analysis revealed that the amino acid sequences of the hisG and hisE gene products were similar to those of other bacteria.
Purification and characterization of a 4-hydroxybenzoate decarboxylase from an anaerobic coculture
Authors: Li Tong , Juteau Pierre , Beaudet Réjean , Lépine François , Villemur Richard , Bisaillon Jean-Guy
Published: April 2011

The oxygen-sensitive 4-hydroxybenzoate decarboxylase (4OHB-DC) activity from a phenol-carboxylating coculture, consisting of Clostridium-like strain 6 and an unidentified strain 7, was studied. Assays done with cell extracts showed that the optimal pH was 5.0-6.5 and the Kmwas 5.4 mM. The activity decreased by 50% in the presence of 5 mM EDTA, and it was restored and even enhanced by the addition of Mg++, Mn++, Zn++, or Ca++. After purification, the molecular mass of the enzyme was estimated as 420 kDa by gel chromatography, and as 119 kDa by SDS-PAGE, suggesting a homotetrameric structure. Its pI was 5.6. The N-terminal amino acid sequence showed 95% and 76% homology with the pyruvate-flavodoxin oxidoreductase (nifJ gene product) from Enterobacter agglomerans and Klebsiella pneumoniae, respectively. The purified enzyme also slowly catalyzed the reverse reaction, that is the phenol carboxylation. These characteristics suggest that this enzyme is different from other known decarboxylases. This includes the 4OHB-DC from Clostridium hydroxybenzoicum, which is the only one that had been purified before.Key words: purification, 4-hydroxybenzoate decarboxylase, coculture, phenol carboxylation, anaerobic conditions.
Differential expression of IL-1 and TNF receptors in murine macrophages infected with virulent vs. avirulentLegionella pneumophila
Authors: McHugh Shannon , Yamamoto Yoshimasa , Klein Thomas W , Friedman Herman
Published: April 2011

Infection of macrophages from genetically susceptible A/J mice with Legionella pneumophila induces high levels of various cytokines in serum as well as in cultures of spleen or peritoneal cells from the mice. However, modulation of receptor expression for these cytokines during infection has not been studied in detail, even though these receptors on macrophages have a critical role in inflammatory responses during the infection. In the present study, the differential expression of mRNA for TNF and IL-1 receptors as well as receptor antigens during infection of macrophages with virulent vs. avirulent L. pneumophila was investigated. Mouse thioglycollate-elicited peritoneal macrophages showed by RT-PCR constitutive steady-state levels of mRNA for TNF-type I and -type II receptors as well as IL-1 type I receptor. However, IL-1 type II receptor mRNA was not expressed in thioglycollate-elicited macrophages. Infection of macrophages with virulent bacteria caused an upregulation of IL-1 type I and TNF type I receptor mRNA, but had no effect on TNF type II receptor message. Avirulent L. pneumophila infection caused much less induction of these receptor mRNAs. The amount of receptor antigen of IL-1 type I on the surface of macrophages was also increased by infection with virulent L. pneumophila determined by flow cytometric analysis. These results indicate that L. pneumophila infection not only causes induction of various cytokines, but also modulation of certain cytokine receptors, which may regulate the susceptibility to infection.Key words: Legionella pneumophila, cytokine receptors, macrophages.
Isolation of a chitinase overproducing mutant of Streptomyces peucetius defective in daunorubicin biosynthesis
Authors: Vetrivel Kuzhandhaivel S , Dharmalingam Kuppamuthu
Published: April 2011

Streptomyces peucetius, producer of the antitumor anthracycline antibiotic daunorubicin, was mutagenized, and mutants defective in daunorubicin biosynthesis were screened. One mutant (SPVI), which failed to produce daunorubicin, was found to overproduce an extracellular chitinase. Time course analyses of chitinase production and of the extracellular protein profile showed that the increase in activity is due to increased synthesis of the enzyme protein. The production of chitinase in SPVI was repressed by glucose as in the case of wild-type S. peucetius. PFGE analysis of VspI restriction fragments of S. peucetius and SPVI showed that there was no major alteration in the mutant genome. The hybridization pattern of S. peucetius and SPVI genomic DNA digested with various restriction enzymes was identical when probed with dnrUVJI genes of the S. peucetius daunorubicin cluster and chiA of Streptomyces lividans 66. The possible step affected in the daunorubicin biosynthetic pathway could be a polyketide synthase, since aklanonic acid, the earliest detectable intermediate in the daunorubicin pathway, was not synthesized in SPVI.Key words: Streptomyces peucetius, chitinase, daunorubicin, NTG mutagenesis.
Methanogens and sulfate-reducing bacteria in oil sands fine tailings waste
Authors: Holowenko Fervone M , MacKinnon Michael D , Fedorak Phillip M
Published: April 2011

In the past decade, the large tailings pond (Mildred Lake Settling Basin) on the Syncrude Canada Ltd. lease near Fort McMurray, Alta., has gone methanogenic. Currently, about 60%-80% of the flux of gas across the surface of the tailings pond is methane. As well as adding to greenhouse gas emissions, the production of methane in the fine tailings zone of this and other settling basins may affect the performance of these settling basins and impact reclamation options. Enumeration studies found methanogens (105-106MPN/g) within the fine tailings zone of various oil sands waste settling basins. SRB were also present (104-105MPN/g) with elevated numbers when sulfate was available. The methanogenic population was robust, and sample storage up to 9 months at 4°C did not cause the MPN values to change. Nor was the ability of the consortium to produce methane delayed or less efficient after storage. Under laboratory conditions, fine tailings samples released 0.10-0.25 mL CH4(at STP)/mL fine tailings. The addition of sulfate inhibited methanogenesis by stimulating bacterial competition.Key words: fine tailings, methanogens, sulfate-reducing bacteria, methane, oil sands.
Glucosidase inhibitor from Umbilicaria esculenta
Authors: Lee Kyung-Ae , Kim Moo-Sung
Published: April 2011

Inhibitory activity toward β-glucosidase was detected in extracts of the lichen, Umbilicaria esculenta. The extract showed strong inhibition of disaccharide hydrolytic enzymes of mold and mammalian origin, but weak or no inhibition of polysaccharide hydrolytic enzymes except glucoamylase and laminarinase. The inhibitor in the extract was very stable, retaining more than 95% of its activity when treated with heat, acid, alkali, and some hydrolytic enzymes. Purified inhibitor was identified as 1-deoxynojirimycin (1,5-dideoxy-1,5-immino-D-glucitol) by NMR spectrometry, which was known to be produced by Streptomyces sp. and the plant Morus sp. Extracts from Parmelia austrosinensis, Parmelia praesorediosa, and an unidentified lichen species, showed glucosidase inhibitory activities similar to U. esculenta.Key words: β-glucosidase inhibitor, 1-deoxynojirimycin, Umbilicaria esculenta, lichen species.
Heat shock protein 80 ofNeurospora crassa: Sequence analysis of the gene and expression during the asexual phase
Authors: Girvitz T L , Ouimet P M , Kapoor M
Published: April 2011

Heat shock protein 80 (Hsp80) of Neurospora crassa, a member of the stress-90 protein family, is a cytosolic molecular chaperone that interacts directly with Hsp70 to form a hetero-oligomeric complex. The complete nucleotide sequence of the gene encoding this protein, along with the 5'- and 3'-flanking DNA, is reported. The coding sequence is interrupted by two introns, 61 and 30 nucleotides, respectively, in length. The deduced amino acid sequence corresponds to a 695-residue polypeptide with a calculated molecular mass of 78 894 Da and an average pI of 4.94. Primer extension experiments demonstrated two transcription start sites, a major and a minor one. No sequence motifs resembling the standard eukaryotic heat shock elements were evident in the putative promoter region. Immunoblot analysis showed Hsp80 protein to be present in the mature, dormant conidia, while the hsp80 transcripts were not detected. Both the transcripts and the protein were present in the germinating conidia in the absence of externally applied stress.Key words: Hsp90, filamentous fungi, sequence, conidia, germination.
Successful strategy for the selection of new strawberry-associated rhizobacteria antagonistic to Verticillium wilt
Authors: Berg Gabriele , Kurze Stefan , Buchner Arno , Wellington Elizabeth M , Smalla Kornelia
Published: April 2011

In order to isolate and characterize new strawberry-associated bacteria antagonistic to the soil-borne pathogenic fungus Verticillium dahliae Kleb., rhizobacterial populations from two different strawberry species, Greenish Strawberry (Fragaria viridis) and Garden Strawberry (F. × ananassa) obtained after plating onto King's B and glycerol-arginine agar, were screened for in vitro antagonism toward V. dahliae. The proportion of isolates with antifungal activity determined in in vitro assay against V. dahliae was higher for the Garden Strawberry than for the Greenish Strawberry. From 300 isolates, 20 isolates with strong antifungal activity were selected characterized by physiological profiling and molecular fingerprinting methods. Diversity among the isolates was characterized with molecular fingerprints using amplified ribosomal DNA restriction analysis (ARDRA) and the more discriminating BOX-PCR fingerprint method. The physiological profiles were well correlated with molecular fingerprinting pattern analysis. Significant reduction of Verticillium wilt by bacterial dipping bath treatment was shown in the greenhouse and in fields naturally infested by V. dahliae. The relative increase of yield ranged from 117% (Streptomyces albidoflavus S1) to 344% (Pseudomonas fluorescens P10) in greenhouse trials, and 113% (Streptomyces albidoflavus S1) to 247% (Pseudomonas fluorescens P6) in field trials. Evaluation resulted in the selection of three effective biocontrol agents (Pseudomonas fluorescens P6, P10, and Streptomyces diastatochromogenes S9) antagonistic to the Verticillium wilt pathogen.Key words: biocontrol, molecular fingerprint, antifungal properties, Pseudomonas, Streptomyces.
Carbon source-dependent transcriptional regulation of the mitochondrial glycerol-3-phosphate dehydrogenase gene,GUT2, fromSaccharomyces cerevisiae
Authors: Grauslund Morten , Rønnow Birgitte
Published: April 2011

Cytosolic glycerol kinase (Gut1p) and mitochondrial glycerol-3-phosphate dehydrogenase (Gut2p) constitute the glycerol utilization pathway in Saccharomyces cerevisiae. Transcriptional analysis of the GUT2 gene showed that it was repressed by glucose and derepressed on the non-fermentable carbon sources, glycerol, lactate and ethanol. Derepression of GUT2 requires the protein kinase Snf1p as well as the heteromeric protein complex, Hap2/3/4/5, and its putative DNA-binding site (UASHAP) located in the promoter region. Furthermore, glucose repression of GUT2 requires the negative regulator, Opi1p.Key words: GUT2, mitochondrial glycerol-3-phosphate dehydrogenase, transcriptional regulation, Saccharomyces cerevisiae.
On the biogeography of yeasts in the Wickerhamiella clade and description of Wickerhamiella lipophila sp. nov., the teleomorph of Candida lipophila
Authors: Lachance Marc-André , Bowles Jane M , Mueller Carmen , Starmer William T
Published: April 2011

We describe the new yeast species Wickerhamiella lipophila, the teleomorph of Candida lipophila, a haploid heterothallic yeast previously isolated from insects associated with morning glories in Hawaii. Both mating types were recovered in the eastern region of Maui, and a single strain was found in the Waimea region of Kauai. We reexamined the mating compatibility of the several strains of Candida lipophila previously collected on the island of Hawaii and found them to be fertile mating types that had been overlooked because of the unpredictability of mating and ascus formation. The type culture of Candida lipophila [UWO(PS)91-681.3 = CBS 8458, h+] is transferred to the genus Wickerhamiella, and strain UWO(PS)00-340.1 (CBS 8812, h-) is designated as isotype. Also found on Maui and Kauai were strains of Candida drosophilae that produced a strong extracellular protease. An update on the global distribution of members of the Wickerhamiella clade is given.Key words: yeasts, Wickerhamiella, Candida, flowers, insects, biogeography.
Phase variation of F1651(Prs-like) fimbriae fromEscherichia colicausing septicaemia in animals
Authors: Harel Josée , Daigle France , Forget Céline , Tessier Marie-Catherine , Crost Cécile , Martin Christine
Published: April 2011

Escherichia coli O115:F165 strains are associated with septicaemia in young pigs and synthesize fimbriae involved in virulence, designated as F1651. F1651fimbriae belong to the P fimbrial family and are encoded by the foo gene cluster. The foo regulatory region of strain 5131 possesses characteristics similar to that of members of the P regulatory family, including papI and papB homologues, and two GATC sites separated by 102 bp, targets of differential Dam methylation. In wild-type strains, the synthesis of F1651is repressed by leucine and the fimbriae undergo phase variation. Immunofluorescence staining showed that phase variation of F1651results in a majority of cells (98%) in the ON phase, in contrast with phase variation of other members of this regulatory family, for which the majority of the cells are in the OFF state. Using a translational fusion in strain 5131 between phoA and fooA, encoding for the major structural subunit of F1651, it was shown that leucine inhibits the OFF to ON switch and modulates the basal transcription of the foo operon.Key words: Escherichia coli, fimbriae, phase variation, regulation, septicaemia.
Identification of DNA sequences that regulate the expression of theEnterobacter cloacaeUW4 1-aminocyclopropane-1-carboxylic acid deaminase gene
Authors: Grichko Varvara P , Glick Bernard R
Published: April 2011

Analysis of the DNA sequence upstream of the previously isolated Enterobacter cloacae UW4 ACC deaminase gene (Shah et al. 1998) suggests that this segment contains several features that are thought to be involved in the transcriptional regulation of this gene. These features include half of a CRP (cAMP receptor protein) binding site, an FNR (fumarate-nitrate reduction) regulatory protein binding site, an LRP (leucine responsive regulatory protein) binding site, and an LRP-like protein coding region. ACC deaminase activity was measured following growth of either various Escherichia coli strains carrying a plasmid that contained the Enterobacter cloacae UW4 ACC deaminase gene, or of Enterobacter cloacae UW4. Variables that were compared include aerobic versus anaerobic conditions, the presence and absence of ACC in the growth medium, addition of leucine to the medium, and bacterial strains that did or did not contain either lrp or fnr genes. The data reported are consistent with the involvement of most, if not all, of the above mentioned potential regulatory regions in the expression of ACC deaminase.Key words: 1-aminocyclopropane-1-carboxylate, ACC, plant growth-promoting rhizobacteria, ACC deaminase, Enterobacter cloacae, leucine responsive regulatory protein.
Evidence for conserved tRNA genes in the 16S-23S rDNA spacer sequence and tworrnoperons ofXylella fastidiosa
Authors: Chen Jianchi , Banks Donna , Jarret Robert L , Jones Jeffrey B
Published: April 2011

The 16S-23S rDNA spacer of the type strain (ATCC 35879) of Xylella fastidiosa was amplified by PCR, cloned, and sequenced. The spacer sequence (455 bp) contains two tRNA (tRNAalaand tRNAile) genes. Identical tRNA genes were also found in the 16S-23S spacer sequences of all the 51 strains of X. fastidiosa retrieved from the GenBank database. At this particular locus, the gene order of tRNAala- tRNAileis conserved among all the studied strains of Xylella and Xanthomonas, and different from those of other bacteria. Sequence analysis showed that Xanthomonas is the most closely related genus. Results from restriction endonuclease analysis suggested the presence of two rrn operons in the genome of a Xylella fastidiosa Pierce's disease strain.Key words: Xylella fastidiosa, rrn operon numbers, 16S-23S rDNA spacer, tRNA.
Isolation and 16S rRNA sequence analysis of the beneficial bacteria from the rhizosphere of rice
Authors: Mehnaz Samina , Mirza M Sajjad , Haurat Jacqueline , Bally Rene , Normand Philippe , Bano Asghari , Malik Kauser A
Published: April 2011

The present study deals with the isolation of plant growth promoting rhizobacteria (PGPR) from rice (variety NIAB IRRI-9) and the beneficial effects of these inoculants on two Basmati rice varieties. Nitrogen-fixing activity (acetylene-reduction activity) was detected in the roots and submerged shoots of field-grown rice variety NIAB IRRI-9. Estimation of the population size of diazotrophic bacteria by ARA-based MPN (acetylene reduction assay-based most probable number) in roots and shoots indicated about 105106counts/g dry weight at panicle initiation and grain filling stages. Four bacterial isolates from rice roots and shoots were obtained in pure culture which produced phytohormone indoleacetic acid (IAA) in the growth medium. Among these, three isolates S1, S4, and R3 reduced acetylene to ethylene in nitrogen-free semi-solid medium. Morphological and physiological characteristics of the isolates indicated that three nitrogen-fixing isolates S1, S4, and R3 belonged to the genus Enterobacter, while the non-fixing isolate R8 belonged to the genus Aeromonas. 16S rRNA sequence of one isolate from root (R8) and one isolate from shoot (S1) was obtained which confirmed identification of the isolates as Aeromonas veronii and Enterobacter cloacae, respectively. The 1517-nucleotide-long sequence of the isolate R8 showed 99% similarity with Aeromonas veronii (accession No. AF099023) while partial 16S rRNA sequence (two stretches of total 1271 nucleotide length) of S1 showed 97% similarity with the sequence of Enterobacter cloacae (accession No. AJ251469). The seedlings of two rice varieties Basmati 385 and Super Basmati were inoculated with the four bacterial isolates from rice and one Azospirillum brasilense strain Wb3, which was isolated from wheat. In the rice variety Basmati 385, maximum increase in root area and plant biomass was obtained in plants inoculated with Enterobacter S1 and Azospirillum Wb3, whereas in the rice variety Super Basmati, inoculation with Enterobacter R3 resulted in maximum increase of root area and plant biomass. Nitrogen fixation was quantified by using15N isotopic dilution method. Maximum fixation was observed in Basmati 385 with the inoculants Azospirillum Wb3 and Enterobacter S1 where nearly 46% and 41% of the nitrogen was derived from atmosphere (%Ndfa), respectively. In general, higher nitrogen fixation was observed in variety Basmati 385 than in Super Basmati, and different bacterial strains were found more effective as inoculants for the rice varieties Basmati 385 and Super Basmati.Key words: phytohormones, nitrogen fixation, Enterobacter, Aeromonas.
Intact soil-core microcosms compared with multi-site field releases for pre-release testing of microbes in diverse soils and climates
Authors: Gagliardi Joel V , Angle J Scott , Germida James J , Wyndham R Campbell , Chanway Christopher P , Watson Robert J , Greer Charles W , McIntyre Terry , Yu Hans H , Levin Morris A , Russek-Cohen Estelle , Rosolen Sarah , Nairn James , Seib Arlette , Martin-Heller Teresa , Wisse Gesine
Published: April 2011

Intact soil-core microcosms were used to compare persistence of Pseudomonas chlororaphis 3732RN-L11 in fallow soil and on wheat roots with field releases at diverse sites. Parallel field and microcosm releases at four sites in 1996 were repeated with addition of one site in 1997. Microcosms were obtained fresh and maintained at 60% soil water holding capacity in a growth chamber at 70% relative humidity, a 12-hour photoperiod, and constant temperature. Persistence of 3732RN-L11 was measured at each site in field plots and microcosms at 721 day intervals, and in duplicate microcosms sampled at an independent laboratory. Linear regression slopes of field plot and microcosm persistence were compared for each site, and between identical microcosms sampled at different sites, using log10transformed plate counts. Microcosm persistence closely matched field plots for wheat roots, but persistence in fallow soil differed significantly in several instances where persistence in field plots was lower than in microcosms. Analysis of weather variations at each site indicated that rainfall events of 3040 mm caused decreased persistence in fallow soil. Cooler temperatures enhanced persistence in field plots at later time points. Inter-laboratory comparison of regression slopes showed good agreement for data generated at different sites, though in two instances, longer sampling periods at one site caused significant differences between the sites. Soil characteristics were compared and it was found that fertility, namely the carbon to nitrogen ratio, and the presence of expanding clays, were related to persistence. These microcosm protocols produced reliable data at low cost, and were useable for pre-release risk analyses for microorganisms.Key words: microcosm, soil, microbiology, risk assessment, 3732RN-L11.
Polyphasic taxonomy of the basidiomycetous yeast genusRhodosporidium:R. azoricumsp. nov.
Authors: Gadanho M , Sampaio J P , Spencer-Martins I
Published: April 2011

This report presents the description of a new heterothallic Rhodosporidium species, R. azoricum sp. nov. The new species is based on two strains previously identified as Rhodotorula glutinis, which were isolated from soil in São Miguel island, Azores, Portugal. Evidence that the two strains were conspecific and distinct from Rhodotorula glutinis was obtained in DNA fingerprinting experiments using the microsatellite-primed PCR approach (MSP-PCR) and the primers M13 and (GTG)5. In order to determine the phylogenetic position of the new species, the nucleotide sequence of the D1/D2 region of the 26S rDNA was analysed and Rhodosporidium azoricum was found to belong to a cluster including R. fluviale, R. lusitaniae, Sporidiobolus microsporus, and S. ruineniae. The life cycle of R. azoricum was investigated and comparisons integrating physiological, morphological, and molecular data were made with related species.Key words: Rhodosporidium azoricum sp. nov., microsatellite-primed PCR, Rhodotorula glutinis, basidiomycetous yeasts.
A common precursor for the three subunits ofL-glutamate oxidase encoded bygoxgene fromStreptomyces platensisNTU3304
Authors: Chen Chien-Yuan , Wu Wen-Tung , Huang Chang-Jen , Lin Mei-Huei , Chang Chen-Kai , Huang Huei-Jen , Liao Jiuan-Miaw , Chen Ling-Yun , Liu Yu-Tien
Published: April 2011

A segment of DNA containing the L-glutamate oxidase (gox) gene from Streptomyces platensis NTU3304 was cloned. The entire nucleotide sequence of the protein-coding portion consisting of 2130 bp (710 codons, including AUG and UGA) of the cloned DNA fragment was determined. The gox gene contained only one open reading frame (ORF) which coded for a 78-kDa polypeptide, the precursor of active extracellular Gox. Mature Gox is composed of three subunits, designated as α, β, and γ, with molecular masses of 39, 19, and 16 kDa, respectively. Analyses of the N-terminal amino acid sequences of the subunits revealed that the order of subunits in the precursor polypeptide encoded by the ORF, from N-terminus to C-terminus, is αγβ. The presence of the flavin adenine dinucleotide (FAD)-binding motif place Gox as a member of the flavoenzyme family. Furthermore, a negative effect of glucose on the biosynthesis of Gox was observed when it was used as carbon source.Key words: L-glutamate oxidase, gox gene, signal peptide, DNA sequence, flavoenzyme, pIJ702 vector.
Pine nuts: the mycobiota and potential mycotoxins
Authors: Weidenbörner Martin
Published: April 2011

The mycobiota of pine nuts was investigated. In total, 1832 fungi belonging to 31 species and 15 genera (Ascomycota, 2; Zygomycota, 3; mitosporic fungi, 10) could be isolated. Cladosporium spp. dominated the mycobiota with 685 isolations followed by Phoma macrostoma with 351 isolations. Overall, 16 potentially mycotoxigenic species were present on pine nuts.Key words: pine nuts, mycobiota, potential mycotoxins.
Presence of double-stranded RNA and virus-like particles in Rhizopus isolates
Authors: Papp Tamás , Nyilasi Ildikó , Fekete Csaba , Ferenczy Lajos , Vágvölgyi Csaba
Published: April 2011

Fungal isolates belonging to four Rhizopus species were screened for the presence of double-stranded RNA (dsRNA) molecules. Five (two R. stolonifer, two R. microsporus, and one R. oryzae) of the 27 isolates examined harboured such genetic elements. Electrophoresis of the nucleic acids revealed five RNA patterns, with 15 discrete dsRNA bands. The molecular sizes corresponding to these bands were 2.214.8 kb. Gel electrophoresis of purified virus-like particles (VLPs) indicated only one capsid of similar size in all virus-harbouring strains; when investigated by electron microscopy, they were found to be polyhedral VLPs 40 nm in diameter. In one of the R. microsporus isolates an uncapsidated large dsRNA segment (14.8 kb) was observed. No phenotypic differences were observed between uninfected and virus-harbouring Rhizopus isolates.Key words: dsRNA, mycovirus, Rhizopus, VLP.
Sulfite oxidation by iron-grown cells ofThiobacillus ferrooxidansat pH 3 possibly involves free radicals, iron, and cytochrome oxidase
Authors: Harahuc Lesia , Suzuki Isamu
Published: April 2011

Thiobacillus ferrooxidans cells grown on ferrous iron oxidized sulfite to sulfate at pH 3, possibly by a free radical mechanism involving iron and cytochrome oxidase. A purely chemical system with low concentrations of Fe3+simulated the T. ferrooxidans system. Metal chelators, ethylenediamine tetraacetic acid (EDTA), 4,5-dihydroxy-1-3-benzene disulfonic acid (Tiron), o-phenanthroline, and 2,2'-dipyridyl, inhibited both sulfite oxidation systems, but the T. ferrooxidans system was inhibited only after the initial brief oxygen consumption. EDTA and Tiron, strong chelators of Fe3+, inhibited the oxidation at lower concentrations than o-phenanthroline and 2,2'-dipyridyl, strong chelators of Fe2+. Inhibition of Fe3+-catalyzed sulfite oxidation by EDTA and Tiron was instant, but the inhibition by o-phenanthroline and dipyridyl was briefly delayed, presumably for the reduction of Fe3+to Fe2+. Mannitol, a free radical scavenger, inhibited both systems to the same extent. Cyanide and azide inhibited only the T. ferrooxidans system, suggesting a role of cytochrome oxidase. It is proposed that sulfite is oxidized by a free radical mechanism initiated by Fe3+on the cell surface of T. ferrooxidans. Cytochrome oxidase is possibly involved in the regeneration of Fe3+from Fe2+by the normal Fe2+-oxidizing system of T. ferrooxidans.Key words: Thiobacillus ferrooxidans, sulfite oxidation, iron, free radical, cytochrome oxidase.
Temporal effects on the composition of a population of Sinorhizobium meliloti associated with Medicago sativa and Melilotus alba
Authors: Bromfield Eden SP , Butler Gail , Barran Leslie R
Published: April 2011

An assessment was made of the impact of temporal separation on the composition of a population of Sinorhizobium meliloti associated with Medicago sativa (alfalfa) and Melilotus alba (sweet clover) grown at a single site that had no known history of alfalfa cultivation. Root nodules were sampled on six occasions over two seasons, and a total of 1620 isolates of S. meliloti were characterized on the basis of phage sensitivity using 16 typing phages. Plant infection tests indicated that symbiotic S. meliloti were deficient in the soil at the time of planting and that these bacteria were present at low density during the first season (<102/g of soil); in the second season numbers increased markedly to about 105/g of soil. Overall, 37 and 51 phage types, respectively, were encountered among the nodule isolates from M. sativa and M. alba. The data indicate significant temporal shifts in the frequency and diversity of types associated with the two legume species. Apparent temporal variation with respect to the frequency of types appeared largely unpredictable and was not attributable to any one sampling time. The results indicate an apparent reduction in phenotypic diversity over the course of the experiment. Differential host plant selection of specific types with respect to nodule occupancy was indicated by significant interactions between legume species and either the frequency or diversity of phage types. Isolates from M. sativa that were resistant to lysis by all typing phages (type 14) were unusual in that they were predominant on this host at all sampling times (between 53% and 82% nodule occupancy) and were relatively homogeneous on the basis of DNA hybridization with 98% of the isolates analysed sharing the same nod EFG hybridization profile. In contrast, those isolates from M. alba comprising type 14 were encountered at low total frequency (2%) and were genetically heterogeneous on the basis of Southern hybridization. The implications of the observed temporal and host plant variation for ecological studies are discussed.Key words: Sinorhizobium meliloti, populations, temporal effects, Medicago sativa, Melilotus alba.
Identification of a system that allows a Rhizobium tropici dctA mutant to grow on succinate, but not on other C4-dicarboxylates
Authors: Batista Silvia , Catalán Ana I , Hernández-Lucas Ismael , Martínez-Romero Esperanza , Aguilar O Mario , Martínez-Drets Gloria
Published: April 2011

A defined insertion mutant of a gene encoding a homolog of the rhizobial C4-dicarboxylate permease (dctA) was constructed in Rhizobium tropici strain CIAT899. This mutant (GA1) was unable to grow on fumarate or malate; however, in contrast with other rhizobial dctA mutants, it retained a limited ability to grow on succinate with ammonia as a nitrogen source. Our results suggest the presence of a novel succinate-specific transport system in R. tropici. Biochemical characterization indicated that this alternative transport system in GA1 is active and dependent on an energized membrane. It was also induced by succinate and aspartate, and was repressed by glucose and glycerol. Bean plants inoculated with GA1 showed a reduced nitrogen-fixing ability, achieving only 29% of the acetylene reduction activity determined in CIAT899 strain nodules, 33 days after inoculation. Also, bean plants inoculated with GA1 had reduced shoot dry weight compared with plants inoculated with the wild-type strain.Key words: succinate transport, Rhizobium tropici, C4-dicarboxylate uptake.
Analysis of the genetic region encoding a novel rhizobiocin fromRhizobium leguminosarumbv.viciaestrain 306
Authors: Venter Alexandra P , Twelker Sunny , Oresnik Ivan J , Hynes Michael F
Published: April 2011

Cross-testing of a number of strains of Rhizobium leguminosarum for bacteriocin production revealed that strain 306 produced at least two distinct bacteriocins. Further analysis involving plasmid transfer to Agrobacterium and other hosts demonstrated that there were bacteriocin determinants on plasmids pRle306b and pRle306c, as well as a third bacteriocin. The bacteriocin encoded by pRle306b was indistinguishable from the bacteriocin encoded by strain 248, whereas the bacteriocin encoded by plasmid pRle306c had a distinctive spectrum of activity against susceptible strains, as well as different physical properties from other bacteriocins that we have studied in our lab. Two mutants altered in production of the pRle306c bacteriocin were generated by transposon Tn5 mutagenesis, and the DNA flanking the transposon inserts in these mutants was cloned and characterized. DNA sequence analysis suggested that the pRle306c bacteriocin was a large protein belonging to the RTX family, and that a type I secretion system involving an ABC type transporter was required for export of the bacteriocin. A mutant unable to produce this bacteriocin was unaltered in its competitive properties, both in broth and in nodulation assays, suggesting that the bacteriocin may not play a major role in determining the ecological success of this strain.Key words: Rhizobium, bacteriocins, RTX proteins, plasmids.
Comparison of the efficacy of different techniques, culture media, and sources of blood in determining the hemolytic activity ofListeriaspp.
Authors: Capita Rosa , Alonso-Calleja Carlos , García-Fernández María Camino , Moreno Benito
Published: April 2011

Hemolytic activity is a fundamental criterion for the differentiation of Listeria species; therefore, a simple and inexpensive procedure to clearly distinguish hemolytic strains from each other and from nonhemolytic strains would be of great aid. We compared the efficacy of several techniques, culture media, and types of blood in demonstrating the hemolysis of Listeria spp. The hemolytic activities of Listeria monocytogenes and Listeria seeligeri were more easily detected with a red blood cell top-layer (RBCTL) technique and with a microplate technique than when the strains were streaked on blood agar (BA). Listeria ivanovii produced a marked hemolysis regardless of the technique employed. In general, the hemolytic activity of these three species was stronger on media containing brain heart infusion (BHI) agar and (or) potassium tellurite (PT). However, Listeria innocua produced questionable hemolytic reactions when nonselective culture media with BHI and PT were utilized, limiting the advantages gained by employing the two compounds. The RBCTL and the BA techniques disclosed greater hemolytic activity for L. monocytogenes, L. seeligeri, and L. ivanovii with sheep and guinea pig blood than with horse and human blood. When the microplate technique was used, all four kinds of blood were equally effective.Key words: Listeria spp., hemolysis, test comparison.
Chitin-supplemented formulations improve biocontrol and plant growth promoting efficiency ofBacillus subtilisAF 1
Authors: Manjula K , Podile A R
Published: April 2011

Formulations of a chitinolytic biocontrol and a plant growth promoting Bacillus subtilis AF 1 were prepared in peat, in peat supplemented with either 0.5% chitin or Aspergillus niger mycelium, or in spent compost obtained from Agaricus bisporus cultivation and were evaluated for biocontrol of two fungal pathogens and plant growth promoting activities on pigeon pea and groundnut. A steady increase in cell numbers of introduced B. subtilis AF 1 was observed in all the formulations at 30°C. The increase in cell numbers was about 5.0 log units. Peat or spent compost inoculated with physiologically active and dormant states of B. subtilis AF 1 showed different time period requirements to attain maximum cell numbers. The presence of chitin or A. niger (in peat) or A. bisporus (in spent compost) as supplement in the carrier material improved the multiplication of B. subtilis AF 1. When used as seed treatments, formulations of AF 1 in peat supplemented with chitin or chitin-containing materials showed better control of A. niger (causing crown rot of groundnut) and Fusarium udum (causing wilt of pigeon pea) than AF 1 culture alone, in both groundnut and pigeon pea. Bacillus subtilis AF 1 formulations promoted seed germination and biomass of both groundnut and pigeon pea even under pathogen pressure. Survival of AF 1 on fresh culture-treated and formulation product-treated plants was similar in pathogen-infested soil.Key words: microbial formulations, Bacillus subtilis AF 1, chitin.
Influence of the zinc hyperaccumulatorThlaspi caerulescensJ. & C. Presl. and the nonmetal accumulatorTrifolium pratenseL. on soil microbial populations
Authors: Delorme T A , Gagliardi J V , Angle J S , Chaney R L
Published: April 2011

Metal hyperaccumulator plants like Thlaspi caerulescens J. & C. Presl. are used for phytoremediation of contaminated soils. Since little is known about the rhizosphere of hyperaccumulators, the influence of T. caerulescens was compared with the effects of Trifolium pratense L. on soil microbes. High- and low-metal soils were collected near a zinc smelter in Palmerton, Penn. Soil pH was adjusted to 5.8 and 6.8 by the addition of Ca(OH)2. Liming increased bacterial populations and decreased metal toxicity to levels allowing growth of both plants. The effects of the plants on total (culturable) bacteria, total fungi, as well as cadmium- and zinc-resistant populations were assessed in nonrhizosphere and rhizosphere soil. Both plants increased microbial populations in rhizosphere soil compared with nonrhizosphere soil. Microbial populations were higher in soils planted with T. pratense, but higher ratios of metal-resistant bacteria were found in the presence of T. caerulescens. We hypothesize that T. caerulescens acidifies its rhizosphere. Soil acidification in the rhizosphere of T. caerulescens would affect metal uptake by increasing available metals around the roots and consequently, increase the selection for metal-resistant bacteria. Soil acidification may be part of the hyperaccumulation process enhancing metal uptake from soil.Key words: phytoremediation, Thlaspi caerulescens J. & C. Presl., Trifolium pratense L., rhizosphere, soil microbial populations.
Involvement ofgacSandrpoSin enhancement of the plant growth-promoting capabilities ofEnterobacter cloacaeCAL2 and UW4
Authors: Saleh Saleema S , Glick Bernard R
Published: April 2011

The plant growth-promoting bacteria Enterobacter cloacae CAL2 and UW4 were genetically transformed with a multicopy plasmid containing an rpoS or gacS gene from Pseudomonas fluorescens. The transformed strains were compared with the nontransformed strains for growth, indoleacetic acid (IAA) production, antibiotic production, 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase activity, siderophore production, cell morphology, and the ability to promote canola root elongation. All transformed strains had a longer lag phase, were slower in reaching stationary phase, and attained a higher cell density than the nontransformed strains. Transformation resulted in cells that were significantly shorter than the nontransformed cells. The transformed strains also produced significantly more IAA than the nontransformed strains. Introduction of rpoS or gacS from Pseudomonas fluorescens was associated with a reduction in the production of both antibiotics, 2,4-diacetylphloroglucinol and mono-acetylphloroglucinol, produced by Enterobacter cloacae CAL2. With Enterobacter cloacae CAL2, plasmid-borne rpoS, but not gacS, increased the level of ACC deaminase activity, while introduction of rpoS in Enterobacter cloacae UW4 caused a decrease in ACC deaminase activity. Neither gacS nor rpoS significantly affected the level of siderophores synthesized by either bacterial strain. Overproduction of either GacA or RpoS in Enterobacter cloacae CAL2 resulted in a significant increase in the root lengths of canola seedlings when seeds were treated with the bacteria, and overproduction of RpoS caused an increase in canola shoot as well as root lengths.Key words: plant growth-promoting bacteria, canola, ethylene, ACC deaminase, GacS, RpoS, indoleacetic acid, siderophores, antibiotics.
A procedure for quantifying adhesion of conidia ofBotrytis cinereato the skin of apple fruit
Authors: Filonow Alexander B
Published: April 2011

Ultrasonication was evaluated as a nonchemical means to quantitatively remove conidia of Botrytis cinerea from the skin of Golden Delicious apple (Malus domestica Borkh.) fruit. A probe immersed in a suspension of conidia and generating 20 kHz at 150 W for 30- or 60-s pulses destroyed 13.3% or 29% of conidia, respectively. Destruction at 150 W for 10 s or at 30120 W for up to 60 s was <2%. The procedure for quantifying adhesion of conidia to the skin of fruit consisted of pipetting a 50-µL water droplet containing 5 × 104conidia onto the skinside of a slice of fruit, incubating the slices inside sealed 500 cm3glass jars, excising a 1 cm diameter piece of skin bearing the droplet, and sonicating the skin in 8 mL of ice-cold water at 150 W for 10 s. The skin was removed, the suspension was centrifuged at 1250 × g for 15 min, and the supernatant was reduced to 1 mL by vacuum suction using a pipet. Conidia were stained with crystal violet and counted in a hemacytometer. Adhesion of conidia to skin was 3.0%, 14.6%, 20.8%, 39.4%, 57.6%, and 73.1% after 0, 2, 4, 8, 12, and 24 h incubation, respectively. Sonication was more effective than two other procedures for recovery of conidia. Conidia on the skin of fruit exposed to 4 µL of butyl acetate in the headspace of glass jars for 4 h at 23°C increased the adhesion of conidia 107% above that for unexposed conidia. Sonication with a programmable power- and time-controlled probe was a simple, rapid, safe, and effective method for quantifying adhesion of B. cinerea conidia to the skin of apple fruit.Key words: ultrasonication, Botrytis cinerea, spore adhesion, butyl acetate, fungal ecology, phyllosphere.
The effects of sequential inoculation of mixed rumen protozoa on the degradation of orchard grass cell walls by anaerobic fungus Anaeromyces mucronatus 543
Authors: Lee Sung S , Ha Jong K , Cheng K -J
Published: April 2011

The effects of protozoa on the degradation of plant cell walls (CW) during different growth stages of the fungus Anaeromyces mucronatus have been investigated. Since fungi show a marked lag in their in vitro cultures and many protozoa rapidly die during a prolonged incubation time, the effects of protozoa may vary according to the growth phase of the fungi. Therefore, the approach adopted was (i) to inoculate CW with fungus monoculture, (ii) to inoculate CW with fungus-protozoa coculture, or (iii) to sequentially inoculate fungal cultures that had been grown in CW for 24 (initial stage of growth), 48, and 72 h (late stage of growth) with mixed protozoa. When a fungus was associated with protozoa, a growth phase dependent effect was observed. Ruminal protozoa adversely affected the growth and activity when introduced in the initial growth stage of A. mucronatus, but a synergetic interaction was detected when added to late growth stage cultures. Although there is no immediate explanation for these results, the data suggested that protozoa can engulf the fungal zoospores, which are in ruminal fluids and (or) attached to small feed particles, but cannot engulf the fungal thallus that is tightly attached to feed particles by a rhizoidal system. Our data indicated that the protozoa did not influence cellulolysis by the fungi in exponential and (or) stationary phase, but they had a marked inhibitory effect on fungi that were in lag phase. Inhibition during lag phase could result from the protozoal predation of fungal zoospores that had failed to attach to substrates.Key words: rumen fungi, rumen protozoa, cellulose digestion, cellulase activity, interactions.