bluuebunny/crossref_metadata_2025 huggingface dataset:
Characterization of major secondary metabolites produced in soilless mix by a formulated strain of the biocontrol fungus Gliocladium virens
Authors: Lumsden R. D. , Ridout C. J. , Vendemia M. E. , Harrison D. J. , Waters R. M. , Walter J. F.
Published: May 2010

A strain of Gliocladium virens (GL-21 = G-20) formulated in alginate prill was grown for 3 days in a commercial soilless potting medium. Chloroform extractions were separated by thin-layer chromatography and individual bands from G-20 cultures in soilless medium were identified by standards, mass spectrometry, and nuclear magnetic resonance as (i) a mixture of fatty acids, (ii) viridin, (iii) gliotoxin, (iv) dimethylgliotoxin, (v) viridiol, and (vi) a mixture of phenolics including ferulic acid. None of these compounds was detected in the medium not amended with prills of isolate G-20 of G. virens. Other isolates of G. virens, including G-3 and G-9, produced gliovirin and did not produce gliotoxin. Of all the metabolites produced by G-20 in soilless medium, only gliotoxin strongly inhibited germination of sporangia and growth of Pythium ultimum (growth prevention at 1.0 μg/mL), growth of Rhizoctonia solani mycelium (0.5 μg/mL), or germination of sclerotia and growth of Sclerotium rolfsii (50 μg/mL). Viridin was more inhibitory to growth of R. solani (1.0 μg/mL, minimum inhibitory concentration) than to P. ultimum (25 μg/mL) or S. rolfsii (50 μg/mL). Growth of G. virens isolate G-20 was inhibited by 25 μg/mL of viridin but not by 100 μg/mL gliotoxin. In contrast, growth of isolate G-3 was inhibited by 50 μg/mL gliotoxin but not by 100 μg/mL of viridin. Dimethylgliotoxin, fatty acids, and phenolics were not inhibitory to any of the fungi tested. These results suggest that gliotoxin is the major antibiotic metabolite inhibitory to Pythium and Rhizoctonia of this formulated strain of G. virens. This information will be useful for monitoring metabolite production by G. virens, determining optimum gliotoxin production in situ, and improving strain performance for biocontrol of plant pathogens. Key words: antibiotic, biological control, antagonist.
Cyanobiont diversity within and among cycads of one field site
Authors: Zimmerman William J. , Rosen Barry H.
Published: May 2010

Limited diversity was found among cyanobionts from a cultivated population of cycads at a field site in Florida. All isolates were classified as Nostoc but were different from the one Nostoc species found in the soil. These cyanobacteria were root endophytes of several plants of Zamia integrifolia and one of Dioon. The isolates were similar morphologically and in their reactions to four fluorescein isothiocyanate conjugated lectins. Electrophoretic protein profiles and zymograms distinguished one cyanobiont and the soil Nostoc. A tenacious Anabaena epiphyte was also discovered inhabiting the surfaces of root nodules. Key words: cyanobacteria, cycad, Nostoc, symbiosis.
Reduction of selenate and selenite to elemental selenium by Wolinella succinogenes
Authors: Tomei Francisco A. , Barton Larry L. , Lemanski Cheryl L. , Zocco Thomas G.
Published: May 2010

Cultures of Wolinella succinogenes were adapted to grow in the presence of 1 mM [Formula: see text] or 10 mM [Formula: see text]. Both selenium salts were reduced to red, amorphous, elemental selenium but only after the culture reached the stationary growth phase. Bacterial cells taken from a culture actively reducing selenium were examined by transmission electron microscopy and were found to have large, electron-dense granules in the cytoplasm. These granules were verified by energy-dispersive X-ray spectroscopy to consist of selenium. Wolinella succinogenes was unable to grow with [Formula: see text] or [Formula: see text] as the final electron acceptor. Key words: Wolinella, selenium, cytology, selenate.
Purification and properties of chlorocatechol 1,2-dioxygenase from Alcaligenes denitrificans BRI 6011
Authors: Miguez Carlos B. , Greer Charles W. , Ingram Jordan M.
Published: May 2010

The specific activity of chlorocatechol 1,2-dioxygenase from Alcaligenes denitrificans BRI 6011 was found to be maximal in the early logarithmic growth phase. The enzyme was purified from cultures at mid-log phase of growth using ammonium sulfate fractionation, and phenyl-Sepharose and DEAE-Sepharose chromatography. The protein gave a single band by SDS polyacrylamide gel electrophoresis with an apparent molecular weight of 33 000, and the temperature and pH optima were 30 °C and 7.5, respectively. Catechol, 3-chlorocatechol (3-CC), 4-CC, 3,4-dichlorocatechol (3,4-DCC), 3,5-DCC, 3,6-DCC, 3-methylcatechol (3-MC), and 4-MC served as substrates for the enzyme. The Vmax values for the dichlorocatechols were similar, while those for the monochlorinated and methylated catechols were higher. The Km values for all the chlorinated catechols were typically below 1 μM, while those for catechol and the methylated catechols were above 10 μM.Key words: chlorocatechol 1,2-dioxygenase, Alcaligenes denitrificans, purification, characterization, chlorobenzoic acid degradation.
Changes in the rumen microbial population and its activities during the refaunation period after the reintroduction of ciliate protozoa into the rumen of defaunated sheep
Authors: Williams Alan G. , Withers Susan E.
Published: May 2010

Changes in the microbial populations, their activities, and the ruminal fermentation were monitored for 50 d following the reintroduction of ciliate protozoa into four defaunated sheep. A protozoal population was reestablished successfully in each recipient, using a washed inoculum containing approximately 103 cells, although there were between-animal differences in the rates of recolonization and genus establishment. Entodinium spp. predominated in the initial stages of the refaunation period and had an apparent maximal generation time of 9–10 h. Bacterial and fungal numbers did not decline following the reintroduction of protozoa and a small transient increase in the numbers of amylolytic and xylanolytic bacteria and fungal zoospores occurred in the early stages of refaunation when the protozoal population was < 105/g ruminal contents, but these subsequently declined as the protozoa established. Although the fibrolytic bacterial population was lowest in period 3 (> 105 protozoa/g), the in sacco ruminal digestion of Lolium perenne hay and polysaccharolytic enzyme activities in the solids-associated populations were either maintained or increased when protozoa were present confirming the important contribution of protozoa to fibre breakdown in the rumen. Significant changes in ruminal microbial activities occurred after protozoal reinoculation but before the rumen had refaunated completely. Arylamidase activities in the liquor-phase population and ruminal ammonia concentrations increased significantly within 48 h of transfaunation; the magnitude of the effects became more pronounced as the protozoal population developed. However, volatile fatty acid formation and ruminal pH were not affected after the reintroduction of protozoa.Key words: rumen, sheep, ciliate protozoa.
Tryptophan conversion to indole-3-acetic acid via indole-3-acetamide in Azospirillum brasilense Sp7
Authors: Bar Tami , Okon Yaacov
Published: May 2010

The phytohormone indole-3-acetic acid is involved in several types of microorganism-plant interactions. In the most widely studied pathway, tryptophan-2-monooxygenase converts tryptophan to the intermediate indole-3-acetamide, and indole-3-acetamide hydrolase catalyzes the conversion of indole-3-acetamide to indole-3-acetic acid. The genetic determinants for these enzymatic conversions are iaaM and iaaH, respectively. This pathway has been observed in many pathogenic and symbiotic soil bacteria. The associative soil bacteria of the genus Azospirillum are known to promote plant growth, probably via the secretion of phytohormones, including indole-3-acetic acid. The following evidence is presented for the existence of the above-described indole-3-acetic acid pathway in Azospirillum brasilense Sp7: the high toxicity of α-methyltryptophan as compared with that of 5-methyltryptophan; indole-3-acetic acid formation in vivo from indole-3-acetamide; the existence of two tryptophan-induced proteins, one of which has a molecular weight similar to that of tryptophan-2-monooxygenase; tryptophan-2-monooxygenase activity observed on nondenaturing gel; the existence of a protein with high tryptophan-2-monooxygenase activity with a molecular weight similar to that of one of the tryptophan-induced proteins on a two-dimensional gel; and the partial homology between the iaaM gene, which encodes tryptophan-2-monooxygenase in Pseudomonas savastanoi, and A. brasilense Sp7 total DNA.Key words: Azospirillum brasilense Sp7, indole-3-acetic acid, tryptophan, indole-3-acetamide.
Seasonal changes in the adherent microflora of the rumen in high-arctic Svalbard reindeer
Authors: Cheng K.-J. , McAllister T. A. , Mathiesen Svein D. , Blix Arnoldus S. , Orpin Colin G. , Costerton J. W.
Published: May 2010

Seasonal changes in bacterial colonization of the epithelial tissue were examined in the rumen of high-arctic Svalbard reindeer. Samples of tissue were collected from eight sites in the rumen of reindeer during summer and winter and bacterial colonization was examined using scanning and transmission electron microscopy. At two of these sites, colonization by adherent bacteria was estimated to cover approximately 30% of the ruminal epithelium in specimens collected from reindeer during summer. Bacteria at these sites resembled Ruminococcus sp. and were surrounded by large amounts of glycocalyx. In winter specimens, less than 10% of the epithelial surface was covered by adherent bacteria. Those bacteria that did colonize the epithelial surface were smaller and had virtually no glycocalyx on their surface. Bacteria attached to plant cell wall material in summer samples of reindeer ingesta contained large intracellular glycogen deposits, whereas feed particle-associated bacteria in ingesta collected in winter contained no intracellular glycogen. These data demonstrate that the ruminal bacterial population responds to seasonal changes in feed intake and quality. It is yet to be determined if these bacterial changes enhance the ability of Svalbard reindeer to survive in the hostile environment of the high Arctic.Key words: ruminal bacteria, attachment, epithelium, reindeer.
Outer membrane proteins from Serratia marcescens
Authors: Puig Marta , Fusté Carme , Viñas Miquel
Published: May 2010

The outer membrane proteins (OMPs) of several strains of Serratia marcescens have been studied by sodium dodecyl sulphate – urea – polyacrylamide gel electrophoresis. Four major OMPs, named Omp1, Omp2, Omp3, and OmpA (42, 40, 39, and 37 kDa, respectively), have been visualized. The relative proportions of Omp2 and Omp3 depend on cultural conditions (temperature of incubation, osmolarity, and nutrient availability).Key words: Serratia marcescens, outer membrane proteins, porin.
Liaisons entre les aptitudes écologiques de différentes souches de Fusarium et leur efficacité dans la protection de plants de lin contre la fusariose vasculaire
Authors: Amir Hamid , Mahdi Nourredine
Published: May 2010

Thirteen strains of Fusarium solani and F. oxysporum were isolated from an initial lot of 96 and submitted to 11 different tests to determine their ecological aptitudes. Results showed a great diversity with respect to performance. Significant total correlations with the strain aptitudes to protect flax against vascular fusariosis in one soil (efficacy) were obtained for three characters: quantity of microconidia produced (negative correlation), respiratory activity, and antagonistic effect in soil in the presence of a pathogenic fungus (positive correlations). Other properties were studied, such as mycelial growth, production of macroconidia, production of chlamydospores, spore germination aptitude, inhibiting effect of secretions against F. oxysporum f.sp. lini, saprophytic development in disinfected soil, and aptitude to maintain their development and activity in soil; all these properties showed generally high partial correlations with respect to efficacy. The multiple correlations obtained for different aptitude associations tested were clearly more significant than simple correlations. The results suggest that competition among Fusarium spp., here illustrated by efficacy, can be interpreted as a complex power corresponding to the sum of different particular aptitudes, notably: global activity, mycelial growth, sporulation, spore germination capacity, and secretion of compounds inhibiting competing germs. If selection of antagonistic Fusarium strains for biological control were based upon a single criterion, this would be arbitrary in part; a multiple-step selection would be recommended. The present work was performed on a single soil and with a limited number of strains; consequently, further experiments are required.Key words: Fusarium, competition, fusariosis, biological control.[Journal translation]not available
Antifungal effect of bean endochitinase on Rhizoctonia solani: ultrastructural changes and cytochemical aspects of chitin breakdown
Authors: Benhamou Nicole , Broglie Karen , Broglie Richard , Chet Ilan
Published: May 2010

A chitinase, purified to homogeneity from ethylene-treated bean leaves, was applied to actively growing mycelial cells of Rhizoctonia solani to evaluate a potential antifungal activity. Light microscopic investigations at 30-min intervals following enzyme exposure revealed the induction of morphological changes such as swelling of hyphal tips and hyphal distortions. More precise information concerning fungal cell alteration was obtained by ultrastructural observation and cytochemical detection of chitin distribution in fungal cell walls. Chitin breakdown was found to be an early event preceding wall disruption and cytoplasm leakage. The large amounts of chitin present in the walls of control R. solani cells and the rapid chitin hydrolysis upon chitinase treatment lead us to suggest that this polysaccharide is one of the main components of this fungal cell wall and is readily accessible to chitinase, especially in the apical zone. By 60 min after enzyme treatment, labeled molecules were observed in the vicinity of some fungal cells, suggesting the release of chitin oligosaccharides from fungal cell walls. The antifungal activity of the bean chitinase on cells of R. solani grown in culture is discussed in relation to the potential of genetically modified transgenic plants to resist attack by R. solani through an antimicrobial activity in planta.Key words: gold labeling, wheat germ agglutinin, cytochemistry, Rhizoctonia solani, bean endochitinase.
Rhizosphere colonization of wheat by selected soil bacteria over diverse environments
Authors: Milus Eugene A. , Rothrock Craig S.
Published: May 2010

The ability to colonize the rhizosphere is essential for bacteria to function as biological control agents for soil-borne plant pathogens. Eight bacterial strains reported to colonize wheat roots, inhibit root pathogens, and (or) improve wheat growth and yield were applied to wheat seeds that were planted in fumigated and nonfumigated soil in the 1990 and 1991 growing seasons at two locations in Arkansas. Rhizosphere population sizes were highly correlated with population sizes on seeds. Bacillus subtilis strain D-39Sr colonized roots as well in nonfumigated as in fumigated soil, and the other seven strains had rhizosphere populations 0.3 to 1.1 log units higher in fumigated soil. Pseudomonas fluorescens strain 2-79R was one of the best colonizers, and Streptomyces strain D-185S was the poorest. The greatest difference among strains was for relative colonization ability of crown roots in the spring. All strains except Streptomyces strain D-185S appear to have broad adaptation to colonize wheat roots and are able to compete with soil microflora for colonization sites. This research indicates that it is possible to select bacteria in the genera Bacillus, Pseudomonas, or Xanthomonas that will colonize roots well over diverse environments. Rhizosphere colonization by these strains was not associated with disease suppression or enhanced plant growth or yield.Key words: biological control, rhizosphere competence, wheat, Triticum aestivum.
Investigations of Entomophthora muscae (Cohn) (Entomophthorales: Entomophthoraceae) conidial infection on housefly Musca domestica (Linn.) by scanning electron microscopy
Authors: Tu C. M. , Singh B. L.
Published: May 2010

An entomogenous fungus, Entomophthora muscae, was studied morphologically with respect to the formation of primary and secondary conidia and germ tubes. The fungus penetrated the insect cuticle by germ tubes that were produced at the base of each conidium that penetrated directly through the cuticle. Fungal germ-tube formation and penetration of host integument were observed. The tough germ-tube penetration point seemed to provide abundant energy for the penetration of the host integument. Conidia not directly on the integument formed secondary conidia but were never observed to form germ tubes. Neither appressorium nor infection cushion was observed on the germ tubes.Key words: Entomophthora muscae, entomogenous fungus, insect mycosis, insect pathology, conidia.
Luminometric analyses of plant root colonization by bioluminescent pseudomonads
Authors: Beauchamp Chantal J. , Kloepper Joseph W. , Lemke Paul A.
Published: May 2010

Bioluminescence was used to study root colonization in nonsterile soil by Tn5-luxAB mutants of four Pseudomonas sp. strains. These bioluminescent strains colonized cucumber, cotton, maize, and soybean roots following seed treatment. In general, there were significant interactions between host and strain. Soybean and cucumber supported the best bacterial colonization, followed by maize and cotton. Across all crops, the best colonizing strains were GR7.4L and GR20.5L, with mean populations of about log 6.0 (colony-forming units + 1)/root system, followed by strains 61.9A.3L and 86.139LR with mean populations of about log 5.0 and 4.5 (colony-forming units + 1)/root system, respectively. The Lux+ strains colonized abundantly the upper root system, while their populations decreased progressively down the root system, and their root-tip populations were independent of mean populations at the crown and middle-root segments. Bioluminescence was observed visually from colonies growing on selective media and photometrically on roots with a luminometer and a charge-coupled device camera. Embedding roots, in situ root luminometer readings, and root-imaging techniques were less sensitive for detecting bacteria on roots than dilution-plating and broth enrichment techniques using selective media. The number of bacteria and competition with other rhizosphere microorganisms affected the sensitivity of the technique used. Selective broth enrichment was less time consuming than the dilution-plating technique. Bioluminescence is a simple and fast method to evaluate colonization of roots by bacteria.Key words: genetic marker, bioluminescence, lux genes, root colonization, Pseudomonas sp.
Cloning, expression, and sequence analysis of a cytolytic enterotoxin gene from Aeromonas hydrophila
Authors: Chopra Ashok K. , Houston Clifford W. , Peterson Johnny W. , Jin Gui-F.
Published: May 2010

The structural gene and regulatory element for a cytolytic enterotoxin of a diarrheal isolate, SSU, of Aeromonas hydrophila was cloned and its DNA sequence was determined. A complementary, mixed synthetic oligonucleotide based on the first 10 NH2-terminal amino acid residues of the Aeromonas cytolytic enterotoxin was used as a probe to screen a genomic library constructed in bacteriophage EMBL3. Cell lysates of Escherichia coli (λCH4), containing the cytolytic enterotoxin gene, lysed rabbit red blood cells and destroyed Chinese hamster ovary cells, caused fluid secretion in rat ileal loops, and were lethal to mice when injected intravenously. All biological activities associated with the cytolytic enterotoxin were neutralized by rabbit homologous polyclonal antibodies. Sodium dodecyl sulfate polyacrylamide gel electrophoresis and subsequent Western blot analysis of the cell lysate of E. coli (λCH4) revealed a protein band of approximately 52 kDa, using antisera to the cytolytic enterotoxin or antibodies generated against a synthetic peptide to the toxin. DNA sequence analysis of a 2.8-kb SalI-BamHI fragment revealed the presence of one large open reading frame (1479 bp) that would encode a protein of 54.5 kDa, a precursor form of the cytolytic enterotoxin, with a 23 amino acid leader peptide. Despite a significant amount of homology at the DNA and amino acid levels between our cytolytic enterotoxin and two aerolysins of Aeromonas species, variation in the restriction maps of these three toxin genes was prominent. Likewise, considerable divergence in DNA sequence was observed upstream of the structural genes for the reported aerolysins and our cytolytic enterotoxin, suggesting that these structurally similar toxin molecules may be regulated differently. Finally, our data showed that the cytolytic enterotoxin from a diarrheal isolate, SSU, of A. hydrophila exhibited characteristics that were unique compared with those of the reported aerolysins.Key words: Aeromonas hydrophila, cytolytic enterotoxin, aerolysin, cholera toxin.
Biodegradation of cycloalkane carboxylic acids in oil sand tailings
Authors: Herman David C. , Fedorak Phillip M. , Costerton J. William
Published: May 2010

The biodegradation of both an n-alkane and several carboxylated cycloalkanes was examined within tailings produced by the extraction of bitumen from the Athabasca oil sands. The carboxylated cycloalkanes examined were structurally similar to naphthenic acids that have been associated with the acute toxicity of oil sand tailings. The biodegradation potential of naphthenic acids was estimated by determining the biodegradation of both the carboxylated cycloalkanes and hexadecane in oil sand tailings. Carboxylated cycloalkanes were biodegraded within oil sand tailings, although compounds with methyl substitutions on the cycloalkane ring were more resistant to microbial degradation. Microbial activity against hexadecane and certain carboxylated cycloalkanes was found to be nitrogen and phosphorus limited.Key words: biodegradation, carboxylated cycloalkanes, oil sand tailings.
Sequence analysis of the lysin gene region of the prolate lactococcal bacteriophage c2
Authors: Ward Lawrence J. H. , Beresford Tom P. J. , Lubbers Mark W. , Jarvis Brion D. W. , Jarvis Audrey W.
Published: May 2010

Approximately 80% of the genome of the prolate-headed lactococcal bacteriophage c2 was cloned into shuttle vectors pSA3 and pFX3 in Escherichia coli and transferred to Lactococcus lactis. A 1.67-kilobase EcoRV fragment containing the gene for the phage lysin was identified and the position and orientation of the phage lysin gene in the physical map of the phage were determined. The phage lysin was expressed in E. coli and its sequence was determined and compared with the sequences of other bacteriophage lytic genes. The sequence was similar, but not identical, to that of the related lactococcal phage ml3, having a number of silent substitutions and an apparent deletion that altered the carboxy terminus of the protein. Possible alternative translation initiation codons for the lysin gene and two possible alternative mechanisms for access of the lysin enzyme to the cell wall are discussed. An open reading frame upstream of the putative lysin gene was found to be 177 base pairs longer than that reported for phage ml3. A codon usage table for the lysin genes of several phages as well as for reported gene sequences from L. lactis and lactococcal bacteriophages is presented.Key words: Lactococcus lactis, codon usage, molecular organization, holin gene.
A medium, solidified with gellan gum, for determining the Na+ requirement of Vibrio species
Authors: Noble Lisa D. , Gow John A.
Published: May 2010

Until now there has not been a satisfactory solid medium for determining the growth responses, to Na+, of marine and other bacteria that have specific growth requirements for Na+. A solid medium would be useful to investigators who would like to take advantage of the efficiency of multipoint inoculation when testing for a Na+ requirement. By using 1% gellan gum (Gel-GroTM) as the solidifying agent a medium was formulated that had a contaminating level of Na+ of slightly less than 2 mM in the basal medium. Two species of Aeromonas, which do not require Na+ for growth, and 31 species of Vibrio, which require Na+, were tested for their growth responses to Na+ on this medium. The Aeromonas strains grew well, within 24 h, at all of the Na+ concentrations tested. Approximately 75% of the Vibrio strains did not grow on the basal medium even after a prolonged incubation period. The remaining species were able to grow on the basal medium, but not without a lag period. These lag periods were as short as 36 h for two of the species and in some instances as long as 312 h. These lag periods were of sufficient duration to determine that Na+ stimulated the growth of the Vibrio strains that were able to grow on the basal medium. Approximately 75% of the strains, representing most species of Vibrio, were able to grow if as little as 25 mM Na+ was present in the medium.Key words: low-sodium medium, Na+ requirement, gellan gum, agar substitute, marine bacteria.
Microorganisms in the gastrointestinal tract of the rat prevent absorption of the mutagen–carcinogen 3-amino-1,4-dimethyl-5H-pyrido(4,3-b)indole
Authors: Zhang Xue Bin , Ohta Yoshiyuki
Published: May 2010

The extent to which lactic acid bacteria, intestinal bacteria, and yeast from the gastrointestinal tract of rats suppress the absorption of 3-amino-1,4-dimethyl-5H-pyrido(4,3-b)indole (Trp-P-1) was investigated. Trp-P-1 was absorbed from the small intestine very rapidly, but in the stomach it was slowly absorbed, requiring 1 or 2 h after administration. When mixtures of Trp-P-1 and freeze-dried microorganisms were administered to rats for 1 h, the amounts of Trp-P-1 absorbed from the small intestine were significantly reduced, and the levels of Trp-P-1 in blood decreased by 40.4–64.7% compared with a control in which only Trp-P-1 was administered. There were no significant differences between the organisms used. In vitro, freeze-dried cells of the strains tested bound 51-97% of Trp-P-1. The Trp-P-1 bound to cells was effectively extracted by aqueous methanol, ethanol, ammonia (50 g/L), and solutions of MgCl2 and CaCl2 (100 mM/mL), but little was extracted by water and solutions of KCl, NaCl, and buffers at various pH values.Key words: Trp-P-1, gastrointestinal absorption, binding, intestinal bacteria, mutagen–carcinogen.
Purification and characterization of an extracellular glucoamylase from the thermophilic fungus Humicola grisea var. thermoidea
Authors: Tosi Luis Ricardo Orsini , Terenzi Héctor Francisco , Jorge Joāo Atílio
Published: May 2010

Humicola grisea var. thermoidea mycelium grown on maltose as the main source of carbon produced at least two amylases. The major amylolytic component was purified to homogeneity and classified as a glucoamylase. The apparent molecular mass of the purified enzyme was estimated to be 63 000 Da by SDS-PAGE and 65 000 Da by Bio-Gel P-100 filtration. The purified enzyme was a glycoprotein with 1.8% carbohydrate content and pH and temperature optima of 5.0 and 55 °C, respectively. The purified glucoamylase was thermostable at 60 °C with a half-life of 16 min at 65 °C. In the presence of starch the purified enzyme retained 75% of its thermostability at 65 °C, while the addition of maltose failed to protect the activity. The purified enzyme hydrolyzed branched substrates more efficiently than linear substrates. Starch and amylopectin were the best substrates utilized and amylose was hydrolyzed faster than maltopentaose, maltotetraose, and maltotriose. Kinetic experiments suggested that maltose and starch were hydrolyzed at the same catalytic site.Key words: glucoamylase, amylase, Humicola grisea.
DNA reassociation and electrokaryotype study of some Candida species and synonymy of Candida terebra, Candida entomaea, and Candida veronae
Authors: Lee Ching-Fu , Lee Fwu-Ling , Hsu Wen-Haw , Hsu Wen Hwei
Published: May 2010

Some Candida species that were similar in phenotypic characteristics were studied by molecular taxonomic comparisons including electrophoretic karyotypes, DNA base composition, and DNA relatedness. Among the species studied, Candida terebra, Candida entomaea, and Candida veronae showed similar electrophoretic karyotypes and a G + C range from 39.7 to 40.2 mol% in DNA base composition, and demonstrated 85–104% DNA similarity on the basis of flourometric hybridization in microdilution wells. As C. terebra has priority, we suggest that C. entomaea and C. veronae should be synonyms of C. terebra. Other species in this study, Candida dendronema, Candida diddensiae, Candida naeodendra, and Candida atmospherica, had low DNA relatedness. Thus, these species should be maintained in their original separate taxa.Key words: DNA reassociation, electrokaryotype, Candida species, synonym.
MutY repair is mutagenic in mutT− strains of Escherichia coli
Authors: Vidmar J. John , Cupples Claire G.
Published: May 2010

We have determined the numbers and types of mutations that occur in strains of Escherichia coli defective in mutT and (or) mutY repair. High rates of C∙G to A∙T mutations in mutY− cells are unaffected by the status of mutT. However, mutT−/mutY− strains have higher rates of A∙T to C∙G mutations than mutT−/mutY− strains. This result indicates that the high rates of A∙T to C∙G mutations seen in mutT− strains of E. coli are due in part to the activity of the mutY repair system. We conclude that mutY repair is mutagenic in a mutT− background.Key words: mutT, mutY, E. coli mutator strains.
Yeasts and coliform bacteria of water accumulated in bromeliads of mangrove and sand dune ecosystems of southeast Brazil
Authors: Hagler Allen N. , Rosa Carlos A. , Morais Paula B. , Mendonça-Hagler Leda C. , Franco Georgia M. O. , Araujo Fabio V. , Soares Carlos A. G.
Published: May 2010

Yeasts and coliform bacteria were isolated from water that accumulated in the central cups and adjacent leaf axilae of two bromeliads, Neoregelia cruenta of a coastal sand dune and Quesnelia quesneliana of a mangrove ecosystem near the city of Rio de Janeiro, Brazil. The mean total coliform counts were above 10 000 per 100 mL for waters of both plants, but the mean fecal coliform counts were only 74 per 100 mL for Q. quesneliana and mostly undetected in water from N. cruenta. Of 90 fecal coliform isolates, 51 were typical of Escherichia coli in colony morphology and indol, methyl red, Volges–Proskauer, and citrate (IMViC) tests. Seven representatives of the typical E. coli cultures were identified as this species, but the identifications of nine other coliform bacteria were mostly dubious. The yeast community of N. cruenta was typical of plant surfaces with basidiomycetous yeasts anamorphs, and the black yeast Aureobasidium pullulans was prevalent. Quesnelia quesneliana had a substantial proportion of ascomycetous yeasts and their anamorphs, including a probable new biotype of Saccharomyces unisporus. Our results suggested that the microbial communities in bromeliad waters are typically autochtonous and not contaminants.Key words: yeasts, fecal coliforms, bromeliad waters, mangrove, Restinga.
Extra- and intra-cellular lipases from a thermophilic Rhizopus oryzae and factors affecting their production
Authors: Salleh A. B. , Musani R. , Basri M. , Ampon K. , Yunus W. M. Z. , Razak C. N. A.
Published: May 2010

A thermophilic Rhizopus oryzae was isolated, and parameters affecting its production of extra- and intra-cellular lipases were investigated. All carbon sources tested with the exception of sucrose generally inhibited the production of extracellular lipase, but enhanced the production of intracellular lipase. Peptone was the best substrate for extracellular enzyme production, but for intracellular lipase production other substrates such as tryptone, tryptic soy digest, polypeptone, and corn steep liquor gave comparable results. Among lipid substrates, glycerol was the only stimulator of extracellular enzyme production, whereas olive oil, triolein, and oleic acid had very positive effects on intracellular enzyme production. Shaking enhanced the production of both types of enzymes; the temperature optima were 45 and 37 °C for extra- and intra-cellular lipases, respectively. A pH of 5.0 was optimal for production of both enzymes.Key words: lipases, Rhizopus oryzae, production.
Specific 16S ribosomal RNA targeted oligonucleotide probe against Clavibacter michiganensis subsp. sepedonicus
Authors: Mirza M. S. , Rademaker J. L. W. , Janse J. D. , Akkermans A. D. L.
Published: May 2010

In this article we report on the polymerase chain reaction amplification of a partial 16S rRNA gene from the plant pathogenic bacterium Clavibacter michiganensis subsp. sepedonicus. A partial sequence (about 400 base pairs) of the gene was determined that covered two variable regions important for oligonucleotide probe development. A specific 24mer oligonucleotide probe targeted against the V6 region of 16S rRNA was designed. Specificity of the probe was determined using dot blot hybridization. Under stringent conditions (60 °C), the probe hybridized with all 16 Cl. michiganensis subsp. sepedonicus strains tested. Hybridization did not occur with 32 plant pathogenic and saprophytic bacteria used as controls under the same conditions. Under less stringent conditions (55 °C) the related Clavibacter michiganensis subsp. insidiosus, Clavibacter michiganensis subsp. nebraskensis, and Clavibacter michiganensis subsp. tesselarius also showed hybridization. At even lower stringency (40 °C), all Cl. michiganensis subspecies tested including Clavibacter michiganensis subsp. michiganensis showed hybridization signal, suggesting that under these conditions the probe may be used as a species-specific probe for Cl. michiganensis.Key words: Clavibacter michiganensis subsp. sepedonicus, PCR, 16S rRNA, oligonucleotide probe.
First year field performance of spruce seedlings inoculated with plant growth promoting rhizobacteria
Authors: Chanway C. P. , Holl F. B.
Published: May 2010

The influence of plant growth promoting rhizobacteria on field performance of hybrid spruce (Picea glauca × engelmannii) was investigated by inoculating seedlings with rhizobacteria capable of stimulating seedling growth in a controlled environment. Two spruce ecotypes (from Mackenzie and Salmon Arm, British Columbia) and two bacterial strains previously isolated from naturally regenerating seedlings of each spruce ecotype were evaluated. Planting trials were conducted in the ecosystem from which each spruce ecotype and associated bacterial strain were originally collected, and at two alternative sites. Hydrogenophaga pseudoflava, which was isolated from Mackenzie spruce seedlings, caused increases in seedling biomass or branch number of up to 49%, but was most effective as a root growth promoter of the Salmon Arm spruce ecotype. Pseudomonas putida, which originated from Salmon Arm spruce seedlings, increased seedling biomass or branch number in two trials, but had inhibitory effects in three others. There was no indication that growth promotion was related to a common ecotypic origin of seedlings and rhizobacteria, or that bacteria were more effective in the ecosystem from which they were originally isolated. However, Salmon Arm spruce growth promotion by H. pseudoflava was greatest at the poorest quality planting site.Key words: field trials, inoculation, PGPR, spruce.
Mutant derivatives of Pseudomonas putida GR12-2R3 defective in nutrient utilization or cell surface structures show reduced ability to promote canola root elongation
Authors: Bayliss Cathy , Lasby Bonnie , Wood Janet M. , Lifshitz Ran , Brown Gerry L.
Published: May 2010

Pseudomonas putida GR12-2R3 is a rifampicin-resistant derivative of a cold-tolerant, nitrogen-fixing bacterium isolated from the roots of grasses growing in the Canadian high arctic. It colonizes canola (Brassica campestris) roots and promotes canola root and shoot growth under gnotobiotic conditions and in the field. TnphoA insertion mutagenesis was used to isolate derivatives of strain GR12-2R3 that had reduced abilities to promote canola root elongation (PRE− mutants). Within a pool of 10 290 TnphoA insertion mutants, 1.4% expressed active PhoA fusion proteins (PhoA+). Among 20 PhoA+ mutants, 6 were PRE− and 4 of those strains secreted PhoA activity into the culture medium. PhoA+ strain PG269 showed PhoA activity, 25% cell associated, that was induced by canola seed exudate. The ability of this strain to promote canola root elongation was similar to that of the parent strain. Like other pseudomonads, strain GR12-2R3 utilizes a wide range of sugars, amino acids, and other compounds as carbon and nitrogen sources. Twenty-one TnphoA insertion mutants, all PhoA−, were unable to utilize a specific nutrient. That group included strains that could not utilize arabinose (Ara−, three mutants) or glycerol and other compounds (Csu−, three mutants) as carbon source and strains that could not utilize glycine (Gut−, eight mutants), histidine (Hut−, three mutants), or proline (Put−, four mutants) as nitrogen source. One Ara− mutant, three Gut− mutants, and one Csu− mutant were PRE−. Five of the mutant strains examined in detail (two PhoA− PRE−, one PhoA− PRE+, one PhoA+ PRE−, and one PhoA+ PRE+) grew less well than strain GR12-2R3 in LB and (or) seed exudate medium. Further characterization of the TnphoA target genes in selected PRE− strains is expected to yield additional insight regarding the molecular basis for the interaction between P. putida GR12-2R3 and canola.Key words: Pseudomonas putida, plant growth promoting rhizobacterium, TnphoA.
Isolation from soil of symbiotic and nonsymbiotic Rhizobium leguminosarum by DNA hybridization
Authors: Laguerre Gisèle , Bardin Marc , Amarger Noëlle
Published: May 2010

A procedure based upon DNA hybridization was developed for the specific detection of Rhizobium leguminosarum and its different biovars among bacteria isolated from soil. DNA colony hybridization and restriction fragment length polymorphism analysis with a R. leguminosarum chromosomal probe were found to be species specific for R. leguminosarum and Rhizobium etli. By using R. leguminosarum nod gene probes, biovar specificity was obtained. Of 302 soil isolates screened for their inability to grow on Luria-Bertani agar medium, 13 strains could be assigned to the R. leguminosarum species on the basis of DNA homology to the chromosomal probe and antibiotic resistance tests. Of these strains, three and two were assigned by colony hybridization and subsequent plant host specificity tests, respectively, to R. leguminosarum biovars viciae and trifolii. The eight other R. leguminosarum soil isolates lacked symbiotic information but were able to gain nodulation capacity with the acquisition of a conjugative symbiotic plasmid. They were thus considered as nonsymbiotic R. leguminosarum.Key words: Rhizobium leguminosarum, DNA hybridization, soil, symbiotic genes.
Nucleotide sequence of the hemG gene involved in the protoporphyrinogen oxidase activity of Escherichia coli K12
Authors: Sasarman Alexandre , Letowski Jaroslav , Czaika Guy , Ramirez Volta , Nead Michael A. , Jacobs Judith M. , Morais Réjean
Published: May 2010

The hemG gene of Escherichia coli K12 is involved in the activity of protoporphyrinogen oxidase, the enzyme responsible for the conversion of protoporphyrinogen IX into protoporphyrin IX during heme and chlorophyll biosynthesis. The gene is located at min 87 on the genetic map of E. coli K12. The hemG gene was isolated by a mini-Mu in vivo cloning procedure. As expected, the hemG gene is able to restore normal growth to the hemG mutant, and the transformed cells display strong protoporphyrinogen oxidase activity. Sequencing of the hemG gene allowed us to identify an open reading frame of 546 nucleotides (181 amino acids), within the minimal fragment able to complement the mutant. The presumed molecular mass of the HemG protein is 21 202 Da, in agreement with values found by SDS-PAGE, in a DNA-directed coupled transcription–translation system. The identity of the first 18 amino acids at the amino-terminal end of the protein was confirmed by microsequencing. To our knowledge, this is the first cloning of a gene involved in the protoporphyrinogen oxidase activity of E. coli.Key words: protoporphyrinogen oxidase (PROTOX), hemG gene, Escherichia coli, DPE herbicides, heme.
MVP interacts with YPEL4 and inhibits YPEL4-mediated activities of the ERK signal pathway
Authors: Liang Pei , Wan Yongqi , Yan Yan , Wang Yuequn , Luo Na , Deng Yun , Fan Xiongwei , Zhou Junmei , Li Yongqing , Wang Zequn , Yuan Wuzhou , Tang Ming , Mo Xiaoyang , Wu Xiushan
Published: May 2010

Human YPEL4 is a member of YPEL family. It contains a Yippee domain, which is a putative zinc-finger-like, metal-binding domain. The human YPEL4 gene maps to chromosome 11q12.1, is ubiquitously expressed in adult tissues, and encodes a nuclear protein of 127 amino acids, the function of which remains unknown. To gain insights into the cellular function of this protein, we searched for YPEL4-interacting proteins using a yeast two-hybrid screen. The major vault protein (MVP), a lung resistance associated protein, was identified as a binding partner of YPEL4. The interaction between YPEL4 and MVP in mammalian cells was further demonstrated by a series of biochemical assays including the mammalian two-hybrid assay, GST pull-down assay, co-immunoprecipitation assay, and immunocytochemistry. Using a reporter system, we found that MVP can inhibit YPEL4’s ability to activate Elk-1 in the MAPK signaling pathway. This study provides new clues for understanding the molecular mechanism of YPEL4 in cell division and signal transduction pathways and should be helpful for understanding molecular functions of the YPEL family.
Endoplasmic reticulum stress inhibits collagen synthesis independent of collagen-modifying enzymes in different chondrocyte populations and dermal fibroblasts
Authors: Vonk Lucienne A. , Doulabi Behrouz Zandieh , Huang Chun-Ling , Helder Marco N. , Everts Vincent , Bank Ruud A.
Published: May 2010

Chondrocytes respond to glucose deprivation with a decreased collagen synthesis due to disruption of a proper functioning of the endoplasmic reticulum (ER): ER stress. Since the mechanisms involved in the decreased synthesis are unknown, we have investigated whether chaperones and collagen-modifying enzymes are affected by glucose deprivation. Chondrocytes obtained from nucleus pulposus, annulus fibrosus, articular cartilage, and meniscus and dermal fibroblasts were cultured under control conditions or exposed to the ER stress-inducing treatments of tunicamycin addition or glucose withdrawal. Both treatments resulted in an up-regulation of the gene expression of the ER stress markers in all cell types, but dermal fibroblasts showed a delayed response to glucose deprivation. Collagen gene expression was down-regulated, and less collagen protein was present in the cells under both ER stress-inducing conditions. The expression levels of the prolyl 4-hydroxylases were either not affected (P4ha3) or increased (P4ha1 and P4ha2), the levels of the lysyl hydroxylases decreased, and the N-propeptidase Adamts2 decreased. Both treatments induced apoptosis. Chondrocytes respond more quickly to glucose deprivation, but it appears that chondrocytes can cope better with tunicamycin-induced ER stress than fibroblasts. Although collagen synthesis was inhibited by the treatments, some collagen-modifying enzymes and chaperones were up-regulated, suggesting that there is no causal relation between them.
Epigenetic analysis of human embryonic carcinoma cells during retinoic acid-induced neural differentiation
Authors: Shahhoseini Maryam , Taei Adeleh , Mehrjardi Narges Zare , Salekdeh Ghasem Hosseini , Baharvand Hossein
Published: May 2010

Differentiation of stem cells from a pluripotent to a committed state involves global changes in genome expression patterns, critically determined by chromatin structure and interactions of chromatin-binding proteins. The dynamics of chromatin structure are tightly regulated by multiple epigenetic mechanisms such as histone modifications and the incorporation of histone variants. In the current work, we induced neural differentiation of a human embryonal carcinoma stem cell line, NTERA2/NT2, by retinoic acid (RA) treatment, primarily according to two different methods of adherent cell culture (rosette formation) and suspension cell culture (EB formation) conditions, and histone modifications and variations were compared through these processes. Western blot analysis of histone extracts showed significant changes in the acetylation and methylation patterns of histone H3, and expression level of the histone variant H2A.Z, after RA treatment in both protocols. Using chromatin immunoprecipitation (ChIP) coupled with real-time PCR, it was shown that these epigenetic changes occurred on the regulatory regions of 4 marker genes (Oct4, Nanog, Nestin, and Pax6) in a culture condition dependent manner. This report demonstrates the dynamic interplay of histone modification and variation in regulating the gene expression profile, during stem cell differentiation and under different culture conditions.
A solvent-free protocol for the green synthesis of arylalkylidene rhodanines in a task-specific ionic liquid
Authors: Alizadeh Abdolhamid , Khodaei Mohammad M. , Eshghi Ali
Published: May 2010

2-Hydroxyethylammonium formate acts as a task-specific ionic liquid (TSIL) for the Knoevenagel condensation of carbonyl compounds with rhodanine to afford arylalkylidene rhodanines under solvent-free conditions and in good-to-excellent yields. Additionally, compared with those in organic solvents, the yields obtained in the presence of our ionic liquid (IL) were significantly increased. The detailed mechanism of the catalytic effect of TSIL is also reported for the first time.
Interactions of cytidine with N2-functionalized guanosines and cytidine – cytidine exchange involving a GC pair — NMR and fluorescence spectroscopic study
Authors: Martic Sanela , Wu Gang , Wang Suning
Published: May 2010

Two N2-functionalized guanosines by diphenylaminobiphenyl and di(2-pyridyl)aminobiphenyl have been found to act as the effective probes for G–C interactions in organic media. Because of the highly emissive nature of the N2-functionalized guanosines in the visible region, the GC base pair formation event accompanied by distinct fluorescence quenching can be readily monitored by fluorescence spectroscopy. NMR and fluorescence results confirm that the N2-arylguanosines form H-bonded pairs with cytidine, selectively. An unusual exchange pathway between non-bound cytidine and bound cytidine, in the GC pair, has been identified and extensively studied by NMR methods.