bluuebunny/crossref_metadata_2025 huggingface dataset:
A truncated β-xylosidase from the anaerobic fungus Neocallimastix patriciarum 27
Authors: Zhu Hong , Cheng K.-J. , Forsberg Cecil W.
Published: February 2010

Two extracellular β-xylosidases, xylosidase I and II, were isolated from the ruminal fungus Neocallimastix patriciarum 27 after growth in a barley straw medium. Xylosidase I was purified 88-fold to apparent homogeneity by ion-exchange, affinity, and gel filtration chromatography. The purified xylosidase I had an isoelectric point (pI) of 4.7 and was a monomelic protein with a molecular mass of 39.5 kDa as estimated by both SDS-PAGE and gel filtration. Xylosidase II was partially purified to approximately 95% purity. Xylosidase II had the same pI (4.7) as xylosidase I, and appeared to be a dimeric enzyme composed of two polypeptides with molecular masses of 85 and 45 kDa, respectively, on SDS-PAGE. Peptide mapping of the three proteins suggested that xylosidase I was a truncated product originating from xylosidase II. Xylosidases I and II had similar pH optima of 6.0, but different temperature optima of 50 and 40 °C, respectively. The Km and Vmax for xylosidase I were 0.59 mM of p-nitrophenyl-β-D-xylopyranoside and 38.04 U∙mg protein−1, respectively, and those for xylosidase II were 0.13 mM and 8.9 U∙mg protein−1, respectively. Both enzymes hydrolysed pNPX and xylobiose with the production of xylose, but only xylosidase I exhibited activity toward p-nitrophenyl-α-L-arabinofuranoside.Key words: xylosidase, Neocallimastix, patriciarum, glycosidase.
Bacteriocin plasmid pMB1 of Enterococcus faecalis: identification of the cell aggregation substance after induction by sex pheromone
Authors: Quirantes R. , Martín I. , Valdivia E. , Gálvez A. , Martínez-Bueno M. , Maqueda M.
Published: February 2010

Strains of Enterococcus faecalis carrying the bacteriocinogenic plasmids pMB1 or pMB 1.1 exhibit a clumping response to culture supernatants of different enterococcal strains. Antibodies raised against cells induced by a homologous pheromone recognize two surface proteins of 152 and 72.5 kDa (the second one is possibly the degradation product of the first), respectively. These antigens are very similar to those found in induced cells of E. faecalis OGIRF(pAM211) as shown by the cross-reaction of the immune sera obtained in this work. We propose that the 152-kDa protein corresponds to the aggregation substance coded by plasmids pMB1 and pMB1.1. Moreover, antibodies raised against induced cells are able to block the pheromone-induced clumping response. When the cells induced to form aggregates by pheromones were examined under a scanning electron microscope they showed a surface layer of hairlike structures.Key words: pMB1 plasmid, sex pheromone, aggregation substance, Enterococcus faecalis.
Isolation and characterization of a Tn5-induced tolQ mutant of Escherichia coli
Authors: Madrid C. , Badenas C. , Juárez A.
Published: February 2010

Transposon mutagenesis was used to isolate an Escherichia coli mutant that released into the external medium a heterologous protein, the Aeromonas hydrophila aerolysin. Genetic mapping and phenotypic characterization of E. coli CM209 showed that Tn5 is inserted in the tolQ gene. This mutant strain released a significant amount of unprocessed (proaerolysin) protein into the external medium, together with other periplasmic proteins. Similar levels of the toxin were detected in the intracellular compartments of the parental and mutant strains. Whereas inactivation of the tolQ gene itself was responsible for some of the phenotypic properties of strain CM209, such as tolerance to group A colicins or to filamentous bacteriophages, the leaky phenotype was associated with a polar effect exerted by Tn5 on distal genes of the tolQRA cluster.Key words: Tn5, tolQ gene, aerolysin, leaky mutants.
Transferrin receptors on ruminant pathogens vary in their interaction with the C-lobe and N-lobe of ruminant transferrins
Authors: Yu Ronghua , Schryvers Anthony B.
Published: February 2010

The interaction between ruminant transferrins and receptor proteins on the surface of the ruminant pathogens Pasteuerella haemolytica, Haemophilus somnus, Pasteurella multocida, Haemophilus agnii, and Moraxella bovis was evaluated by a combination of binding assays and affinity isolation procedures. Membranes isolated from P. haemolytica, P. multocida, and H. agnii were capable of binding sheep, goat, and cattle transferrins whereas binding by membranes from H. somnus and M. bovis was specific for bovine transferrin. Proteolytically derived bovine transferrin C-lobe was capable of inhibiting the interaction between bovine transferrin and both Tbpl and Tbp2 from P. haemolytica and M. bovis but only Tbpl from H. somnus and P. multocida. Proteolytically derived N-lobe inhibited the binding of P. multocida and H. somnus Tbp2 to bovine transferrin and the binding of bovine transferrin to the single receptor protein identified in H. agnii. The implications of these results regarding the nature of the ligand–receptor interaction and similarities of this interaction with ligand–receptor interactions in different species are discussed.Key words: iron acquisition, transferrin receptor, binding specificity, Pasteurella, ruminants.
Identification of variability of ribosomal DNA spacer from Pseudomonas soil isolates
Authors: Gill Santokh , Belles-Isles Julie , Brown Gerry , Gagné Serge , Lemieux Claude , Mercier Jean-Patrick , Dion Patrice
Published: February 2010

The polymerase chain reaction was used to amplify the spacer region located between the 16S and 23S ribosomal RNA genes of strains of Pseudomonas fluorescens and Pseudomonas putida isolated from peat bog, canola field, or arctic plants. Some of these amplified spacer regions were used to probe Southern blots of total DNA digests from the various strains of P. fluorescens and P. putida. Differences were observed in the patterns of hybridization of the various bacterial DNAs. The ribosomal DNA spacer region of four of the P. fluorescens strains examined, strains 64-3, 63-28, QP5, and R17-FP2, was about 515 base pairs (bp) in length, and contained the genes for tRNAIle and tRNAAla. The DNA sequences of two strains from canola, 64-3 and 63-28, differed at only two positions. The sequences of the peat bog strains QP5 and R17-FP2 were identical. However, differences were noted between the DNA sequence common to the pair of strains 64-3 and 63-28 and the corresponding common sequence for strains QP5 and R17-FP2. These differences were mainly concentrated in two DNA segments of 10 and 19 bp, respectively. A probe for the 19-bp variable segment that occurs in the ribosomal spacer of strains QP5 and R17-FP2 recognized total DNA from these two strains, but not DNA from other bacteria of different origins. These results suggest the existence of a limited degree of variability within the 16S-23S ribosomal DNA spacer region, and that this variability may be useful to the recognition of particular Pseudomonas strains from environmental samples.Key words: ribosomal RNA genes, intergenic spacer, Pseudomonas soil isolates, specific DNA probe.
Description of bacteria able to degrade isoquinoline in pure culture
Authors: Aislabie J. , Richards N. K. , Lyttle T. C.
Published: February 2010

Isoquinoline is a nitrogen heterocyclic compound that is associated with coal- and oil-derived wastes. Four strains of bacteria able to degrade isoquinoline in pure culture were isolated from sites known to be contaminated with oil. Isoquinoline was used as the sole source of carbon and nitrogen by these isolates. Isoquinoline was initially transformed to 1-hydroxyisoquinoline, which accumulated in the broth culture, and then disappeared. The four strains isolated were Gram negative, aerobic, rod-shaped bacteria with polar flagella. The strains have been presumptively identified as members of the family Comamonadaceae.Key words: isoquinoline degradation, Comamonadaceae.not available
Catalase released from beneficial and plant-pathogenic pseudomonads by water and chloroform treatments
Authors: Pounder J. I. , Anderson A. J.
Published: February 2010

Survival of pseudomonads during plant colonization may involve bacterial catalases to degrade the hydrogen peroxide produced by the plant. The specific activities of catalases in lysates from two saprophytic isolates of Pseudomonas putida and Pseudomonas fluorescens and three races of Pseudomonas syringae pv. glycinea were similar. To explore the location of the bacterial catalases, cells of the pathogenic and saprophytic pseudomonads were treated with chloroform, which is reported to release periplasmic proteins. Although catalase was released by chloroform treatment, the cytoplasmic enzymes isocitrate dehydrogenase, superoxide dismutase, and glucose-6-phosphate dehydrogenase were also detected. These proteins may have come from lysis of a small proportion of the cells rather than the periplasm. Water treatment of cells also released amounts of protein similar to those derived from chloroform treatment. Similar responses were found from both pathogenic and saprophytic strains. The release of catalase and proteins from the leaf pathogen P. syringae pv. glycinea race 0 and the root-associated saprophyte P. putida decreased as the cultures aged. With P. putida and P. syringae pv. glycinea race 0, the single isozyme of catalase released by water and chloroform treatment also was detected in lysates. Additional catalase isozymes were present in lysates as the cultures aged.Key words: periplasmic proteins, survival.
Sterile fungi from zoysiagrass rhizosphere as plant growth promoters in spring wheat
Authors: Shivanna Manchanahally B. , Meera Manchanahally S. , Hyakumachi Mitsuro
Published: February 2010

Eleven out of 18 sterile fungal isolates and an isolate each of Penicillium sp. and Trichoderma sp. from the zoysiagrass rhizosphere were effective in enhancing the growth of two wheat varieties in greenhouse conditions. They enhanced the top length and top dry biomass of plants significantly and induced the plants to produce long earheads and more seeds. Notable among isolates were GS6-1, GS6-2, GS7-3, GS7-4, GS8-6, GS10-1, GS10-2, and GU23-3, which enhanced the growth by several times, resulting in a conspicuous growth promotion effect that differed depending on the variety. Penicillium and Trichoderma species were less effective than sterile isolates in enhancing growth. Seven of the 11 effective sterile isolates from the zoysiagrass rhizosphere (as determined under greenhouse conditions) and a wheat rhizosphere isolate (K-17) were further tested under field conditions. Most of the isolates except K-17 enhanced the growth of one variety, whereas a few isolates enhanced the growth of the other variety. However, at least four isolates each increased yields of both varieties. Isolate GS6-1, which was very effective under greenhouse conditions in promoting overall growth, was less effective under field conditions; however, the yield components were not affected. The efficiency of the plant growth promoting isolates depended upon the wheat variety and soil nutrient conditions in addition to their inherent growth promotion abilities.Key words: plant growth promoting fungi (PGPF), sterile fungi, wheat growth promotion, yield components.
Lipopolysaccharides of Legionella erythra and Legionella oakridgensis
Authors: Sonesson Anders , Jantzen Erik , Tangen Torill , Zähringer Ulrich
Published: February 2010

The chemical composition of lipopolysaccharides from Legionella erythra and Legionella oakridgensis was analysed. Sodium dodecyl sulfate polyacrylamide gel electrophoresis showed both lipopolysaccharides to have a smooth-type character. The polysaccharide part of both lipopolysaccharides contained D-mannose, D-glucose, D-glycero-D-mannoheptose, L-glycero-D-manno-heptose, 2-keto-3-deoxyoctonic acid, L-fucosamine, D-glucosamine, and glucosamine phosphate. In addition, L-rhamnose, glycerol phosphate, and glucose phosphate were identified in the polysaccharide part of L. erythra lipopolysaccharide. The main sugar identified in the lipid A part of both lipopolysaccharides, 2,3-diamino-2,3-dideoxy-D-glucose, was found to be substituted with a complex fatty acid composition including at least 16 different amide-linked 3-hydroxy fatty acids. Both lipopolysaccharides contained nonhydroxy fatty acids and the uncommon 27-oxo-octacosanoic acid, 29-oxotriacontanoic acid, and 27-hydroxyoctacosanoic acid. The lipopolysaccharide of L. oakridgensis also contained 29-hydroxytriacontanoic acid. The dioic long-chain acids heptacosane-1,27-dioic acid and nonacosane-1,29-dioic acid were only present in the lipopolysaccharide of L. erythra.Key words: taxonomy, long-chain fatty acids, chemical analysis, 2,3-diamino-2,3-dideoxy-D-glucose.
Numerical taxonomy of heterotrophic sulfur-oxidizing bacteria isolated from southwestern Pacific hydrothermal vents
Authors: Durand Pascale , Benyagoub Afeda , Prieur Daniel
Published: February 2010

Sulfur-oxidizing bacteria (n = 161) were enriched and isolated from samples of vent water, invertebrates, and chimney rocks collected at two deep-sea hydrothermal vents (2000 m) in back-arc basins from the southwestern Pacific: the North Fiji Basin and the Lau Basin. Several types of heterotrophic sulfur-oxidizing bacteria were repeatedly isolated. They oxidized thiosulfate either to sulfate (acid producing) or to polythionate (base producing). In most of the acid-producing cultures, thiosulfate was transitorily oxidized to polythionate. All of the bacteria were Gram negative, 37% were fermentative, and 88% were denitrifiers or nitrate reducers. Numerical taxonomy and analysis of the G+C content showed that they belong to several genera including Pseudomonas, Acinetobacter, and Vibrio.Key words: hydrothermal vent, culturable thiosulfate-oxidizing bacteria, numerical taxonomy.
Degradation of morpholine by Mycobacterium aurum MO1
Authors: Mazure N. , Truffaut N.
Published: February 2010

When Mycobacterium aurum MO1 was grown with morpholine, the release of ammonia into the supernatant was proportional to the disappearance of morpholine, showing that this compound was mineralized. MO1 was able to grow in high concentrations of morpholine but accumulation of ammonia inhibited growth and degradation of morpholine. Immobilization of bacterial cells in carrageenan gel beads showed that morpholine degradation in these conditions began earlier and was faster than in free culture. One of the two branches of the lower pathway of morpholine biodegradation was induced while the other branch was inhibited in the presence of morpholine. Strain MO1 grew on heterocyclic compounds similar to morpholine, demonstrating that MO1 is able to degrade heterocyclic compounds containing nitrogen atoms (piperidine and pyrrolidine). Compounds containing sulphur or oxygen atoms or compounds with double bonds were not degraded.Key words: Mycobacterium aurum, morpholine, biodegradation.
Isolation and characterization of conditional lethal mutants of Beauveria bassiana
Authors: Hegedus Dwayne D. , Khachatourians George G.
Published: February 2010

Temperature-sensitive mutants of Beauveria bassiana GK2016 were isolated and characterized. Heat-sensitive mutants that grew at 20 °C but not at 30 °C were generated using mutagenesis with ultraviolet light and several rounds of selection. After 2160 colonies from a heat-sensitive mutant enriched population were screened, 11 heat-sensitive strains were isolated for further study. Five mutant strains, HS1, HS2, HS6, HS9, and HS11, were stable and closely resembled the wild-type strain with respect to morphology, growth rate, and enzyme synthesis at 20 °C. Characterization of macromolecular synthesis at 30 °C using a radiolabelled precursor uptake assay indicated that three mutants, HS6, HS9, and HS11, had reduced levels of DNA, RNA, and protein synthesis at the nonpermissive temperature. Mutants affected in cell division and cell wall synthesis were characterized by microscopy. At 30 °C, mutant HS6 was defective in cellular compartmentalization and formed long, branched, aseptate mycelia that fragmented easily. Mutant HS1 was defective in cell wall biosynthesis and at 30 °C ceased to maintain cell wall integrity and lysed. Mutants HS2 and HS9 possessed temperature-sensitive lesions that could not be specified. None of the mutants were directly affected in either nuclear division or mitosis as evidenced by the accumulation of Giemsa-stainable mitotic nuclei at 30 °C. Bioassays conducted with grasshoppers showed the efficacy of the mutant strains, with the exception of HS11, to be comparable to that of the parent strain, at 20 °C. Conversely, at 32 °C the mutant strains were uninfective whereas the parental strain GK2016 was infective. It is our intention to use these strains to examine fundamental aspects of entomopathology by dissecting fungal growth and development in vitro and extending these observations to pathogenesis in insects.Key words: Beauveria bassiana, temperature-sensitive, mutants, characterization, infection.
Molecular cloning and sequencing of the capsid and the nuclear inclusion protein genes of a North American PVYN isolate
Authors: Dhar A. K. , Singh R. P. , Boucher A.
Published: February 2010

The sequence of the 3′-terminal 3258 nucleotides of a tobacco veinal necrosis strain of a potato virus Y (PVYN) isolate from North America was determined. The sequence revealed an open reading frame of 2931 nucleotides, of which the start codon was not identified. The nontranslated region contains 327 nucleotides upstream of a poly(A) tract. The open reading frame encodes a large polyprotein containing 976 amino acids. The data indicate that the coat protein (CP) and the nuclear inclusion protein (NIb) are derived from the large polypeptide by proteolytic cleavage similar to many other potyviruses. The putative cleavage sites of NIa/NIb and NIb/CP correspond to Q/A and Q/G sequences, respectively, as in other PVYN isolates. The CP and NIb contain 267 and 519 amino acid residues, respectively. Higher sequence homology of CP was observed with other PVYN isolates than with the PVYO isolates.Key words: potato virus Y, nucleotide sequence.
Purification and characterization of a protease from Pseudomonas pseudomallei
Authors: Sexton M. M. , Jones A. L. , Chaowagul W. , Woods D. E.
Published: February 2010

Pseudomonas pseudomallei is the causative agent of melioidosis, a glanders-like disease of humans and animals. The pathogenesis of melioidosis is not well understood, and the role of various extracellular enzymes produced by P. pseudomallei in the development of this disease is not known. The present studies were designed to purify and characterize an extracellular protease produced by P. pseudomallei isolates and to test the hypothesis that this protease may play a role in melioidosis. The protease was present in culture supernatants as an enzyme with a molecular weight of 36 000 that was optimally active at 60 °C and at pH 8.0. The P. pseudomallei protease was shown to be a metalloenzyme requiring iron for maximal activity, and activity was inhibited in the presence of ethylenediaminetetraacetic acid (150 mM). Antibodies directed against an alkaline protease produced by Pseudomonas aeruginosa cross-reacted with the P. pseudomallei protease. These data indicate that the P. pseudomallei protease belongs to the family of alkaline proteases sensitive to metal chelators. Purified P. pseudomallei protease was capable of digesting a variety of eucaryotic protein substrates including immunoglobulins. A P. pseudomallei strain deficient in protease production was shown to be less virulent than the parental strain in an animal model of lung infection. These data suggest that this protease may be a significant pathogenic determinant in infections caused by P. pseudomallei.Key words: protein, pathogenesis, substrates, disease.
1-Aminocyclopropane-1-carboxylic acid deaminase mutants of the plant growth promoting rhizobacterium Pseudomonas putida GR12-2 do not stimulate canola root elongation
Authors: Glick Bernard R. , Jacobson Christian B. , Schwarze Melinda M. K. , Pasternak J. J.
Published: February 2010

The plant growth promoting rhizobacterium Pseudomonas putida GR12-2 was mutagenized with nitrosoguanidine and three separate mutants that were unable to utilize 1-aminocyclopropane-1-carboxylic acid (ACC) as a sole nitrogen source were selected. These mutants are devoid of the ACC deaminase activity that is present in wild-type P. putida GR12-2 cells. Only wild-type cells, but not any of the ACC deaminase mutants, promoted root elongation of developing canola seedlings under gnotobiotic conditions. These results are interpreted in terms of a model in which P. putida GR12-2 promotes root elongation by binding to germinating seeds and sequesters and hydrolyzes some of the unbound ACC, thereby lowering the level of ACC and hence the endogenous ethylene concentration, allowing the roots to grow longer.Key words: 1-aminocyclopropane-1-carboxylate, ACC, plant growth promoting rhizobacteria, PGPR, ACC deaminase, bacterial fertilizer.
Cysteine protease activity of streptococcal pyrogenic exotoxin B
Authors: Ohara-Nemoto Yuko , Sasaki Minoru , Kaneko Masaru , Nemoto Takayuki , Ota Minoru
Published: February 2010

Streptococcal pyrogenic exotoxin B (SPE B) was purified and its protease and mitogenic activities were investigated. The apparent molecular mass of SPE B purified in the presence of iodoacetic acid was 42 kDa, whereas 29-kDa SPE B was predominant without the reagent. A polyclonal antibody raised against the 29-kDa species reacted with both species, indicating that the 42-kDa species was a precursor of the 29-kDa entity. Both the 42- and 29-kDa species enhanced [3H]thymidine incorporation into human peripheral blood mononuclear cells, whereas neither had any effect on T cell depleted mononuclear cells. The 29-kDa SPE B possessed caseinolytic activity, with an optimal pH of 8, and the activity was specifically suppressed by the antibody. A group of cysteine protease inhibitors, but no serine-, metallo-, or acidic-protease inhibitors, limited the protease activity, whereas dithiothreitol increased the activity. The DNA sequence around a putative active cysteine residue was identical among the speB genes from Streptococcus pyogenes R70, NY-5, and T 19. Taken together, these results indicate that SPE B is identical to a cysteine protease, streptopain (EC 3.4.22.10).Key words: streptococcal pyrogenic exotoxin B, streptopain, Streptococcus pyogenes, cysteine protease.
Analysis of bacterial populations in a basalt aquifer
Authors: Zheng M. , Kellogg S. T.
Published: February 2010

Basalt aquifers are common in many parts of Canada and the United States; however, little is known about their microbial ecology. Microbial populations were characterized at a basalt aquifer with 7 m of clay and silt in the vadose zone and a saturated zone extending from 7 to 500 m. Planktonic total counts and viable counts were 1.7 × 105 and 2.3 × 104 cells/mL, while sessile total counts and viable counts were 1.9 × 107 and 1.7 × 105 cells/g, respectively. Some of the physical and chemical habitat conditions measured were pH (7.1), Eh (135 mV), dissolved O2 (4.1 mg/L), temperature (10 °C), NH4+ (4 μM), NO3− (253 μM), Fe (0.17 μM), and Mn (36 nM). Variation in physical and chemical variables correlated with depth and the region of the aquifer sampled. One hundred forty-nine random isolates of the dominant culturable bacteria from groundwater and subsurface cores were submitted to phenotypic analysis on the basis of antibiotic responses. The resultant dominant bacteria were Gram-negative, mesophilic heterotrophs. Cluster analysis revealed at least 14 discernible clusters containing representatives from the genera Pseudomonas, Bacillus, Acinetobacter, Arthrobacter, Micrococcus, and Clavibacter. Centrotype representatives were selected from each cluster and tentatively identified.Key words: basalt aquifer bacterial ecology.
The transfer of Wolbachia pipientis and reproductive incompatibility between infected and uninfected strains of the flour beetle, Tribolium confusum, by microinjection
Authors: Chang Nancy W. , Wade Michael J.
Published: February 2010

We report a protocol for transferring the cytoplasmic endosymbiont Wolbachia pipientis from infected to uninfected hosts in the flour beetle, Tribolium confusum, by microinjection of cytoplasm from infected donor eggs into uninfected host eggs. Postinjection survivorship was limited by the halocarbon oil, which was lethal to hatching larvae but necessary to stem the backflow of cytoplasm out of injected eggs. In test crosses, approximately 39% of surviving and maturing larvae exhibited the complete reproductive incompatibility typically associated with W. pipientis in this species. However, some transfected females failed to transmit the W. pipientis to their progeny and the incompatibility could be lost in the generation following transfection. Conversely, in some cases, females that did not themselves exhibit the incompatibility had some offspring that did exhibit partial incompatibilities.Key words: microinjection, Wolbachia, flour beetle, Tribolium confusum.
Frequency of genes in aromatic and aliphatic hydrocarbon biodegradation pathways within bacterial populations from Alaskan sediments
Authors: Sotsky J. B. , Greer C. W. , Atlas R. M.
Published: February 2010

A significant proportion of the naturally occurring hydrocarbon-degrading populations within Alaskan sediments affected by the Exxon Valdez oil spill had both the xylE and alkB genes and could convert hexadecane and naphthalene to carbon dioxide; a greater proportion of the population had xylE than had alkB, reflecting the composition of the residual oil at the time of sampling; nearly equal populations with xylE alone, alkB alone, and xylE + alkB genes together were found after exposure to fresh crude oil; populations with xylE lacking alkB increased after enrichment on naphthalene. Thus, the genotypes of hydrocarbon-degrading populations reflected the composition of the hydrocarbons to which they were exposed.Key words: hydrocarbon biodegradation, aromatic hydrocarbon biodegradation, aliphatic hydrocarbon biodegradation, alkB, xylE.
Degradation of methyl parathion by Pseudomonas putida
Authors: Rani N. Lakshmi , Lalithakumari D.
Published: February 2010

Pseudomonas putida utilized methyl parathion as sole carbon and (or) phosphorus source. The bacterium elaborated the enzyme organophosphorus acid anhydrase, which hydrolyzed methyl parathion to p-nitrophenol. p-Nitrophenol was further degraded to hydroquinone and 1, 2, 4-benzenetriol. The final ring compound, 1, 2, 4-benzenetriol, was cleaved by benzenetriol oxygenase to maleyl acetate.Key words: Pseudomonas putida, organophosphorus pesticides, degradation, methyl parathion.
Cellular effects of misscheduled brlA, abaA, and wetA expression in Aspergillus nidulans
Authors: Sewall Tommy C.
Published: February 2010

Asexual sporulation in the fungus Aspergillus nidulans is controlled, in part, by a central regulatory pathway composed of the brlA, abaA, and wetA genes. The coding region of each of these genes was fused, in frame, to the threonine-inducible alcohol dehydrogenase promoter then stably incorporated into the A. nidulans genome at the argB locus. Misscheduled expression of each of these developmental genes interfered with normal growth and sporulation. Expression of brlA or abaA terminated vegetative growth and led to anomalous deposition of wall material at the hyphal apex and septa. brlA alone induced the formation of phialide-like structures and the production of conidia.Key words: Aspergillus nidulans, conidiogenesis, sporulation, gene regulation.
Demonstration of the indolepyruvate decarboxylase gene homologue in different auxin-producing species of the Enterobacteriaceae
Authors: Zimmer Wolfgang , Hundeshagen Barbara , Niederau Edith
Published: February 2010

Different Enterobacteriaceae were assayed for their ability to produce the plant hormone indole-3-acetate with the aim to study the distribution of the indole-3-pyruvate pathway, which is known to be involved in the production of indole-3-acetate in a root-associated Enterobacter cloacae strain. Other E. cloacae strains, and also Enterobacter agglomerans strains, Pantoea agglomerans, Klebsiella aerogenes, and Klebsiella oxytoca were found to convert tryptophan into indole-3-acetate. As it was also intended to identify the conserved regions of the indole-3-pyruvate decarboxylase, which is involved in producing indole-3-acetate in the E. cloacae strain, oligonucleotide primers were synthesized for different regions of the corresponding gene. One pair of these primers allowed us to amplify a segment of the predicted size by the polymerase chain reaction with DNA of the seven different Enterobacteriaceae that produce indole-3-acetate. Segments of five strains were cloned and sequenced. All sequences showed significant homology to the indole-3-pyruvate decarboxylase gene. As in addition a positive DNA–DNA hybridization signal was detected in the seven strains using the E. cloacae or E. agglomerans segments as a probe, indole-3-acetate biosynthesis is suggested to be catalyzed via the indole-3-pyruvate pathway not only in E. cloacae but also in the other soil-living Enterobacteriaceae. Conserved regions were detected in the indole-3-decarboxylase by alignment of the now-available five different partial sequences. These regions should enable identification of the gene in other bacterial families or even in plants.Key words: indole-3-pyruvate decarboxylase, indole-3-acetic acid production, auxin, polymerase chain reaction, Enterobacteriaceae.
Purification and characterization of a β-glucosidase from solid-state cultures ofHumicola griseavar.thermoidea
Authors: Filho Edivaldo Ximenes Ferreira
Published: February 2010

The thermophilic fungus Humicola grisea var. thermoidea produced β-glucosidase activity when grown in a solid-state culture on wheat bran as carbon source. A β-glucosidase was purified to apparent homogeneity by ultrafiltration, gel filtration chromatography on Sephacryl S-100, and ion-exchange chromatography on S-Sepharose, as judged by sodium dodecyl sulfate – polyacrylamide gel electrophoresis (SDS–PAGE) on a 12.5% (w/v) slab gel. The enzyme had a molecular mass of 82 and 156 kDa, as estimated by SDS–PAGE and gel filtration on a high performance liquid chromatographic column, respectively, suggesting that the native enzyme may consist of two identical subunits. The purified enzyme was thermostable at 60 °C for 1 h with a half-life of 15 min at 65 °C, and displayed optimum activity at 60 °C and a pH range of 4.0–4.5. The Kmand Vmaxvalues for p-nitrophenyl β-D-glucopyranoside were determined to be 0.316 mM and 0.459 IU∙mL−1, respectively. D-Glucose, D-gluconic acid lactone, Hg2+, Cu2+, and Mn2+inhibited β-glucosidase activity. The enzyme activity was competitively inhibited by D-glucose (Ki = 0.6 mM). The purified enzyme was very active against cellobiose and p-nitrophenyl β-D-glucopyranoside.Key words: Humicola, β-glucosidase, purification, characterization.
Galactose induces inSaccharomyces cerevisiaesensitivity of the utilization of hexoses to inhibition byD-glucosamine
Authors: Nevado Julián , Heredia Claudio F.
Published: February 2010

Inhibition by glucosamine of the utilization of hexoses by Saccharomyces cerevisiae is induced by growing the cells in media with galactose as carbon source. The intensity of inhibition parallels the induction of the galactose pathway. These findings contrast with the fact that glucosamine is a substrate of the constitutive glucose but not of the inducible galactose transport and phosphorylation systems. The inhibition by glucosamine is pH dependent; the extent seems to be related with the phosphorylation of the hexosamine, as shown by its greater effect with substrates or with conditions that less interfere with the phosphorylation of the inhibitor. Inhibition is not a consequence of ATP depletion of the cell. Intracellular accumulated glucosamine derivatives impair the transport of glucose and mannose in yeast cells grown in galactose-supplemented media but not those grown with glucose or ethanol supplements (i.e., under conditions in which the utilization of these sugars is inhibited). However, impairment of the transport is not enough to explain the characteristics of the observed inhibition. The changes induced by growing the yeast in galactose that render the cells sensitive to glucosamine are under the control of the gal80 and gal4 genes.Key words: yeast, glycolysis inhibition, glucosamine, Saccharomyces cerevisiae.
Influence of disease-suppressive strains of Streptomyces on the native Streptomyces community in soil as determined by the analysis of cellular fatty acids
Authors: Bowers John H. , Kinkel Linda L. , Jones Roger K.
Published: February 2010

Analysis of cellular fatty acid profiles was used to distinguish among introduced pathogen- suppressive strains and indigenous strains of Streptomyces spp. isolated from soil of field plots established to test the efficacy of Streptomyces strains PonSSII and PonR in the biological control of potato scab. Reference libraries of fatty acid profiles were developed for a collection of known pathogenic strains and the introduced suppressive strains. Population densities of pathogen-related, suppressive, and saprophytic Streptomyces strains were determined from the relationship of field isolates to mean library profiles using cluster analysis and the unweighted pair-group method using arithmetic averages. Community diversity was similarly determined. Streptomyces strains PonSSII and PonR were distinguished from each other and from the pathogen group (which clustered together) based on fatty acid profiles. The introduced, suppressive strains successfully colonized the soil and represented 2–19% of the isolates sampled over 2 years. The introduction of the suppressive strains inhibited the population of strains related to the pathogen library at each sample date; the pathogen population was substantially lower in soil from treatments where the suppressive strains were introduced compared with the nonamended control. At harvest, the pathogen-related population was suppressed 85–93 and 36–44% in 1991 and 1992, respectively, in treatments with the suppressive strains compared with the nonamended control. Diversity of the community was not affected by the introduced strains, and diversity and equitability indices were similar among treatments at any sample time. The inhibition of the pathogen-related population was correlated with a reduction of scab symptoms observed in the field plots into which the suppressive strains were introduced. Implications of a fundamental shift in the pathogen-related population in response to the introduction of the suppressive strains for long-term biological control of potato scab are encouraging.Key words: Streptomyces, fatty acid analysis, biological control, community ecology.
Influence of peracetic acid onEscherichia coliH10407 strain in laboratory microcosms
Authors: Jolivet-Gougeon A. , Braux A. S. , Sauvager F. , Arturo-Schaan M. , Cormier M.
Published: February 2010

The bactericidal properties of peracetic acid (PAA) were tested using Escherichia coli H10407, in sterilized artificial seawater, sewage effluent water, and distilled water microcosms. No LT enterotoxin synthesis was detected by GM1 enzyme-linked immunosorbent assay of the water supernatants, but a specific fragment of the eltB gene was always amplified by polymerase chain reaction for 21 days after PAA treatment. The resuscitation capacity of starved cells was assayed in rich medium and their inability to overcome the effects of PAA stress was observed, despite the emergence of viable but nonculturable cells in microcosms 24 or 48 h after treatment. Moreover, no obvious differences in response were obtained, concerning enterotoxigenesis, between bacteria subjected to osmotic and (or) nutrient starvation-induced stress with or without PAA treatment.Key words: peracetic acid, disinfection, enterotoxigenic Escherichia coli.
Changes in glutathione metabolic enzymes during yeast-to-mycelium conversion of Candida albicans
Authors: Manavathu Merlin , Manavathu Elias , Gunasekaran Suresh , Porte Quallyna , Gunasekaran Muthukumaran
Published: February 2010

Candida albicans is a dimorphic yeast capable of producing alternate morphological forms (yeast or mycelium) in response to environmental changes. The intracellular level of glutathione, which helps to maintain the redox potential of the cell, is decreased significantly during the thermal induction of yeast-to-mycelium conversion. The reason for the decline of glutathione in the mycelial form is not understood. We have, therefore, investigated the levels of glutathione reductase, glutathione S-transferase, γ-glutamyltranspeptidase, and glutathione peroxidase, four key enzymes involved in glutathione metabolism, in the yeast and mycelial forms. Yeast cells of C. albicans 3153A were induced in Lee's medium (pH 6.5) at 37 °C for 3 h to produce germ tubes. Cell lysates were prepared from yeast and mycelial cells, and glutathione reductase, glutathione S-transferase, γ-glutamyltranspeptidase, and glutathione peroxidase were assayed spectrophotometrically. There was a 640% increase of the level of γ-glutamyltranspeptidase in the germ tubes as compared with the yeast cells. No other significant alteration of the levels of enzymes was noted. This increased activity of γ-glutamyltranspeptidase, which cleaves the glutamic acid residue of glutathione (Glu-Cys-Gly) appears to be, at least in part, responsible for the rapid decrease of the intracellular glutathione in C. albicans during the yeast-to-mycelium conversion.Key words: Candida albicans, dimorphism, glutathione, glutathione reductase, glutathione peroxidase, γ-glutamyltranspeptidase.
Ultrastructure and seasonal growth patterns of microbial mats in a temperate climate saline-alkaline lake: Goodenough Lake, British Columbia, Canada
Authors: Schultze-Lam S. , Ferris F. G. , Sherwood-Lollar B. , Gerits J. P.
Published: February 2010

Goodenough Lake is a small (ca. 1 by 0.6 km) saline-alkaline lake on the Caribou Plateau in British Columbia, Canada, which has a mean depth of < 1 m, allowing the growth of substantial prokaryote-dominated benthic microbial mats over the entire lake bottom. Microbial mats were investigated on both a detailed ultrastructural level and at a macroscopic level to determine, on a lakewide scale, the seasonally influenced changes that occurred over the course of a spring to fall growth season. The acceleration of phototrophic activity with warming of the lake water in spring could be followed by increases in dissolved oxygen levels, which quickly reached supersaturation early in the summer. The grazing activity of brine fly larvae (Ephydra hians) was found to have a significant influence on mat growth and development patterns. On an ultrastructural level, several types of filamentous cyanobacteria were the main phototrophic organisms present, even in anaerobic parts of the mats, where they were accompanied by purple bacteria. The dominant cyanobacterium present was a large oscillatoriacean form which, in certain parts of the lake, formed unusual columnar structures that had a very low diversity of microbial species in comparison to mats from other regions of the lake. The microbial community also differed in areas of the lake where less saline groundwater springs emerged from the lake bed.Key words: microbial mats, saline-alkaline lakes, cyanobacteria, ultrastructure.
The repB gene required for production of extracellular enzymes and fluorescent siderophores in Pseudomonas viridiflava is an analog of the gacA gene of Pseudomonas syringae
Authors: Liao C.-H. , McCallus D. E. , Wells J. M. , Tzean S.-S. , Kang G.-Y.
Published: February 2010

Two genes, designated repA and repB, are involved in the regulation of the synthesis of extracellular pectate lyase, protease, and alginate in Pseudomonas viridiflava. The repA gene has been shown to encode a protein highly homologous to several bacterial sensors in the two-component regulator family including the LemA of Pseudomonas syringae. In this study, the repB locus, initially identified in a 6.3-kb EcoRI genomic fragment of P. viridiflava, was further characterized. Results obtained from restriction mapping, deletion subclonings, and mini-Mu-LacZ fusions indicated that the repB gene was contained within a 0.8-kb HindIII–PstI region. Sequence analysis of this repB region revealed the presence of an open reading frame, which was predicted to encode a protein similar or identical to the gacA response regulator found in P. syringae and Pseudomonas fluorescens. The repB gene of P. viridiflava also regulated the production of fluorescent siderophores, in addition to the aforementioned extracellular enzymes and alginate. The repB or gacA homologs were detected in the genomes of nine other strains of P. viridiflava, P. fluorescens, and P. syringae included in the study. The data presented here and earlier indicate that the repA/repB gene regulatory system of P. viridiflava is analogous to the lemA/gacA system of P. syringae and P. fluorescens.Key words: response regulator, signal transduction, soft-rot bacteria, enzyme production.
Heat sterilization of archaeal liposomes
Authors: Choquet Christian G. , Patel Girishchandra B. , Sprott G. Dennis
Published: February 2010

Thermal stabilities were compared between liposomes prepared from the ether lipids extracted from various archaeobacteria and liposomes composed of ester lipids. Leakage of entrapped carboxyfluorescein from the liposomes exposed to 121 °C indicated a marked stability of certain ether liposomes, comparable or superior to cholesterol-stabilized liposomes prepared from the saturated synthetic lipids dimyristoyl phosphatidylcholine and dimyristoyl phosphatidylglycerol. The heat stability of diether liposomes could be increased by the inclusion of tetraether lipids.Key words: archaeal liposomes, ether liposomes, heat sterilization, thermal stability, archaeobacteria ("Archaea").
Antifungal properties of 3-methyl-5-aminoisoxazole-4-thiocyanate on four yeasts: an unusual dose-dependent effect on Rhodotorula glutinis Harrison
Authors: Mares D. , Romagnoli C. , Donini A. , Bruni A.
Published: February 2010

The antifungal activity of 3-methyl-5-aminoisoxazole-4-thiocyanate was studied on Candida albicans, Saccharomyces cerevisiae, Schizosaccharomyces pombe, and Rhodotorula glutinis. A significant inhibition of growth was evident in all yeasts. The ultrastructural modifications have demonstrated that the compound at half-minimum inhibitory concentration (MIC) dose (1.25 μg∙mL−1) causes the transformation of R. glutinis from yeast to mycelium form by the formation of a germ tube. The accumulation of abnormal amounts of wall substances after treatment with the MIC is discussed in relation to ergosterol and chitin production.Key words: 3-methyl-5-aminoisoxazole-4-thiocyanate, Rhodotorula, Candida, Saccharomyces, Schizosaccharomyces.
Restriction analysis of actinomycetes chromosomal DNA
Authors: Novella Isabel S. , Sánchez Jesus , Marín Irma
Published: February 2010

Actinomycetes DNAs were digested with restriction enzymes to study the presence of methylated bases. Analysis showed that the enterobacterial Dam and Dcm systems are absent. Methylation at the internal cytosine in CCGG sequences, typical of eukaryotes, was also absent. We also tested 18 restriction endonucleases recognizing six base pair sequences (all of which were inhibited by methylation). Results showed a higher number of restriction sites for enzymes recognizing CG-rich sequences (CG endonucleases) than for enzymes recognizing AT-rich sequences (AT endonucleases). Restriction patterns with CG endonucleases were quite uniform, with the remarkable exception of XhoI, which yielded a small number of DNA bands. The study performed with AT endonucleases allowed differentiation of three groups of enzymes based on different degrees of chromosomal sensitivity. One group (BelI and BglII) produced restriction patterns with more abundant restriction sites than expected, a second group (ClaI, EcoRI, and EcoKV) yielded the predicted number of DNA fragments, and the third group (HpaI, HindIII, XbaI, and DraI) produced an unexpectedly low number of fragments. Some individual cases of resistance to particular enzymes could be explained by the presence of restriction-modification systems with the same specificity.Key words: Streptomyces, DNA methylation, restriction modification, sequence counterselection, pulsed-field gel electrophoresis.
Physicochemical properties of PM-factor, a surface-active agent produced byPseudomonas marginalis
Authors: Burd G. , Ward O. P.
Published: February 2010

An extracellular surface-active agent, PM-factor, was obtained by high-speed centrifugation from the culture broth of Pseudomonas marginalis PD-14B. PM-factor exhibited emulsifying activity on a broad spectrum of hydrocarbon liquids, including aromatics, aliphatics, crude oil, and creosote. The factor appeared as ball-shaped particles of varying diameter when examined by electron microscopy (0.16–1.4 μm). Gel filtration chromatography demonstrated a high molecular mass of the factor (> 106 Da). The ultraviolet absorption spectrum manifested a peak in the region 200 nm rather in the region 260–280 nm. Amino acid analysis showed a very low amount of aromatic amino acids residues in the protein moiety of PM-factor. The presence of 3-deoxy-D-mannooctulosonic acid, heptose, hexosamine, phosphorus, and 3-hydroxy fatty acid indicated that PM-factor contained lipopolysaccharide. The emulsifying activity of PM-factor was inhibited strongly by mercuric chloride and moderately by EDTA. Polymyxin B, Ca2+, and Mg2+markedly stimulated the factors emulsifying activity. Roles of the bioemulsifier in the functioning of P. marginalis as a plant pathogen and in bioremediation are discussed.Key words: bioemulsifier, Pseudomonas marginalis, polycyclic aromatic hydrocarbons, plant pathogenesis, bioremediation.
Chitinases of Bacillus licheniformis B-6839: isolation and properties
Authors: Trachuk Lesya A. , Revina Lyudmila P. , Shemyakina Tatyana M. , Chestukhina Galina G. , Stepanov Valentin M.
Published: February 2010

Five chitinases were isolated from culture filtrates of Bacillus licheniformis B-6839 R and S variants by combination of hydrophobic, ion-exchange, and gel permeation chromatography. The enzymes had molecular masses of 66, 62, 53, 49, and 42 kDa. The chitinases revealed two activity optima against colloidal chitin at pH 4.5–5.5 and 9.0–9.5 and they were rather stable at pH 4.0–9.5. The temperature optimum of activity was 90 °C for the 62-kDa chitinase and 70 °C for the other enzymes. The 66-, 53-, and 42-kDa chitinases showed pronounced similarities in their N-terminal sequences and apparently belonged to the same group, which might be related to Bacillus circulans chitinase A1. The 49- and 62-kDa enzymes did not reveal structural similarities with other chitinases produced by the studied B. licheniformis strain. No relationship was found with the 89- and 76-kDa chitinases isolated earlier from B. licheniformis X-7u.Key words: Bacillus licheniformis, chitinase, multiplicity.