bluuebunny/crossref_metadata_2025 huggingface dataset:
Growth of group B streptococci in human serum leads to increased cell surface sialic acid and decreased activation of the alternative complement pathway
Authors: Platt Mark W. , Correa Jr. Norberto , Mold Carolyn
Published: February 2010

Group B streptococcus type III is a major cause of neonatal death. The terminal sialic acid moiety of the group B streptococcus type specific capsule has been shown to be an important virulence factor. We demonstrate here that bacteria grown in human serum have increased cell surface sialic acid content compared with cells grown in common laboratory media. This sialic acid was removed by incubation with neuraminidase, showing that it was on the bacterial surface. Serum-dependent sialylation was dependent on metabolic activity, as the addition of chloramphenicol reduced the amount of added sialic acid by more than 90%. Probing the cell surface with an antibody specific for group B streptococcus type III capsular sialic acid showed an increase in antibody binding after growth in human serum. This effect could be lowered by incubating serum-grown cells in neuraminidase prior to antibody exposure. A group B streptococcus mutant that when grown in laboratory media lacks cell surface sialic acid showed significant cell surface sialic acid when grown in human serum. This increase was associated with a significantly decreased ability to bind C3 and hence activate the alternative complement pathway.Key words: group B streptococcus, capsule, human serum.
Identification of Clavibacter michiganensis subsp. sepedonicus using the polymerase chain reaction
Authors: Firrao Giuseppe , Locci Romano
Published: February 2010

From the sequence of a subcloned DNA fragment of a highly conserved plasmid of Clavibacter michiganensis subsp. sepedonicus a pair of oligonucleotides were devised for use as polymerase chain reaction primers. The primer sequences do not show significant homology with any other sequence deposited in public databases. Polymerase chain reactions carried out using this primer pair and untreated cells of all strains of C. michiganensis sepedonicus tested resulted in the amplification of a DNA fragment of about 670 base pairs. No amplification was observed when bacteria belonging to other species were submitted to polymerase chain reaction under the same conditions. The detection limit of the assay was 4 × 103 bacteria.Key words: detection, potato ring rot, Corynebacterium sepedonicum.
Characterization and identification of Pseudomonas strain WRF
Authors: Fulghum Robert S.
Published: February 2010

Phenotypic characterization of Pseudomonas strain WRF shows it to be a member of Pseudomonas putida most closely resembling biovar A. The organism has been deposited in the American Type Culture Collection as ATCC 35546.Key words: Pseudomonas WRF characterization, Pseudomonas putida, one-carbon metabolism, siderophores, peptide-bound flavin enzymes.
A proposed helical mechanism for gliding motility in three gliding bacteria (order Cytophagales)
Authors: Beatson Peter J. , Marshall Kevin C.
Published: February 2010

Observations of a typical Cytophaga sp. and two unclassified gliding bacteria, using phase-contrast microscopy and videotaping techniques, revealed that the bacteria rotated sinistrally at strain-specific rates as they glided. Microscopic spheres attached to the bacteria moved in helical paths around them. The machinery of gliding is conjectured, therefore, to be orientated helically on the cell surface, and probably acts on a substratum via an adhesive polymer network. Cells and surfaces often appear to be connected at some distance by an invisible elastic material, which might be equated to fibrous structures observed extending from cells. Variations in motility behaviour between strains could be explained by differences in the orientation of moving cell surface elements, in the amount and type of adhesive polymer surrounding cells, and in the distance at which attached bacteria lie from their substrata.Key words: Cytophaga, gliding motility, mechanism of gliding, adhesion.
Microbial conversion of arabinose and xylose to hydrogen by a newly isolated Clostridium sp. No. 2.
Authors: Taguchi Fumiaki , Mizukami Naoki , Hasegawa Katsushige , Saito-Taki Tatsuo
Published: February 2010

For 24 h at pH 6.0 and 36 °C, Clostridium sp. No. 2, isolated from termites, converted arabinose and xylose to hydrogen with yields of 14.55 and 13.73 mmol/g of substrate consumed, respectively. These yields were greater than the maximum value of 11.07 mmol/g of glucose consumed in 1 L of batch culture with 10 g of each substrate. The organism produced hydrogen from 45 g of glucose and xylose in 1 L of culture with a total accumulation of 380.6 mmol from glucose and 479.0 mmol from xylose, at a conversion rate of 270.9 mmol/g of dry cells from glucose and 405.9 mmol/g of dry cells from xylose. Hydrogen was produced at a maximum evolution rate of 27.2 mmol/h after 8 h of incubation with glucose and of 28.6 mmol/h after 6 h of incubation with xylose.Key words: hydrogen production, xylose, arabinose, Clostridium sp. No. 2.
Dependence of H+ exchange and oxygen evolution on K+ in the marine cyanobacterium Synechococcus sp. strain UTEX 2380
Authors: Spiller Hart , Stallings Jr. William , Tu Chingkuang , Gunasekaran Muthukumaran
Published: February 2010

Light-induced net H+ efflux, photosynthetic oxygen evolution, and medium alkalization during the steady state of photosynthesis were specifically stimulated by K+ from pH 6 through 8.5 in the marine cyanobacterium Synechococcus sp. strain UTEX 2380. Net proton efflux in the light was completely abolished by the uncoupler carbonylcyanide m-chlorophenylhydrazone and the membrane ATPase inhibitor diethylstilbestrol or partially abolished by the membrane ATPase inhibitor N,N′-dicyclohexyl-carbodiimide and by orthovanadate. H+ extrusion in the light was accompanied by K+ uptake at rates of 30 μmol∙mg chlorophyll−1∙h−1. During the steady state of CO2 fixation, potassium was excreted simultaneously with medium alkalization. The K+ content of the cells was 432 mM for air-grown cells. K+ in the cells was displaced by diethanolamine, which was inhibited by m-chlorophenylhydrazone. Na+-loaded cells showed nearly complete inactivation of oxygen evolution and medium alkalization. Both activities were reactivated by the addition of K+ or Rb+. A fivefold increase in inorganic carbon uptake in the light was observed in the presence of K+. The pH of the cytoplasm in the light increased from 7.2 to 8.04 in the pH range 6.6–8.6. These results suggest that a light-dependent, proton-excreting ATPase is active in conjunction with a K+ uptake system in a marine Synechococcus species, while a K+–H+ antiporter may function as a regulator of cytoplasmic pH during photosynthesis.Key words: potassium transport, oxygen, cyanobacterium, Synechococcus, proton extrusion.
Effect of condensed tannins from birdsfoot trefoil on endoglucanase activity and the digestion of cellulose filter paper by ruminal fungi
Authors: McAllister T. A. , Bae H. D. , Yanke L. J. , Cheng K.-J. , Muir A.
Published: February 2010

The ruminal fungi Neocallimastix frontalis RE1, Neocallimastix patriciarum 27, Piromyces communis 22, and Orpinomyces joyonii 19-2 were examined for their ability to digest filter paper in the presence of condensed tannins from birdsfoot trefoil (Lotus corniculatus L.). For all four fungi, inhibition of endoglucanases was evident at 100 μg condensed tannins∙mL−1 with nearly complete inhibition at 300 μg condensed tannins∙mL−1. At 100 and 200 μg condensed tannins∙mL−1, the endoglucanase activity of N. frontalis RE1 was greater (P < 0.01) than that of the other three fungal species. Exposure to 100 μg condensed tannins∙mL−1 did not affect the ability of N. frontalis RE1 or N. patriciarum 27 to digest filter paper, and although digestion was reduced, N. frontalis RE1 and P. communis 22 solubilized more than 20% of the filter paper at 500 μg condensed tannins∙mL−1. In contrast, O. joyonii 19-2 was virtually unable to digest filter paper at 300 μg condensed tannins∙mL−1. Mycelia of fungi grown with condensed tannins were covered by filamentous material, which may have arisen from the formation of condensed tannin–protein complexes. Less than 86% of the condensed tannins (as measured by the H2SO4 method) were recovered after 120 h of incubation with N. frontalis RE1, P. communis 22, and N. patriciarum 27. The need for detailed studies to examine the ability of ruminal fungi to metabolize condensed tannins is evident.Key words: ruminal fungi, condensed tannins, cellulose digestion, ruminant.
Coordinate derepression of alkaline phosphatase, and binding protein dependent transport systems for phosphate and sn-glycerol-3-phosphate by phosphate starvation in Erwinia carotovora subsp. carotovora
Authors: Schweizer Herbert P.
Published: February 2010

In Erwinia carotovora subsp. carotovora EC14 grown under conditions of phosphate starvation, alkaline phosphatase, and binding protein dependent transport systems for inorganic phosphate andsn-glycerol-3-phosphate were simultaneously derepressed. Analysis of periplasmic protein profiles from phosphate-starved cells revealed four starvation-inducible proteins. Three of these proteins corresponded to alkaline phosphatase, a binding protein for phosphate, and a binding protein for.sn-glycerol-3-phosphate, respectively. The coordinate derepression of these proteins demonstrates the existence of a phosphate regulon similar to that found in other members of the family Enterobacteriaceae.Key words: Erwinia, phosphate, regulon, alkaline phosphatase, binding protein.
Short-term recovery of Bradyrhizobium japonicum during an inoculation process using mineral microgranules
Authors: Fouilleux Georges , Revellin Cécile , Catroux Gérard
Published: February 2010

Soybean (Glycine max (L.) Merr.) can be inoculated by delivering inoculant mixed with mineral microgranules to the seedbed. An average decline of 0.61 log units in the number of viable Bradyrhizobium japonicum was found during the 1sth, using available commercial inoculants and granules. These losses were shown to be influenced by inoculant type (peat based, liquid, lyophilized) and granule type. When mixed with granules, a peat-based inoculant was found to be desiccated immediately and subjected to a water potential as low as −170 MPa. Bradyrhizobium japonicum recovery was improved when the moisture content of the granules increased. It was concluded that water suction by granules can subject the bacteria to rapid desiccation and significantly decrease their number. Thus, the nature of inoculant and granular material and their relative water retention characteristics must be taken into account to improve the efficiency of this inoculation process.Key words: Bradyrhizobium japonicum, granular inoculant, inoculation, soybean, survival.
Survey of cytotoxin production among Escherichia coli strains characterized as enteropathogenic (EPEC) by serotyping and presence of EPEC adherence factor (EAF) sequences
Authors: Guth Beatriz E. C. , Giraldi Regina , Gomes Tania A. T. , Marques Lilian R. M.
Published: February 2010

A total of 108 Escherichia coli strains characterized as enteropathogenic (EPEC) by serotyping and the presence of EPEC adherence factor (EAF) sequences were examined for cytotoxin production by cell line assays and colony hybridization with Shiga-like toxin (SLT) probes. Cytolethal distending toxin (CLDT) production was found in three (2.8%) strains belonging to serotype 086:H34, while one O11 lab:NM strain hybridized with a SLT-II probe but did not express any cytotoxic activity. All four strains showed localized adherence to HeLa cells and hybridized to an E. coli attaching–effacing gene (eae) probe. The CLDT-producing strains had multiple plasmids and some were present in all strains, including a plasmid of ~54 MDa that hybridized with the EAF probe.Key words: enteropathogenic Escherichia coli, EPEC adherence factor sequences, Shiga-like toxin, cytolethal distending toxin.
Genotypic and phenotypic diversity of Rhizobium isolated from chickpea (Cicer arietinum L.)
Authors: Nour Sarah M. , Cleyet-Marel Jean-Claude , Beck Douglas , Effosse Aline , Fernandez Maria P.
Published: February 2010

The diversity of 16 strains of chickpea-infective rhizobia from various geographical origins was analysed using genotypic and phenotypic approaches. Multilocus enzyme electrophoresis was performed, and restriction fragment length polymorphisms of the amplified 16S+IGS (intergenic spacer) rRNA gene, assimilation of 147 carbon sources, antibiotic resistance, and tolerance to NaCl and extreme pH values and temperatures were tested. These approaches had different discriminating powers. Esterase polymorphisms gave a unique pattern for each strain, allowing this method to be used for strain fingerprinting. Genetic distances between strains were estimated. The three approaches used in this study yielded consistent results. They evidenced high heterogeneity among the strains, and made it possible to classify the strains into two clusters. Isozyme patterns for superoxide dismutase were particularly interesting, since they delineated the same two groups. The phenotypic tests clearly confirmed the existence of two genetic groups on the basis of 11 phenotypic characters. Owing to the large phylogenetic distance between the two groups of strains, the taxonomic status of chickpea-infective strains is discussed.Key words: Rhizobium sp. (Cicer arietinum L.), genetic diversity, multilocus enzyme electrophoresis, restriction fragment length polymorphisms, phenotypic diversity.
Cloning of two xylanase genes from the newly isolated actinomycete Actinomadura sp. strain FC7 and characterization of the gene products
Authors: Éthier Jean-François , Harpin Serge , Girard Christine , Beaulieu Carole , Déry Claude V. , Brzezinski Ryszard
Published: February 2010

A screening program was established to find actinomycetes that produce xylanases able to hydrolyze xylan chains in a hemicellulose liquor (a by-product of steam treatment of the lignocellulosic biomass) at moderately acidic pH (4.0) and high temperature (70 °C). The first step involved a selection for xylanolytic actinomycetes having optimal growth at 50–60 °C. In the second step, the crude enzymes produced by the selected actinomycetes were compared for their xylan hydrolysis rates at pH 5 and 60 °C versus pH 4 and 70 °C. The crude enzymes produced by strain FC7 retained 65% of their activity under the more stringent of the two conditions. FC7 was identified as a member of the genus Actinomadura by chemotaxonomic procedures. A gene bank of FC7 was constructed using the shuttle vector pFD666. The plasmids were introduced into a periplasmic-leaky Escherichia coli host to rapidly detect xylanolytic recombinants. Two classes of recombinants were isolated. The plasmids pJFl and pJF6 were introduced into a xylanase- and cellulase-negative mutant of Streptomyces lividans, allowing overproduction and subsequent purification of two different xylanases, Xyl I and Xyl II. Both enzymes were classified in the low isoelectric point group of xylanases. In the presence of 100 μg/mL of bovine serum albumin, the half-life of Xyl I at pH 4 and 70 °C was 22 h.Key words: xylanase, thermophilic actinomycetes, Actinomadura, thermostability.
Effect of ionizing dose rate on the radioresistance of some food pathogenic bacteria
Authors: Dion Paule , Charbonneau Raymond , Thibault Chantal
Published: February 2010

Food pathogenic bacteria including Listeria monocytogenes (1A1 and ATCC 19111), Staphylococcus aureus (GD13 and ATCC 13565), Escherichia coli 0157:H7 (ATCC 35150), Salmonella typhimurium, Yersinia enterocolitica, Vibrio parahaemolyticus, and Campylobacter jejuni were exposed to various rates of ionizing radiation (0.78, 2.6, and 22 kGy/h) emitted by three different 60Co irradiators. D10 values (D10 is the radiation dose required to eliminate 90% of a bacterial population (one logarithmic cycle reduction)) were calculated for the various strains and growth conditions tested. A covariance analysis of these results revealed that the dose rates studied had no significant influence on the radiosensitivity of these bacteria. At all dose rates, the bacteria were more radiosensitive when irradiated in a saline solution (0.85% NaCl) than in a chicken breast meat suspension. The growth phase of the bacterial population had a variable influence on its radioresistance. For L. monocytogenes 1A1, Staphylococcus aureus ATCC 13565, E. coli 0157:H7, Y. enterocolitica, and V. parahaemolyticus, radioresistance was not significantly different in the exponential and stationary phases. Populations of L. monocytogenes ATCC 19111 and Staphylococcus aureus GD13 were significantly more resistant in the stationary phase (D10 = 0.23 and 0.12 kGy, respectively) than in the exponential phase (D10 = 0.17 and 0.09 kGy, respectively). Among the pathogenic bacteria investigated in this study, the most radioresistant was L. monocytogenes (D10 = 0.16–0.38 kGy, Gram-positive bacilli) and the most radiosensitive was V. parahaemolyticus (D10 = 0.03–0.04 kGy, halophilic Gram-negative bacilli).Key words: ionization, food pathogenic bacteria, dose rate effect, radioresistance.
The role of catalase isozymes in the culturability of the root colonizer Pseudomonas putida after exposure to hydrogen peroxide and antibiotics
Authors: Klotz Martin G. , Anderson Anne J.
Published: February 2010

The culturability of Pseudomonas putida cells after exposure to hydrogen peroxide and antibiotics was correlated with growth-dependent expression of catalase isozymes. Exponential phase wild-type cells, which contained catalase isozyme A, survived a 15-min treatment with less than 4 mM hydrogen peroxide, but were killed by higher concentrations. The culturability of P. putida mutant JIM, which lacked any functional catalase in exponential phase, was reduced by more than 75% after a 15-min exposure to ≥ 0.25 mM hydrogen peroxide. Because submillimolar concentrations of hydrogen peroxide are physiologically relevant in the bacterial cell, our results demonstrate that catalase isozyme A has essential housekeeping functions for growing cultures of P. putida. The accumulation of catalase isozymes B and C during growth into stationary phase coincided with a decrease in the sensitivity of wild-type and JIM cells of P. putida to hydrogen peroxide. Late stationary phase wild-type cells survived a 15-min exposure to even 50 mM hydrogen peroxide and mutant J1M cells survived exposure to 20 mM but not 50 mM hydrogen peroxide. The antibiotics tetracycline and kanamycin, which inhibit protein synthesis, were used to study the role of catalase induction in resistance to hydrogen peroxide. More than 40 and 80% of exponential phase cells of P. putida wild-type and J1M strains, respectively, were rendered nonculturable after a 20-min exposure to 45 μM tetracycline. Surprisingly, stationary phase cells of both P. putida strains were culturable after a 20-min exposure to tetracycline but remained sensitive to kanamycin. Exposure to tetracycline of stationary phase cells did not reduce the resistance of these cells to hydrogen peroxide. Tetracycline but not kanamycin increased the activity of catalase in lysates prepared from P. putida wild-type and mutant cells in early stationary growth phase. At this growth phase, only catalase isozyme B is operational in both strains, which suggests that tetracycline affects the activity of this enzyme.Key words: Pseudomonas putida, antibiotics, catalase, culturability, growth phase.
Effects of a chromated-copper-arsenate wood preservative on the bacterial degradation of pentachlorophenol
Authors: Wall A. James , Stratton Glenn W.
Published: February 2010

The effect of a chromated-copper-arsenate wood preservative on the degradation of pentachlorophenol by Flavobacterium sp. strain ATCC 53874 was examined in liquid culture. Both a commercially available and a laboratory-prepared formulation were tested. Each increased the lag time required for measurable pentachlorophenol degradation and the time required for complete degradation to nondetectable levels. This response was noted at all pentachlorophenol concentrations examined (10, 25, 50, 75, and 100 μg∙mL−1). The commercial formulation of chromated-copper-arsenate had the more significant impact on pentachlorophenol degradation. Inhibitory effects were evident at chromated-copper-arsenate component metal concentrations 0.1–0.5 mg∙L−1. These levels are thousands of times below those used commercially.Key words: pentachlorophenol, biodegradation, chromated-copper-arsenate, toxicity.
Genes encoding osmoregulatory proline/glycine betaine transporters and the proline catabolic system are present and expressed in diverse clinical Escherichia coli isolates
Authors: Culham Doreen E. , Emmerson Katherine S. , Lasby Bonnie , Mamelak Daniel , Steer Brian A. , Gyles Carlton L. , Villarejo Merna , Wood Janet M.
Published: February 2010

Sixty-three clinical isolates identified as Escherichia coli, 30 from the human urinary tract and 33 derived from other human origins, were screened for proline/glycine betaine transporters similar to those that support proline catabolism and proline- or glycine betaine-based osmoregulation in E. coli K-12. Both molecular (DNA- and protein-based) analyses and physiological tests were performed. All tests were calibrated with E. coli K-12 derivatives from which genetic loci putP (encoding a proline transporter required for proline catabolism), proP, and (or) proU (loci encoding osmoregulatory proline/glycine betaine transporters) had been deleted. All clinical isolates showed both enhanced sensitivity to the toxic proline analogue azetidine-2-carboxylate on media of high osmolality and growth stimulation by glycine betaine in an artificial urine preparation of high osmolality. DNA sequences similar to the putP, proP, and proU loci of E. coli K-12 were detected by DNA amplification and (or) hybridization and protein specifically reactive with antibodies raised against the ProX protein of E. coli K-12 (a ProU constituent) was detected by western blotting in over 95% of the isolates. Two anomalous isolates were reclassified as non-E. coli on the basis of the API 20E series of tests. A protein immunochemically cross-reactive with the ProP protein of E. coli K-12 was also expressed by the clinical isolates. Since all three transporters were ubiquitous, no particular correlation between clinical origin and PutP, ProP, or ProU activity was observed. These data suggest that the transporters encoded in loci putP, proP, and proU perform housekeeping functions essential for the survival of E. coli cells in diverse habitats.Key words: osmoregulation, betaine transport, urinary tract infection, Escherichia coli.
Using fura-2 to measure intracellular free calcium in Propionibacterium acnes
Authors: Futsaether C. M. , Johnsson A.
Published: February 2010

The fluorescent probe fura-2 was used to measure the intracellular free calcium concentration [Ca2+]i in the Gram-positive skin bacterium Propionibacterium acnes. Three methods for loading the probe into P. acnes were used. Two consisted of using the membrane permeable acetoxymethyl ester of fura-2, fura-2/AM, whereas in the third method, fura-2 was loaded directly into the bacteria by means of an acid shock procedure. One method using fura-2/AM was found to be satisfactory. In this case, a statistical experimental design [Formula: see text] was used to devise a suitable loading procedure. [Ca2+]i of P. acnes was dependent upon the external calcium concentration. At low external concentrations (0.2 < [Ca2+]o < 5 μM), [Ca2+]i was 135 ± 13 nM (n = 20). An increase in [Ca2+]o to 1 mM resulted in an increase in [Ca2+]i to approximately 280 ± 40 nM whereas in a Ca-free solution ([Ca2+]o < 5 nM), [Ca2+]i decreased to a lower resting value of 70 ± 7 nM. The time constants for calcium regulation upon step changes in the external concentration were of the order of 10 min. Intracellular alkalinization induced by changes in the external pH (pHo) resulted in an increase in [Ca2+]i when [Ca2+]o was in the range 0.2–5 μM. [Ca2+]i was approximately constant in the range pHo 5.5–7.7 but increased when pHo > 8.0. This increase was partially reversed when pHo was again lowered to below 8.0. In a Ca-free solution, little or no increase in [Ca2+]i was observed when pHo > 8.0.Key words: Propionibacterium acnes, fura-2, calcium.
Characterization of N-acetyl-D-glucosamine uptake in Aphanocladium album
Authors: Vasseur Valérie , Barchietto Thierry , Bompeix Gilbert
Published: February 2010

A mutant of the fungus Aphanocladium album, characterized as chitinase-overproducing, does not grow on medium containing N-acetyl-D-glucosamine as sole carbon source. The lack of growth may indicate a defect either in the uptake system or in any subsequent step in the metabolism of N-acetyl-D-glucosamine. Since transport is the first step at which control of metabolism could operate, the mutant strain was studied for its ability to transport N-acetyl-D-glucosamine. The results were compared with those obtained with the wild-type strain. Labelling experiments, designed to characterize N-acetyl-D-glucosamine uptake, indicated that uptake was due to the simultaneous operation of two transport systems, one of high affinity and one of low affinity. Wild-type and mutant strains displayed different Km values for the low-affinity system. The Km value of the mutant's low-affinity system was very high compared with that of the wild-type strain.Key words: deuteromycete, filamentous fungus, Aphanocladium album, uptake, N-acetyl-D-glucosamine.
Squalene metabolism in two species of Lagenidium
Authors: Domnas Aristotle , Biswas Shankha S. , Gallagher Patricia A.
Published: February 2010

Squalene metabolism of the sterol auxotroph Lagenidium giganteum was studied and compared with that of the positive control Lagenidium callinectes. Application of experimentally derived precautions ensured both the stability and the purity of squalene during incubations. Under these conditions mycelia of L. giganteum converted squalene to squalene oxide and to a sterol-like compound. Cell-free and microsomal preparations also converted squalene to the oxide, which was identified by thin layer chromatography with five different solvent systems, by co-chromatography with authentic oxide, and by conversion to the glycol. Supporting evidence for the production of squalene oxide was obtained by gas–liquid chromatography, high performance liquid chromatography, and autoradiography. The squalene oxide produced was identified by mass spectrometry.Key words: Lagenidium giganteum, Lagenidium callinectes, squalene, 2,3-squalene epoxide.
Construction of a DNA probe to detect isoquinoline-degrading bacteria
Authors: Richards N. K. , Mahanty H. K. , Aislabie J.
Published: February 2010

To facilitate the cloning of DNA encoding isoquinoline degradation an assay was developed that allowed the rapid visual scoring of the isoquinoline degradation phenotype of single colonies. Transposon mutagenesis of one of the isolates, Comamonas acidovorans IQ3, was performed using Tn 5, and nine Isq− mutants deficient in the ability to utilise isoquinoline as the sole nitrogen source were isolated. These mutants were also incapable of utilising the first metabolite of the isoquinoline degradation pathway, 1-hydroxyisoquinoline, as the sole carbon source. For each Isq− mutant, the EcoRI fragment containing the Tn5 insertion was cloned into pBR322. Restriction and Southern analyses of the cloned DNA revealed that of the nine Isq− mutants, six contained Tn5 insertions in a common 8.9-kb EcoRI fragment derived from the wild type, C. acidovorans IQ3. The cloned DNA thought to be involved in the degradation of isoquinoline proved to be specific when used as a probe in colony hybridization to some bacteria possessing the ability to degrade isoquinoline.Key words: DNA probe, isoquinoline degradation, transposon mutagenesis, Comamonas acidovorans.
Cellular regulation of nitrate uptake in denitrifying Flexibacter canadensis
Authors: Wu Qitu , Knowles Roger
Published: February 2010

Nitrate uptake and its regulation were investigated using an ion-specific nitrate electrode for denitrifying Flexibacter canadensis under anaerobic conditions. Glucose supported a greater rate of nitrate uptake than did glycerol, glutamate, lactose, cellobiose, or ethanol. Nitrate uptake closely approximated Michaelis–Menten kinetics; the estimated Ks(glucose) and apparent Km(nitrate) for nitrate uptake were 21 and 44 μM, respectively. Nitrate disappearance was correlated with nitrite accumulation, and nitrate had an inhibitory effect on nitrite reduction. Oxygen inhibition of nitrate uptake increased as the percent air saturation increased, and reversed readily as the percent air saturation decreased. The minimal air saturation showing inhibition of nitrate uptake was about 2–4%. Azide and cyanide completely inhibited nitrate uptake. No nitrate uptake was observed in cells grown in the presence of 1 or 5 mM tungstate (no added molybdate). When molybdate (100–200μM) was present in the medium, nitrate uptake was exhibited by organisms grown with 1 mM, but not with 5 mM, tungstate, indicating that nitrate uptake was dependent on the presence of an active nitrate reductase, and that competition between tungsten and molybdenum occurred during the formation of nitrate reductase. Nitrite production from nitrate by whole cells but not cell-free extracts was inhibited by 2,4-dinitrophenol and carbonyl cyanide m-chlorophenylhydrazone, indicating that nitrate and (or) nitrite transport depended upon the electrochemical proton gradient.Key words: denitrification, nitrate uptake, Flexibacter canadensis.
Excision and transposition of Tn5 upon insertion in the hha gene of Escherichia coli
Authors: Badenas C. , Madrid C. , Juárez A.
Published: February 2010

We report the occurrence of Tn5 secondary transpositions in Escherichia coli HB101 hha::Tn5(pANN202-312). Tn5-induced hha mutants (hemolysin oveiproducers) segregated at high frequency either hemolysin-negative clones or clones showing the parental hemolytic phenotype (reduced hemolysin production). Secondary transpositions of Tn5 appeared to be responsible for the phenotypes of both types of derivatives. The hemolysin-negative clones no longer harboured Tn5 in the hha gene, but rather Tn5 was in the hly genes of the hemolytic plasmid pANN202-312. The derivatives that recovered the parental hemolytic phenotype contained a single copy of Tn5 in a chromosomal location different from that of hha. Both kinds of transpositional events appeared to restore the function of the hha gene.Key words: Tn5, transposition, Escherichia coli, excision.
Effect of carbon and nitrogen metabolism on nitrate reductase activity of Rhodobacter capsulatus E1F1
Authors: Dobao María M. , Martínez-Luque Manuel , Moreno-Vivián Conrado , Castillo Francisco
Published: February 2010

The phototrophic bacterium Rhodobacter capsulatus E1F1 possesses an assimilatory, inducible nitrate reductase that is regulated by carbon and nitrogen metabolism. Nitrate reductase activity was detected in cells cultured with amino acids and nitrate as simultaneous nitrogen source but it required an additional carbon source such as D,L-malate. A significant rise in nitrate reductase activity was observed in media with increasing nitrate concentrations up to 10 mM KNO3, although higher nitrate concentrations had an inhibitory effect. Growth yield, generation time, and nitrate reductase activity were also dependent on the concentration of D,L-malate in cells growing with 10 mM nitrate. In carbon-starved cells, nitrate reductase activity dropped even in the presence of nitrate. The intracellular concentration of keto acids such as oxaloacetate or 2-oxoglutarate fluctuated widely depending on the presence of nitrogen and carbon sources in the culture medium. The increase in the intracellular concentration of oxaloacetate or 2-oxoglutarate in R. capsulatus E1F1 correlated well with a rise in nitrate reductase activity. These results suggest that the intracellular carbon–nitrogen balance regulates nitrate uptake in R. capsulatus E1F1, thus affecting the expression of nitrate reductase.Key words: carbon–nitrogen balance, nitrate reductase, Rhodobacter capsulatus.
Relationship between valine, fatty acids, and spiramycin biosynthesis in Streptomyces ambofaciens
Authors: Laakel Mohamed , Lebrihi Ahmed , Khaoua Saida , Schneider François , Lefebvre Gérard , Germain Pierre
Published: February 2010

Spiramycin biosynthesis in Streptomyces ambofaciens was stimulated in the presence of valine or by sequential addition of some short-chain fatty acids to a culture medium containing an ammonium salt as source of nitrogen. Acetate kinase and acetyl-CoA carboxylase, enzymes that catalysed the formation of precursors of spiramycin biosynthesis (acetyl-CoA and malonyl-CoA), were detected during the active growth and antibiotic production phases. In this latter phase a higher level of acetyl-CoA carboxylase activity was observed with valine (1.02 μmol∙min−1∙mg protein−1) than with ammonium (0.05 μmol∙min−1∙mg protein−1) as nitrogen source, while the evolution and the level of acetate kinase activity were the same in both media. Successive addition of acetate and isobutyrate stimulated highly and weakly the acetyl-CoA carboxylase and acetate kinase activity, respectively.Key words: spiramycin, Streptomyces ambofaciens, valine.
Entomopathogenic fungi of the order Entomophthorales (Zygomycotina) in adult black fly populations (Diptera: Simuliidae) in Quebec
Authors: Nadeau Martin P. , Dunphy Gary B. , Boisvert Jacques L.
Published: February 2010

Infections by insect-pathogenic fungi of the order Entomophthorales in two adult black fly populations were studied in the Réserve Faunique du Saint-Maurice (Quebec). Entomophaga near limoniae infected Simulium verecundum/rostratum in June whereas Erynia curvispora predominantly parasitized Simulium decorum from the 2nd week of July to September and Erynia conica attacked the Simulium venustum complex, Simulium verecundum/rostratum, and the Simulium vittatum complex from May to September. The fungi did not infect the adult stage of Prosimulium species. Possible evidence of host specificity for Erynia conica is discussed.Key words: Entomophthorales, black fly, host specificity, Entomophaga, Erynia, Simulium.
Differential increase in cytoplasmic pH at bud and germ tube formation in Candida albicans: studies of a nongerminative variant
Authors: Kaur Simminder , Mishra Prashant
Published: February 2010

The changes in cytoplasmic pH (pHi) accompanying the morphological transition of Candida albicans were studied using a nongerminative variant. A transient cytoplasmic alkalinization at the time of evagination was observed in both the variant and its parent. Under zinc-deficient conditions the wild-type cells that formed buds at pH 4.5 and mycelia at pH 6.5 showed an increase in pHi of 0.58 and 0.12 pH units, respectively. Under similar conditions the pHi of the nongerminative variant that formed buds at both pH 4.5 and 6.5 increased by 0.66 and 0.40 pH units, respectively, suggesting that a greater magnitude of increase in pHi at the time of evagination is probably needed for bud formation. These results provide evidence for a correlation between intracellular alkalinization and time of cell differentiation in C. albicans.Key words: C. albicans, cytoplasmic pH, morphogenesis, nongerminative variant.
Flunarizine affects F-actin pattern in Mucor rouxii germlings
Authors: Hašek Jiří , Streiblová Eva
Published: February 2010

The effect of the Ca2+ antagonist flunarizine on Mucor rouxii germlings was analyzed. In sporangiospores cultivated in aerobiosis, the drug blocked germ tube emergence, but enlarged spheres continued to grow isodiametrically. The formation of broad protuberances and irregular buds was observed. Calcofluor staining indicated aberrations on the surface of the flunarizine-treated cells. This was not a simple switch to the yeast form, since labelling with rhodarmine-tagged phalloidin revealed various ring-shaped structures of F-actin, indicating furrowing of the plasma membrane. Flunarizine apparently did not interfere with nuclear divisions or with the integrity of microtubules. In young germlings, flunarizine treatment induced cessation of tip growth and promoted septation associated with the rearrangement of F-actin. Apical growth was reestablished after either the drug was removed or an excess of Ca2+ ions was added to the medium. The results suggest that tip-related arrangement of F-actin requires flunarizine-sensitive Ca2+ entry.Key words: Mucor rouxii, F-actin, Ca2+ antagonist, flunarizine, germination.
Identification and mapping of the gene translation products involved in the first steps of the Comamonas testosteroni B-356 biphenyl/chlorobiphenyl biodegradation pathway
Authors: Bergeron Janique , Ahmad Darakhshan , Barriault Diane , Larose Angèle , Sylvestre Michel , Powlowski Justin
Published: February 2010

In this study, we have mapped Comamonas testosteroni B-356 genes encoding enzymes for the conversion of biphenyl and 4-chlorobiphenyl into the corresponding meta-cleavage compounds onto a 6.3-kb DNA fragment, and we have determined the subunit composition of the enzymes involved in this pathway. The various proteins encoded by this 6.3-kb DNA fragment and by subclones derived from it were overexpressed and selectively labelled using the T7 polymerase promoter system in Escherichia coli. They were then analyzed using SDS-PAGE, which allowed the encoding locus of each polypeptide to be mapped. Despite apparent dissimilarity in the congener selectivity patterns of the initial oxygenase of strain B-356 with those of Pseudomonas sp. strain LB400, the number and sizes of the polypeptides involved in the enzymatic conversion of biphenyl or 4-chlorobiphenyl into the meta-cleavage product appear to be similar in the two strains. In both strains, the bph operon encodes the following: the large (51-kDa polypeptide encoded by bphA) and the small (22-kDa polypeptide encoded by bphE) subunits of the iron sulphur protein, which is thought to interact directly with the substrate to introduce the oxygen molecule; the ferredoxin (12-kDa polypeptide encoded by bphF) involved in electron transfer from the reduced ferredoxin reductase to the oxidized iron sulphur protein; the 29-kDa polypeptide of the 2,3-dihydro-2,3-dihydroxybiphenyl dehydrogenase encoded by bphB; and the 32-kDa polypeptide of the 2,3-dihydroxybiphenyl-1,2-dioxygenase encoded by bphC, which catalyzes meta-1,2 fission of the aromatic ring. A major difference between strain B-356 and strain LB400 is that the bphG gene encoding biphenyl dioxygenase ferredoxin reductase is located outside the bph gene cluster in strain B-356. Several lines of evidence indicate that bphG is absent in clones carrying the bph operon from strain B-356.Key words: PCB, gene expression, biphenyl oxygenase, bph.
Optimization of culture conditions for indole-3-pyruvic acid production by Streptomyces griseoflavus
Authors: El-Abyad Mohamed S. , El-Sayed Mostafa A. , El-Shanshoury Abdel-Reheem , Farid Mervat
Published: February 2010

Thirty strains of actinomycetes were isolated from fertile soils in Egypt on starch–nitrate agar. The isolated organisms were recognized as spore formers with aerial mycelia containing L-DAP (diaminopimelic acid) and glycine. The inability of the vegetative mycelia to fragment into bacillary or coccoid forms and the presence of spores borne on sporophores, placed these organisms in the family Streptomycetaceae. The isolates also proved to be aerobic and Gram positive. The absence of DL-DAP and the presence of L-DAP in their cell hydrolysates, and the production of a wide range of pigments in the aerial and substrate mycelia, assigned these isolates to the genus Streptomyces. The isolated organisms were screened for production of plant growth regulators, and the most active producer of indole-3-pyruvic acid was examined further. Using the described morphological, physiological, and biochemical criteria for identification, this organism was identified as Streptomyces griseoflavus. The optimal culture conditions for maximal production of both cellular and extracellular indole-3-pyruvic acid by S. griseoflavus were investigated and shown to be as follows: a culture medium composed of (g∙100 mL−1) NH4NO3, 0.55; DL-tryptophan, 0.4; NaCl, 0.7; at pH 7 (buffered); with an incubation period of 6 days at 28 °C in the dark and under shaking conditions.Key words: actinomycetes, plant growth regulators, Streptomyces griseoflavus, indole-3-pyruvic acid, culture conditions.
Alkaline phosphatase and a cellulase reporter protein are not exported from the cytoplasm when fused to large N-terminal portions of the Caulobacter crescentus surface (S)-layer protein
Authors: Bingle Wade H. , Smit John
Published: February 2010

Using a gene fusion approach, hybrid proteins were created by linking alkaline phosphatase (PhoA) or a cellulase reporter (ΔCenA) to four large N-terminal portions of the Caulobacter crescentus surface (S)-layer protein (RsaA; 1026 amino acids). Three of the sites (amino acids 189, 220, 315) were selected on the basis of TnphoA experiments that suggested the first 250–350 amino acids of RsaA could mediate export of PhoA from the cytoplasm while the fourth lay only 21 amino acids from the C-terminus. Expression of all fusions except rsaA(315):ΔcenA and rsaA(315):phoA was toxic to C. crescentus. None of the gene fusions were toxic when expressed by Escherichia coli DH5α, where all the hybrid proteins accumulated as inclusion bodies. The toxicity of hybrid proteins encoding 189, 220, and 1005 RsaA-derived amino acids was related to the nature of the hybrid protein itself because truncated RsaA peptides lacking their reporter domains were nontoxic. Further study of RsaA(ΔC21) showed that this and presumably other truncated RsaA derivatives were neither secreted nor prone to intracellular accumulation. Although C. crescentus tolerated the expression of rsaA(3l5):ΔcenA and rsaA(315):phoA, the encoded hybrid proteins were not exported in significant quantities from the cytoplasm. These results extend and confirm earlier work that large portions of the S-layer protein N-terminus cannot mediate export of passenger proteins from the cytoplasm and that the entire native S-layer protein may be required to properly interact with the RsaA secretion machinery.Key words: surface layer, secretion, reporter protein, Caulobacter crescentus.
Incidence of Norwalk virus infections during a prospective epidemiological study of drinking water related gastrointestinal illness
Authors: Payment Pierre , Franco Eduardo , Fout G. Shay
Published: February 2010

To determine the seroprevalence of Norwalk virus and whether Norwalk virus contributed to an observed increase in illness in tap water drinkers participating in a prospective epidemiological study, sera collected during the study were examined for changes in Norwalk virus antibody titer, using a specific enzyme immunoassay. Antibodies to Norwalk virus were measured in sera collected in March, June, and September 1988 and in June 1989, and antibodies were found in 79% of the individuals. Seroprevalence increased with age, being 55% (ages 9–19), 79% (20–39), 87% (40–49), 84% (50–59), and 100% (60 and older). Norwalk infections occurred in 33% of the individuals during the course of the study. The highest rate of infection (expressed as a monthly rate) was observed during the summer of 1988. These results confirm that a large number of infections owing to Norwalk viruses occur throughout the year. A previous seroconversion or a high serum titer were not always protective. Finally, there was no detectable difference in infection rate between consumers of tap water and consumers of water treated by reverse-osmosis units, suggesting that Norwalk virus infections were not responsible for the excess of gastrointestinal illness observed in tap water drinkers during this epidemiological study.Key words: drinking water, Norwalk virus, gastroenteritis, seroprevalence.
An asexual variant of Phytophthora insolita
Authors: Ann P. J. , Ko W. H.
Published: February 2010

All tested isolates of Phytophthora insolita and an unknown asexual Phytophthora species found in soil, ditch water, and diseased plant tissues in Taiwan produced ovoid, nonpapillate, nondeciduous sporangia on sporangiophores proliferating through an empty sporangium or in a nesting fashion, and formed irregular as well as spherical hyphal swellings. All tested Phytophthora isolates grew at an unusually high temperature of 39 °C, displayed similar or identical electrophoretic patterns of soluble proteins, and produced α1 hormone. The ability of one isolate of P. insolita to produce oospores was decreased and that of another was lost completely during storage. The results suggest that all isolates of the asexual Phytophthora sp. tested were Phytophthora insolita and were unable to produce oospores owing to a defect in the physiological process of sexual reproduction.Key words: asexual Phytophthora, Phytophthora insolita, high-temperature Phytophthora, sexual reproduction.
Thiosulfate oxidation by sulfur-grown Thiobacillus thiooxidans cells, cell-free extracts, and thiosulfate-oxidizing enzyme
Authors: Chan C. W. , Suzuki Isamu
Published: February 2010

The oxidation of thiosulfate by Thiobacillus thiooxidans grown on sulfur was studied in cells, cell-free extracts, and a thiosulfate-oxidizing enzyme system. Thiosulfate was oxidized to tetrathionate by cells treated with N-ethylmaleimide with a pH optimum at 2.3. The cell-free extracts also oxidized thiosulfate with the same pH optimum and O2 consumption. Untreated cells, on the other hand, oxidized thiosulfate to sulfite in the presence of 2-n-heptyl-4-hydroxyquinoline N-oxide, an inhibitor of sulfite oxidation. The cells treated with N-ethylmaleimide showed two Km values for thiosulfate while the cell-free system showed only one Km. The Km value for thiosulfate generally increased with the increasing pH. A soluble thiosulfate-oxidizing enzyme system was extracted from the cells at pH 2.5 in the presence of 1 M ammonium sulfate by passage through a French pressure cell. The system contained a native cytochrome c that was reduced by thiosulfate at pH 2.5 and a thiosulfate-ferricyanide oxidoreductase activity with a pH optimum around 2.0. The acidic extract also contained a component that reduced horse heart cytochrome c at a neutral pH. The reduction at an acidic pH required sulfite.Key words: thiosulfate oxidation, Thiobacillus thiooxidans, tetrathionate, sulfite.
The ether lipids of Methanosarcina mazei and other Methanosarcina species, compared by fast atom bombardment mass spectrometry
Authors: Sprott G. D. , Dicaire C. J. , Patel G. B.
Published: February 2010

The total lipids of Methanosarcina mazei accounted for 4.0% of the cell dry mass, and 90% of these were polar lipids. Nearly all of the polar fraction consisted of diether (2,3-di-O-phytanyl-sn-glycerol) and 3-hydroxydiether analogs of phosphatidylglycerol, phosphatidylinositol, phosphatidylserine, and phosphatidylethanolamine. Several highly fluorescent trace components in the lipid extracts were purified and partially characterized by mass spectrometry. Fast atom bombardment mass spectrometry of total lipid extracts provided data to establish a close relationship among the polar lipids present in Methanosarcina mazei, Methanosarcina thermophila, Methanosarcina acetivorans, Methanosarcina barkeri Fusaro, and Methanosarcina barkeri MS.Key words: diether lipids, hydroxydiether lipids, fast atom bombardment mass spectrometry, Methanosarcina, archaeobacteria.