bluuebunny/crossref_metadata_2025 huggingface dataset:
Adherence of Legionella pneumophila to U-937 cells, guinea-pig alveolar macrophages, and MRC-5 cells by a novel, complement-independent binding mechanism
Authors: Gibson III Frank C. , Tzianabos Arthur O. , Rodgers Frank G.
Published: February 2010

In the absence of serum, Legionella pneumophila demonstrated wash-resistant adherence to U-937 cells, primary guinea-pig alveolar macrophages, and MRC-5 cells. Neither complement nor antibody was required for binding. The dynamics of adherence following inoculation of L. pneumophila at increasing 10-fold multiplicities of infection to each of the three host cell types resulted in a first-order kinetic relationship of binding, indicative of one bacterial adhesin molecule recognized by one host cell receptor moiety. Host cell receptor saturation studies showed that depending on the cell type, 2–8% of the bacterial inoculum adhered to cells under these nonopsonic conditions. Preliminary adhesin and receptor characterization studies were preformed to define the chemical composition of the binding structures on both the organism and the three different host cell surfaces. The adherence phenomenon was investigated using competitive binding assays in the presence of putative adhesin analogs as well as following treatments modifying the microbial and host cell surface membranes. Attachment was evaluated both by viable bacterial cell colony counts and by indirect immunofluorescent assay. With the exception of aldehyde treatments, the various membrane-modifying regimes and the presence of the adhesin analogs were shown to have no effect on organism or host cell viability. Data suggested that the L. pneumophila adhesin responsible for opsonin-independent binding to these host cells was a protein structure with lectin-like properties. Furthermore, this protein would appear to be intimately associated with carbohydrate or lipid structures located on the bacterial outer membrane. The receptor moiety present on all host cells responsible for binding L. pneumophila had properties consistent with a carbohydrate or complex saccharide structure. To evaluate the role of complement receptors as the structures necessary for L. pneumophila infection of macrophages, a battery of monoclonal antibodies were used to block the complement receptor (CR) types 1 (CD35), CR3 (CD 18, CD11b), and CR4 (CD18, CD11c). Blocking studies with CR-specific monoclonal antibodies indicated that CR1 and the integrin receptors CR3 and CR4 were not involved in the opsonin-independent binding of L. pneumophila to macrophage-like cells.Key words: Legionella, opsonin-independent attachment, bacterial adherence, complement receptors, adhesion–receptor interactions.
Expression of the symbiotic plasmid from Rhizobium leguminosarum biovar trifolii in Sphingobacterium multivorum
Authors: Fenton M. , Jarvis B. D. W.
Published: February 2010

An inoculant strain of Rhizobium leguminosarum biovar trifolii containing a Tn5 marked symbiotic plasmid transferred this plasmid by conjugation to Sphingobacterium multivorum, an organism that can be found in soil. The transconjugant bacteria nodulated the roots of white clover (Trifolium repens) seedlings but did not fix atmospheric nitrogen. Microscopic examination revealed abnormal nodule structures. Bacteria isolated from the nodules were shown to be closely related to the recipient S. multivorum and Southern blots of genomic digests probed with nodA DNA confirmed that the transconjugants contained symbiotic genes. This is the first report of the spontaneous transfer, by conjugation, of a symbiotic plasmid from R. leguminosarum biovar trifolii to S. multivorum.Key words: Rhizobium, Sphingobacterium, nodulation, nod gene transfer.
Scanning electron microscopy of the septal pore cap of the basidiomycete Schizophyllum commune
Authors: Müller Wally H. , van Aelst Adriaan C. , van der Krift Theo P. , Boekhout Teun
Published: February 2010

As part of a comparative study of the structure and function of pore structures in heterobasidiomycetous yeasts, dikaryotic hyphae of Schizophyllum commune were subjected to chemical fixation, freeze fracturing, maceration, and freeze substitution, and were subsequently prepared for scanning electron microscopy. The interior of the hyphal cell was visualized and revealed the perforated septal pore cap or parenthesome, mitochondria, vacuoles, and tubular endoplasmic reticulum. The septal pore cap showed connections with tubular endoplasmic reticulum. This tubular endoplasmic reticulum covered the dolipore septal surface. The results presented here complement and extend the ultrastructural image of the septal pore cap obtained from transmission electron micrographs.Key words: septal pore cap, Schizophyllum commune, freeze fracture, maceration, scanning electron microscopy.
Detection of naturally occurring enteroviruses and hepatitis A virus in undigested and anaerobically digested sludge using the polymerase chain reaction
Authors: Straub Timothy M. , Pepper Ian L. , Gerba Charles P.
Published: February 2010

Four undigested and four anaerobically digested sewage sludge samples were analyzed for enteroviruses and hepatitis A virus using seminested and double polymerase chain reaction (PCR), respectively. For enteroviruses, all eight samples were positive when detection was by seminested PCR. Using cell culture all samples except two digested sludge samples were positive. For hepatitis A virus, seven out of eight samples were positive by PCR detection. In all samples, PCR inhibitory substances were removed by passage through Sephadex G-50 and Chelex 100 columns. Overall the PCR methodology was highly successful in identifying the presence of both viruses; however, with this methodology, there was no indication as to whether enteroviruses or hepatitis A viruses not confirmed in cell culture were infectious.Key words: RT-PCR, seminested PCR, sewage sludge, enteroviruses, hepatitis A virus.
O2 regulation of denitrification in Flexibacter canadensis
Authors: Wu Qitu , Knowles Roger , Niven Donald F.
Published: February 2010

We studied the sensitivity to oxygen of the reductases involved in denitrification by whole cells and membrane fractions of Flexibacter canadensis. All of the nitrate reductase activity was found in the membrane fraction, suggesting that the nitrate reductase of F. canadensis is largely or entirely a membrane-bound enzyme. Methyl viologen and benzyl viologen were good electron donors to nitrate reductase in both whole cells and membrane fractions, whereas glucose and glycerol were effective in whole cells but, as expected, not in membrane fractions. Oxygen, generated by means of H2O2 plus catalase, inhibited the production of nitrite from nitrate by intact cells but not by membrane fractions, suggesting that O2 exerts its inhibitory effect at the level of nitrate transport rather than nitrate reduction. In intact cells, the rates of nitric oxide accumulation during reduction of nitrite in the presence of 20 μM carbonyl cyanide m-chlorophenylhydrazone, and consumption of nitric oxide and nitrous oxide, decreased as the concentration of H2O2 was increased. The concentrations of H2O2 giving 50% inhibition of reduction of nitrate and nitrite were 0.34 and 0.12 mM, respectively. In contrast, the rates of nitric oxide and nitrous oxide consumption were inhibited by only 36 and 32% at a concentration of H2O2 of 3.99 mM. These results indicate that the reduction of both nitric oxide and nitrous oxide is relatively tolerant to oxygen, and that nitrite reductase is much more sensitive to oxygen than the other reductases.Key words: nitrate reductase, nitrate transport, denitrification, O2 inhibition, Flexibacter canadensis.
Construction of composite transposons for halophilic Archaea
Authors: Dyall-Smith Michael L. , Doolittle W. Ford
Published: February 2010

Transposons with selectable marker genes (e.g., antibiotic resistance) have been extremely useful tools in bacterial genetics but have not been found naturally in Archaea. We constructed synthetic transposons consisting of halobacterial ISH elements (ISH2, ISH26, or ISH28) flanking a mevinolin resistance determinant. Introduction of these constructs into Haloferax volcanii cells can produce drug-resistant transformants through homologous recombination between the plasmid hmgA gene and the chromosomal hmgA locus. This problem was overcome by using another host, Haloarcula hispanica, the hmgA gene of which shares little homology with that from Haloferax volcanii. Introduction of an ISH28-based transposon (ThD28) into Haloarcula hispanica cells produced numerous transformants. Each of these was shown to contain an ISH-flanked mevinolin resistance determinant integrated into the cellular DNA. Integration was not obviously site specific. Transposon ThD26 (based on ISH26a), was less mobile, relative to ThD28, and the ISH2-based construct (ThD22) did not transpose at all in these cells. The further development of halobacterial transposons may provide useful genetic tools allowing rapid isolation and analysis of halobacterial genes, particularly those with no selectable phenotype.Key words: archaebacteria, Archaea, transposon, halobacteria, insertion sequence.
Interactions between Lentinula edodes and pseudomonads
Authors: Tsuneda Akihiko , Thorn Greg
Published: February 2010

Interactions between four dikaryotic strains of Lentinula edodes and seven strains of Pseudomonas species were investigated in vitro. Two strains of Pseudomonas tolaasii were strongly pathogenic to fruiting bodies of L. edodes, causing browning and lysis of the inoculated tissues. The mode and outcome of interactions on agar varied with the combination between strains of L. edodes and bacteria. Pseudomonas cepacia and two strains of Pseudomonas tolaasii inhibited hyphal growth of L. edodes strains, whereas Pseudomonas fluorescens and one strain of Pseudomonas tolaasii were generally susceptible to bacteriolysis by L. edodes. Strains of L. edodes differed markedly in their ability to attack and lyse bacteria. In general, hyphal tips were the most potent in bacteriolysis. Ultrastructural variations were recognized in the process of bacterial wall degradation by L. edodes hyphae, but walls were eventually disintegrated to minute granules that ranged in diameter from 0.05 μm to smaller than measurable by scanning electron microscopy.Key words: bacteriolysis, cell wall, Pseudomonas, Lentinula, SEM.
Occurrence of legionellae in hot water distribution systems of Finnish apartment buildings
Authors: Zacheus Outi M. , Martikainen Pertti J.
Published: February 2010

The occurrence of legionellae in the hot water distribution systems of 67 buildings located in different parts of Finland was studied. Most of the buildings were apartment buildings. They had different hot water temperatures, and some received their cold potable water from surface water plants and some from ground water plants. Hot water samples were taken from taps, showers, and water mains just before and after the heat exchanger. Legionella pneumophila was isolated from 30% of the distribution systems. In the legionella-positive samples the legionella concentration varied from < 50 to 3.2 × 105 colony-forming units (cfu)/L (mean 2.7 × 103 cfu/L). The highest concentration of legionellae was found in the shower water. Legionellae appeared more often and with higher concentrations in hot water systems using cold water processed in surface water plants than in hot water systems associated with ground water plants. A high organic matter content in surface waters might favor the occurrence of legionellae and also the growth of other heterotrophic microbes. Mean water temperature just after heating was slightly higher in the legionella-negative systems than in the legionella-positive systems (53.5 vs. 51.5 °C).Key words: Legionella, organic carbon, hot water distribution system, water temperature.
Detection and identification of Clavibacter michiganensis subsp. sepedonicus and Clavibacter michiganensis subsp. michiganensis by nonradioactive hybridization, polymerase chain reaction, and restriction enzyme analysis
Authors: Rademaker J. L. W. , Janse J. D.
Published: February 2010

To develop a rapid and reliable detection and identification method for Clavibacter michiganensis subsp. sepedonicus and C. michiganensis subsp. michiganensis, two biotinylated probes and derived primer sets were evaluated for specificity using a large number of bacterial strains. Detection in dot blot analysis using the Diagen probe against C. michiganensis subsp. sepedonicus was possible with all 32 C. michiganensis subsp. sepedonicus strains tested. Cross-hybridization occurred with all nine C. michiganensis subsp. insidiosus strains tested. No hybridization occurred with any of 54 other related and unrelated bacterial strains including C. michiganensis subsp. michiganensis, C. michiganensis subsp. nebraskensis, C. michiganensis subsp. tessellarius, C. iranicus, C. rathayi, and C. tritici and potato saprophytes. Hybridization of the MIC 1 probe against C. michiganensis subsp. michiganensis was obtained with 22 out of 24 C. michiganensis subsp. michiganensis strains. A weak hybridization signal occurred only with two strains of C. michiganensis subsp. insidiosns. No hybridization occurred with any of the 71 other related and unrelated bacterial strains tested including tomato saprophytes. Restriction fragment length polymorphisms detected with the Diagen probe allowed differentiation between C. michiganensis subsp. sepedonicus and the related C. michiganensis subsp. insidiosus. Restriction fragment length polymorphism analysis using the MIC 1 probe and BamH1 showed at least two groups of patterns within C. michiganensis subsp. michiganensis. By using a primer set derived from the Diagen probe, a DNA sequence could be amplified with all C. michiganensis subsp. sepedonicus strains tested. Only the nontarget organism C. michiganensis subsp. insidiosus yielded a similar polymerase chain reaction product. Restriction enzyme analysis of the polymerase chain reaction product enabled rapid distinction between the subspecies. With a CMM primer set derived from the MIC 1 probe a DNA sequence was amplified from the same 22 out of 24 C. michiganensis subsp. michiganensis strains that showed hybridization with the MIC 1 probe. The polymerase chain reaction product could be verified by restriction enzyme analysis. The Diagen and MIC 1 probes and derived primer sets were shown to be useful for the detection and identification of C. michiganensis subsp. sepedonicus and C. michiganensis subsp. michiganensis. The MIC 1 probe, however, failed to detect two strains of the latter subspecies.Key words: biotin, PCR, REA, potato bacterial ring rot, bacterial canker of tomato, RFLP, Clavibacter michiganensis subsp. insidiosus.
Influence of some selected ions on system water activity and on ethanol vapour pressure and its inhibitory action on Saccharomyces cerevisiae
Authors: Guerzoni Maria Elisabetta , Lanciotti Rosalba , Sinigaglia Milena , Anese Monica , Lerici Carlo Raffaele
Published: February 2010

The individual and interactive effects of some ionic species on the ethanol vapour pressure in equilibrium with the system, the water activity, and the fermentative performance of Saccharomyces cerevisiae were investigated. The concentrations of the ions Ca2+, Mg2+, and NH4+ were modulated according to a central composite design, within a range of values that can be individually considered optimal for S. cerevisiae. Although the temperature and ethanol and glucose concentrations were kept constant, the initial water activity and ethanol vapour pressure of the systems were remarkably affected by the interactions between the three ions studied. The data concerning the fermentative activity emphasize the role of the physical state of ethanol. In fact, when the ethanol concentration was kept constant, the highest fermentative performances of yeast were obtained when the ethanol vapour pressure, depending particularly on the Ca2+ and Mg2+ interaction, was the lowest in the matrix of values considered.Key words: Saccharomyces cerevisiae, water activity, ethanol.
Salt stress in a halophilic bacterium: alterations in oxidative metabolism and oxy-intermediate scavenging systems
Authors: Brown-Peterson Nancy J. , Salin Marvin L.
Published: February 2010

Halobaeterium halobium, an archaebacterium that grows optimally in 4 M NaCl, was subjected to hypo-osmotic stress by growth in media containing either 1, 1.25, 1.65, or 2 M NaCl. Exponential growth occurred over a 48-h period in 2 M NaCl as well as in the standard 4 M NaCl growth medium. In growth media containing NaCl in a concentration below 2 M, exponential growth was observed after a 6-h lag. After 12 h of growth in the lowest NaCl concentrations, extracts from H. halobium showed nearly a 100-fold increase in the specific activity of catalase compared with bacteria grown at higher NaCl concentrations; superoxide dismutase activity increased 10-fold, whereas peroxidase activity increased 4- to 5-fold compared with the activities at the higher salt levels. Specific activities remained elevated throughout the 48-h period. In contrast, malate dehydrogenase activity was not affected by alterations in the salt environment. Respiration of H. halobium growing in the most hyposaline medium was significantly elevated at 12 h and remained high over the ensuing 36 h. Metabolic rates correlated with increases observed in oxy-intermediate scavenging enzymes. No additional isozymes of superoxide dismutase were induced by salt stress. However, the catalase activity emanated from a newly induced mesohalic catalase distinct from the constitutive catalase–peroxidase.Key words: Halobaeterium halobium, metabolism, oxy-intermediate, superoxide dismutase, catalase.
Production of 2,4-diacetylphloroglucinol by the biocontrol agent Pseudomonas fluorescens Pf-5
Authors: Nowak-Thompson Brian , Gould Steven J. , Kraus Jennifer , Loper Joyce E.
Published: February 2010

2,4-Diacetylphloroglucinol was detected in and isolated from culture extracts of the biological control bacterium Pseudomonas fluorescens Pf-5. Its structure was identified using a combination of chromatographic techniques and NMR spectroscopic methods. Carbon source influenced 2,4-diacetylphloroglucinol production by Pf-5 in culture media. 2,4-Diacetylphloroglucinol inhibited growth of the plant pathogenic fungi Pythium ultimum and Rhizoctonia solani, and the plant pathogenic bacterium Erwinia carotovora subsp. atroceptica, which cause diseases that are suppressed by strain Pf-5. The results of this study provide further evidence for the prevalence of 2,4-diacetylphloroglucinol production among strains of Pseudomonas fluorescens that suppress plant diseases.Key words: 2,4-diacetylphloroglucinol, Pseudomonas fluorescens, biocontrol.
Evaluation of Biolog MT plates for aromatic and chloroaromatic substrate utilization tests
Authors: Fulthorpe Roberta R. , Allen D. Grant
Published: February 2010

Biolog MT microplates are miniaturized substrate utilization test plates that exploit the colorimetric change in tetrazolium dyes as they are reduced in the presence of respiring bacteria. The test plates are attractive for the rapid screening of culture collections for unusual degradative activities. We evaluated these plates for the screening for the use of aromatic and chloroaromatic compounds by a culture collection of bacteria isolated from the waste water treatment system of a pulp and paper mill. We found that cell densities between 106 and 109 colony-forming units/mL could be used with the kits. However, the substrate concentration required to allow utilization detection lay between 0.25 and 2.5 mM, a level not significantly different from that required for more classical tests. Bacterial utilization of aromatic substrates such as phenol, vanillin, and benzoate could not be detected when the manufacturer's instructions were followed, because all proved to be toxic to test strains at the recommended test concentrations. We assessed the toxicity of monochlorophenols, 5-chlorovanillin, and 3-chlorobenzoate to representative species from our collection and found that only the latter two were sufficiently nontoxic to be tested with these microplates. The value of the tetrazolium dye reduction assay used in these kits to screen for bacterial utilization of more toxic substrates is limited by the high substrate concentrations required.Key words: substrate utilization, bacteria, aromatic, chloroaromatic, test kit.
Halopicolinic acids, novel products arising through the degradation of chloro- and bromo-biphenyl bySphingomonas paucimobilisBPSI-3
Authors: Davison Annette D. , Veal Duncan A. , Karuso Peter , Jardine Daniel R.
Published: February 2010

Sphingomonas paucimobilis BPSI-3 was previously isolated from a mixed microbial consortium growing on biphenyl as the sole source of carbon and energy. Transformation of 4 -chlorobiphenyl (4CBP) was demonstrated by this strain, although little or no growth was observed. In minimal salts medium supplemented with 4CBP or bromobiphenyl and dextrose, yellow coloured product(s) were rapidly formed. Gas chromatography – mass spectrometry (GC–MS) revealed single ring N-heterocyclic compounds that were identified as halopicolinic acids. We believe this to be the first report of such compounds being formed via biological transformation of halobiphenyls. A mechanism is proposed for their formation.Key words: halobiphenyl degradation, halopicolinic acid, Sphingomonas paucimobilis BPSI-3, bioremediation.
Isolation and description of carbazole-degrading bacteria
Authors: Shotbolt-Brown Joanna , Hunter David W. F. , Aislabie Jackie
Published: February 2010

Carbazole is a nitrogen hetrocyclic compound associated with fossil fuels and their products. Enrichment cultures were established to isolate bacteria able to degrade carbazole and a plate assay was developed to select carbazole-degrading microorganisms from the enrichments. Three different bacterial isolates capable of mineralizing carbazole were obtained. No intermediates of carbazole degradation were detected. The bacteria had a limited substrate specificity; all used benzoate for growth but were unable to utilize the analogues of carbazole, fluorene, or dibenzothiophene.Key words: carbazole, biodegradation, bacteria.
Serogroup conversion of Vibrio cholerae non-O1 to Vibrio cholerae O1: effect of growth state of cells, temperature, and salinity
Authors: Montilla Rafael , Chowdhury M. A. R. , Huq A. , Xu B. , Colwell Rita R.
Published: February 2010

Recently, we reported the occurrence of seroconversion from Vibrio cholerae non-O1 to V. cholerae O1, but little is known about the environmental and physiological factors influencing seroconversion. We investigated effects of temperature (4, 25, and 35 °C) and salinity (<0.5 and 10‰), as well as the stage of growth of cells, on serogroup conversion. Seroconversion of V. cholerae occurred under various environmental conditions. However, the rate of seroconversion in natural water (<0.5‰ salinity) and synthetic seawater microcosms (10‰ salinity), employing cells harvested from stationary phase culture, was approximately 2 logs higher than cells harvested from cultures in the logarithmic phase (i.e., 105 versus 103 per 1010 cells). Thus, the physiological state of the cells, and to a lesser degree, temperature and salinity, is an important factor in the conversion of V. cholerae from non-O1 to O1 serogroup.Key words: Vibrio cholerae O1, Vibrio cholerae non-O1, growth stage, serogroup conversion.
Restriction fragment profiles of genome segment A of infectious bursal disease virus correlate with serotype and geographical origin of avibirnaviruses
Authors: Qian B. , Kibenge F. S. B.
Published: February 2010

Previous analyses of the two serotypes of infectious bursal disease virus (IBDV) have demonstrated the correlation between antigenicity and similarities of nucleotide and amino acid sequences of the VP2 coding region in genome segment A. Restriction fragment profiles of genomic segment A cDNA of five IBDV isolates (QC-2 and QT-1 of serotype 1, SK9, and Nos. 39 and 52 of serotype 2) were determined in order to establish the genetic relationship of these viruses to other avibirnaviruses. The restriction fragment profiles using three of seven restriction enzymes (Sad which cuts in the VP2 region, DraIII which cuts in the VP3 region, and EcoRI which cuts in the VP4 region) were used to place QC-2, QT-1, SK9, No. 39, and No. 52 within the phylogenetic tree among seven other avibirnaviruses of known sequence. The two IBDV serotypes corresponded to two genotypes on the basis of the presence or absence of the SacI restriction site. The serotype 1 cluster of strains was further differentiated into five minor clusters on the basis of the PstI, EcoRI, BamHI, HindIII, DraIII, and Bsu36I restriction sites, which emphasized the geographical origins of the strains. It is concluded that restriction analysis of cDNA of the whole viral genomic segment A allows differentiation of IBDV isolates on the basis of their antigenicity and geographical origin.Key words: phylogeny, avibirnavirus, geographical origin.
Distribution of diaminopropane and acetylspermidine inEnterobacteriaceae
Authors: Hamana Koei
Published: February 2010

Polyamines of 97 strains (60 species) belonging to 18 genera of the family Enterobacteriaceae were determined by high performance liquid chromatographic analysis. In addition to putrescine and cadavarine, diaminopropane was widely distributed in Enterobacteriaceae and almost ubiquitously within Enterobacter, Pantoea, Erwinia, Leminorella, Proteus, Leclercia, Morganella, Klebsiella, Hafnia, Rahnella, Serratia, and Tatumella species and sporadically within Citrobacter, Escherichia, Moellerella, Providencia, Yokenella, and Yersinia species. Histamine was detected in some cultures of Proteus and Morganella. Agmatine was sporadically spread. Heterogeneity in the occurrence of spermidine was observed within the 18 genera. Acetylated spermidine was found concomitantly in the spermidine-containing cultures. Distribution profiles of diaminopropane, spermidine, and acetylspermidine in Enterobacteriaceae can serve as a chemotaxonomic marker to distinguish this family from other taxa of the gamma subclass of the class Proteobacteria.Key words: polyamine, diaminopropane, acetylspermidine, Enterobacteriaceae.
Chloride requirement for reproduction by Pythium ultimum
Authors: Saunders G. A. , Hancock J. G.
Published: February 2010

Chloride (≥ 0.1 mM) was essential for asexual and sexual reproduction, but not mycelial growth, by Pythium ultimum in synthetic culture media. Bromide partially substituted for chloride in support of oogonia formation. The production of gemmae (sporangia or hyphal swellings) increased in proportion to concentrations of KCl in culture media between 0.2 and about 0.5 mM but leveled off between 0.5 and 4 mM. Chloride contents of mycelia after 3 days incubation were proportional to the number of gemmae produced when the fungus was grown in low concentrations of KCl. Under the culture conditions of this study, production of oogonia and gemmae commenced in about 70 and 95 h, respectively, in complete media. When 0.2 mM KCl was added to cultures 95 h or older that were grown in chloride deficient media, oogonium or gemma production was initiated in 20–25 or 10–17 h, respectively. Germination of gemmae, mycelial growth (gain in dry matter), and culture pH were not influenced significantly by the chloride deficiences that prevented sexual and asexual reproduction.Key words: chloride requirement, soilborne plant pathogen, reproduction of fungi, sporangia, hyphal swellings.
Phenotypic characterization and virulence of a sae−agr− mutant of Staphylococcus aureus
Authors: Giraudo Ana T. , Rampone Horacio , Calzolari Aldo , Nagel Rosa
Published: February 2010

A sae::Tn551 agr::tetM double mutant was constructed and characterized. The production of several exoproteins (e.g., β-hemolysin, DNase, and proteases) by this mutant was determined and found to be lower than the already diminished production of either isogenic single mutant sae− or agr−. The double mutant also showed, like the agr− mutant, null production of α- and δ-hemolysins and diminished levels of lipase. The reduced levels of many exoproteins in the double mutant as compared with their already diminished levels in either single mutant suggest that there is an additive or synergistic interaction between the two mutations involved, sae− and agr−. However, inactivation of both loci, sae and agr, had a different effect on the two exoproteins that are up regulated in the agr− mutant; thus, coagulase dropped to levels close to the null levels of the sae− parental strain, while extracellular protein A displayed the high levels characteristic of the agr− single mutant. The virulence of the sae−agr− double mutant, determined by intraperitoneal injection in mice, was found to be significantly diminished as compared with that of the sae+agr+ parental strain or the sae−agr+ single mutant.Key words: Staphylococcus aureus, exoprotein expression, sae− mutant, agr− mutant.
Molecular characterization of Spiroplasma citri BR3 lines that differ in transmissibility by the leafhopper Circulifer tenellus
Authors: Fletcher Jacqueline , Shaw Mary E. , Baker Ginger R. , Dugan Karla J. , Ye Fengchun , Sha YeHsiung , Zuck Paul D. , Myers G. Douglas
Published: February 2010

Four lines of Spiroplasma citri strain BR3, derived from different maintenance conditions over several years and differing in their ability to be transmitted by the beet leafhopper Circulifer tenellus were characterized. The lines included BR3-T (transmissible), maintained in turnip by leafhopper transmission; BR3-G (now nontransmissible), maintained in plants by periodic graft transmission; BR3-P (now rarely transmissible), subcultured in artificial medium over 130 times; and BR3-M (transmissible), subcultured 43 times. Although all four lines had similar overall protein profiles, the two transmissible lines each contained two proteins missing in the non- or rarely transmitted lines. In addition, one protein was unique to BR3-M and another was unique to BR3-P. Spiralin, a major S. citri membrane protein, had dual mobility in line BR3-G only. Patterns of extrachromosomal DNA and restricted total DNA also were similar, although differences occurred among the four lines. The genome of line BR3-G was larger than those of the other lines and unique restriction bands occurred in this line. Protein and DNA profiles of six to eight individual clones of each line also were compared. Protein patterns within each clone were indistinguishable except for a difference in the migration rate of spiralin in clones of BR3-G. Restricted total DNA showed differential patterns among clones of each line, possibly reflecting differences in extrachromosomal DNA. Molecular differences among the spiroplasma lines may reflect the selection pressures of the different environments in which they were maintained and suggest genes and proteins that may be involved in the biological phenotypes of these lines.Key words: spiroplasma, Mollicutes, insect transmission.
Purification and characterization of urease fromSchizosaccharomyces pombe
Authors: Lubbers Mark W. , Rodriguez Susan B. , Honey Neville K. , Thornton Roy J.
Published: February 2010

The urease from the ascomycetous fission yeast Schizosaccharomyces pombe was purified about 4000-fold (34% yield) to homogeneity by acetone precipitation, ammonium sulfate precipitation, DEAE-Sepharose ion-exchange column chromatography, and if required, Mono-Q ion-exchange fast protein liquid chromatography. The enzyme was intracellular and only one species of urease was detected by nondenaturing polyacrylamide gel electrophoresis (PAGE). The native enzyme had a Mrof 212 kDa (Sepharose CL6B-200 gel filtration) and a single subunit was detected with a Mrof 102 kDa (PAGE with sodium dodecyl sulfate). The subunit stoichiometry was not specifically determined, but the molecular mass estimations indicate that the undissociated enzyme may be a dimer of identical subunits. The specific activity was 700–800 μmol urea∙min−1∙mg protein−1, the optimum pH for activity was 8.0, and the Kmfor urea was 1.03 mM. The sequence of the amino terminus was Met-Gln-Pro-Arg-Glu-Leu-His-Lys-Leu-Thr-Leu-His-Gln-Leu-Gly-Ser-Leu-Ala and the sequence of two tryptic peptides of the enzyme were Phe-Ile-Glu-Thr-Asn-Glu-Lys and Leu-Tyr-Ala-Pro-Glu-Asn-Ser-Pro-Gly-Phe-Val-Glu-Val-Leu-Glu-Gly-Glu-Ile-Glu-Leu-Leu-Pro-Asn-Leu-Pro. The N-terminal sequence and physical and kinetic properties indicated that S. pombe urease was more like the plant enzymes than the bacterial ureases.Key words: urease, Schizosaccharomyces pombe, fission yeast, ascomycetous yeast.
Distribution, activity, and diversity of heterotrophic nitrifiers originating from East Pacific deep-sea hydrothermal vents
Authors: Mével Genevieve , Faidy Claudette , Prieur Daniel
Published: February 2010

Nitrifiers (bacteria, n = 160) were enriched and isolated from samples of hydrothermal waters, sediments, invertebrate tissues, and chimney rocks collected from two East Pacific deep-sea hydrothermal vents (2000 m): the 13°N site and the Guaymas Basin. They were nitrite producers and seemed be widely and uniformly distributed in various parts of hydrothermal ecosystem. These bacteria grew and nitrified better heterotrophically than autotrophically and they possessed characteristics of heterotrophic nitrifiers. All were aerobic, mesophilic gram-negative rods with a unfermentative metabolism and 88% were nitrate reducers or denitrifiers. They were characterized by a high physiological and nutritional diversity, and because of their ability to ammonify, nitrify, and reduce nitrate, they could largely contribute to the nitrogen cycle in hydrothermal sites.Key words: hydrothermal vents, heterotrophic bacteria, nitrifying activity.
Factors affecting yields of α hormones of Phytophthora parasitica obtained by adsorption
Authors: Chern L. L. , Ko W. H. , Tang C. S.
Published: February 2010

The conditions for improved yields of the α hormones of Phytophthora parasitica compared with those of the original technique established by Ko are described. Both hormones al and α2 were adsorbed on the Millipore filter by exposing both sides of the filter directly to freshly inoculated 40% V-8 agar blocks for 4 days. Usage of polycarbonate membrane in the original method to maintain sterility of the filter was replaced by sterilization of the filter with ethanol. Among five organic solvents tested, 95% ethanol was the most effective in extracting both α hormones from the Millipore filter. A 1-min shaking of the hormone-loaded filter with 95% ethanol was sufficient to extract ca. 95% of hormones α1 and α2. Results on the solubility properties suggest that the α hormones of Phytophthora parasitica are lipid-like natural products and that hormone α2 is more polar than hormone α1.Key words: sex hormone, hormone production, sexual reproduction, Phytophthora parasitica.
Modulation of carbohydrate-binding capacities and attachment ability ofBradyrhizobiumsp. (Lupinus) to white lupin roots
Authors: Wisniewski J.-P. , Delmotte F. M.
Published: February 2010

The attachment of Bradyrhizobium sp. (Lupinus) strain MSDJ718 to excised roots of white lupin was examined. Maximal attachment occurred at early to middle log phases of bacterial growth. This binding was pH dependent, with an optimal value reached at 6.6. Irrespective of the culture age, the attachment was strongly affected by the calcium concentration of the growth medium: a Ca2+limitation in the Bergersen medium led to optimal attachment of the bacteria. When L-fucose was added during the attachment assay, a significant inhibition was observed. The binding was stimulated when bacteria were cultivated with lupin root extracts, genistein and genistin (two lupin isoflavonoids), and some monosaccharides. In addition, with a spectrofluorimetric method using fluoresceinylated neoglycoproteins, it was shown that the increase of the attachment of bacteria to host cells was correlated to the increase of the L-fucoside binding capacity of the rhizobial cells. Taken together, the results obtained in the present study evidenced a possible role of the L-fucose specific bacterial lectin previously described in the Rhizobium – lupin host cell recognition.Key words: rhizobia, lupin, attachment, bacterial lectin, root extracts.
Selective enumeration of aromatic and aliphatic hydrocarbon degrading bacteria by a most-probable-number procedure
Authors: Wrenn Brian A. , Venosa Albert D.
Published: February 2010

A most-probable-number (MPN) procedure was developed to separately enumerate aliphatic and aromatic hydrocarbon degrading bacteria, because most of the currently available methods are unable to distinguish between these two groups. Separate 96-well microtiter plates are used to estimate the sizes of these two populations. The alkane-degrader MPN method uses hexadecane as the selective growth substrate and positive wells are detected by reduction of iodonitrotetrazolium violet, which is added after incubation for 2 weeks at 20 °C. Polycyclic aromatic hydrocarbon degraders are grown on a mixture of phenanthrene, anthracene, fluorene, and dibenzothiophene in a second plate. Positive wells turn yellow to greenish-brown from accumulation of the partial oxidation products of the aromatic substrates and they can be scored after a 3-week incubation period. These MPN procedures are accurate and selective. For pure cultures, heterotrophic plate counts on a nonselective medium and the appropriate MPN procedure provide similar estimates of the population density. Bacteria that cannot grow on the selective substrates do not produce false positive responses even when the inoculum density is very high. Thus, this method, which is simple enough for use in the field, provides reliable estimates for the density and composition of hydrocarbon-degrading microbial populations.Key words: most probable number, polycyclic aromatic hydrocarbon, alkane, hydrocarbon, bacteria.
Isolation and characterization of a gene encoding a chymotrypsin-like serine protease fromStreptomyces lividans66
Authors: Binnie Craig , Liao Linda , Walczyk Eva , Malek Lawrence T.
Published: February 2010

A gene encoding a Streptomyces lividans homologue of the chymotrypsin-like serine protease (SAM-P20) of Streptomyces albogriseolus was isolated using the Streptomyces griseus prtB gene as a hybridization probe. Nucleotide sequence analysis of a representative clone uncovered the possible presence of a sequence of 900 nucleotides encoding 300 amino acids, including a putative "prepro" region of 115 amino acids. Alignment of the predicted amino acid sequence of this putative gene with other members of the family of Streptomyces extracellular chymotrypsin-like proteases indicated a high degree of homology in all cases, especially with the SAM-P20 protease. This gene product has been identified as the second member of a potentially larger family of SAL (SAM-P20-like) proteases in S. lividans 66. Keywords: Streptomyces, protease, chymotrypsin.
Purification and properties of aromatic amino acid aminotransferases fromAzospirillum brasilenseUAP 14 strain
Authors: Soto-Urzua Lucia , Xochinua-Corona Yolanda G. , Flores-Encarnacion Marcos , Baca Beatriz E.
Published: February 2010

The purification and characterization of AAT1, one of two aromatic amino acid aminotransferase (EC 2.6.1.57) in Azospirillum brasilense, is described. Purified AAT1 had a subunit mass of 33 kDa and a nondenatured molecular mass of 66 kDa, suggesting a dimeric structure. Other properties include a pI of 5.04, an optimum temperature of 45 °C, and optimum pH of 8.5. AAT1 utilized all aromatic amino acids, the L-tryptophan derivatives such as L-5-methyl tryptophan and L-flourtryptophan, and L-histidine. The apparent Kmvalues for L-tyrosine, L-phenylalanine, and L-tryptophan were 0.19, 0.43, and 1.05 mM, respectively. The enzyme was competive inhibited by indole-3-pyruvic acid with a Kiof 0.17 mM.Key words: aromatic aminotransferase, Azospirillum brasilense, indole acetic acid production.
A comparative study on the conditions of growth and sporulation of three strains ofClostridium petfringenstype A
Authors: Decaudin Mathilde , Tholozan Jean-Luc
Published: February 2010

Different conditions of growth and sporulation of a strain of Clostridium perfringens type A (NCTC 8798) and two derived mutant strains, the lysozyme-germination dependent strain 8-6 and the revertant strain R3, have been determined. No sporulation was detected for the three strains in the Duncan and Strong (DS) medium; 100% sporulation was routinely obtained for the two mutant strains in the defined (D) medium. Factors promoting in vitro sporulation of C. perfringens type A were assayed: the volume of the culture, the type of preculture, and the addition of lysozyme in precultures. The paper also provides additional information on growth and sporulation of the mutant strains 8-6 and R3. Glucose concentrations up to 11 mM produced high percentages of sporulation. However, strain R3 still sporulated at 20% with 56 mM of glucose. A high volume of D medium led to slow growth kinetics and favoured sporulation. Faster kinetics of growth and the best percentage of sporulation were obtained with a young inoculum of the two mutant strains. On the other hand, the type of medium in the preculture (fluid thioglycollate (FTG) or basal carbonate yeast trypticase (BCYT)) did not influence the percentage of sporulation. However, while strain R3 was not affected by the addition of lysozyme in D medium, kinetics of growth were strongly influenced by this addition in strain 8-6, and the percentage of sporulation increased with a preculture in FTG medium and decreased when BCYT medium was used.Key words: Clostridium perfringens, medium, growth, sporulation.
Physical mapping of several heat-shock genes inPseudomonas aeruginosaand the cloning of themopA(GroEL) gene
Authors: Farinha Mark A. , Mockett Robin , Went Catherine J. , Jardine Stephanie , Naczynski Lina M. , Kropinski Andrew M.
Published: February 2010

Using a series of oligonucleotides synthesized on the basis of conserved nucleotide or amino acid motifs in heat-shock genes/proteins, we have physically mapped the dnaK, lon, and hptG genes of Pseudomonas aeruginosa. Hybridization data suggest that there is a single copy of the mopBA (GroES/GroEL) operon but several additional copies of mopA. In addition, the map coordinates for the rpoD, rpoS, and rpoH genes were determined. The mopA gene from the mopBA operon was cloned and sequenced. The protein product of this gene showed 79% amino acid identity to the Escherichia coli GroEL and 98% identity to the GroEL sequence from P. aeruginosa ATCC 27853. A number of discrepancies were found with the latter sequence.Key words: Pseudomonas, heat shock, GroEL, DnaK, pulsed-field gel electrophoresis.
Glycoconjugate and lipid components ofRhizobium"hedysari" IS123, a root-nodule symbiont of the stress-tolerant legumeHedysarum coronarium
Authors: Orgambide Guy G. , Philip-Hollingsworth Saleela , Dazzo Frank B. , Tola Elisabetta , Squartini Andrea , Nuti Marco P. , Cedergren Robert A. , Hollingsworth Rawle I.
Published: February 2010

We have examined the diversity of glycoconjugates and cellular lipids of Rhizobium "hedysari" IS123, a bacterial symbiont that specifically nodulates the drought-tolerant forage legume Hedysarum coronarium. IS123 develops a complete capsule consisting of a loose fibrillar network of ruthenium-red-staining acidic polymers and produces two different exopolysaccharides (EPS). EPS-A contains glucose, galactose, mannose, and a noncarbohydrate substitution tentatively identified as a lactyl ester. The composition of EPS-B, which includes glucose and galactose, as well as O-acetyl, pyruvyl, and succinyl substituents, is very similar to that of the EPS-II described in Rhizobium meliloti. IS123 also makes an O-acetylated heterooligosaccharide and unsubstituted β-1,2-glucans. The cellular fatty acid composition of IS123 is dominated by 18:1 and also includes 14:0, 16:0, 16:1, 3OH-16:0, 17:0Δ, 18:0, 3OH-18:0, and 19:0Δ. Phospholipids of IS123 include phosphatidylethanolamine, N-methyl phosphatidylethanolamine, N,N-dimethyl phosphatidylethanolamine, phosphatidylcholine, and phosphatidylglycerol.Key words: Rhizobium, capsule, exopolysaccharides, oligosaccharides, lipids.
An overview of microbiologically influenced corrosion of metals and alloys used in the storage of nuclear wastes
Authors: Little Brenda , Wagner Patricia
Published: February 2010

Microbiologically influenced corrosion has been documented for all metals and alloys currently in use or proposed for packaging nuclear wastes, with the possible exception of titanium. Selection of a material that will provide a reliable physical barrier to prevent leakage of radionuclides for 103–106 years will depend on a thorough understanding of mechanisms for microbiologically influenced corrosion; the geological, microbiological, and chemical environments of storage sites; and their evolution over time.Key words: nuclear waste storage, microbiologically influenced corrosion, biofilms.
Isolation and characterization of resin acid degrading bacteria found in effluent from a bleached kraft pulp mill
Authors: Morgan C. A. , Wyndham R. C.
Published: February 2010

Thirteen resin acid degrading bacteria enriched on abietic or dehydroabietic acids were isolated from waste water from the aerated stabilization basin of a bleached kraft pulp mill. Standard biochemical tests were used to characterize each isolate. Each isolate was tested for its ability to degrade six abietane- and pimarane-type resin acids. Resin acid concentrations were determined by high pressure liquid chromatography and UV absorbance. Cluster analysis based on phenotypic characteristics identified two distinct clusters of degraders that differed in their ability to utilize carbohydrates as carbon sources. Fatty acid methyl ester analysis of representative isolates from each cluster identified A19-6a and D11-13 as Comamonas and Alcaligenes species, respectively. To determine genotypic relatedness, enterobacterial repetitive intergenic consensus sequences were used to amplify genomic DNA fragments from 10 isolates. These results supported the phenotypic analysis for all isolates tested except A19-5 and A19-6b. These two organisms were clustered closely together based on phenotype but had distinctly different banding patterns, suggesting that they are not related genotypically. All isolates degraded a subset of the six resin acid congeners. Isolates A19-3, A19-6a, A19-6b, and D11-37 were the most effective at degrading all six congeners.Key words: biodegradation, resin acids, metabolism, Comamonas, Alcaligenes.
Use of continuous culture to screen for lipase-producing microorganisms and interesterification of butter fat by lipase isolates
Authors: Pabai F. , Kermasha S. , Morin A.
Published: February 2010

The continuous cultivation technique was used to investigate the screening for lipase-producing microorganisms from four commercial starters suitable for the degradation of domestic wastes. Using this technique, three strains of lipase-producing bacteria were isolated and identified: Pantoea agglomerons (BB96CC1, BB168CC2) and Pseudomonas fluorescens (BW96CC1). In addition, butter fat induced more lipase production when present in the growth medium. Interesterification of butter fat triacylglycerols by enzymatic extracts of the isolated strains of microorganisms resulted in an appreciable interesterification yield, implying that hydrolysis was suppressed and interesterification of butter fat triacylglycerols was maximized in a microemulsion free-cosurfactant system.Key words: continuous culture, screening, interesterification, lipases, butter fat.