bluuebunny/crossref_metadata_2025_split huggingface dataset:
Cultivation of Denitrifying Bacteria: Optimization of Isolation Conditions and Diversity Study
Authors: Heylen Kim , Vanparys Bram , Wittebolle Lieven , Verstraete Willy , Boon Nico , De Vos Paul 
Published: April 2006

ABSTRACTAn evolutionary algorithm was applied to study the complex interactions between medium parameters and their effects on the isolation of denitrifying bacteria, both in number and in diversity. Growth media with a pH of 7 and a nitrogen concentration of 3 mM, supplemented with 1 ml of vitamin solution but not with sodium chloride or riboflavin, were the most successful for the isolation of denitrifiers from activated sludge. The use of ethanol or succinate as a carbon source and a molar C/N ratio of 2.5, 20, or 25 were also favorable. After testing of 60 different medium parameter combinations and comparison with each other as well as with the standard medium Trypticase soy agar supplemented with nitrate, three growth media were highly suitable for the cultivation of denitrifying bacteria. All evaluated isolation conditions were used to study the cultivable denitrifier diversity of activated sludge from a municipal wastewater treatment plant. One hundred ninety-nine denitrifiers were isolated, the majority of which belonged to theBetaproteobacteria(50.4%) and theAlphaproteobacteria(36.8%). Representatives ofGammaproteobacteria(5.6%),Epsilonproteobacteria(2%), andFirmicutes(4%) and one isolate of theBacteroideteswere also found. This study revealed a much more diverse denitrifying community than that previously described in cultivation-dependent research on activated sludge.
Assessment of the Roles of LuxS,
S
-Ribosyl Homocysteine, and Autoinducer 2 in Cell Attachment during Biofilm Formation by
Listeria monocytogenes
EGD-e
Authors: Challan Belval Sylvain , Gal Laurent , Margiewes Sylvain , Garmyn Dominique , Piveteau Pascal , Guzzo Jean 
Published: April 2006

ABSTRACT

LuxS is responsible for the production of autoinducer 2 (AI-2), which is involved in the quorum-sensing response of
Vibrio harveyi
. AI-2 is found in several other gram-negative and gram-positive bacteria and is therefore considered a good candidate for an interspecies communication signal molecule. In order to determine if this system is functional in the gastrointestinal pathogen
Listeria monocytogenes
EGD-e, an AI-2 bioassay was performed with culture supernatants. The results indicated that this bacterium produces AI-2 like molecules. A potential ortholog of
V. harveyi luxS
, lmo1288, was found by performing sequence similarity searches and complementation experiments with
Escherichia coli
DH5α, a
luxS
null strain. lmo1288 was found to be a functional
luxS
ortholog involved in AI-2 synthesis. Indeed, interruption of lmo1288 resulted in loss of the AI-2 signal. Although no significant differences were observed between Lux1 and EGD-e with regard to planktonic growth (at 10°C, 15°C, 25°C, and 42°C), swimming motility, and phospholipase and hemolytic activity, biofilm culture experiments showed that under batch conditions between 25% and 58% more Lux1 cells than EGD-e cells were attached to the surface depending on the incubation time. During biofilm growth in continuous conditions after 48 h of culture, Lux1 biofilms were 17 times denser than EGD-e biofilms. Finally, our results showed that Lux1 accumulates more
S
-adenosyl homocysteine (SAH) and
S
-ribosyl homocysteine (SRH) in culture supernatant than the parental strain accumulates and that SRH, but not SAH or AI-2, is able to modify the number of attached cells.
The Locus Coding for the 3-Nitrobenzoate Dioxygenase of
Comamonas
sp. Strain JS46 Is Flanked by IS
1071
Elements and Is Subject to Deletion and Inversion Events
Authors: Providenti Miguel A. , Shaye Rachel E. , Lynes Krista D. , McKenna Neil T. , O'Brien Jason M. , Rosolen Sarah , Wyndham R. Campbell , Lambert Iain B. 
Published: April 2006

ABSTRACT

In
Comamonas
sp. strain JS46, 3-nitrobenzoate (3Nba) is initially oxidized at the 3,4 position by a dioxygenase, which results in release of nitrite and production of protocatechuate. The locus coding for the 3Nba dioxygenase (designated
mnb
, for
m-n
itro
b
enzoate) was mobilized from strain JS46 using a plasmid capture method, cloned, and sequenced. The 3Nba dioxygenase (MnbA) is a member of the phthalate family of aromatic oxygenases. An open reading frame designated
mnbB
that codes for an NAD(P)H-dependent class IA aromatic oxidoreductase is downstream of
mnbA
. MnbB is tentatively identified as the oxidoreductase that transfers reducing equivalents to MnbA in strain JS46. The
mnb
locus is flanked by IS
1071
elements. The upstream element is interrupted by a novel insertion sequence designated IS
Csp1
, and the transposase genes of the flanking insertion elements are transcribed in the direction opposite the direction of
mnbA
transcription. Spontaneous deletion of
mnb
occurs because of homologous recombination between the directly repeated flanking IS
1071
elements. In addition, in ∼0.007 to 0.2% of any population of JS46 cells growing on 3Nba, alternative orientations of
mnb
relative to the flanking IS
1071
elements are detected. These alternative forms are the result of inversions of
mnb
and the flanking IS
1071
elements. Inversions appear to occur because of homologous recombination between the inverted repeats that flank the IS
1071
elements.
Genes ofEscherichia coliO157:H7 That Are Involved in High-Pressure Resistance
Authors: Malone Aaron S. , Chung Yoon-Kyung , Yousef Ahmed E. 
Published: April 2006

ABSTRACTSeventeenEscherichia coliO157:H7 strains were treated with ultrahigh pressure at 500 MPa and 23 ± 2°C for 1 min. This treatment inactivated 0.6 to 3.4 log CFU/ml, depending on the strain. The diversity of these strains was confirmed by pulsed-field gel electrophoresis (PFGE) analysis, and there was no apparent association between PFGE banding patterns and pressure resistance. The pressure-resistant strainE. coliO157:H7 EC-88 (0.6-log decrease) and the pressure-sensitive strain ATCC 35150 (3.4-log decrease) were treated with a sublethal pressure (100 MPa for 15 min at 23 ± 2°C) and subjected to DNA microarray analysis using anE. coliK-12 antisense gene chip. High pressure affected the transcription of many genes involved in a variety of intracellular mechanisms of EC-88, including the stress response, the thiol-disulfide redox system, Fe-S cluster assembly, and spontaneous mutation. Twenty-fourE. coliisogenic pairs with mutations in the genes regulated by the pressure treatment were treated with lethal pressures at 400 MPa and 23 ± 2°C for 5 min. The barotolerance of the mutants relative to that of the wild-type strains helped to explain the results obtained by DNA microarray analysis. This study is the first report to demonstrate that the expression of Fe-S cluster assembly proteins and the fumarate nitrate reductase regulator decreases the resistance to pressure, while sigma factor (RpoE), lipoprotein (NlpI), thioredoxin (TrxA), thioredoxin reductase (TrxB), a trehalose synthesis protein (OtsA), and a DNA-binding protein (Dps) promote barotolerance.
Comparison of Vertical Distributions of Prokaryotic Assemblages in the Anoxic Cariaco Basin and Black Sea by Use of Fluorescence In Situ Hybridization
Authors: Lin Xueju , Wakeham Stuart G. , Putnam Isabell F. , Astor Yrene M. , Scranton Mary I. , Chistoserdov Andrei Y. , Taylor Gordon T. 
Published: April 2006

ABSTRACTIndividual prokaryotic cells from two major anoxic basins, the Cariaco Basin and the Black Sea, were enumerated throughout their water columns using fluorescence in situ hybridization (FISH) with the fluorochrome Cy3 or horseradish peroxidase-modified oligonucleotide probes. For both basins, significant differences in total prokaryotic abundance and phylogenetic composition were observed among oxic, anoxic, and transitional (redoxcline) waters. Epsilon-proteobacteria,Crenarchaeota, andEuryarchaeotawere more prevalent in the redoxclines, where previous studies reported high rates of chemoautotrophic production relative to those in waters above and below the redoxclines. Relative abundances ofArchaeain both systems varied between 1% and 28% of total prokaryotes, depending on depth. The prokaryotic community composition varied between the two anoxic basins, consistent with distinct geochemical and physical conditions. In the Black Sea, the relative contributions of group ICrenarchaeota(median, 5.5%) to prokaryotic communities were significantly higher (P< 0.001;n= 20) than those of group IIEuryarchaeota(median, 2.9%). In contrast, their proportions were nearly equivalent in the Cariaco Basin. Beta-proteobacteria were unexpectedly common throughout the Cariaco Basin's water column, accounting for an average of 47% of 4′,6′-diamidino-2-phenylindole (DAPI)-stained cells. This group was below the detection limit (<1%) in the Black Sea samples. Compositional differences between basins may reflect temporal variability in microbial populations and/or systematic differences in environmental conditions and the populations for which they select.
Pseudomonad Cyclopentadecanone Monooxygenase Displaying an Uncommon Spectrum of Baeyer-Villiger Oxidations of Cyclic Ketones
Authors: Iwaki Hiroaki , Wang Shaozhao , Grosse Stephan , Bergeron Hélène , Nagahashi Ayako , Lertvorachon Jittiwud , Yang Jianzhong , Konishi Yasuo , Hasegawa Yoshie , Lau Peter C. K. 
Published: April 2006

ABSTRACTBaeyer-Villiger monooxygenases (BVMOs) are biocatalysts that offer the prospect of high chemo-, regio-, and enantioselectivity in the organic synthesis of lactones or esters from a variety of ketones. In this study, we have cloned, sequenced, and overexpressed inEscherichia colia new BVMO, cyclopentadecanone monooxygenase (CpdB or CPDMO), originally derived fromPseudomonassp. strain HI-70. The 601-residue primary structure of CpdB revealed only 29% to 50% sequence identity to those of known BVMOs. A new sequence motif, characterized by a cluster of charged residues, was identified in a subset of BVMO sequences that contain an N-terminal extension of ∼60 to 147 amino acids. The 64-kDa CPDMO enzyme was purified to apparent homogeneity, providing a specific activity of 3.94 μmol/min/mg protein and a 20% yield. CPDMO is monomeric and NADPH dependent and contains ∼1 mol flavin adenine dinucleotide per mole of protein. A deletion mutant suggested the importance of the N-terminal 54 amino acids to CPDMO activity. In addition, a Ser261Ala substitution in a Rossmann fold motif resulted in an improved stability and increased affinity of the enzyme towards NADPH compared to the wild-type enzyme (Km= 8 μM versusKm= 24 μM). Substrate profiling indicated that CPDMO is unusual among known BVMOs in being able to accommodate and oxidize both large and small ring substrates that include C11to C15ketones, methyl-substituted C5and C6ketones, and bicyclic ketones, such as decalone and β-tetralone. CPDMO has the highest affinity (Km= 5.8 μM) and the highest catalytic efficiency (kcat/Kmratio of 7.2 × 105M−1s−1) toward cyclopentadecanone, hence the Cpd designation. A number of whole-cell biotransformations were carried out, and as a result, CPDMO was found to have an excellent enantioselectivity (E> 200) as well as 99%S-selectivity toward 2-methylcyclohexanone for the production of 7-methyl-2-oxepanone, a potentially valuable chiral building block. Although showing a modest selectivity (E= 5.8), macrolactone formation of 15-hexadecanolide from the kinetic resolution of 2-methylcyclopentadecanone using CPDMO was also demonstrated.
Effects of Preculturing Conditions on Lag Time and Specific Growth Rate of
Enterobacter sakazakii
in Reconstituted Powdered Infant Formula
Authors: Kandhai M. C. , Reij M. W. , Grognou C. , van Schothorst M. , Gorris L. G. M. , Zwietering M. H. 
Published: April 2006

ABSTRACT

Enterobacter sakazakii
can be present, although in low levels, in dry powdered infant formulae, and it has been linked to cases of meningitis in neonates, especially those born prematurely. In order to prevent illness, product contamination at manufacture and during preparation, as well as growth after reconstitution, must be minimized by appropriate control measures. In this publication, several determinants of the growth of
E. sakazakii
in reconstituted infant formula are reported. The following key growth parameters were determined: lag time, specific growth rate, and maximum population density. Cells were harvested at different phases of growth and spiked into powdered infant formula. After reconstitution in sterile water,
E. sakazakii
was able to grow at temperatures between 8 and 47°C. The estimated optimal growth temperature was 39.4°C, whereas the optimal specific growth rate was 2.31 h
−1
. The effect of temperature on the specific growth rate was described with two secondary growth models. The resulting minimum and maximum temperatures estimated with the secondary Rosso equation were 3.6°C and 47.6°C, respectively. The estimated lag time varied from 83.3 ± 18.7 h at 10°C to 1.73 ± 0.43 h at 37°C and could be described with the hyperbolic model and reciprocal square root relation. Cells harvested at different phases of growth did not exhibit significant differences in either specific growth rate or lag time. Strains did not have different lag times, and lag times were short given that the cells had spent several (3 to 10) days in dry powdered infant formula. The growth rates and lag times at various temperatures obtained in this study may help in calculations of the period for which reconstituted infant formula can be stored at a specific temperature without detrimental impact on health.
A Suppression Subtractive Hybridization Approach Reveals Niche-Specific Genes That May Be Involved in Predator Avoidance in Marine
Synechococcus
Isolates
Authors: Jones H. , Ostrowski M. , Scanlan D. J. 
Published: April 2006

ABSTRACT

Picocyanobacteria of the genus
Synechococcus
are important contributors to marine primary production and are ubiquitous in the world's oceans. This genus is genetically diverse, and at least 10 discrete lineages or clades have been identified phylogenetically. However, little if anything is known about the genetic attributes which characterize particular lineages or are unique to specific strains. Here, we used a suppression subtractive hybridization (SSH) approach to identify strain- and clade-specific genes in two well-characterized laboratory strains,
Synechococcus
sp. strain WH8103 (clade III) and
Synechococcus
sp. strain WH7803 (clade V). Among the genes that were identified as potentially unique to each strain were genes encoding proteins that may be involved in specific predator avoidance, including a glycosyltransferase in strain WH8103 and a permease component of an ABC-type polysaccharide/polyol phosphate export system in WH7803. During this work the genome of one of these strains, WH7803, became available. This allowed assessment of the number of false-positive sequences (i.e., sequences present in the tester genome) present among the SSH-enriched sequences. We found that approximately 9% of the WH8103 sequences were potential false-positive sequences, which demonstrated that caution should be used when this technology is used to assess genomic differences in genetically similar bacterial strains.
Specific Bacterial, Archaeal, and Eukaryotic Communities in Tidal-Flat Sediments along a Vertical Profile of Several Meters
Authors: Wilms Reinhard , Sass Henrik , Köpke Beate , Köster Jürgen , Cypionka Heribert , Engelen Bert 
Published: April 2006

ABSTRACT

The subsurface of a tidal-flat sediment was analyzed down to 360 cm in depth by molecular and geochemical methods. A community structure analysis of all three domains of life was performed using domain-specific PCR followed by denaturing gradient gel electrophoresis analysis and sequencing of characteristic bands. The sediment column comprised horizons easily distinguishable by lithology that were deposited in intertidal and salt marsh environments. The pore water profile was characterized by a subsurface sulfate peak at a depth of about 250 cm. Methane and sulfate profiles were opposed, showing increased methane concentrations in the sulfate-free layers. The availability of organic carbon appeared to have the most pronounced effect on the bacterial community composition in deeper sediment layers. In general, the bacterial community was dominated by fermenters and syntrophic bacteria. The depth distribution of methanogenic archaea correlated with the sulfate profile and could be explained by electron donor competition with sulfate-reducing bacteria. Sequences affiliated with the typically hydrogenotrophic
Methanomicrobiales
were present in sulfate-free layers. Archaea belonging to the
Methanosarcinales
that utilize noncompetitive substrates were found along the entire anoxic-sediment column. Primers targeting the eukaryotic 18S rRNA gene revealed the presence of a subset of archaeal sequences in the deeper part of the sediment cores. The phylogenetic distance to other archaeal sequences indicates that these organisms represent a new phylogenetic group, proposed as “tidal-flat cluster 1.” Eukarya were still detectable at 360 cm, even though their diversity decreased with depth. Most of the eukaryotic sequences were distantly related to those of grazers and deposit feeders.
Geobacter lovleyi
sp. nov. Strain SZ, a Novel Metal-Reducing and Tetrachloroethene-Dechlorinating Bacterium
Authors: Sung Youlboong , Fletcher Kelly E. , Ritalahti Kirsti M. , Apkarian Robert P. , Ramos-Hernández Natalia , Sanford Robert A. , Mesbah Noha M. , Löffler Frank E. 
Published: April 2006

ABSTRACT

A bacterial isolate, designated strain SZ, was obtained from noncontaminated creek sediment microcosms based on its ability to derive energy from acetate oxidation coupled to tetrachloroethene (PCE)-to-
cis
-1,2-dichloroethene (
cis
-DCE) dechlorination (i.e., chlororespiration). Hydrogen and pyruvate served as alternate electron donors for strain SZ, and the range of electron acceptors included (reduced products are given in brackets) PCE and trichloroethene [
cis
-DCE], nitrate [ammonium], fumarate [succinate], Fe(III) [Fe(II)], malate [succinate], Mn(IV) [Mn(II)], U(VI) [U(IV)], and elemental sulfur [sulfide]. PCE and soluble Fe(III) (as ferric citrate) were reduced at rates of 56.5 and 164 nmol min
−1
mg of protein
−1
, respectively, with acetate as the electron donor. Alternate electron acceptors, such as U(VI) and nitrate, did not inhibit PCE dechlorination and were consumed concomitantly. With PCE, Fe(III) (as ferric citrate), and nitrate as electron acceptors, H
2
was consumed to threshold concentrations of 0.08 ± 0.03 nM, 0.16 ± 0.07 nM, and 0.5 ± 0.06 nM, respectively, and acetate was consumed to 3.0 ± 2.1 nM, 1.2 ± 0.5 nM, and 3.6 ± 0.25 nM, respectively. Apparently, electron acceptor-specific acetate consumption threshold concentrations exist, suggesting that similar to the hydrogen threshold model, the measurement of acetate threshold concentrations offers an additional diagnostic tool to delineate terminal electron-accepting processes in anaerobic subsurface environments. Genetic and phenotypic analyses classify strain SZ as the type strain of the new species,
Geobacter lovleyi
sp. nov., with
Geobacter
(formerly
Trichlorobacter
)
thiogenes
as the closest relative. Furthermore, the analysis of 16S rRNA gene sequences recovered from PCE-dechlorinating consortia and chloroethene-contaminated subsurface environments suggests that
Geobacter lovleyi
belongs to a distinct, dechlorinating clade within the metal-reducing
Geobacter
group. Substrate versatility, consumption of electron donors to low threshold concentrations, and simultaneous reduction of electron acceptors suggest that strain SZ-type organisms have desirable characteristics for bioremediation applications.
Chlorophenol Hydroxylases Encoded by Plasmid pJP4 Differentially Contribute to Chlorophenoxyacetic Acid Degradation
Authors: Ledger T. , Pieper D. H. , González B. 
Published: April 2006

ABSTRACT

Phenoxyalkanoic compounds are used worldwide as herbicides.
Cupriavidus necator
JMP134(pJP4) catabolizes 2,4-dichlorophenoxyacetate (2,4-D) and 4-chloro-2-methylphenoxyacetate (MCPA), using
tfd
functions carried on plasmid pJP4. TfdA cleaves the ether bonds of these herbicides to produce 2,4-dichlorophenol (2,4-DCP) and 4-chloro-2-methylphenol (MCP), respectively. These intermediates can be degraded by two chlorophenol hydroxylases encoded by the
tfdB
I
and
tfdB
II
genes to produce the respective chlorocatechols. We studied the specific contribution of each of the TfdB enzymes to the 2,4-D/MCPA degradation pathway. To accomplish this, the
tfdB
I
and
tfdB
II
genes were independently inactivated, and growth on each chlorophenoxyacetate and total chlorophenol hydroxylase activity were measured for the mutant strains. The phenotype of these mutants shows that both TfdB enzymes are used for growth on 2,4-D or MCPA but that TfdB
I
contributes to a significantly higher extent than TfdB
II
. Both enzymes showed similar specificity profiles, with 2,4-DCP, MCP, and 4-chlorophenol being the best substrates. An accumulation of chlorophenol was found to inhibit chlorophenoxyacetate degradation, and inactivation of the
tfdB
genes enhanced the toxic effect of 2,4-DCP on
C. necator
cells. Furthermore, increased chlorophenol production by overexpression of TfdA also had a negative effect on 2,4-D degradation by
C. necator
JMP134 and by a different host,
Burkholderia xenovorans
LB400, harboring plasmid pJP4. The results of this work indicate that codification and expression of the two
tfdB
genes in pJP4 are important to avoid toxic accumulations of chlorophenols during phenoxyacetic acid degradation and that a balance between chlorophenol-producing and chlorophenol-consuming reactions is necessary for growth on these compounds.
Quantitative Real-Time
Legionella
PCR for Environmental Water Samples: Data Interpretation
Authors: Joly Philippe , Falconnet Pierre-Alain , André Janine , Weill Nicole , Reyrolle Monique , Vandenesch François , Maurin Max , Etienne Jerome , Jarraud Sophie 
Published: April 2006

ABSTRACT

Quantitative
Legionella
PCRs targeting the 16S rRNA gene (specific for the genus
Legionella
) and the
mip
gene (specific for the species
Legionella pneumophila
) were applied to a total of 223 hot water system samples (131 in one laboratory and 92 in another laboratory) and 37 cooling tower samples (all in the same laboratory). The PCR results were compared with those of conventional culture. 16S rRNA gene PCR results were nonquantifiable for 2.8% of cooling tower samples and up to 39.1% of hot water system samples, and this was highly predictive of
Legionella
CFU counts below 250/liter. PCR cutoff values for identifying hot water system samples containing >10
3
CFU/liter legionellae were determined separately in each laboratory. The cutoffs differed widely between the laboratories and had sensitivities from 87.7 to 92.9% and specificities from 77.3 to 96.5%. The best specificity was obtained with
mip
PCR. PCR cutoffs could not be determined for cooling tower samples, as the results were highly variable and often high for culture-negative samples. Thus, quantitative
Legionella
PCR appears to be applicable to samples from hot water systems, but the positivity cutoff has to be determined in each laboratory.