bluuebunny/crossref_metadata_2025_split huggingface dataset:
Phylogenetic Relationships of Butyrate-Producing Bacteria from the Human Gut
Authors: Barcenilla Adela , Pryde Susan E. , Martin Jennifer C. , Duncan Sylvia H. , Stewart Colin S. , Henderson Colin , Flint Harry J. 
Published: July 2002

ABSTRACT

Butyrate is a preferred energy source for colonic epithelial cells and is thought to play an important role in maintaining colonic health in humans. In order to investigate the diversity and stability of butyrate-producing organisms of the colonic flora, anaerobic butyrate-producing bacteria were isolated from freshly voided human fecal samples from three healthy individuals: an infant, an adult omnivore, and an adult vegetarian. A second isolation was performed on the same three individuals 1 year later. Of a total of 313 bacterial isolates, 74 produced more than 2 mM butyrate in vitro. Butyrate-producing isolates were grouped by 16S ribosomal DNA (rDNA) PCR-restriction fragment length polymorphism analysis. The results indicate very little overlap between the predominant ribotypes of the three subjects; furthermore, the flora of each individual changed significantly between the two isolations. Complete sequences of 16S rDNAs were determined for 24 representative strains and subjected to phylogenetic analysis. Eighty percent of the butyrate-producing isolates fell within the XIVa cluster of gram-positive bacteria as defined by M. D. Collins et al. (Int. J. Syst. Bacteriol. 44:812–826, 1994) and A. Willems et al. (Int. J. Syst. Bacteriol. 46:195–199, 1996), with the most abundant group (10 of 24 or 42%) clustering with
Eubacterium rectale
,
Eubacterium ramulus
, and
Roseburia cecicola
. Fifty percent of the butyrate-producing isolates were net acetate consumers during growth, suggesting that they employ the butyryl coenzyme A-acetyl coenzyme A transferase pathway for butyrate production. In contrast, only 1% of the 239 non-butyrate-producing isolates consumed acetate.
Structural Changes and Interactions Involved in the Ca
2+
-Triggered Stabilization of the Cell-Bound Cell Envelope Proteinase in
Lactococcus lactis
subsp.
cremoris
SK11
Authors: Exterkate Fred A. 
Published: July 2002

ABSTRACT

The cell-bound cell envelope proteinase (CEP) of the mesophilic cheese-starter organism
Lactococcus lactis
subsp.
cremoris
SK11 is protected from rapid thermal inactivation at 25°C by calcium bound to weak binding sites. The interactions with calcium are believed to trigger reversible structural rearrangements which are coupled with changes in specific activity (F. A. Exterkate and A. C. Alting, Appl. Env. Microbiol. 65:1390–1396, 1999). In order to determine the significance of the rearrangements for CEP stability and the nature of the interactions involved, the effects of the net charge present on the enzyme and of different neutral salts were studied with the stable Ca-loaded CEP, the unstable so-called “Ca-free” CEP and with the Ca-free CEP which was stabilized nonspecifically and essentially in its native conformation by the nonionic additive sucrose. The results suggest that strengthening of hydrophobic interactions is conducive to stabilization of the Ca-free CEP. On the other hand, a hydrophobic effect contributes significantly to the stability of the Ca-loaded CEP; a phased salting-in effect by a chaotropic salt suggests a complex inactivation process of this enzyme due to weakening of hydrophobic interactions and involving an intermediate enzyme species. Moreover, a Ca-triggered increase of a relatively significant hydrophobic effect in the sucrose-stabilized Ca-free CEP occurs. It is suggested that in the Ca-free CEP the absence of both local calcium-mediated backbone rigidification and neutralization of negative electrostatic potentials in the weak Ca-binding sites, and in addition the lack of significant hydrophobic stabilization, increase the relative effectiveness of electrostatic repulsive forces on the protein to an extent that causes the observed instability. The conditions in cheese seem to confer stability upon the cell-bound enzyme; its possible involvement in proteolysis throughout the ripening period is discussed.
Role of Volatile Fatty Acids in Development of the Cecal Microflora in Broiler Chickens during Growth
Authors: van der Wielen Paul W. J. J. , Biesterveld Steef , Notermans Servé , Hofstra Harm , Urlings Bert A. P. , van Knapen Frans 
Published: July 2002

ABSTRACT

It is known that volatile fatty acids can inhibit growth of species of the family
Enterobacteriaceae
in vitro. However, whether these volatile fatty acids affect bacterial populations in the ceca of chickens is unknown. Therefore, a study was conducted to investigate if changes in volatile fatty acids in ceca of broiler chickens during growth affect bacterial populations. Results showed that members of the
Enterobacteriaceae
and enterococci are present in large numbers in 3-day-old broilers and start to decrease when broilers grow older. Lactobacilli are present in large numbers as well in 3-day-old broilers, but they remain stable during the growth of broilers. Acetate, butyrate, and propionate increase from undetectable levels in 1-day-old broilers to high concentrations in 15-day-old broilers, after which they stabilize. Significant negative correlations could be calculated between numbers of
Enterobacteriaceae
and concentrations of undissociated acetate, propionate, and butyrate. Furthermore, pure cultures of
Enterobacteriaceae
isolated from the ceca were grown in the presence of volatile fatty acids. Growth rates and maximal optical density decreased when these strains grew in the presence of increasing volatile fatty acid concentrations. It is concluded that volatile fatty acids are responsible for the reduction in numbers of
Enterobacteriaceae
in the ceca of broiler chickens during growth.
Bacillus anthracis
,
Bacillus cereus
, and
Bacillus thuringiensis
—One Species on the Basis of Genetic Evidence
Authors: Helgason Erlendur , Økstad Ole Andreas , Caugant Dominique A. , Johansen Henning A. , Fouet Agnes , Mock Michéle , Hegna Ida , Kolstø Anne-Brit 
Published: July 2002

ABSTRACT

Bacillus anthracis
,
Bacillus cereus
, and
Bacillus thuringiensis
are members of the
Bacillus cereus
group of bacteria, demonstrating widely different phenotypes and pathological effects.
B. anthracis
causes the acute fatal disease anthrax and is a potential biological weapon due to its high toxicity.
B. thuringiensis
produces intracellular protein crystals toxic to a wide number of insect larvae and is the most commonly used biological pesticide worldwide.
B. cereus
is a probably ubiquitous soil bacterium and an opportunistic pathogen that is a common cause of food poisoning. In contrast to the differences in phenotypes, we show by multilocus enzyme electrophoresis and by sequence analysis of nine chromosomal genes that
B. anthracis
should be considered a lineage of
B. cereus
. This determination is not only a formal matter of taxonomy but may also have consequences with respect to virulence and the potential of horizontal gene transfer within the
B. cereus
group.
Archaeal Population Dynamics during Sequential Reduction Processes in Rice Field Soil
Authors: Lueders Tillmann , Friedrich Michael 
Published: July 2002

ABSTRACT

The population dynamics of
Archaea
after flooding of an Italian rice field soil were studied over 17 days. Anoxically incubated rice field soil slurries exhibited a typical sequence of reduction processes characterized by reduction of nitrate, Fe
3+
, and sulfate prior to the initiation of methane production. Archaeal population dynamics were followed using a dual approach involving molecular sequence retrieval and fingerprinting of small-subunit (SSU) rRNA genes. We retrieved archaeal sequences from four clone libraries (30 each) constructed for different time points (days 0, 1, 8, and 17) after flooding of the soil. The clones could be assigned to known methanogens (i.e.,
Methanosarcinaceae
,
Methanosaetaceae
,
Methanomicrobiaceae
, and
Methanobacteriaceae
) and to novel euryarchaeotal (rice clusters I, II, and III) and crenarchaeotal (rice clusters IV and VI) lineages previously detected in anoxic rice field soil and on rice roots (R. Grosskopf, S. Stubner, and W. Liesack, Appl. Environ. Microbiol. 64:4983–4989, 1998). During the initiation of methanogenesis (days 0 to 17), we detected significant changes in the frequency of individual clones, especially of those affiliated with the
Methanosaetaceae
and
Methanobacteriaceae
. However, these findings could not be confirmed by terminal restriction fragment length polymorphism (T-RFLP) analysis of SSU rDNA amplicons. Most likely, the fluctuations in sequence composition of clone libraries resulted from cloning bias. Clonal SSU rRNA gene sequences were used to define operational taxonomic units (OTUs) for T-RFLP analysis, which were distinguished by group-specific
Taq
I restriction sites. Sequence analysis showed a high degree of conservation of
Taq
I restriction sites within the different archaeal lineages present in Italian rice field soil. Direct T-RFLP analysis of archaeal populations in rice field soil slurries revealed the presence of all archaeal lineages detected by cloning with a predominance of terminal restriction fragments characteristic of rice cluster I (389 bp),
Methanosaetaceae
(280 bp), and
Methanosarcinaceae
/rice cluster VI (182 bp). In general, the relative gene frequency of most detected OTUs remained rather constant over time during the first 17 days after flooding of the soil. Most minor OTUs (e.g.,
Methanomicrobiaceae
and rice cluster III) and
Methanosaetaceae
did not change in relative frequency. Rice cluster I (37 to 30%) and to a lesser extent rice cluster IV as well as
Methanobacteriaceae
decreased over time. Only the relative abundance of
Methanosarcinaceae
(182 bp) increased, roughly doubling from 15 to 29% of total archaeal gene frequency within the first 11 days, which was positively correlated to the dynamics of acetate and formate concentrations. Our results indicate that a functionally dynamic ecosystem, a rice field soil after flooding, was linked to a relatively stable archaeal community structure.
Variable Expressions of
Staphylococcus aureus
Bicomponent Leucotoxins Semiquantified by Competitive Reverse Transcription-PCR
Authors: Bronner Stéphane , Stoessel Patricia , Gravet Alain , Monteil Henri , Prévost Gilles 
Published: July 2002

ABSTRACT

A competitive reverse transcription-PCR method was developed for the semiquantitation of the expression of genes encoding bicomponent leucotoxins of
Staphylococcus aureus
, e.g., Panton-Valentine leucocidin (
lukPV
), gamma-hemolysin (
hlgA
and
hlgCB
), and LukE-LukD (
lukED
). The optimization procedure included RNA preparation; reverse transcription; the use of various amounts of enzymes, antisense primer, and RNA; and the final amplification chain reaction. Reproducible results were obtained, with sensitivity for detection of cDNA within the range of 1 mRNA/10
4
CFU to 10
2
mRNA/CFU, depending on the gene. Both specific mRNAs were more significantly expressed at the late-exponential phase of growth. Expression was about 100-fold higher in yeast extract-Casamino Acids-pyruvate medium than in heart infusion medium. Expression of the widely distributed gamma-hemolysin locus in the NTCC 8178 strain was around 10-fold diminished compared with that in the ATCC 49775 strain. Because of the lower level of
hlgA
expression, the corresponding protein, which is generally not abundant in culture supernatant, should be investigated for its contribution to the leucotoxin-associated virulence. The
agr
,
sar
, and
agr sar
mutant strains revealed a great dependence with regard to leucotoxin expression on the global regulatory system in
S. aureus
, except that expression of
hlgA
was not affected in the
agr
mutant.
A Novel Gene Encoding Xanthan Lyase of
Paenibacillus alginolyticus
Strain XL-1
Authors: Ruijssenaars Harald J. , Hartmans Sybe , Verdoes Jan C. 
Published: July 2002

ABSTRACT

Xanthan-modifying enzymes are powerful tools in studying structure-function relationships of this polysaccharide. One of these modifying enzymes is xanthan lyase, which removes the terminal side chain residue of xanthan. In this paper, the cloning and sequencing of the first xanthan lyase-encoding gene is described, i.e., the
xalA
gene, encoding pyruvated mannose-specific xanthan lyase of
Paenibacillus alginolyticus
XL-1. The
xalA
gene encoded a 100,823-Da protein, including a 36-amino-acid signal sequence. The 96,887-Da mature enzyme could be expressed functionally in
Escherichia coli
. Like the native enzyme, the recombinant enzyme showed no activity on depyruvated xanthan. Compared to production by
P. alginolyticus
, a 30-fold increase in volumetric productivity of soluble xanthan lyase was achieved by heterologous production in
E. coli
. The recombinant xanthan lyase was used to produce modified xanthan, which showed a dramatic loss of the capacity to form gels with locust bean gum.
Cellular Microcystin Content in N-Limited
Microcystis aeruginosa
Can Be Predicted from Growth Rate
Authors: Long Benedict M. , Jones Gary J. , Orr Philip T. 
Published: July 2002

ABSTRACT

Cell quotas of microcystin (
Q
MCYST
; femtomoles of MCYST per cell), protein, and chlorophyll
a
(Chl
a
), cell dry weight, and cell volume were measured over a range of growth rates in N-limited chemostat cultures of the toxic cyanobacterium
Microcystis aeruginosa
MASH 01-A19. There was a positive linear relationship between
Q
MCYST
and specific growth rate (μ), from which we propose a generalized model that enables
Q
MCYST
at any nutrient-limited growth rate to be predicted based on a single batch culture experiment. The model predicts
Q
MCYST
from μ, μ
max
(maximum specific growth rate),
Q
MCYSTmax
(maximum cell quota), and
Q
MCYSTmin
(minimum cell quota). Under the conditions examined in this study, we predict a
Q
MCYSTmax
of 0.129 fmol cell
−1
at μ
max
and a
Q
MCYSTmin
of 0.050 fmol cell
−1
at μ = 0. Net MCYST production rate (
R
MCYST
) asymptotes to zero at μ = 0 and reaches a maximum of 0.155 fmol cell
−1
day
−1
at μ
max
. MCYST/dry weight ratio (milligrams per gram [dry weight]) increased linearly with μ, whereas the MCYST/protein ratio reached a maximum at intermediate μ. In contrast, the MCYST/Chl
a
ratio remained constant. Cell volume correlated negatively with μ, leading to an increase in intracellular MCYST concentration at high μ. Taken together, our results show that fast-growing cells of N-limited
M. aeruginosa
are smaller, are of lower mass, and have a higher intracellular MCYST quota and concentration than slow-growing cells. The data also highlight the importance of determining cell MCYST quotas, as potentially confusing interpretations can arise from determining MCYST content as a ratio to other cell components.
The Enterococcal Surface Protein, Esp, Is Involved in
Enterococcus faecalis
Biofilm Formation
Authors: Toledo-Arana Alejandro , Valle Jaione , Solano Cristina , Arrizubieta Marı́a Jesús , Cucarella Carme , Lamata Marta , Amorena Beatriz , Leiva José , Penadés José Rafael , Lasa Iñigo 
Published: July 2002

ABSTRACT

The enterococcal surface protein, Esp, is a high-molecular-weight surface protein of unknown function whose frequency is significantly increased among infection-derived
Enterococcus faecalis
isolates. In this work, a global structural similarity was found between Bap, a biofilm-associated protein of
Staphylococcus aureus
, and Esp. Analysis of the relationship between the presence of the Esp-encoding gene (
esp
) and the biofilm formation capacity in
E. faecalis
demonstrated that the presence of the
esp
gene is highly associated (
P
< 0.0001) with the capacity of
E. faecalis
to form a biofilm on a polystyrene surface, since 93.5% of the
E. faecalis esp
-positive isolates were capable of forming a biofilm. Moreover, none of the
E. faecalis esp
-deficient isolates were biofilm producers. Depending on the
E. faecalis
isolate, insertional mutagenesis of
esp
caused either a complete loss of the biofilm formation phenotype or no apparent phenotypic defect. Complementation studies revealed that Esp expression in an
E. faecalis esp
-deficient strain promoted primary attachment and biofilm formation on polystyrene and polyvinyl chloride plastic from urine collection bags. Together, these results demonstrate that (i) biofilm formation capacity is widespread among clinical
E. faecalis
isolates, (ii) the biofilm formation capacity is restricted to the
E. faecalis
strains harboring
esp
, and (iii) Esp promotes primary attachment and biofilm formation of
E. faecalis
on abiotic surfaces.
Characterization of the Reduction of Selenate and Tellurite by Nitrate Reductases
Authors: Sabaty Monique , Avazeri Cécile , Pignol David , Vermeglio André 
Published: July 2002

ABSTRACTPreliminary studies showed that the periplasmic nitrate reductase (Nap) ofRhodobacter sphaeroidesand the membrane-bound nitrate reductases ofEscherichia coliare able to reduce selenate and tellurite in vitro with benzyl viologen as an electron donor. In the present study, we found that this is a general feature of denitrifiers. Both the periplasmic and membrane-bound nitrate reductases ofRalstonia eutropha, Paracoccus denitrificans, andParacoccus pantotrophuscan utilize potassium selenate and potassium tellurite as electron acceptors. In order to characterize these reactions, the periplasmic nitrate reductase ofR. sphaeroidesf. sp.denitrificansIL106 was histidine tagged and purified. TheVmaxandKmwere determined for nitrate, tellurite, and selenate. For nitrate, values of 39 μmol · min−1· mg−1and 0.12 mM were obtained forVmaxandKm, respectively, whereas theVmaxvalues for tellurite and selenate were 40- and 140-fold lower, respectively. These low activities can explain the observation that depletion of the nitrate reductase inR. sphaeroidesdoes not modify the MIC of tellurite for this organism.
Improvement and Optimization of Two Engineered Phage Resistance Mechanisms in
Lactococcus lactis
Authors: McGrath Stephen , Fitzgerald Gerald F. , van Sinderen Douwe 
Published: July 2002

ABSTRACT

Homologous replication module genes were identified for four P335 type phages. DNA sequence analysis revealed that all four phages exhibited more than 90% DNA homology for at least two genes, designated

rep
2009

and
orf17
. One of these genes,

rep
2009

, codes for a putative replisome organizer protein and contains an assumed origin of phage DNA replication (

ori
2009

), which was identical for all four phages. DNA fragments representing the

ori
2009

sequence confer a phage-encoded resistance (Per) phenotype on lactococcal hosts when they are supplied on a high-copy-number vector. Furthermore, cloning multiple copies of the

ori
2009

sequence was found to increase the effectiveness of the Per phenotype conferred. A number of antisense plasmids targeting specific genes of the replication module were constructed. Two separate plasmids targeting

rep
2009

and
orf17
were found to efficiently inhibit proliferation of all four phages by interfering with intracellular phage DNA replication. These results represent two highly effective strategies for inhibiting bacteriophage proliferation, and they also identify a novel gene,
orf17
, which appears to be important for phage DNA replication. Furthermore, these results indicate that although the actual mechanisms of DNA replication are very similar, if not identical, for all four phages, expression of the replication genes is significantly different in each case.
Diversity of Sulfur Isotope Fractionations by Sulfate-Reducing Prokaryotes
Authors: Detmers Jan , Brüchert Volker , Habicht Kirsten S. , Kuever Jan 
Published: July 2002

ABSTRACT

Batch culture experiments were performed with 32 different sulfate-reducing prokaryotes to explore the diversity in sulfur isotope fractionation during dissimilatory sulfate reduction by pure cultures. The selected strains reflect the phylogenetic and physiologic diversity of presently known sulfate reducers and cover a broad range of natural marine and freshwater habitats. Experimental conditions were designed to achieve optimum growth conditions with respect to electron donors, salinity, temperature, and pH. Under these optimized conditions, experimental fractionation factors ranged from 2.0 to 42.0‰. Salinity, incubation temperature, pH, and phylogeny had no systematic effect on the sulfur isotope fractionation. There was no correlation between isotope fractionation and sulfate reduction rate. The type of dissimilatory bisulfite reductase also had no effect on fractionation. Sulfate reducers that oxidized the carbon source completely to CO
2
showed greater fractionations than sulfate reducers that released acetate as the final product of carbon oxidation. Different metabolic pathways and variable regulation of sulfate transport across the cell membrane all potentially affect isotope fractionation. Previous models that explained fractionation only in terms of sulfate reduction rates appear to be oversimplified. The species-specific physiology of each sulfate reducer thus needs to be taken into account to understand the regulation of sulfur isotope fractionation during dissimilatory sulfate reduction.
Investigating Microbial (Micro)colony Heterogeneity by Vibrational Spectroscopy
Authors: Choo-Smith L.-P. , Maquelin K. , van Vreeswijk T. , Bruining H. A. , Puppels G. J. , Thi N. A. Ngo , Kirschner C. , Naumann D. , Ami D. , Villa A. M. , Orsini F. , Doglia S. M. , Lamfarraj H. , Sockalingum G. D. , Manfait M. , Allouch P. , Endtz H. P. 
Published: July 2002

ABSTRACT
Fourier transform infrared and Raman microspectroscopy are currently being developed as new methods for the rapid identification of clinically relevant microorganisms. These methods involve measuring spectra from microcolonies which have been cultured for as little as 6 h, followed by the nonsubjective identification of microorganisms through the use of multivariate statistical analyses. To examine the biological heterogeneity of microorganism growth which is reflected in the spectra, measurements were acquired from various positions within (micro)colonies cultured for 6, 12, and 24 h. The studies reveal that there is little spectral variance in 6-h microcolonies. In contrast, the 12- and 24-h cultures exhibited a significant amount of heterogeneity. Hierarchical cluster analysis of the spectra from the various positions and depths reveals the presence of different layers in the colonies. Further analysis indicates that spectra acquired from the surface of the colonies exhibit higher levels of glycogen than do the deeper layers of the colony. Additionally, the spectra from the deeper layers present with higher RNA levels than the surface layers. Therefore, the 6-h colonies with their limited heterogeneity are more suitable for inclusion in a spectral database to be used for classification purposes. These results also demonstrate that vibrational spectroscopic techniques can be useful tools for studying the nature of colony development and biofilm formation.