bluuebunny/crossref_metadata_2025_split huggingface dataset:
Analysis of Facultative Lithotroph Distribution and Diversity on Volcanic Deposits by Use of the Large Subunit of Ribulose 1,5-Bisphosphate Carboxylase/Oxygenase
Authors: Nanba K. , King G. M. , Dunfield K. 
Published: April 2004

ABSTRACTA 492- to 495-bp fragment of the gene coding for the large subunit of the form I ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) (rbcL) was amplified by PCR from facultatively lithotrophic aerobic CO-oxidizing bacteria, colorless and purple sulfide-oxidizing microbial mats, and genomic DNA extracts from tephra and ash deposits from Kilauea volcano, for which atmospheric CO and hydrogen have been previously documented as important substrates. PCR products from the mats and volcanic sites were used to constructrbcLclone libraries. Phylogenetic analyses showed that therbcLsequences from all isolates clustered with form ICrbcLsequences derived from facultative lithotrophs. In contrast, the microbial mat clone sequences clustered with sequences from obligate lithotrophs representative of form IArbcL. Clone sequences from volcanic sites fell within the form IC clade, suggesting that these sites were dominated by facultative lithotrophs, an observation consistent with biogeochemical patterns at the sites. Based on phylogenetic and statistical analyses, clone libraries differed significantly among volcanic sites, indicating that they support distinct lithotrophic assemblages. Although some of the clone sequences were similar to knownrbcLsequences, most were novel. Based on nucleotide diversity and average pairwise difference, a forested site and an 1894 lava flow were found to support the most diverse and least diverse lithotrophic populations, respectively. These indices of diversity were not correlated with rates of atmospheric CO and hydrogen uptake but were correlated with estimates of respiration and microbial biomass.
Species Diversity of and Toxin Production by
Gibberella fujikuroi
Species Complex Strains Isolated from Native Prairie Grasses in Kansas
Authors: Leslie John F. , Zeller Kurt A. , Logrieco Antonio , Mulè Giuseppina , Moretti Antonio , Ritieni Alberto 
Published: April 2004

ABSTRACT

Fusarium
species from agricultural crops have been well studied with respect to toxin production and genetic diversity, while similar studies of communities from nonagricultural plants are much more limited. We examined 72
Fusarium
isolates from a native North American tallgrass prairie and found that
Gibberella intermedia
(
Fusarium proliferatum
),
Gibberella moniliformis
(
Fusarium verticillioides
), and
Gibberella konza
(
Fusarium konzum
) dominated.
Gibberella thapsina
(
Fusarium thapsinum
) and
Gibberella subglutinans
(
Fusarium subglutinans
) also were recovered, as were seven isolates that could not be assigned to any previously described species on the basis of either morphological or molecular characters. In general, isolates from the prairie grasses produced the same toxins in quantities similar to those produced by isolates of the same species recovered from agricultural hosts. The
G. konza
isolates produce little or no fumonisins (up to 120 μg/g by one strain), and variable but generally low to moderate amounts of beauvericin (4 to 320 μg/g) and fusaproliferin (50 to 540 μg/g). Toxicity to
Artemia salina
larvae within most species was correlated with the concentration of either beauvericin or fusaproliferin produced. Organic isolates from some cultures of
G. moniliformis
were highly toxic towards
A. salina
even though they produced little, if any, beauvericin or fusaproliferin. Thus, additional potentially toxigenic compounds may be synthesized by
G. moniliformis
strains isolated from prairie grasses. The
Fusarium
community from these grasses appears to contain some species not found in surrounding agricultural communities, including some that probably are undescribed, and could be capable of serving as a reservoir for strains of potential agricultural importance.
GC Fractionation Enhances Microbial CommunityDiversity Assessment and Detection of Minority Populations ofBacteria by Denaturing Gradient GelElectrophoresis
Authors: Holben William E. , Feris Kevin P. , Kettunen Anu , Apajalahti Juha H. A. 
Published: April 2004

ABSTRACT
Effectively
and accurately assessing total microbial community diversity is one of
the primary challenges in modern microbial ecology. This is
particularly true with regard to the detection and characterization of
unculturable populations and those present only in low abundance. We
report a novel strategy, GC fractionation combined with denaturing
gradient gel electrophoresis (GC-DGGE), which combines mechanistically
different community analysis approaches to enhance assessment of
microbial community diversity and detection of minority populations of
microbes. This approach employs GC fractionation as an initial step to
reduce the complexity of the community in each fraction. This reduced
complexity facilitates subsequent detection of diversity in individual
fractions. DGGE analysis of individual fractions revealed bands that
were undetected or only poorly represented when total bacterial
community DNA was analyzed. Also, directed cloning and sequencing of
individual bands from DGGE lanes corresponding to individual
G+C fractions allowed detection of numerous phylotypes that
were not recovered using a traditional random cloning and sequencing
approach.
Sodium Chloride Reduces Production of Curvacin A, a Bacteriocin Produced by
Lactobacillus curvatus
Strain LTH 1174, Originating from Fermented Sausage
Authors: Verluyten Jurgen , Messens Winy , De Vuyst Luc 
Published: April 2004

ABSTRACT

Lactobacillus curvatus
LTH 1174, a strain originating in fermented sausage, produces the antilisterial bacteriocin curvacin A. Its biokinetics of cell growth and bacteriocin production as a function of various concentrations of salt (sodium chloride) were investigated in vitro during laboratory fermentations using modified MRS medium. A model was set up to describe the effects of different NaCl concentrations on microbial behavior. Both cell growth and bacteriocin activity were affected by changes in the salt concentration. Sodium chloride clearly slowed down the growth of
L. curvatus
LTH 1174, but more importantly, it had a detrimental effect on specific curvacin A production (

k
B

) and hence on overall bacteriocin activity. Even a low salt concentration (2%, wt/vol) decreased bacteriocin production, while growth was unaffected at this concentration. The inhibitory effect of NaCl was mainly due to its role as an a
w
-lowering agent. Further, it was clear that salt interfered with bacteriocin induction. Additionally, when 6% (wt/vol) sodium chloride was added, the minimum biomass concentration necessary to start the production of curvacin A (

X
B

) was 0.90 g (cell dry mass) per liter. Addition of the cell-free culture supernatant or a protein solution as a source of induction factor resulted in a decrease in

X
B

, an increase in

k
B

, and hence an increase in the maximum attainable bacteriocin activity.
Characterization of Azo Reduction Activity in a Novel Ascomycete Yeast Strain
Authors: Ramalho Patrícia A. , Cardoso M. Helena , Cavaco-Paulo A. , Ramalho M. Teresa 
Published: April 2004

ABSTRACTSeveral model azo dyes are reductively cleaved by growing cultures of an ascomycete yeast species,Issatchenkia occidentalis. In liquid media containing 0.2 mM dye and 2% glucose in a mineral salts base, more than 80% of the dyes are removed in 15 h, essentially under microaerophilic conditions. Under anoxic conditions, decolorization does not occur, even in the presence of pregrown cells. Kinetic assays of azo reduction activities in quasi-resting cells demonstrated the following: (i) while the optimum pH depends on dye structure, the optimum pH range was observed in the acidic range; (ii) the maximum decolorizing activity occurs in the late exponential phase; and (iii) the temperature profile approaches the typical bell-shaped curve. These results indirectly suggest the involvement of an enzyme activity in azo dye reduction. The decolorizing activity ofI. occidentalisis still observed, although at a lower level, when the cells switch to aerobic respiration at the expense of ethanol after glucose exhaustion in the culture medium. Decolorization ceased when all the ethanol was consumed; this observation, along with other lines of evidence, suggests that azo dye reduction depends on cell growth. Anthraquinone-2-sulfonate, a redox mediator, enhances the reduction rates of theN,N-dimethylaniline-based dyes and reduces those of the 2-naphthol-based dyes, an effect which seems to be compatible with a thermodynamic factor. The dye reduction products were tested as carbon and nitrogen sources. 1-Amino-2-naphthol was used as a carbon and nitrogen source, andN,N-dimethyl-p-phenylenediamine was used only as a nitrogen source. Sulfanilic and metanilic acids did not support growth either as a carbon or nitrogen source.
Pulsed Electric Field Alters Molecular Chaperone Expression and Sensitizes
Listeria monocytogenes
to Heat
Authors: Lado Beatrice H. , Bomser Joshua A. , Dunne C. Patrick , Yousef Ahmed E. 
Published: April 2004

ABSTRACT

Pulsed electric field (PEF)-resistant and PEF-sensitive
Listeria monocytogenes
strains were sublethally treated with electric pulses at 15 kV/cm for 29 μs and held at 25°C for 5 to 30 min prior to protein extraction. The levels of the molecular chaperones GroEL, GroES, and DnaJ were determined by immunoblotting. After 10 to 20 min after sublethal PEF treatment, a transient decrease in molecular chaperone expression was observed in the PEF-sensitive strain (Scott A). The levels of GroEL and DnaJ increased back to the basal expression level within 30 min. A substantial decrease in GroES expression persisted for at least 30 min after PEF treatment. Chaperone expression was suppressed after PEF treatment to a smaller extent in the PEF-resistant (OSY-8578) than in the PEF-sensitive strain, and no clear expression pattern was identified in OSY-8578. Inactivation of Scott A and OSY-8578 in phosphate buffer was compared when lethal PEF (27.5 kV/cm, 144 μs) and heat (55°C, 10 min) were applied in sequence. When PEF and heat treatments were applied separately, the populations of
L. monocytogenes
Scott A and OSY-8578 decreased 0.5 to 0.6 log CFU/ml. Cells treated first with PEF and incubated at 25°C for 10 min showed substantial sensitivity to subsequent heat treatment; the decrease in counts for Scott A and OSY-8578 was 6.1 and 2.8 log CFU/ml, respectively. The sequence and time lapse between the two treatments were crucial for achieving high inactivation rates. It is concluded that PEF sensitized
L. monocytogenes
to heat and that maximum heat sensitization occurred when chaperone expression was at a minimum level.
Molecular Basis for Anaerobic Growth of
Saccharomyces cerevisiae
on Xylose, Investigated by Global Gene Expression and Metabolic Flux Analysis
Authors: Sonderegger Marco , Jeppsson Marie , Hahn-Hägerdal Bärbel , Sauer Uwe 
Published: April 2004

ABSTRACT

Yeast xylose metabolism is generally considered to be restricted to respirative conditions because the two-step oxidoreductase reactions from xylose to xylulose impose an anaerobic redox imbalance. We have recently developed, however, a
Saccharomyces cerevisiae
strain that is at present the only known yeast capable of anaerobic growth on xylose alone. Using transcriptome analysis of aerobic chemostat cultures grown on xylose-glucose mixtures and xylose alone, as well as a combination of global gene expression and metabolic flux analysis of anaerobic chemostat cultures grown on xylose-glucose mixtures, we identified the distinguishing characteristics of this unique phenotype. First, the transcript levels and metabolic fluxes throughout central carbon metabolism were significantly higher than those in the parent strain, and they were most pronounced in the xylose-specific, pentose phosphate, and glycerol pathways. Second, differential expression of many genes involved in redox metabolism indicates that increased cytosolic NADPH formation and NADH consumption enable a higher flux through the two-step oxidoreductase reaction of xylose to xylulose in the mutant. Redox balancing is apparently still a problem in this strain, since anaerobic growth on xylose could be improved further by providing acetoin as an external NADH sink. This improved growth was accompanied by an increased ATP production rate and was not accompanied by higher rates of xylose uptake or cytosolic NADPH production. We concluded that anaerobic growth of the yeast on xylose is ultimately limited by the rate of ATP production and not by the redox balance per se, although the redox imbalance, in turn, limits ATP production.
Seasonal Dynamics of Shallow-Hyporheic-Zone Microbial Community Structure along a Heavy-Metal Contamination Gradient
Authors: Feris Kevin P. , Ramsey Philip W. , Frazar Chris , Rillig Matthias , Moore Johnnie N. , Gannon James E. , Holben William E. 
Published: April 2004

ABSTRACT

Heavy metals contaminate numerous freshwater streams and rivers worldwide. Previous work by this group demonstrated a relationship between the structure of hyporheic microbial communities and the fluvial deposition of heavy metals along a contamination gradient during the fall season. Seasonal variation has been documented in microbial communities in numerous terrestrial and aquatic environments, including the hyporheic zone. The current study was designed to assess whether relationships between hyporheic microbial community structure and heavy-metal contamination vary seasonally by monitoring community structure along a heavy-metal contamination gradient for more than a year. No relationship between total bacterial abundance and heavy metals was observed (
R
2
= 0.02,
P
= 0.83). However, denaturing gradient gel electrophoresis pattern analysis indicated a strong and consistent linear relationship between the difference in microbial community composition (populations present) and the difference in the heavy metal content of hyporheic sediments throughout the year (
R
2
= 0.58,
P
< 0.001). Correlations between heavy-metal contamination and the abundance of four specific phylogenetic groups (most closely related to the α, β, and γ-proteobacteria and cyanobacteria) were apparent only during the fall and early winter, when the majority of organic matter is deposited into regional streams. These seasonal data suggest that the abundance of susceptible populations responds to heavy metals primarily during seasons when the potential for growth is highest.
Oxygen-Controlled Bacterial Growth in the Sponge
Suberites domuncula
: toward a Molecular Understanding of the Symbiotic Relationships between Sponge and Bacteria
Authors: Müller Werner E. G. , Grebenjuk Vladislav A. , Thakur Narsinh L. , Thakur Archana N. , Batel Renato , Krasko Anatoli , Müller Isabel M. , Breter Hans J. 
Published: April 2004

ABSTRACT

Sponges (phylum Porifera), known to be the richest producers among the metazoans of bioactive secondary metabolites, are assumed to live in a symbiotic relationship with microorganisms, especially bacteria. Until now, the molecular basis of the mutual symbiosis, the exchange of metabolites for the benefit of the other partner, has not been understood. We show with the demosponge
Suberites domuncula
as a model that the sponge expresses under optimal aeration conditions the enzyme tyrosinase, which synthesizes diphenols from monophenolic compounds. The cDNA isolated was used as a probe to determine the steady-state level of gene expression. The gene expression level parallels the level of specific activity in sponge tissue, indicating that without aeration the tyrosinase level drops drastically; this effect is reversible. The SB2 bacterium isolated from the sponge surface grew well in M9 minimal salt medium supplemented with the dihydroxylated aromatic compound protocatechuate; this carbon source supported growth more than did glucose. From the SB2 bacterium the protocatechuate gene cluster was cloned and sequenced. This cluster comprises all genes coding for enzymes involved in the conversion of protocatechuate to acetyl coenzyme A. Expression is strongly induced if the bacteria are cultivated on M9-protocatechuate medium; the genes
pcaQ
(encoding the putative transcriptional activator of the
pca
operon) and
pcaDC
were used for quantitative PCR analyses. We conclude that metabolites, in this case diphenols, which might be produced by the sponge
S. domuncula
are utilized by the sponge surface-associated bacterium for energy generation. This rationale will help to further uncover the symbiotic pathways between sponges and their associated “nonculturable” microorganisms; our approach is flanked by the establishment of an EST (expressed sequence tags) database in our laboratory.
Urease-Encoding Genes in Ammonia-Oxidizing Bacteria
Authors: Koper Teresa E. , El-Sheikh Amal F. , Norton Jeanette M. , Klotz Martin G. 
Published: April 2004

ABSTRACT

Many but not all ammonia-oxidizing bacteria (AOB) produce urease (urea amidohydrolase, EC 3.5.1.5) and are capable of using urea for chemolithotrophic growth. We sequenced the urease operons from two AOB, the β-proteobacterium
Nitrosospira
sp. strain NpAV and the γ-proteobacterium
Nitrosococcus oceani
. In both organisms, all seven urease genes were contiguous: the three structural urease genes
ureABC
were preceded and succeeded by the accessory genes
ureD
and
ureEFG
, respectively. Green fluorescent protein reporter gene fusions revealed that the
ure
genes were under control of a single operon promoter upstream of the
ureD
gene in
Nitrosococcus oceani
. Southern analyses revealed two copies of
ureC
in the
Nitrosospira
sp. strain NpAV genome, while a single copy of the
ure
operon was detected in the genome of
Nitrosococcus oceani.
The
ureC
gene encodes the alpha subunit protein containing the active site and conserved nickel binding ligands; these conserved regions were suitable primer targets for obtaining further
ureC
sequences from additional AOB. In order to develop molecular tools for detecting the ureolytic ecotype of AOB,
ureC
genes were sequenced from several β-proteobacterial AOB. Pairwise identity values ranged from 80 to 90% for the UreC peptides of AOB within a subdivision. UreC sequences deduced from AOB urease genes and available UreC sequences in the public databases were used to construct alignments and make phylogenetic inferences. The UreC proteins from β-proteobacterial AOB formed a distinct monophyletic group. Unexpectedly, the peptides from AOB did not group most closely with the UreC proteins from other β-proteobacteria. Instead, it appears that urease in β-proteobacterial autotrophic ammonia oxidizers is the product of divergent evolution in the common ancestor of γ- and β-proteobacteria that was initiated before their divergence during speciation. Sequence motifs conserved for the proteobacteria and variable regions possibly discriminatory for
ureC
from β-proteobacterial AOB were identified for future use in environmental analysis of ureolytic AOB. These gene sequences are the first publicly available for
ure
genes from autotrophic AOB.
Purification and Characterization of a Feruloyl Esterase from the Intestinal BacteriumLactobacillus acidophilus
Authors: Wang Xiaokun , Geng Xin , Egashira Yukari , Sanada Hiroo 
Published: April 2004

ABSTRACTDietary ferulic acid (FA), a significant antioxidant substance, is currently the subject of extensive research. FA in cereals exists mainly as feruloylated sugar ester. To release FA from food matrices, it is necessary to cleave ester cross-linking by feruloyl esterase (FAE) (hydroxycinnamoyl esterase; EC 3.1.1.73). In the present study, the FAE from a human typical intestinal bacterium,Lactobacillus acidophilus, was isolated, purified, and characterized for the first time. The enzyme was purified in successive steps including hydrophobic interaction chromatography and anion-exchange chromatography. The purified FAE appeared as a single band in sodium dodecyl sulfate-polyacrylamide gel electrophoresis, with an apparent molecular mass of 36 kDa. It has optimum pH and temperature characteristics (5.6 and 37°C, respectively). The metal ions Cu2+and Fe3+(at a concentration of 5 mmol liter−1) inhibited FAE activity by 97.25 and 94.80%, respectively. Under optimum pH and temperature with 5-O-feruloyl-l-arabinofuranose (FAA) as a substrate, the enzyme exhibited aKmof 0.0953 mmol liter−1and aVmaxof 86.27 mmol liter−1min−1mg−1of protein. Furthermore, the N-terminal amino acid sequence of the purified FAE was found to be A R V E K P R K V I L V G D G A V G S T. The FAE released FA fromO-(5-O-feruloyl-α-l-arabinofuranosyl)-(1→3)-O-β-d-xylopyranosyl-(1→4)-d-xylopyranose (FAXX) and FAA obtained from refined corn bran. Moreover, it released two times more FA from FAXX in the presence of added xylanase.
Real-Time Quantitative PCR for Assessment of Abundance of
Pseudoalteromonas
Species in Marine Samples
Authors: Skovhus Torben L. , Ramsing Niels B. , Holmström Carola , Kjelleberg Staffan , Dahllöf Ingela 
Published: April 2004

ABSTRACT

A real-time quantitative PCR (RTQ-PCR) method for measuring the abundance of
Pseudoalteromonas
species in marine samples is presented. PCR primers targeting a
Pseudoalteromonas
-specific region of the 16S rRNA gene were tested at three different levels using database searches (in silico), a selection of pure cultures (in vitro), and a combined denaturing gradient gel electrophoresis and cloning approach on environmental DNA (in situ). The RTQ-PCR method allowed for the detection of SYBR Green fluorescence from double-stranded DNA over a linear range spanning six orders of magnitude. The detection limit was determined as 1.4 fg of target DNA (1,000 gene copies) measured in the presence of 20 ng of nontarget DNA from salmon testes. In this study, we discuss the importance of robust post-PCR analyses to overcome pitfalls in RTQ-PCR when samples from different complex marine habitats are analyzed and compared on a nonroutine basis. Representatives of the genus
Pseudoalteromonas
were detected in samples from all investigated habitats, suggesting a widespread distribution of this genus across many marine habitats (e.g., seawater, rocks, macroalgae, and marine animals). Three sample types were analyzed by RTQ-PCR to determine the relative abundance of
Pseudoalteromonas
ribosomal DNA (rDNA) compared to the total abundance of eubacterial rDNA. The rDNA fractions of
Pseudoalteromonas
compared to all
Eubacteria
were 1.55% on the green alga
Ulva lactuca
, 0.10% on the tunicate
Ciona intestinalis
, and 0.06% on the green alga
Ulvaria fusca
.