bluuebunny/crossref_metadata_2025_split huggingface dataset:
Sequence and Expression Analyses of
Cytophaga
-Like Hydrolases in a Western Arctic Metagenomic Library and the Sargasso Sea
Authors: Cottrell Matthew T. , Yu Liying , Kirchman David L. 
Published: December 2005

ABSTRACT

Sequence analysis of environmental DNA promises to provide new insights into the ecology and biogeochemistry of uncultured marine microbes. In this study we used the Sargasso Sea Whole Genome Sequence (WGS) data set to search for hydrolases used by
Cytophaga
-like bacteria to degrade biopolymers such as polysaccharides and proteins. Analysis of the Sargasso WGS data for contigs bearing both the 16S rRNA genes of
Cytophaga
-like bacteria and hydrolase genes revealed a cellulase gene (
celM
) most similar to the gene found in
Cytophaga hutchinsonii
. A BLAST search of the entire Sargasso Sea WGS data set indicated that
celM
was the most abundant cellulase-like gene in the Sargasso Sea. However, the similarity between CelM-like cellulases and peptidases belonging to metalloprotease family M42 led us to question whether CelM is involved in the degradation of polysaccharides or proteins. PCR primers were designed for the
celM
genes in the Sargasso Sea WGS data set and used to identify
celM
in a fosmid library constructed with prokaryotic DNA from the western Arctic Ocean. Expression analysis of the
Cytophaga
-like Arctic CelM, which is 63% identical and 77% similar to CelM in
C. hutchinsonii
, indicated that there was peptidase activity, whereas cellulase activity was not detected. Our analysis suggests that the
celM
gene plays a role in the degradation of protein by
Cytophaga
-like bacteria. The abundance of peptidase genes in the
Cytophaga
-like fosmid clone provides further evidence for the importance of
Cytophaga
-like bacteria in the degradation of protein in high-molecular-weight dissolved organic matter.
Enantioselective Transformation of α-Hexachlorocyclohexane by the Dehydrochlorinases LinA1 and LinA2 from the Soil Bacterium
Sphingomonas paucimobilis
B90A
Authors: Suar Mrutyunjay , Hauser Andrea , Poiger Thomas , Buser Hans-Rudolf , Müller Markus D. , Dogra Charu , Raina Vishakha , Holliger Christof , van der Meer Jan Roelof , Lal Rup , Kohler Hans-Peter E. 
Published: December 2005

ABSTRACT

Sphingomonas paucimobilis
B90A contains two variants, LinA1 and LinA2, of a dehydrochlorinase that catalyzes the first and second steps in the metabolism of hexachlorocyclohexanes (R. Kumari, S. Subudhi, M. Suar, G. Dhingra, V. Raina, C. Dogra, S. Lal, J. R. van der Meer, C. Holliger, and R. Lal, Appl. Environ. Microbiol.
68:
6021-6028, 2002). On the amino acid level, LinA1 and LinA2 were 88% identical to each other, and LinA2 was 100% identical to LinA of
S. paucimobilis
UT26. Incubation of chiral α-hexachlorocyclohexane (α-HCH) with
Escherichia coli
BL21 expressing functional LinA1 and LinA2
S
-glutathione transferase fusion proteins showed that LinA1 preferentially converted the (+) enantiomer, whereas LinA2 preferred the (−) enantiomer. Concurrent formation and subsequent dissipation of β-pentachlorocyclohexene enantiomers was also observed in these experiments, indicating that there was enantioselective formation and/or dissipation of these enantiomers. LinA1 preferentially formed (3
S
,4
S
,5
R
,6
R
)-1,3,4,5,6-pentachlorocyclohexene, and LinA2 preferentially formed (3
R
,4
R
,5
S
,6
S
)-1,3,4,5,6-pentachlorocyclohexene. Because enantioselectivity was not observed in incubations with whole cells of
S. paucimobilis
B90A, we concluded that LinA1 and LinA2 are equally active in this organism. The enantioselective transformation of chiral α-HCH by LinA1 and LinA2 provides the first evidence of the molecular basis for the changed enantiomer composition of α-HCH in many natural environments. Enantioselective degradation may be one of the key processes determining enantiomer composition, especially when strains that contain only one of the
linA
genes, such as
S. paucimobilis
UT26, prevail.
Waterborne Pathogen Detection by Use of Oligonucleotide-Based Microarrays
Authors: Maynard Christine , Berthiaume Frédéric , Lemarchand Karine , Harel Josée , Payment Pierre , Bayardelle Paul , Masson Luke , Brousseau Roland 
Published: December 2005

ABSTRACT

A small-oligonucleotide microarray prototype was designed with probes specific for the universal 16S rRNA and
cpn60
genes of several pathogens that are usually encountered in wastewaters. In addition to these two targets,
wecE
-specific oligonucleotide probes were included in the microarray to enhance its discriminating power within the
Enterobacteriaceae
family. Universal PCR primers were used to amplify variable regions of 16S rRNA,
cpn60
, and
wecE
genes directly in
Escherichia coli
and
Salmonella enterica
serovar Typhimurium genomic DNA mixtures (binary);
E. coli
,
S. enterica
serovar Typhimurium, and
Yersinia enterocolitica
genomic DNA mixtures (ternary); or wastewater total DNA. Amplified products were fluorescently labeled and hybridized on the prototype chip. The detection sensitivity for
S. enterica
serovar Typhimurium was estimated to be on the order of 0.1% (10
4
S. enterica
genomes) of the total DNA for the combination of PCR followed by microarray hybridization. The sensitivity of the prototype could be increased by hybridizing amplicons generated by PCR targeting genes specific for a bacterial subgroup, such as
wecE
genes, instead of universal taxonomic amplicons. However, there was evidence of PCR bias affecting the detection limits of a given pathogen as increasing amounts of a different pathogen were spiked into the test samples. These results demonstrate the feasibility of using DNA microarrays in the detection of waterborne pathogens within mixed populations but also raise the problem of PCR bias in such experiments.
Effects of Elevated Atmospheric CO
2
on Soil Microbial Biomass, Activity, and Diversity in a Chaparral Ecosystem
Authors: Lipson David A. , Wilson Richard F. , Oechel Walter C. 
Published: December 2005

ABSTRACT

This study reports the effects of long-term elevated atmospheric CO
2
on root production and microbial activity, biomass, and diversity in a chaparral ecosystem in southern California. The free air CO
2
enrichment (FACE) ring was located in a stand dominated by the woody shrub
Adenostoma fasciculatum
. Between 1995 and 2003, the FACE ring maintained an average daytime atmospheric CO
2
concentration of 550 ppm. During the last two years of operation, observations were made on soil cores collected from the FACE ring and adjacent areas of chaparral with ambient CO
2
levels. Root biomass roughly doubled in the FACE plot. Microbial biomass and activity were related to soil organic matter (OM) content, and so analysis of covariance was used to detect CO
2
effects while controlling for variation across the landscape. Extracellular enzymatic activity (cellulase and amylase) and microbial biomass C (chloroform fumigation-extraction) increased more rapidly with OM in the FACE plot than in controls, but glucose substrate-induced respiration (SIR) rates did not. The metabolic quotient (field respiration over potential respiration) was significantly higher in FACE samples, possibly indicating that microbial respiration was less C limited under high CO
2
. The treatments also differed in the ratio of SIR to microbial biomass C, indicating a metabolic difference between the microbial communities. Bacterial diversity, described by 16S rRNA clone libraries, was unaffected by the CO
2
treatment, but fungal biomass was stimulated. Furthermore, fungal biomass was correlated with cellulase and amylase activities, indicating that fungi were responsible for the stimulation of enzymatic activity in the FACE treatment.
Amplified Expression of Fructose 1,6-Bisphosphatase in
Corynebacterium glutamicum
Increases In Vivo Flux through the Pentose Phosphate Pathway and Lysine Production on Different Carbon Sources
Authors: Becker Judith , Klopprogge Corinna , Zelder Oskar , Heinzle Elmar , Wittmann Christoph 
Published: December 2005

ABSTRACT

The overexpression of fructose 1,6-bisphosphatase (FBPase) in
Corynebacterium glutamicum
leads to significant improvement of lysine production on different sugars. Amplified expression of FBPase via the promoter of the gene encoding elongation factor TU (EFTU) increased the lysine yield in the feedback-deregulated lysine-producing strain
C. glutamicum
lysC
fbr
by 40% on glucose and 30% on fructose or sucrose. Additionally formation of the by-products glycerol and dihydroxyacetone was significantly reduced in the P
EFTU
fbp mutant. As revealed by
13
C metabolic flux analysis on glucose the overexpression of FBPase causes a redirection of carbon flux from glycolysis toward the pentose phosphate pathway (PPP) and thus leads to increased NADPH supply. Normalized to an uptake flux of glucose of 100%, the relative flux into the PPP was 56% for
C. glutamicum
lysC
fbr
P
EFTU
fbp and 46% for
C. glutamicum
lysC
fb

r

. The flux for NADPH supply was 180% in the P
EFTU
fbp strain and only 146% in the parent strain. Amplification of FBPase increases the production of lysine via an increased supply of NADPH. Comparative studies with another mutant containing the
sod
promoter upstream of the
fbp
gene indicate that the expression level of FBPase relates to the extent of the metabolic effects. The overexpression of FBPase seems useful for starch- and molasses-based industrial lysine production with
C. glutamicum
. The redirection of flux toward the PPP should also be interesting for the production of other NADPH-demanding compounds as well as for products directly stemming from the PPP.
Proteome Analysis of
Streptococcus thermophilus
Grown in Milk Reveals Pyruvate Formate-Lyase as the Major Upregulated Protein
Authors: Derzelle Sylviane , Bolotin Alexander , Mistou Michel-Yves , Rul Françoise 
Published: December 2005

ABSTRACT

We investigated the adaptation to milk of
Streptococcus thermophilus
LMG18311 using a proteomic approach. Two-dimensional electrophoresis of cytosolic proteins were performed after growth in M17 medium or in milk. A major modification of the proteome concerned proteins involved in the supply of amino acids, like the peptidase PepX, and several enzymes involved in amino acid biosynthesis. In parallel, we observed the upregulation of the synthesis of seven enzymes directly involved in the synthesis of purines, as well as formyl-tetrahydrofolate (THF) synthetase and serine hydroxy-methyl transferase, two enzymes responsible for the synthesis of compounds (THF and glycine, respectively) feeding the purine biosynthetic pathway. The analysis also revealed a massive increase in the synthesis of pyruvate formate-lyase (PFL), the enzyme which converts pyruvate into acetyl coenzyme A and formate. PFL has been essentially studied for its role in mixed-acid product formation in lactic acid bacteria during anaerobic fermentation. However, formate is an important methyl group donor for anabolic pathway through the formation of folate derivates. We hypothesized that PFL was involved in purine biosynthesis during growth in milk. We showed that PFL expression was regulated at the transcriptional level and that
pfl
transcription occurred during the exponential growth phase in milk. The complementation of milk with formate or purine bases was shown to reduce
pfl
expression, to suppress PFL synthesis, and to stimulate growth of
S. thermophilus.
These results show a novel regulatory mechanism controlling the synthesis of PFL and suggest an unrecognized physiological role for PFL as a formate supplier for anabolic purposes.
Transcriptional Activation of Quinoline Degradation Operons of
Pseudomonas putida
86 by the AraC/XylS-Type Regulator OxoS and Cross-Regulation of the P
qorM
Promoter by XylS
Authors: Carl Birgit , Fetzner Susanne 
Published: December 2005

ABSTRACT

The quinoline-degradative gene cluster (
oxoO
, open reading frames 1 to 6 [ORF1 to -6],
qorMSL
, ORF7 to -9,
oxoR
) of
Pseudomonas putida
86 consists of several overlapping operons controlled in response to quinoline by the master promoter P
oxoO
and internal promoters Porf3, P
qorM
, and P
oxoR
. ORF7 to -9, presumed to be important for maturation of the molybdenum hydroxylase quinoline 2-oxidoreductase, are also weakly transcribed independently of quinoline. Expression of the
oxoS
gene, located upstream of
oxoO
, is not influenced by the carbon source. OxoS shows 26% amino acid sequence identity to XylS, the transcriptional regulator of the
meta
pathway promoter Pm of TOL plasmid pWW0, and is required for quinoline-dependent transcription from P
oxoO
, Porf3, P
qorM
, and P
oxoR
. 5′ deletion analysis of P
oxoO
and P
qorM
suggested that a 5′-TGCPuCT-N
3
-GGGATA-3′ motif, which resembles the distal 5′-TGCA-N
6
-GGNTA-3′ half-site of the tandem XylS binding site, is essential for
oxoS
-dependent transcriptional activation. P
qorM
, which shows similarity to the tandem XylS recognition site of Pm, was cross-activated by the
xylS
gene product in response to benzoate. The distal half-site of P
qorM
is necessary, but probably not sufficient, for transcriptional activation by XylS. Despite conservation in P
oxoO
of a distal 5′-TGCA-N
6
-GGNTA-3′ sequence, cross-activation of P
oxoO
by XylS and benzoate was not observed. The
oxoS
gene product in the presence of quinoline weakly stimulated transcription from the Pm promoter. Involvement of an XylS-type protein in the regulation of genes encoding synthesis of a molybdenum hydroxylase is without precedent and may reflect the evolutionary origin of this pathway in the metabolism of aromatic compounds.
Evaluation of Gel-Pad Oligonucleotide Microarray Technology by Using Artificial Neural Networks
Authors: Pozhitkov Alex , Chernov Boris , Yershov Gennadiy , Noble Peter A. 
Published: December 2005

ABSTRACTPast studies have suggested that thermal dissociation analysis of nucleic acids hybridized to DNA microarrays would improve discrimination among duplex types by scanning through a broad range of stringency conditions. To more fully constrain the utility of this approach using a previously described gel-pad microarray format, artificial neural networks (NNs) were trained to recognize noisy or low-quality data, as might derive from nonspecific fluorescence, poor hybridization, or compromised data collection. The NNs were trained to classify dissociation profiles (melts) into groups based on selected characteristics (e.g., initial signal intensity, area under the curve) using a data set of 21,044 profiles derived from 186 probes hybridized to a study set of RNA extracted from 32 microbes common to the human oral cavity. Three melt profile groups were identified: one group consisted mostly of ideal melt profiles; another group consisted mostly of poor melt profiles; and, the remainder were difficult to classify. Screening of melting profiles of perfect-match hybrids revealed inconsistencies in the form of melting profiles even for identical probes on the same microarray hybridized to same target rRNA. Approximately 18% of perfect-match duplex types were correctly classified as poor. Experimental variability and deviation from ideal melt behavior were shown to be attributable primarily to a method of local background subtraction that was very sensitive to displacement of the grid frames used for image capture (both determined by the image analysis system) and duplexes with low binding constants. Additional results showed that long RNA fragments limit the discriminating power among duplex types.
Microcolony Cultivation on a Soil Substrate Membrane System Selects for Previously Uncultured Soil Bacteria
Authors: Ferrari Belinda C. , Binnerup Svend J. , Gillings Michael 
Published: December 2005

ABSTRACT
Traditional microbiological methods of cultivation recover less than 1% of the total bacterial species, and the culturable portion of bacteria is not representative of the total phylogenetic diversity. Classical cultivation strategies are now known to supply excessive nutrients to a system and therefore select for fast-growing bacteria that are capable of colony or biofilm formation. New approaches to the cultivation of bacteria which rely on growth in dilute nutrient media or simulated environments are beginning to address this problem of selection. Here we describe a novel microcultivation method for soil bacteria that mimics natural conditions. Our soil slurry membrane system combines a polycarbonate membrane as a growth support and soil extract as the substrate. The result is abundant growth of uncharacterized bacteria as microcolonies. By combining microcultivation with fluorescent in situ hybridization, previously “unculturable” organisms belonging to cultivated and noncultivated divisions, including candidate division TM7, can be identified by fluorescence microscopy. Successful growth of soil bacteria as microcolonies confirmed that the missing culturable majority may have a growth strategy that is not observed when traditional cultivation indicators are used.
Cooccurrence of
ScDSP
Gene Expression, Cell Death, and DNA Fragmentation in a Marine Diatom,
Skeletonema costatum
Authors: Chung Chih-Ching , Hwang Sheng-Ping L. , Chang Jeng 
Published: December 2005

ABSTRACT

A novel death-specific gene,
ScDSP
, was obtained from a death stage subtraction cDNA library of the diatom
Skeletonema costatum
. The full length of
ScDSP
cDNA was 921 bp in length, containing a 699-bp open reading frame encoding 232 amino acids and two stretches of 66 and 156 bp in the 5′ and 3′ untranslated regions, respectively. Analysis of the peptide structure revealed that ScDSP contained a signal peptide domain, a transmembrane domain, and a pair of EF-hand motifs. When
S. costatum
grew exponentially at a rate of 1.3 day
−1
, the
ScDSP
mRNA level was at 2 μmol · mole 18S rRNA
−1
. In contrast, when the culture entered the death phase with a growth rate decreasing to 0.5 day
−1
,
ScDSP
mRNA increased dramatically to 668 μmol · mole 18S rRNA
−1
, and a high degree of DNA fragmentation was simultaneously observed. Under the influence of a light-dark cycle,
ScDSP
expression in both exponential and stationary phases clearly showed a diel rhythm, but the daily mean mRNA level was significantly higher in the stationary phase. Our results suggest that
ScDSP
may play a role in the molecular mechanism of self-destructive autolysis in phytoplankton under stress.
Transcriptional Response of
Saccharomyces cerevisiae
to Desiccation and Rehydration
Authors: Singh Jatinder , Kumar Deept , Ramakrishnan Naren , Singhal Vibha , Jervis Jody , Garst James F. , Slaughter Stephen M. , DeSantis Andrea M. , Potts Malcolm , Helm Richard F. 
Published: December 2005

ABSTRACT

A transcriptional analysis of the response of
Saccharomyces cerevisiae
strain BY4743 to controlled air-drying (desiccation) and subsequent rehydration under minimal glucose conditions was performed. Expression of genes involved in fatty acid oxidation and the glyoxylate cycle was observed to increase during drying and remained in this state during the rehydration phase. When the BY4743 expression profile for the dried sample was compared to that of a commercially prepared dry active yeast, strikingly similar expression changes were observed. The fact that these two samples, dried by different means, possessed very similar transcriptional profiles supports the hypothesis that the response to desiccation is a coordinated event independent of the particular conditions involved in water removal. Similarities between “stationary-phase-essential genes” and those upregulated during desiccation were also noted, suggesting commonalities in different routes to reduced metabolic states. Trends in extracellular and intracellular glucose and trehalose levels suggested that the cells were in a “holding pattern” during the rehydration phase, a concept that was reinforced by cell cycle analyses. Application of a “redescription mining” algorithm suggested that sulfur metabolism is important for cell survival during desiccation and rehydration.
SelectListeria monocytogenesSubtypes Commonly Found in Foods Carry Distinct Nonsense Mutations ininlA, Leading to Expression of Truncated and Secreted Internalin A, and Are Associated with a Reduced Invasion Phenotype for Human Intestinal Epithelial Cells
Authors: Nightingale K. K. , Windham K. , Martin K. E. , Yeung M. , Wiedmann M. 
Published: December 2005

ABSTRACTThe surface protein internalin A (InlA) contributes to the invasion of human intestinal epithelial cells byListeria monocytogenes. Screening ofL. monocytogenesstrains isolated from human clinical cases (n= 46), foods (n= 118), and healthy animals (n= 58) in the United States revealed mutations ininlAleading to premature stop codons (PMSCs) inL. monocytogenesribotypes DUP-1052A and DUP-16635A (PMSC mutation type 1), DUP-1025A and DUP-1031A (PMSC mutation type 2), and DUP-1046B and DUP-1062A (PMSC mutation type 3). While all DUP-1046B, DUP-1062A, DUP-16635A, and DUP-1031A isolates (n= 76) containedinlAPMSCs, ribotypes DUP-1052A and DUP-1025A (n= 72) contained isolates with and withoutinlAPMSCs. Western immunoblotting showed that all threeinlAPMSCs result in the production of truncated and secreted InlA. Searches of the Pathogen Tracker database, which contains subtype and source information for more than 5,000L. monocytogenesisolates, revealed that the six ribotypes shown to contain isolates withinlAPMSCs were overall more commonly isolated from foods than from human listeriosis cases.L. monocytogenesstrains carryinginlAPMSCs also showed significantly (P= 0.0004) reduced invasion of Caco-2 cells compared to isolates with homologous 3′inlAsequences without PMSCs. Invasion assays with an isogenic PMSC mutant further supported the observation thatinlAPMSCs lead to reduced invasion of Caco-2 cells. Our data show that specificL. monocytogenessubtypes which are common among U.S. food isolates but rare among human listeriosis isolates carryinlAmutations that are associated with, and possibly at least partially responsible for, an attenuated invasion phenotype.
Symbiosis and Insect Diversification: an Ancient Symbiont of Sap-Feeding Insects from the Bacterial Phylum
Bacteroidetes
Authors: Moran Nancy A. , Tran Phat , Gerardo Nicole M. 
Published: December 2005

ABSTRACT

Several insect groups have obligate, vertically transmitted bacterial symbionts that provision hosts with nutrients that are limiting in the diet. Some of these bacteria have been shown to descend from ancient infections. Here we show that the large group of related insects including cicadas, leafhoppers, treehoppers, spittlebugs, and planthoppers host a distinct clade of bacterial symbionts. This newly described symbiont lineage belongs to the phylum
Bacteroidetes
. Analyses of 16S rRNA genes indicate that the symbiont phylogeny is completely congruent with the phylogeny of insect hosts as currently known. These results support the ancient acquisition of a symbiont by a shared ancestor of these insects, dating the original infection to at least 260 million years ago. As visualized in a species of spittlebug (Cercopoidea) and in a species of sharpshooter (Cicadellinae), the symbionts have extraordinarily large cells with an elongate shape, often more than 30 μm in length; in situ hybridizations verify that these correspond to the phylum
Bacteroidetes
. “
Candidatus
Sulcia muelleri” is proposed as the name of the new symbiont.