bluuebunny/crossref_metadata_2025_split huggingface dataset:
Dissimilatory Arsenate Reduction with Sulfide as Electron Donor: Experiments with Mono Lake Water and Isolation of Strain MLMS-1, a Chemoautotrophic Arsenate Respirer
Authors: Hoeft Shelley E. , Kulp Thomas R. , Stolz John F. , Hollibaugh James T. , Oremland Ronald S. 
Published: May 2004

ABSTRACT

Anoxic bottom water from Mono Lake, California, can biologically reduce added arsenate without any addition of electron donors. Of the possible in situ inorganic electron donors present, only sulfide was sufficiently abundant to drive this reaction. We tested the ability of sulfide to serve as an electron donor for arsenate reduction in experiments with lake water. Reduction of arsenate to arsenite occurred simultaneously with the removal of sulfide. No loss of sulfide occurred in controls without arsenate or in sterilized samples containing both arsenate and sulfide. The rate of arsenate reduction in lake water was dependent on the amount of available arsenate. We enriched for a bacterium that could achieve growth with sulfide and arsenate in a defined, mineral medium and purified it by serial dilution. The isolate, strain MLMS-1, is a gram-negative, motile curved rod that grows by oxidizing sulfide to sulfate while reducing arsenate to arsenite. Chemoautotrophy was confirmed by the incorporation of H
14
CO
3
−
into dark-incubated cells, but preliminary gene probing tests with primers for ribulose-1,5-biphosphate carboxylase/oxygenase did not yield PCR-amplified products. Alignment of 16S rRNA sequences indicated that strain MLMS-1 was in the δ-
Proteobacteria
, located near sulfate reducers like
Desulfobulbus
sp. (88 to 90% similarity) but more closely related (97%) to unidentified sequences amplified previously from Mono Lake. However, strain MLMS-1 does not grow with sulfate as its electron acceptor.
Effect of Adaptation to Ethanol on Cytoplasmic and Membrane Protein Profiles of
Oenococcus oeni
Authors: Silveira M. Graça , Baumgärtner Maja , Rombouts Frank M. , Abee Tjakko 
Published: May 2004

ABSTRACT

The practical application of commercial malolactic starter cultures of
Oenococcus oeni
surviving direct inoculation in wine requires insight into mechanisms of ethanol toxicity and of acquired ethanol tolerance in this organism. Therefore, the site-specific location of proteins involved in ethanol adaptation, including cytoplasmic, membrane-associated, and integral membrane proteins, was investigated. Ethanol triggers alterations in protein patterns of
O. oeni
cells stressed with 12% ethanol for 1 h and those of cells grown in the presence of 8% ethanol. Levels of inosine-5′-monophosphate dehydrogenase and phosphogluconate dehydrogenase, which generate reduced nicotinamide nucleotides, were decreased during growth in the presence of ethanol, while glutathione reductase, which consumes NADPH, was induced, suggesting that maintenance of the redox balance plays an important role in ethanol adaptation. Phosphoenolpyruvate:mannose phosphotransferase system (PTS) components of mannose PTS, including the phosphocarrier protein HPr and EII
Man
, were lacking in ethanol-adapted cells, providing strong evidence that mannose PTS is absent in ethanol-adapted cells, and this represents a metabolic advantage to
O. oeni
cells during malolactic fermentation. In cells grown in the presence of ethanol, a large increase in the number of membrane-associated proteins was observed. Interestingly, two of these proteins, dTDT-glucose-4,6-dehydratase and
d
-alanine:
d
-alanine ligase, are known to be involved in cell wall biosynthesis. Using a proteomic approach, we provide evidence for an active ethanol adaptation response of
O. oeni
at the cytoplasmic and membrane protein levels.
Subpopulation Characteristics of Egg-Contaminating
Salmonella enterica
serovar Enteritidis as Defined by the Lipopolysaccharide O Chain
Authors: Guard-Bouldin Jean , Gast Richard K. , Humphrey Thomas J. , Henzler David J. , Morales Cesar , Coles Karen 
Published: May 2004

ABSTRACT

Characterization of
Salmonella enterica
serovar Enteritidis was refined by incorporating new data from isolates obtained from avian sources, from the spleens of naturally infected mice, and from the United Kingdom into an existing lipopolysaccharide (LPS) O-chain compositional database. From least to greatest, the probability of avian isolates producing high-molecular-mass LPS O chain ranked as follows: pooled kidney, liver, and spleen; intestine; cecum; ovary and oviduct; albumen; yolk; and whole egg. Mouse isolates were most like avian intestinal samples, whereas United Kingdom isolates were most like those from the avian reproductive tract and egg. Non-reproductive tract organ isolates had significant loss of O chain. Isogenic isolates that varied in ability to make biofilm and to be orally invasive produced different O-chain structures at 25°C but not at 37°C. Hens infected at a 91:9 biofilm-positive/-negative colony phenotype ratio yielded only the negative phenotype from eggs. These results indicate that the environment within the hen applies stringent selection pressure on subpopulations of
S. enterica
serovar Enteritidis at certain points in the infection pathway that ends in egg contamination. The avian cecum, rather than the intestines, is the early interface between the environment and the host that supports emergence of subpopulation diversity. These results suggest that diet and other factors that alter cecal physiology should be investigated as a means to reduce egg contamination.
Cloning, Overexpression, and Characterization of a Novel Thermostable Penicillin G Acylase from
Achromobacter xylosoxidans
: Probing the Molecular Basis for Its High Thermostability
Authors: Cai Gang , Zhu Songcheng , Yang Sheng , Zhao Guoping , Jiang Weihong 
Published: May 2004

ABSTRACT

The gene encoding a novel penicillin G acylase (PGA), designated
pgaW
, was cloned from
Achromobacter xylosoxidans
and overexpressed in
Escherichia coli
. The
pgaW
gene contains an open reading frame of 2,586 nucleotides. The deduced protein sequence encoded by
pgaW
has about 50% amino acid identity to several well-characterized PGAs, including those of
Providencia rettgeri
,
Kluyvera cryocrescens
, and
Escherichia coli
. Biochemical studies showed that the optimal temperature for this novel PGA (PGA650) activity is greater than 60°C and its half-life of inactivation at 55°C is four times longer than that of another previously reported thermostable PGA from
Alcaligenes faecalis
(R. M. D. Verhaert, A. M. Riemens, J. V. R. Laan, J. V. Duin, and W. J. Quax, Appl. Environ. Microbiol. 63:3412-3418, 1997). To our knowledge, this is the most thermostable PGA ever characterized. To explore the molecular basis of the higher thermostability of PGA650, homology structural modeling and amino acid composition analyses were performed. The results suggested that the increased number of buried ion pair networks, lower N and Q contents, excessive arginine residues, and remarkably high content of proline residues in the structure of PGA650 could contribute to its high thermostability. The unique characteristic of higher thermostability of this novel PGA provides some advantages for its potential application in industry.
Two Distinct Pathways for Trehalose Assimilation in the YeastSaccharomyces cerevisiae
Authors: Jules Matthieu , Guillou Vincent , François Jean , Parrou Jean-Luc 
Published: May 2004

ABSTRACTThe yeastSaccharomyces cerevisiaecan synthesize trehalose and also use this disaccharide as a carbon source for growth. However, the molecular mechanism by which extracellular trehalose can be transported to the vacuole and degraded by the acid trehalase Ath1p is not clear. By using an adaptation of the assay of invertase on whole cells with NaF, we showed that more than 90% of the activity of Ath1p is extracellular, splitting of the disaccharide into glucose. We also found that Agt1p-mediated trehalose transport and the hydrolysis of the disaccharide by the cytosolic neutral trehalase Nth1p are coupled and represent a second, independent pathway, although there are several constraints on this alternative route. First, theAGT1/MAL11gene is controlled by theMALsystem, and Agt1p was active in neither non-maltose-fermenting nor maltose-inducible strains. Second, Agt1p rapidly lost activity during growth on trehalose, by a mechanism similar to the sugar-induced inactivation of the maltose permease. Finally, both pathways are highly pH sensitive and effective growth on trehalose occurred only when the medium was buffered at around pH 5.0. The catabolism of trehalose was purely oxidative, and since levels of Ath1p limit the glucose flux in the cells, batch cultures on trehalose may provide a useful alternative to glucose-limited chemostat cultures for investigation of metabolic responses in yeast.
Acquisition of an
Agrobacterium
Ri Plasmid and Pathogenicity by Other α-
Proteobacteria
in Cucumber and Tomato Crops Affected by Root Mat
Authors: Weller S. A. , Stead D. E. , Young J. P. W. 
Published: May 2004

ABSTRACT

Root mat of cucumbers and tomatoes has previously been shown to be caused by
Agrobacterium radiobacter
strains harboring a root-inducing Ri plasmid (pRi). Nine other pRi-harboring α-
Proteobacteria
have subsequently been isolated from root mat-infected crops. Fatty acid profiling and partial 16S rRNA sequence analysis identified three of these strains as being in the genus
Ochrobactrum
, five as being in the genus
Rhizobium
, and one as being in the genus
Sinorhizobium
. An in vitro pathogenicity test involving inoculation of cucumber cotyledons was developed. All pRi-harboring α-
Proteobacteria
induced typical root mat symptoms from the cotyledons. Average transformation rates for rhizogenic
Ochrobactrum
(46%) and
Rhizobium
(44%) strains were lower than those observed for rhizogenic
A. radiobacter
strains (64%). However, individual strains from these three genera all had transformation rates comparable to those observed from cotyledons inoculated with a rhizogenic
Sinorhizobium
strain (75%).
Detection of Molecular Diversity in
Bacillus atrophaeus
by Amplified Fragment Length Polymorphism Analysis
Authors: Burke S. A. , Wright J. D. , Robinson M. K. , Bronk B. V. , Warren R. L. 
Published: May 2004

ABSTRACT

Phenotypically,
Bacillus atrophaeus
is indistinguishable from the type strain of
Bacillus subtilis
except by virtue of pigment production on certain media. Several pigmented variants of
B. subtilis
have been reclassified as
B. atrophaeus
, but several remain ambiguous in regard to their taxonomic placement. In this study, we examined strains within the American Type Culture Collection originally deposited as
Bacillus globigii
,
B. subtilis
var. niger, or
Bacillus niger
using 16S rRNA gene sequencing and amplified fragment length polymorphism (AFLP) analysis to determine the level of molecular diversity among these strains and their relationship with closely related taxa. The 16S rRNA gene sequences revealed little variation with one base substitution between the
B. atrophaeus
type strain ATCC 49337 and the other pigmented bacilli. AFLP analysis produced high-quality DNA fingerprints with sufficient polymorphism to reveal strain-level variation. Cluster analysis of Dice similarity coefficients revealed that three strains, ATCC 31028, ATCC 49760, and ATCC 49822, are much more closely related to
B. atrophaeus
than to
B. subtilis
and should be reclassified as
B. atrophaeus
. A very closely related cluster of
B. atrophaeus
strains was also observed; this cluster was genetically distinct from the type strain. The level of variation between the two groups was approximately the same as the level of variation observed between members of the two
B. subtilis
subspecies,
subtilis
and
spizizenii
. It is proposed that the cluster of strains typified by ATCC 9372 be designated a new subspecies,
B. atrophaeus
subsp.
globigii
.
Spatial Distribution and Stability of the Eight Microbial Species of the Altered Schaedler Flora in the Mouse Gastrointestinal Tract
Authors: Sarma-Rupavtarm Ramahi B. , Ge Zhongming , Schauer David B. , Fox James G. , Polz Martin F. 
Published: May 2004

ABSTRACT

The overall complexity of the microbial communities in the gastrointestinal (GI) tracts of mammals has hindered observations of dynamics and interactions of individual bacterial populations. However, such information is crucial for understanding the diverse disease-causing and protective roles that gut microbiota play in their hosts. Here, we determine the spatial distribution, interanimal variation, and persistence of bacteria in the most complex defined-flora (gnotobiotic) model system to date, viz., mice colonized with the eight strains of the altered Schaedler flora (ASF). Quantitative PCR protocols based on the 16S rRNA sequence of each ASF strain were developed and optimized to specifically detect as few as 10 copies of each target. Total numbers of the ASF strains were determined in the different regions of the GI tracts of three C.B-17 SCID mice. Individual strain abundance was dependent on oxygen sensitivity, with microaerotolerant
Lactobacillus murinus
ASF361 present at 10
5
to 10
7
cells/g of tissue in the upper GI tract and obligate anaerobic ASF strains being predominant in the cecal and colonic flora at 10
8
to 10
10
cells/g of tissue. The variation between the three mice was small for most ASF strains, except for
Clostridium
sp. strain ASF502 and
Bacteroides
sp. strain ASF519 in the cecum. A comparison of the relative distribution of the ASF strains in feces and the colon indicated large differences, suggesting that fecal bacterial levels may provide a poor approximation of colonic bacterial levels. All ASF strains were detected by PCR in the feces of C57BL/6 restricted flora mice, which had been maintained in an isolator without sterile food, water, or bedding for several generations, providing evidence for the stability of these strains in the face of potential competition by bacteria introduced into the gut.
Double-Layer Plaque Assay for Quantification of Enteroviruses
Authors: Mocé-Llivina Laura , Lucena Francisco , Jofre Juan 
Published: May 2004

ABSTRACTWe describe here a double-layer plaque assay for the quantification of enteroviruses, combining a monolayer plaque assay and a suspended-cell plaque assay. The double-layer assay provides significantly greater counts than other methods of virus quantification of both suspensions of pure culture viruses and naturally occurring viruses. The counts obtained by this method are approximately one order of magnitude greater than those obtained with the more commonly used method, the monolayer plaque assay. We conclude that the methods available for quantifying viruses rank in efficiency as follows: double-layer plaque assay ≥ suspended-cell plaque assay > counting cytopathogenic virus adsorbed to cellulose nitrate membrane filters ≥ most probable number of cytopathogenic units > monolayer plaque assay. Moreover, the double-layer plaque assay allows the use of two different cell lines in the two layers. Using the human colonic carcinoma cell line CaCo2 facilitates the recovery of a greater number and diversity of naturally occurring enteroviruses in water than the monolayer agar method. In addition, the pretreatment of cells with 5-iodo-2′-deoxyuridine (IDU) prior to the quantification of enteroviruses by the double-layer plaque assay provides significantly higher recoveries than the use of IDU does with the other methods of quantification.
New Recombination Methods for
Sinorhizobium meliloti
Genetics
Authors: House Brent L. , Mortimer Michael W. , Kahn Michael L. 
Published: May 2004

ABSTRACT

The availability of bacterial genome sequences has created a need for improved methods for sequence-based functional analysis to facilitate moving from annotated DNA sequence to genetic materials for analyzing the roles that postulated genes play in bacterial phenotypes. A powerful cloning method that uses lambda integrase recombination to clone and manipulate DNA sequences has been adapted for use with the gram-negative α-proteobacterium
Sinorhizobium meliloti
in two ways that increase the utility of the system. Adding plasmid
oriT
sequences to a set of vehicles allows the plasmids to be transferred to
S. meliloti
by conjugation and also allows cloned genes to be recombined from one plasmid to another in vivo by a pentaparental mating protocol, saving considerable time and expense. In addition, vehicles that contain yeast Flp recombinase target recombination sequences allow the construction of deletion mutations where the end points of the deletions are located at the ends of the cloned genes. Several deletions were constructed in a cluster of 60 genes on the symbiotic plasmid (pSymA) of
S. meliloti
, predicted to code for a denitrification pathway. The mutations do not affect the ability of the bacteria to form nitrogen-fixing nodules on
Medicago sativa
(alfalfa) roots.
Transposon Mutagenesis of the Obligate Intracellular Pathogen
Rickettsia prowazekii
Authors: Qin Aiping , Tucker Aimee M. , Hines Andria , Wood David O. 
Published: May 2004

ABSTRACT

Genetic analysis of
Rickettsia prowazekii
has been hindered by the lack of selectable markers and efficient mechanisms for generating rickettsial gene knockouts. We have addressed these problems by adapting a gene that codes for rifampin resistance for expression in
R. prowazekii
and by incorporating this selection into a transposon mutagenesis system suitable for generating rickettsial gene knockouts. The
arr-2
gene codes for an enzyme that ADP-ribosylates rifampin, thereby destroying its antibacterial activity. Based on the published sequence, this gene was synthesized by PCR with overlapping primers that contained rickettsial codon usage base changes. This
R. prowazekii
-adapted
arr-2
gene (
Rparr-2
) was placed downstream of the strong rickettsial
rpsL
promoter (

rpsL
P

), and the entire construct was inserted into the Epicentre EZ::TN transposome system. A purified transposon containing

rpsL
P
-Rparr-2

was combined with transposase, and the resulting DNA-protein complex (transposome) was electroporated into competent rickettsiae. Following selection with rifampin, rickettsiae with transposon insertions in the genome were identified by PCR and Southern blotting and the insertion sites were determined by rescue cloning and inverse PCR. Multiple insertions into widely spaced areas of the
R. prowazekii
genome were identified. Three insertions were identified within gene coding sequences. Transposomes provide a mechanism for generating random insertional mutations in
R. prowazekii
, thereby identifying nonessential rickettsial genes.
Germination of
Penicillium paneum
Conidia Is Regulated by 1-Octen-3-ol, a Volatile Self-Inhibitor
Authors: Chitarra Gilma S. , Abee Tjakko , Rombouts Frank M. , Posthumus Maarten A. , Dijksterhuis Jan 
Published: May 2004

ABSTRACT

Penicillium paneum
is an important contaminant of cereal grains which is able to grow at low temperature, low pH, high levels of carbon dioxide, and under acid conditions.
P. paneum
produces mycotoxins, which may be harmful to animals and humans. We found that conidia in dense suspensions showed poor germination, suggesting the presence of a self-inhibitor. A volatile compound(s) produced by these high-density conditions also inhibited mycelial growth of different species of fungi belonging to a variety of genera, suggesting a broad action range. The heat-stable compound was isolated by successive centrifugation of the supernatant obtained from spore suspensions with a density of 10
9
conidia ml
−1
. By using static headspace analyses, two major peaks were distinguished, with the highest production of these metabolites after 22 h of incubation at 25°C and shaking at 140 rpm. Gas chromatography coupled with mass spectra analysis revealed the compounds to be 3-octanone and 1-octen-3-ol. Notably, only the latter compound appeared to block the germination process at different developmental stages of the conidia (swelling and germ tube formation). In this study, 1-octen-3-ol influenced different developmental processes during the
P. paneum
life cycle, including induction of microcycle conidiation and inhibition of spore germination. Therefore, the compound can be considered a fungal hormone during fungal development.
Addition of Aromatic Substrates Restores Trichloroethylene Degradation Activity in
Pseudomonas putida
F1
Authors: Morono Yuki , Unno Hajime , Tanji Yasunori , Hori Katsutoshi 
Published: May 2004

ABSTRACT

The rate of trichloroethylene (TCE) degradation by toluene dioxygenase (TDO) in resting cells of
Pseudomonas putida
F1 gradually decreased and eventually stopped within 1.5 h, as in previous reports. However, the subsequent addition of toluene, which is the principal substrate of TDO, resulted in its immediate degradation without a lag phase. After the consumption of toluene, degradation of TCE restarted at a rate similar to its initial degradation, suggesting that this degradation was mediated by TDO molecules that were present before the cessation of TCE degradation. The addition of benzene and cumene, which are also substrates of TDO, also caused restoration of TCE degradation activity: TCE was degraded simultaneously with cumene, and a larger amount of TCE was degraded after cumene was added than after toluene or benzene was added. But substrates that were expected to supply the cells with NADH or energy did not restore TCE degradation activity. This cycle of pseudoinactivation and restoration of TCE degradation was observed repeatedly without a significant decrease in the number of viable cells, even after six additions of toluene spread over 30 h. The results obtained in this study demonstrate a new type of restoration of TCE degradation that has not been previously reported.
Prevalence of the
Chloroflexi
-Related SAR202 Bacterioplankton Cluster throughout the Mesopelagic Zone and Deep Ocean
Authors: Morris R. M. , Rappé M. S. , Urbach E. , Connon S. A. , Giovannoni S. J. 
Published: May 2004

ABSTRACT

Since their initial discovery in samples from the north Atlantic Ocean, 16S rRNA genes related to the environmental gene clone cluster known as SAR202 have been recovered from pelagic freshwater, marine sediment, soil, and deep subsurface terrestrial environments. Together, these clones form a major, monophyletic subgroup of the phylum
Chloroflexi
. While members of this diverse group are consistently identified in the marine environment, there are currently no cultured representatives, and very little is known about their distribution or abundance in the world's oceans. In this study, published and newly identified SAR202-related 16S rRNA gene sequences were used to further resolve the phylogeny of this cluster and to design taxon-specific oligonucleotide probes for fluorescence in situ hybridization. Direct cell counts from the Bermuda Atlantic time series study site in the north Atlantic Ocean, the Hawaii ocean time series site in the central Pacific Ocean, and along the Newport hydroline in eastern Pacific coastal waters showed that SAR202 cluster cells were most abundant below the deep chlorophyll maximum and that they persisted to 3,600 m in the Atlantic Ocean and to 4,000 m in the Pacific Ocean, the deepest samples used in this study. On average, members of the SAR202 group accounted for 10.2% (±5.7%) of all DNA-containing bacterioplankton between 500 and 4,000 m.