bluuebunny/crossref_metadata_2025_split huggingface dataset:
Bacillus anthracis
Multiplication, Persistence, and Genetic Exchange in the Rhizosphere of Grass Plants
Authors: Saile Elke , Koehler Theresa M. 
Published: May 2006

ABSTRACT

Bacillus anthracis
, the causative agent of anthrax, is known for its rapid proliferation and dissemination in mammalian hosts. In contrast, little information exists regarding the lifestyle of this important pathogen outside of the host. Considering that
Bacillus
species, including close relatives of
B. anthracis
, are saprophytic soil organisms, we investigated the capacity of
B. anthracis
spores to germinate in the rhizosphere and to establish populations of vegetative cells that could support horizontal gene transfer in the soil. Using a simple grass plant-soil model system, we show that
B. anthracis
strains germinate on and around roots, growing in characteristic long filaments. From 2 to 4 days postinoculation, approximately one-half of the
B. anthracis
CFU recovered from soil containing grass seedlings arose from heat-sensitive organisms, while
B. anthracis
CFU retrieved from soil without plants consisted of primarily heat-resistant spores. Coinoculation of the plant-soil system with spores of a fertile
B. anthracis
strain carrying the tetracycline resistance plasmid pBC16 and a selectable
B. anthracis
recipient strain resulted in transfer of pBC16 from the donor to the recipient as early as 3 days postinoculation. Our findings demonstrate that
B. anthracis
can survive as a saprophyte outside of the host. The data suggest that horizontal gene transfer in the rhizosphere of grass plants may play a role in the evolution of the
Bacillus cereus
group species.
Isolation and Characterization of Acid-Tolerant, Thermophilic Bacteria for Effective Fermentation of Biomass-Derived Sugars to Lactic Acid
Authors: Patel Milind A. , Ou Mark S. , Harbrucker Roberta , Aldrich Henry C. , Buszko Marian L. , Ingram Lonnie O. , Shanmugam K. T. 
Published: May 2006

ABSTRACT

Biomass-derived sugars, such as glucose, xylose, and other minor sugars, can be readily fermented to fuel ethanol and commodity chemicals by the appropriate microbes. Due to the differences in the optimum conditions for the activity of the fungal cellulases that are required for depolymerization of cellulose to fermentable sugars and the growth and fermentation characteristics of the current industrial microbes, simultaneous saccharification and fermentation (SSF) of cellulose is envisioned at conditions that are not optimal for the fungal cellulase activity, leading to a higher-than-required cost of cellulase in SSF. We have isolated bacterial strains that grew and fermented both glucose and xylose, major components of cellulose and hemicellulose, respectively, to
l
(+)-lactic acid at 50°C and pH 5.0, conditions that are also optimal for fungal cellulase activity. Xylose was metabolized by these new isolates through the pentose-phosphate pathway. As expected for the metabolism of xylose by the pentose-phosphate pathway, [
13
C]lactate accounted for more than 90% of the total
13
C-labeled products from [
13
C]xylose. Based on fatty acid profile and 16S rRNA sequence, these isolates cluster with
Bacillus coagulans
, although the
B. coagulans
type strain, ATCC 7050, failed to utilize xylose as a carbon source. These new
B. coagulans
isolates have the potential to reduce the cost of SSF by minimizing the amount of fungal cellulases, a significant cost component in the use of biomass as a renewable resource, for the production of fuels and chemicals.
Characterization of Shiga Toxin Gene (
stx
)-Positive and Intimin Gene (
eae
)-Positive
Escherichia coli
Isolates from Wastewater of Slaughterhouses in France
Authors: Loukiadis Estelle , Kérourédan Monique , Beutin Lothar , Oswald Eric , Brugère Hubert 
Published: May 2006

ABSTRACT

Wastewater samples from 12 slaughterhouses located in different regions in France were tested for the presence of
stx
-positive and
eae
-positive
Escherichia coli
isolates, and characteristics of the isolates obtained were determined. A total of 224 wastewater samples were collected in wastewater treatment plants at different stages of wastewater processing. Altogether, 5,001
E. coli
isolates were obtained by colony counting and screened for the presence of
stx
and
eae
genes by multiplex PCR.
stx
-positive and
eae
-positive
E. coli
isolates were detected in 25% of the samples collected; they were found in 13% and 3% of the samples obtained from treated effluent and sludge, respectively, suggesting that they could be spread into the environment. Screening of the samples collected by immunomagnetic separation allowed us to isolate 31 additional
E. coli
serogroup O157 isolates. Four of these isolates harbored
stx
and
eae
genes. All
stx
-positive and
eae
-positive
E. coli
isolates were analyzed for
eae
and
stx
genetic variants, as well as for additional virulence factors and serotypes. Our results suggest that the majority of the
stx
- and
eae
-positive
E. coli
isolates from wastewater have low virulence for humans. However, the diversity of the enterohemorrhagic
E. coli
-associated virulence factors in the strains indicates that the environment may play an important role in the emergence of new pathogenic enterohemorrhagic
E. coli
strains.
Quantitative Microbial Risk Assessment Models for Consumption of Raw Vegetables Irrigated with Reclaimed Water
Authors: Hamilton Andrew J. , Stagnitti Frank , Premier Robert , Boland Anne-Maree , Hale Glenn 
Published: May 2006

ABSTRACT

Quantitative microbial risk assessment models for estimating the annual risk of enteric virus infection associated with consuming raw vegetables that have been overhead irrigated with nondisinfected secondary treated reclaimed water were constructed. We ran models for several different scenarios of crop type, viral concentration in effluent, and time since last irrigation event. The mean annual risk of infection was always less for cucumber than for broccoli, cabbage, or lettuce. Across the various crops, effluent qualities, and viral decay rates considered, the annual risk of infection ranged from 10
−3
to 10
−1
when reclaimed-water irrigation ceased 1 day before harvest and from 10
−9
to 10
−3
when it ceased 2 weeks before harvest. Two previously published decay coefficients were used to describe the die-off of viruses in the environment. For all combinations of crop type and effluent quality, application of the more aggressive decay coefficient led to annual risks of infection that satisfied the commonly propounded benchmark of ≤10
−4
, i.e., one infection or less per 10,000 people per year, providing that 14 days had elapsed since irrigation with reclaimed water. Conversely, this benchmark was not attained for any combination of crop and water quality when this withholding period was 1 day. The lower decay rate conferred markedly less protection, with broccoli and cucumber being the only crops satisfying the 10
−4
standard for all water qualities after a 14-day withholding period. Sensitivity analyses on the models revealed that in nearly all cases, variation in the amount of produce consumed had the most significant effect on the total uncertainty surrounding the estimate of annual infection risk. The models presented cover what would generally be considered to be worst-case scenarios: overhead irrigation and consumption of vegetables raw. Practices such as subsurface, furrow, or drip irrigation and postharvest washing/disinfection and food preparation could substantially lower risks and need to be considered in future models, particularly for developed nations where these extra risk reduction measures are more common.
Environmental Whole-Genome Amplification To Access Microbial Populations in Contaminated Sediments
Authors: Abulencia Carl B. , Wyborski Denise L. , Garcia Joseph A. , Podar Mircea , Chen Wenqiong , Chang Sherman H. , Chang Hwai W. , Watson David , Brodie Eoin L. , Hazen Terry C. , Keller Martin 
Published: May 2006

ABSTRACT

Low-biomass samples from nitrate and heavy metal contaminated soils yield DNA amounts that have limited use for direct, native analysis and screening. Multiple displacement amplification (MDA) using φ29 DNA polymerase was used to amplify whole genomes from environmental, contaminated, subsurface sediments. By first amplifying the genomic DNA (gDNA), biodiversity analysis and gDNA library construction of microbes found in contaminated soils were made possible. The MDA method was validated by analyzing amplified genome coverage from approximately five
Escherichia coli
cells, resulting in 99.2% genome coverage. The method was further validated by confirming overall representative species coverage and also an amplification bias when amplifying from a mix of eight known bacterial strains. We extracted DNA from samples with extremely low cell densities from a U.S. Department of Energy contaminated site. After amplification, small-subunit rRNA analysis revealed relatively even distribution of species across several major phyla. Clone libraries were constructed from the amplified gDNA, and a small subset of clones was used for shotgun sequencing. BLAST analysis of the library clone sequences showed that 64.9% of the sequences had significant similarities to known proteins, and “clusters of orthologous groups” (COG) analysis revealed that more than half of the sequences from each library contained sequence similarity to known proteins. The libraries can be readily screened for native genes or any target of interest. Whole-genome amplification of metagenomic DNA from very minute microbial sources, while introducing an amplification bias, will allow access to genomic information that was not previously accessible.
Short-Term Metabolome Dynamics and Carbon, Electron, and ATP Balances in Chemostat-Grown
Saccharomyces cerevisiae
CEN.PK 113-7D following a Glucose Pulse
Authors: Wu Liang , van Dam Jan , Schipper Dick , Kresnowati M. T. A. Penia , Proell Angela M. , Ras Cor , van Winden Wouter A. , van Gulik Walter M. , Heijnen Joseph J. 
Published: May 2006

ABSTRACT

The in vivo kinetics in
Saccharomyces cerevisiae
CEN.PK 113-7D was evaluated during a 300-second transient period after applying a glucose pulse to an aerobic, carbon-limited chemostat culture. We quantified the responses of extracellular metabolites, intracellular intermediates in primary metabolism, intracellular free amino acids, and in vivo rates of O
2
uptake and CO
2
evolution. With these measurements, dynamic carbon, electron, and ATP balances were set up to identify major carbon, electron, and energy sinks during the postpulse period. There were three distinct metabolic phases during this time. In phase I (0 to 50 seconds after the pulse), the carbon/electron balances closed up to 85%. The accumulation of glycolytic and storage compounds accounted for 60% of the consumed glucose, caused an energy depletion, and may have led to a temporary decrease in the anabolic flux. In phase II (50 to 150 seconds), the fermentative metabolism gradually became the most important carbon/electron sink. In phase III (150 to 300 seconds), 29% of the carbon uptake was not identified in the measurements, and the ATP balance had a large surplus. These results indicate an increase in the anabolic flux, which is consistent with macroscopic balances of extracellular fluxes and the observed increase in CO
2
evolution associated with nonfermentative metabolism. The identified metabolic processes involving major carbon, electron, and energy sinks must be taken into account in in vivo kinetic models based on short-term dynamic metabolome responses.
Genetic Population Structure of the Soil BacteriumMyxococcusxanthusat the Centimeter Scale
Authors: Vos Michiel , Velicer Gregory J. 
Published: May 2006

ABSTRACTMyxococcus xanthusis a gram-negative soil bacterium best known for its remarkable life history of social swarming, social predation, and multicellular fruiting body formation. Very little is known about genetic diversity within this species or how social strategies might vary among neighboring strains at small spatial scales. To investigate the small-scale population structure ofM. xanthus, 78 clones were isolated from a patch of soil (16 by 16 cm) in Tübingen, Germany. Among these isolates, 21 genotypes could be distinguished from a concatemer of three gene fragments:csgA(developmental C signal),fibA(extracellular matrix-associated zinc metalloprotease), andpilA(the pilin subunit of type IV pili). Accumulation curves showed that most of the diversity present at this scale was sampled. ThepilAgene contains both conserved and highly variable regions, and two frequency-distribution tests provide evidence for balancing selection on this gene. The functional domains in thecsgAgene were found to be conserved. Three instances of lateral gene transfer could be inferred from a comparison of individual gene phylogenies, but no evidence was found for linkage equilibrium, supporting the view thatM. xanthusevolution is largely clonal. This study shows thatM. xanthusis surrounded by a variety of distinct conspecifics in its natural soil habitat at a spatial scale at which encounters among genotypes are likely.
Overflow Metabolism in
Escherichia coli
during Steady-State Growth: Transcriptional Regulation and Effect of the Redox Ratio
Authors: Vemuri G. N. , Altman E. , Sangurdekar D. P. , Khodursky A. B. , Eiteman M. A. 
Published: May 2006

ABSTRACT

Overflow metabolism in the form of aerobic acetate excretion by
Escherichia coli
is an important physiological characteristic of this common industrial microorganism. Although acetate formation occurs under conditions of high glucose consumption, the genetic mechanisms that trigger this phenomenon are not clearly understood. We report on the role of the NADH/NAD ratio (redox ratio) in overflow metabolism. We modulated the redox ratio in
E. coli
through the expression of
Streptococcus pneumoniae
(water-forming) NADH oxidase. Using steady-state chemostat cultures, we demonstrated a strong correlation between acetate formation and this redox ratio. We furthermore completed genome-wide transcription analyses of a control
E. coli
strain and an
E. coli
strain overexpressing NADH oxidase. The transcription results showed that in the control strain, several genes involved in the tricarboxylic acid (TCA) cycle and respiration were repressed as the glucose consumption rate increased. Moreover, the relative repression of these genes was alleviated by expression of NADH oxidase and the resulting reduced redox ratio. Analysis of a promoter binding site upstream of the genes which correlated with redox ratio revealed a degenerate sequence with strong homology with the binding site for ArcA. Deletion of
arcA
resulted in acetate reduction and increased the biomass yield due to the increased capacities of the TCA cycle and respiration. Acetate formation was completely eliminated by reducing the redox ratio through expression of NADH oxidase in the
arcA
mutant, even at a very high glucose consumption rate. The results provide a basis for studying new regulatory mechanisms prevalent at reduced NADH/NAD ratios, as well as for designing more efficient bioprocesses.
Extracellular Production of Neoculin, a Sweet-Tasting Heterodimeric Protein with Taste-Modifying Activity, by
Aspergillus oryzae
Authors: Nakajima Ken-ichiro , Asakura Tomiko , Maruyama Jun-ichi , Morita Yuji , Oike Hideaki , Shimizu-Ibuka Akiko , Misaka Takumi , Sorimachi Hiroyuki , Arai Soichi , Kitamoto Katsuhiko , Abe Keiko 
Published: May 2006

ABSTRACT

Neoculin (NCL), a protein with sweetness approximately 500-fold that of sugar, can be utilized as a nonglycemic sweetener. It also has taste-modifying activity to convert sourness to sweetness. NCL is a heterodimer composed of an
N
-glycosylated acidic subunit (NAS) and a basic subunit (NBS), which are conjugated by disulfide bonds. For the production of recombinant NCL (rNCL) by
Aspergillus oryzae
, α-amylase with a KEX2 cleavage site, -K-R-, was fused upstream of each of NAS and NBS and the resulting fusion proteins were simultaneously expressed. For accurate and efficient cleavage of the fusion construct by KEX2-like protease, a triglycine motif was inserted after the KEX2 cleavage site. As NBS showed lower production efficiency than did NAS, a larger amount of the NBS expression plasmid than of NAS expression plasmid was introduced during cotransformation, resulting in successful production of rNCL in the culture medium. Moreover, to obtain a higher production yield of rNCL, the active form of
hacA
cDNA encoding a transcription factor that induces an unfolded protein response was cloned and expressed constitutively. This resulted in a 1.5-fold increase in the level of rNCL production (2.0 mg/liter). rNCL was purified by chromatography, and its NAS was found to be
N
-glycosylated as expected. The original sweetness and taste-modifying activity of rNCL were comparable to those of native NCL when confirmed by calcium imaging with human embryonic kidney cells expressing the human sweet taste receptor and by sensory tests.
High-Throughput Transposon Mutagenesis of
Corynebacterium glutamicum
and Construction of a Single-Gene Disruptant Mutant Library
Authors: Suzuki Nobuaki , Okai Naoko , Nonaka Hiroshi , Tsuge Yota , Inui Masayuki , Yukawa Hideaki 
Published: May 2006

ABSTRACT

A simple and high-throughput transposon-mediated mutagenesis system employing two different types of transposons in combination with direct genomic DNA amplification and thermal asymmetric interlaced PCR (TAIL-PCR) was developed. Each of the two minitransposons based on IS
31831
(ISL3 family) and Tn
5
(IS4 family) was integrated into the
Corynebacterium glutamicum
R genome. By using BLAST and Perl, transposon insertion locations were automatically identified based on the sequences of TAIL-PCR products of mutant cells. Insertion locations of 18,000 mutants were analyzed, and a comprehensive insertion library covering nearly 80% of the 2,990 open reading frames of
C. glutamicum
R was generated. Eight thousand of the mutants, exhibiting disruption in 2,330 genes, survived on complex medium under normal laboratory conditions, indicating that the genes were not essential for cell survival. Of the 2,330 genes, 30 exhibited high similarity to essential genes of
Escherichia coli
or
Bacillus subtilis
. This approach could be useful in furthering genetic understanding of cellular life and facilitating the functional analysis of microorganisms.
Salmonella
Serovars in the Herpetofauna of Indiana County, Pennsylvania
Authors: Chambers David L. , Hulse Arthur C. 
Published: May 2006

ABSTRACT

Herpetofaunal
Salmonella enterica
serovars have not been fully examined in any U.S. region. Thirty-three
Salmonella
serovars were isolated from 156 samples from 34 species, all within Indiana County, Pennsylvania. Results suggest that herpetofaunas could potentially pose a threat to humans. Further understanding of
Salmonella
in herpetofaunas may prevent future human cases.
Effect of Biofilm Growth on Expression of Surface Proteins of
Actinomyces naeslundii
Genospecies 2
Authors: Paddick James S. , Brailsford Susan R. , Rao Susmitha , Soares Renata F. , Kidd Edwina A. M. , Beighton David , Homer Karen A. 
Published: May 2006

ABSTRACT

The predominant surface proteins of biofilm and planktonic
Actinomyces naeslundii
, a primary colonizer of the tooth surface, were examined. Seventy-nine proteins (the products of 52 genes) were identified in biofilm cells, and 30 of these, including adhesins, chaperones, and stress-response proteins, were significantly up-regulated relative to planktonic cells.
Viable Bacteria Present within Oral Squamous Cell Carcinoma Tissue
Authors: Hooper Samuel J. , Crean St John , Lewis Michael A. O. , Spratt David A. , Wade William G. , Wilson Melanie J. 
Published: May 2006

ABSTRACTDespite increasing interest in the possible relationships between bacteria and the different stages of cancer development, the association of bacteria with cancer of the oral cavity has yet to be adequately examined. With that in mind, the primary objective of this study was to identify any bacterial species within oral squamous cell carcinoma tissue using a standard microbiological culture approach. At the time of surgery, a 1-cm3portion of tissue was harvested from deep within the tumor mass using a fresh blade for each cut. Whenever possible, “superficial” portions from the mucosa overlying the tumor and nontumorous control specimens from at least 5 cm away from the primary tumor site were also obtained. Surface contamination was eliminated by immersion in Betadine and washing with phosphate-buffered saline. Each specimen was aseptically macerated and cultured on nonselective media under both aerobic and anaerobic conditions. Isolates were identified by 16S rRNA gene sequencing. Twenty deep-tissue specimens, 19 with corresponding superficial tissues and 12 with control tissues, were successfully processed. A diversity of bacterial taxa were isolated and identified, including several putatively novel species. Most isolates were found to be saccharolytic and acid-tolerant species. Notably, some species were isolated only from either the tumorous or nontumorous tissue type, indicating a degree of restriction. Successful surface decontamination of the specimens indicates that the bacteria detected were from within the tissue. A diversity of bacterial groups have been isolated from within oral squamous cell carcinoma tissue. The significance of these bacteria within the tumor warrants further study.
Human Papillomavirus Type 16 Integration in Cervical Carcinoma In Situ and in Invasive Cervical Cancer
Authors: Arias-Pulido Hugo , Peyton Cheri L. , Joste Nancy E. , Vargas Hernan , Wheeler Cosette M. 
Published: May 2006

ABSTRACT

Integration of human papillomavirus type 16 (HPV-16) into the host DNA has been proposed as a potential marker of cervical neoplastic progression. In this study, a quantitative real-time PCR (qRT-PCR) was used to examine the physical status of HPV-16 in 126 cervical carcinoma in situ and 92 invasive cervical cancers. Based on criteria applied to results from this qRT-PCR assay, HPV-16 was characterized in carcinoma in situ cases as episomal (61.9%), mixed (i.e., episomal and integrated; 29.4%), and integrated (8.7%) forms. In invasive cervical cancer samples, HPV-16 was similarly characterized as episomal (39.1%), mixed (45.7%), and integrated (15.2%) forms. The difference in the frequency of integrated or episomal status estimated for carcinoma in situ and invasive cervical cancer cases was statistically significant (
P
= 0.003). Extensive mapping analysis of HPV-16 E1 and E2 genes in 37 selected tumors demonstrated deletions in both E1 and E2 genes with the maximum number of losses (78.4%) observed within the HPV-16 E2 hinge region. Specifically, deletions within the E2 hinge region were detected most often between nucleotides (nt) 3243 and 3539. The capacity to detect low-frequency HPV-16 integration events was highly limited due to the common presence and abundance of HPV episomal forms. HPV-16 E2 expressed from intact episomes may act in trans to regulate integrated genome expression of E6 and E7.