bluuebunny/crossref_metadata_2025 huggingface dataset:
Viewing angle performance of medical liquid crystal displays
Authors: Samei Ehsan , Wright Steven L.
Published: March 2006

Cathode‐ray tube (CRT) and liquid crystal display (LCD) are currently two main technologies for displaying medical images. LCDs possess a number of advantages, but their performance varies as a function of viewing angle. Our purpose in this study was to characterize the angular response performance of five medical‐grade LCDs, and to substantiate their impact on their compliance with the DICOM gray scale display function (GSDF). Furthermore, the study aimed to test a framework to define an angular acceptance range for medical LCDs based on the recent AAPM TG18 guidelines. Measurements were made on five calibrated dual‐domain LCDs, including two 3 megapixel monochrome LCDs, two 5 megapixel monochrome LCDs, and one 9 megapixel color LCD. The luminance performance of each display device was measured as a function of the viewing angle at 17 discrete levels using TG18‐LN test patterns and a Fourier‐optics‐based luminance meter. The luminance data were analyzed according to the AAPM TG18 methodology. The displays showed notable variation in luminance and contrast performance as a function of the viewing angle, particularly in diagonal viewing orientations. Overall, the luminance ratio remained greater than 175 within and viewing angle cones . Aiming to maintain a maximum deviation from the GSDF contrast less than 0.3, i.e., , acceptable viewing angle cones of and were indicated . The findings demonstrate the significant impact of angular response on image contrast, and the utility of and quantities for defining the viewing angle cones within which a medical LCD device can be effectively utilized.
An inversion formula for the exponential Radon transform in spatial domain with variable focal‐length fan‐beam collimation geometry
Authors: Wen Junhai , Liang Zhengrong
Published: March 2006

Inverting the exponential Radon transform has a potential use for SPECT (single photon emission computed tomography) imaging in cases where a uniform attenuation can be approximated, such as in brain and abdominal imaging. Tretiak and Metz derived in the frequency domain an explicit inversion formula for the exponential Radon transform in two dimensions for parallel‐beam collimator geometry. Progress has been made to extend the inversion formula for fan‐beam and varying focal‐length fan‐beam (VFF) collimator geometries. These previous fan‐beam and VFF inversion formulas require a spatially variant filtering operation, which complicates the implementation and imposes a heavy computing burden. In this paper, we present an explicit inversion formula, in which a spatially invariant filter is involved. The formula is derived and implemented in the spatial domain for VFF geometry (where parallel‐beam and fan‐beam geometries are two special cases). Phantom simulations mimicking SPECT studies demonstrate its accuracy in reconstructing the phantom images and efficiency in computation for the considered collimator geometries.
Multispectral breast imaging using a ten‐wavelength, source/detector channels silicon photodiode‐based diffuse optical tomography system
Authors: Li Changqing , Zhao Hongzhi , Anderson Bonnie , Jiang Huabei
Published: March 2006

We describe a compact diffuse optical tomography system specifically designed for breast imaging. The system consists of 64 silicon photodiode detectors, 64 excitation points, and 10 diode lasers in the near‐infrared region, allowing multispectral, three‐dimensional optical imaging of breast tissue. We also detail the system performance and optimization through a calibration procedure. The system is evaluated using tissue‐like phantom experiments and an in vivo clinic experiment. Quantitative two‐dimensional (2D) and three‐dimensional (3D) images of absorption and reduced scattering coefficients are obtained from these experiments. The ten‐wavelength spectra of the extracted reduced scattering coefficient enable quantitative morphological images to be reconstructed with this system. From the in vivo clinic experiment, functional images including deoxyhemoglobin, oxyhemoglobin, and water concentration are recovered and tumors are detected with correct size and position compared with the mammography.
Uncertainties of exposure‐related quantities in mammographic x‐ray unit quality control
Authors: Gregory Kent J. , Pattison John E. , Bibbo Giovanni
Published: March 2006

Breast screening programs operate in many countries with mammographic x‐ray units subject to stringent quality control tests. These tests include the evaluation of quantities based on exposure measurements, such as half value layer, automatic exposure control reproducibility, average glandular dose, and radiation output rate. There are numerous error sources that contribute to the uncertainty of these exposure‐related quantities, some of which are unique to the low energy x‐ray spectrum produced by mammographic x‐ray units. For each of these exposure‐related quantities, the applicable error sources and their magnitudes vary, depending on the test equipment used to make the measurement, and whether or not relevant corrections have been applied. This study has identified and quantified a range of error sources that may be used to estimate the combined uncertainty of these exposure‐related quantities, given the test equipment used and corrections applied. The uncertainty analysis uses methods described by the International Standards Organization's Guide to the Expression of Uncertainty in Measurement. Examples of how these error sources combine to give the uncertainty of the exposure‐related quantities are presented. Using the best test equipment evaluated in this study, uncertainties of the four exposure‐related quantities at the 95% confidence interval were found to be (half value layer), (automatic exposure control reproducibility), (average glandular dose), and (radiation output rate). In some cases, using less precise test equipment or failing to apply corrections, resulted in uncertainties more than double in magnitude.
Probing Proton Dynamics in Molecules on an Attosecond Time Scale
Authors: Baker S. , Robinson J. S. , Haworth C. A. , Teng H. , Smith R. A. , Chirilă C. C. , Lein M. , Tisch J. W. G. , Marangos J. P.
Published: March 2006

We demonstrate a technique that uses high-order harmonic generation in molecules to probe nuclear dynamics and structural rearrangement on a subfemtosecond time scale. The chirped nature of the electron wavepacket produced by laser ionization in a strong field gives rise to a similar chirp in the photons emitted upon electron-ion recombination. Use of this chirp in the emitted light allows information about nuclear dynamics to be gained with 100-attosecond temporal resolution, from excitation by an 8-femtosecond pulse, in a single laser shot. Measurements on molecular hydrogen and deuterium agreed well with calculations of ultrafast nuclear dynamics in the H
2
+
molecule, confirming the validity of the method. We then measured harmonic spectra from CH
4
and CD
4
to demonstrate a few-femtosecond time scale for the onset of proton rearrangement in methane upon ionization.
Naïve and Memory CD4
+
T Cell Survival Controlled by Clonal Abundance
Authors: Hataye Jason , Moon James J. , Khoruts Alexander , Reilly Cavan , Jenkins Marc K.
Published: March 2006

Immunity to a plethora of microbes depends on a diverse repertoire of naïve lymphocytes and the production of long-lived memory cells. We present evidence here that low clonal abundance in a polyclonal repertoire favors the survival and activation of naïve CD4
+
T cells as well as the survival of their memory cell progeny. The inverse relation between clonal frequency and survival suggests that intraclonal competition could help maintain an optimally diverse repertoire of T cells and an optimal environment for the generation of long-lived memory cells.
Sedolisins, a New Class of Secreted Proteases fromAspergillus fumigatuswith Endoprotease or Tripeptidyl-Peptidase Activity at Acidic pHs
Authors: Reichard Utz , Léchenne Barbara , Asif Abdul R. , Streit Frank , Grouzmann Eric , Jousson Olivier , Monod Michel
Published: March 2006

The secreted proteolytic activity ofAspergillus fumigatusis of potential importance as a virulence factor and in the industrial hydrolysis of protein sources. TheA. fumigatusgenome contains sequences that could encode a five-member gene family that produces proteases in the sedolisin family (MEROPS S53). Four putative secreted sedolisins with a predicted 17- to 20-amino-acid signal sequence were identified and termed SedA to SedD. SedA produced heterologously inPichia pastoriswas an acidic endoprotease. Heterologously produced SedB, SedC, and SedD were tripeptidyl-peptidases (TPP) with a common specificity for tripeptide-p-nitroanilide substrates at acidic pHs. Purified SedB hydrolyzed the peptide Ala-Pro-Gly-Asp-Arg-Ile-Tyr-Val-His-Pro-Phe to Arg-Pro-Gly, Asp-Arg-Ile, and Tyr-Val-His-Pro-Phe, thereby confirming TPP activity of the enzyme. SedB, SedC, and SedD were detected by Western blotting in culture supernatants ofA. fumigatusgrown in a medium containing hemoglobin as the sole nitrogen source. A degradation product of SedA also was observed. A search for genes encoding sedolisin homologues in other fungal genomes indicates that sedolisin gene families are widespread among filamentous ascomycetes.
Transcriptional Analysis of the Toxin-Coding Plasmid pBtoxis from
Bacillus thuringiensis
subsp.
israelensis
Authors: Stein Claudia , Jones Gareth W. , Chalmers Tanya , Berry Colin
Published: March 2006

In
Bacillus thuringiensis
subsp.
israelensis
all of the insecticidal toxins are encoded on a single, large plasmid, pBtoxis. Sequencing of this plasmid revealed 125 potential coding sequences, many of which have predicted functions in gene regulation and physiological processes, such as germination. As a first step in understanding the possible role of pBtoxis in its host bacterium, a survey of the transcription of genes with predicted functions was carried out. Whereas many coding sequences, including those previously identified as probable pseudogenes, were not transcribed, mRNA was detected for 29 of the 40 sequences surveyed. Several of these sequences, including eight with similarities to the sequences of known transcriptional regulators, may influence wider gene regulation and thus may alter the phenotype of the host bacterium.
Freeze-Thaw Tolerance and Clues to the Winter Survival of a Soil Community
Authors: Walker Virginia K. , Palmer Gerald R. , Voordouw Gerrit
Published: March 2006

Although efforts have been made to sample microorganisms from polar regions and to investigate a few of the properties that facilitate survival at freezing or subzero temperatures, soil communities that overwinter in areas exposed to alternate freezing and thawing caused by Foehn or Chinook winds have been largely overlooked. We designed and constructed a cryocycler to automatically subject soil cultures to alternating freeze-thaw cycles. After 48 freeze-thaw cycles, control
Escherichia coli
and
Pseudomonas chlororaphis
isolates were no longer viable. Mixed cultures derived from soil samples collected from a Chinook zone showed that the population complexity and viability were reduced after 48 cycles. However, when bacteria that were still viable after the freeze-thaw treatments were used to obtain selected cultures, these cultures proved to be >1,000-fold more freeze-thaw tolerant than the original consortium. Single-colony isolates obtained from survivors after an additional 48 freeze-thaw cycles were putatively identified by 16S RNA gene fragment sequencing. Five different genera were recognized, and one of the cultures,
Chryseobacterium
sp. strain C14, inhibited ice recrystallization, a property characteristic of antifreeze proteins that prevents the growth of large, potentially damaging ice crystals at temperatures close to the melting temperature. This strain was also notable since cell-free medium derived from cultures of it appeared to enhance the multiple freeze-thaw survival of another isolate,
Enterococcus
sp. strain C8. The results of this study and the development of a cryocycler should allow further investigations into the biochemical and soil community adaptations to the rigors of a Chinook environment.
Characterization of Two Polyketide Synthase Genes Involved in Zearalenone Biosynthesis in
Gibberella zeae
Authors: Gaffoor Iffa , Trail Frances
Published: March 2006

Zearalenone, a mycotoxin produced by several
Fusarium
spp., is most commonly found as a contaminant in stored grain and has chronic estrogenic effects on mammals. Zearalenone is a polyketide derived from the sequential condensation of multiple acetate units by a polyketide synthase (PKS), but the genetics of its biosynthesis are not understood. We cloned two genes, designated
ZEA1
and
ZEA2
, which encode polyketide synthases that participate in the biosynthesis of zearalenone by
Gibberella zeae
(anamorph
Fusarium graminearum
). Disruption of either gene resulted in the loss of zearalenone production under inducing conditions.
ZEA1
and
ZEA2
are transcribed divergently from a common promoter region. Quantitative PCR analysis of both PKS genes and six flanking genes supports the view that the two polyketide synthases make up the core biosynthetic unit for zearalenone biosynthesis. An appreciation of the genetics of zearalenone biosynthesis is needed to understand how zearalenone is synthesized under field conditions that result in the contamination of grain.
Control of Substrate Specificity by Active-Site Residues in Nitrobenzene Dioxygenase
Authors: Ju Kou-San , Parales Rebecca E.
Published: March 2006

Nitrobenzene 1,2-dioxygenase from
Comamonas
sp. strain JS765 catalyzes the initial reaction in nitrobenzene degradation, forming catechol and nitrite. The enzyme also oxidizes the aromatic rings of mono- and dinitrotoluenes at the nitro-substituted carbon, but the basis for this specificity is not understood. In this study, site-directed mutagenesis was used to modify the active site of nitrobenzene dioxygenase, and the contribution of specific residues in controlling substrate specificity and enzyme performance was evaluated. The activities of six mutant enzymes indicated that the residues at positions 258, 293, and 350 in the α subunit are important for determining regiospecificity with nitroarene substrates and enantiospecificity with naphthalene. The results provide an explanation for the characteristic specificity with nitroarene substrates. Based on the structure of nitrobenzene dioxygenase, substitution of valine for the asparagine at position 258 should eliminate a hydrogen bond between the substrate nitro group and the amino group of asparagine. Up to 99% of the mononitrotoluene oxidation products formed by the N258V mutant were nitrobenzyl alcohols rather than catechols, supporting the importance of this hydrogen bond in positioning substrates in the active site for ring oxidation. Similar results were obtained with an I350F mutant, where the formation of the hydrogen bond appeared to be prevented by steric interference. The specificity of enzymes with substitutions at position 293 varied depending on the residue present. Compared to the wild type, the F293Q mutant was 2.5 times faster at oxidizing 2,6-dinitrotoluene while retaining a similar

K
m

for the substrate based on product formation rates and whole-cell kinetics.
Effect of pH on Isolation and Distribution of Members of Subdivision 1 of the Phylum
Acidobacteria
Occurring in Soil
Authors: Sait Michelle , Davis Kathryn E. R. , Janssen Peter H.
Published: March 2006

The pH strongly influenced the development of colonies by members of subdivision 1 of the phylum
Acidobacteria
on solid laboratory media. Significantly more colonies of this group formed at pH 5.5 than at pH 7.0. At pH 5.5, 7 to 8% of colonies that formed on plates that were incubated for 4 months were formed by subdivision 1 acidobacteria. These colonies were formed by bacteria that spanned almost the entire phylogenetic breadth of the subdivision, and there was considerable congruence between the diversity of this group as determined by the cultivation-based method and by surveying 16S rRNA genes in the same soil. Members of subdivision 1 acidobacteria therefore appear to be readily culturable. An analysis of published libraries of 16S rRNAs or 16S rRNA genes showed a very strong correlation between the abundance of subdivision 1 acidobacteria in soil bacterial communities and the soil pH. Subdivision 1 acidobacteria were most abundant in libraries from soils with pHs of <6, but rare or absent in libraries from soils with pHs of >6.5. This, together with the selective cultivation of members of the group on lower-pH media, indicates that growth of many members of subdivision 1 acidobacteria is favored by slightly to moderately acidic growth conditions.
Composition and Dynamics of Bacterial Communities of a Drinking Water Supply System as Assessed by RNA- and DNA-Based 16S rRNA Gene Fingerprinting
Authors: Eichler Stefan , Christen Richard , Höltje Claudia , Westphal Petra , Bötel Julia , Brettar Ingrid , Mehling Arndt , Höfle Manfred G.
Published: March 2006

Bacterial community dynamics of a whole drinking water supply system (DWSS) were studied from source to tap. Raw water for this DWSS is provided by two reservoirs with different water characteristics in the Harz mountains of Northern Germany. Samples were taken after different steps of treatment of raw water (i.e., flocculation, sand filtration, and chlorination) and at different points along the supply system to the tap. RNA and DNA were extracted from the sampled water. The 16S rRNA or its genes were partially amplified by reverse transcription-PCR or PCR and analyzed by single-strand conformation polymorphism community fingerprints. The bacterial community structures of the raw water samples from the two reservoirs were very different, but no major changes of these structures occurred after flocculation and sand filtration. Chlorination of the processed raw water strongly affected bacterial community structure, as reflected by the RNA-based fingerprints. This effect was less pronounced for the DNA-based fingerprints. After chlorination, the bacterial community remained rather constant from the storage containers to the tap. Furthermore, the community structure of the tap water did not change substantially for several months. Community composition was assessed by sequencing of abundant bands and phylogenetic analysis of the sequences obtained. The taxonomic compositions of the bacterial communities from both reservoirs were very different at the species level due to their different limnologies. On the other hand, major taxonomic groups, well known to occur in freshwater, such as
Alphaproteobacteria
,
Betaproteobacteria
, and
Bacteroidetes
, were found in both reservoirs. Significant differences in the detection of the major groups were observed between DNA-based and RNA-based fingerprints irrespective of the reservoir. Chlorination of the drinking water seemed to promote growth of nitrifying bacteria. Detailed analysis of the community dynamics of the whole DWSS revealed a significant influence of both source waters on the overall composition of the drinking water microflora and demonstrated the relevance of the raw water microflora for the drinking water microflora provided to the end user.
In Vivo
Himar1
-Based Transposon Mutagenesis of
Francisella tularensis
Authors: Maier Tamara M. , Pechous Roger , Casey Monika , Zahrt Thomas C. , Frank D. W.
Published: March 2006

Francisella tularensis
is the intracellular pathogen that causes human tularemia. It is recognized as a potential agent of bioterrorism due to its low infectious dose and multiple routes of entry. We report the development of a
Himar1
-based random mutagenesis system for
F. tularensis
(
HimarFT
). In vivo mutagenesis of
F. tularensis
live vaccine strain (LVS) with
HimarFT
occurs at high efficiency. Approximately 12 to 15% of cells transformed with the delivery plasmid result in transposon insertion into the genome. Results from Southern blot analysis of 33 random isolates suggest that single insertions occurred, accompanied by the loss of the plasmid vehicle in most cases. Nucleotide sequence analysis of rescued genomic DNA with
HimarFT
indicates that the orientation of integration was unbiased and that insertions occurred in open reading frames and intergenic and repetitive regions of the chromosome. To determine the utility of the system, transposon mutagenesis was performed, followed by a screen for growth on Chamberlain's chemically defined medium (CDM) to isolate auxotrophic mutants. Several mutants were isolated that grew on complex but not on the CDM. We genetically complemented two of the mutants for growth on CDM with a newly constructed plasmid containing a nourseothricin resistance marker. In addition, uracil or aromatic amino acid supplementation of CDM supported growth of isolates with insertions in
pyrD
,
carA
, or
aroE1
supporting the functional assignment of genes within each biosynthetic pathway. A mutant containing an insertion in
aroE1
demonstrated delayed replication in macrophages and was restored to the parental growth phenotype when provided with the appropriate plasmid in
trans
. Our results suggest that a comprehensive library of mutants can be generated in
F. tularensis
LVS, providing an additional genetic tool to identify virulence determinants required for survival within the host.
Chromosomal Dynamism in Progeny of Outbreak-Related Sorbitol-Fermenting EnterohemorrhagicEscherichia coliO157:NM
Authors: Bielaszewska Martina , Prager Rita , Zhang Wenlan , Friedrich Alexander W. , Mellmann Alexander , Tschäpe Helmut , Karch Helge
Published: March 2006

Sorbitol-fermenting (SF) enterohemorrhagicEscherichia coli(EHEC) O157:NM (nonmotile) is a unique clone that causes outbreaks of hemorrhagic colitis and hemolytic-uremic syndrome. In well-defined clusters of cases, we have observed significant variability in pulsed-field gel electrophoresis (PFGE) patterns which could indicate coinfection by different strains. An analysis of randomly selected progeny colonies of an outbreak strain after subcultivation demonstrated that they displayed either the cognate PFGE outbreak pattern or one of four additional patterns and were <89% similar. These profound alterations were associated with changes in the genomic position of one of two Shiga toxin 2-encoding genes (stx2) in the outbreak strain or with the loss of this gene. The twostx2alleles in the outbreak strain were identical but were flanked with phage-related sequences with only 77% sequence identity. Neither of these phages produced plaques, but one lysogenizedE. coliK-12 and integrated inyecEin the lysogens and the wild-type strain. The presence of twostx2genes which correlated with increased production of Stx2 in vitro but not with the clinical outcome of infection was also found in 14 (21%) of 67 SF EHEC O157:NM isolates from sporadic cases of human disease. The variability of PFGE patterns for the progeny of a single colony must be considered when interpreting PFGE patterns in SF EHEC O157-associated outbreaks.
Conservation of the Pho regulon in
Pseudomonas fluorescens
Pf0-1
Authors: Monds Russell D. , Newell Peter D. , Schwartzman Julia A. , O'Toole George A.
Published: March 2006

The Pho regulon integrates the sensing of environmental inorganic phosphate (P
i
) availability with coregulation of gene expression, mediating an adaptive response to P
i
limitation. Many aspects of the Pho regulon have been addressed in studies of
Escherichia coli
; however, it is unclear how transferable this knowledge is to other bacterial systems. Here, we report work to discern the conservation of the Pho regulon in
Pseudomonas fluorescens
Pf0-1. We demonstrate by mutational studies that PhoB/PhoR and the Pst system have conserved functions in the regulation of P
i
-induced phosphatase activities, as well as expression of other P
i
-regulated genes. A genetic screen was carried out to isolate factors that affect Pho-regulated phosphatase activity. We identified the Pho-regulated phosphatases PhoX and PhoD and present evidence that these enzymes are exported via the Tat system. The
phoX
and
phoD
genes were shown to be members of the Pho regulon by reverse transcription-PCR, as well as by functional assessment of putative PhoB binding sites (Pho boxes). Our data also suggested that at least one other non-Tat-secreted Pho-regulated phosphatase exists. From the genetic screen, numerous siderophore mutants that displayed severe defects in Pho-activated phosphatase activity were isolated. Subsequently, iron was shown to be important for modulating the activity of Pho-regulated phosphatases, but it does not regulate this activity at the level of transcription. We also identify and demonstrate a novel role in siderophore production and Pho-regulated phosphatase activity for ApaH, the hydrolase for the nucleotide-signaling molecule AppppA. Finally, numerous mutations in multiple cellular pathways were recovered that may be required for maximal induction of the Pho regulon under P
i
-limiting conditions.
Genome-Based Metabolic Engineering of
Mannheimia succiniciproducens
for Succinic Acid Production
Authors: Lee Sang Jun , Song Hyohak , Lee Sang Yup
Published: March 2006

Succinic acid is a four-carbon dicarboxylic acid produced as one of the fermentation products of anaerobic metabolism. Based on the complete genome sequence of a capnophilic succinic acid-producing rumen bacterium,
Mannheimia succiniciproducens
, gene knockout studies were carried out to understand its anaerobic fermentative metabolism and consequently to develop a metabolically engineered strain capable of producing succinic acid without by-product formation. Among three different CO
2
-fixing metabolic reactions catalyzed by phosphoenolpyruvate (PEP) carboxykinase, PEP carboxylase, and malic enzyme, PEP carboxykinase was the most important for the anaerobic growth of
M. succiniciproducens
and succinic acid production. Oxaloacetate formed by carboxylation of PEP was found to be converted to succinic acid by three sequential reactions catalyzed by malate dehydrogenase, fumarase, and fumarate reductase. Major metabolic pathways leading to by-product formation were successfully removed by disrupting the
ldhA
,
pflB
,
pta
, and
ackA
genes. This metabolically engineered LPK7 strain was able to produce 13.4 g/liter of succinic acid from 20 g/liter glucose with little or no formation of acetic, formic, and lactic acids, resulting in a succinic acid yield of 0.97 mol succinic acid per mol glucose. Fed-batch culture of
M. succiniciproducens
LPK7 with intermittent glucose feeding allowed the production of 52.4 g/liter of succinic acid, with a succinic acid yield of 1.16 mol succinic acid per mol glucose and a succinic acid productivity of 1.8 g/liter/h, which should be useful for industrial production of succinic acid.
Metabolic Engineering of
Corynebacterium glutamicum
for Trehalose Overproduction: Role of the TreYZ Trehalose Biosynthetic Pathway
Authors: Carpinelli Jorge , Krämer Reinhard , Agosin Eduardo
Published: March 2006

Trehalose has many potential applications in biotechnology and the food industry due to its protective effect against environmental stress. Our work explores microbiological production methods based on the capacity of
Corynebacterium glutamicum
to excrete trehalose. We address here raising trehalose productivity through homologous overexpression of maltooligosyltrehalose synthase and the maltooligosyltrehalose trehalohydrolase genes. In addition, heterologous expression of the UDP-glucose pyrophosphorylase gene from
Escherichia coli
improved the supply of glycogen. Gene expression effects were tested on enzymatic activities and intracellular glycogen content, as well as on accumulated and excreted trehalose. Overexpression of the
treY
gene and the
treY/treZ
synthetic operon significantly increased maltooligosyltrehalose synthase activity, the rate-limiting step, and improved the specific productivity and the final titer of trehalose. Furthermore, a strong decrease was noted in glycogen accumulation. Expression of
galU/treY
and
galU/treYZ
synthetic operons showed a partial recovery in the intracellular glycogen levels and a significant improvement in both intra- and extracellular trehalose content.
Unexpected Diversity of Bacteria Capable of Carbon Monoxide Oxidation in a Coastal Marine Environment, and Contribution of the
Roseobacter
-Associated Clade to Total CO Oxidation
Authors: Tolli J. D. , Sievert S. M. , Taylor C. D.
Published: March 2006

The species diversity, phylogenetic affiliations, and physiological activity rates of carbon monoxide-oxidizing microorganisms were investigated, using new isolates from surface waters collected from the coast of New England and type strains from established collections. A direct isolation method allowed the simultaneous recovery of organisms with different growth rates and nutritional requirements and the identification of marine microorganisms that oxidize CO at an environmentally relevant concentration (42 nM CO). Isolates that oxidized CO at environmentally relevant rates (>4.5 × 10
−11
nmol CO oxidized cell
−1
h
−1
) were taxonomically diverse, with representatives in the alpha and gamma subclasses of the
Proteobacteria
and the phylum
Bacteroidetes
, and represent a hitherto unreported metabolic function for several diverse microbial types. Isolates and type strains having the greatest specific rates of CO metabolism (1.1 × 10
−10
to 2.3 × 10
−10
nmol CO oxidized cell
−1
h
−1
) belonged to the
Roseobacter
-associated clade (RAC) of the alpha subclass of the
Proteobacteria
. By using triple-labeled slide preparations, differential counts of active CO-oxidizing RAC cells, total RAC cells, and total bacterial cell counts in environmental samples were obtained. RAC organisms were a major component of total cell numbers (36%). Based on the density of active CO-oxidizing RAC cells in natural samples and RAC-specific metabolic activities determined for pure cultures, active CO-oxidizing RAC cells may contribute up to 15% of the total CO oxidation occurring in coastal waters.
Bacteriophage Migration via Nematode Vectors: Host-Parasite-Consumer Interactions in Laboratory Microcosms
Authors: Dennehy John J. , Friedenberg Nicholas A. , Yang Yul W. , Turner Paul E.
Published: March 2006

Pathogens vectored by nematodes pose serious agricultural, economic, and health threats; however, little is known of the ecological and evolutionary aspects of pathogen transmission by nematodes. Here we describe a novel model system with two trophic levels, bacteriophages and nematodes, each of which competes for bacteria. We demonstrate for the first time that nematodes are capable of transmitting phages between spatially distinct patches of bacteria. This model system has considerable advantages, including the ease of maintenance and manipulation at the laboratory bench, the ability to observe many generations in short periods, and the capacity to freeze evolved strains for later comparison to their ancestors. More generally, experimental studies of complex multispecies interactions, host-pathogen coevolution, disease dynamics, and the evolution of virulence may benefit from this model system because current models (e.g., chickens, mosquitoes, and malaria parasites) are costly to maintain, are difficult to manipulate, and require considerable space. Our initial explorations centered on independently assessing the impacts of nematode, bacterium, and phage population densities on virus migration between host patches. Our results indicated that virus transmission increases with worm density and host bacterial abundance; however, transmission decreases with initial phage abundance, perhaps because viruses eliminate available hosts before migration can occur. We discuss the microbial growth dynamics that underlie these results, suggest mechanistic explanations for nematode transmission of phages, and propose intriguing possibilities for future research.
“CandidatusThiobios zoothamnicoli,” an Ectosymbiotic Bacterium Covering the Giant Marine CiliateZoothamnium niveum
Authors: Rinke Christian , Schmitz-Esser Stephan , Stoecker Kilian , Nussbaumer Andrea D. , Molnár Dávid A. , Vanura Katrina , Wagner Michael , Horn Matthias , Ott Jörg A. , Bright Monika
Published: March 2006

Zoothamnium niveumis a giant, colonial marine ciliate from sulfide-rich habitats obligatorily covered with chemoautotrophic, sulfide-oxidizing bacteria which appear as coccoid rods and rods with a series of intermediate shapes. Comparative 16S rRNA gene sequence analysis and fluorescence in situ hybridization showed that the ectosymbiont ofZ. niveumbelongs to only one pleomorphic phylotype. TheZ. niveumectosymbiont is only moderately related to previously identified groups of thiotrophic symbionts within theGammaproteobacteria, and shows highest 16S rRNA sequence similarity with the free-living sulfur-oxidizing bacterial strain ODIII6 from shallow-water hydrothermal vents of the Mediterranean Sea (94.5%) and an endosymbiont from a deep-sea hydrothermal vent gastropod of the Indian Ocean Ridge (93.1%). A replacement of this specific ectosymbiont by a variety of other bacteria was observed only for senescent basal parts of the host colonies. The taxonomic status “CandidatusThiobios zoothamnicoli” is proposed for the ectosymbiont ofZ. niveumbased on its ultrastructure, its 16S rRNA gene, the intergenic spacer region, and its partial 23S rRNA gene sequence.
Sulfide Oxidation Coupled to Arsenate Reduction by a Diverse Microbial Community in a Soda Lake
Authors: Hollibaugh James T. , Budinoff Charles , Hollibaugh Ryan A. , Ransom Briana , Bano Nasreen
Published: March 2006

We characterized the arsenate-reducing, sulfide-oxidizing population of Mono Lake, California, by analyzing the distribution and diversity of
rrnA
,
cbbL
, and dissimilatory arsenate reductase (
arrA
) genes in environmental DNA, arsenate-plus sulfide-amended lake water, mixed cultures, and isolates. The arsenate-reducing community was diverse. An organism represented by an
rrnA
sequence previously retrieved from Mono Lake and affiliated with the
Desulfobulbaceae
(
Deltaproteobacteria
) appears to be an important member of the arsenate-reducing, sulfide-oxidizing community. Sulfide oxidation coupled with arsenate reduction appears to proceed via a two-electron transfer, resulting in the production of arsenite and an intermediate S compound that is subsequently disproportionated. A realgar-like As/S mineral was formed in some experiments.
Genome Sequence of the Chemolithoautotrophic Nitrite-Oxidizing Bacterium
Nitrobacter winogradskyi
Nb-255
Authors: Starkenburg Shawn R. , Chain Patrick S. G. , Sayavedra-Soto Luis A. , Hauser Loren , Land Miriam L. , Larimer Frank W. , Malfatti Stephanie A. , Klotz Martin G. , Bottomley Peter J. , Arp Daniel J. , Hickey William J.
Published: March 2006

The alphaproteobacterium
Nitrobacter winogradskyi
(ATCC 25391) is a gram-negative facultative chemolithoautotroph capable of extracting energy from the oxidation of nitrite to nitrate. Sequencing and analysis of its genome revealed a single circular chromosome of 3,402,093 bp encoding 3,143 predicted proteins. There were extensive similarities to genes in two alphaproteobacteria,
Bradyrhizobium japonicum
USDA110 (1,300 genes) and
Rhodopseudomonas palustris
CGA009 CG (815 genes). Genes encoding pathways for known modes of chemolithotrophic and chemoorganotrophic growth were identified. Genes encoding multiple enzymes involved in anapleurotic reactions centered on C
2
to C
4
metabolism, including a glyoxylate bypass, were annotated. The inability of
N. winogradskyi
to grow on C
6
molecules is consistent with the genome sequence, which lacks genes for complete Embden-Meyerhof and Entner-Doudoroff pathways, and active uptake of sugars. Two gene copies of the nitrite oxidoreductase, type I ribulose-1,5-bisphosphate carboxylase/oxygenase, cytochrome
c
oxidase, and gene homologs encoding an aerobic-type carbon monoxide dehydrogenase were present. Similarity of nitrite oxidoreductases to respiratory nitrate reductases was confirmed. Approximately 10% of the
N. winogradskyi
genome codes for genes involved in transport and secretion, including the presence of transporters for various organic-nitrogen molecules. The
N. winogradskyi
genome provides new insight into the phylogenetic identity and physiological capabilities of nitrite-oxidizing bacteria. The genome will serve as a model to study the cellular and molecular processes that control nitrite oxidation and its interaction with other nitrogen-cycling processes.
Denitrifier Community Composition along a Nitrate and Salinity Gradient in a Coastal Aquifer
Authors: Santoro Alyson E. , Boehm Alexandria B. , Francis Christopher A.
Published: March 2006

Nitrogen flux into the coastal environment via submarine groundwater discharge may be modulated by microbial processes such as denitrification, but the spatial scales at which microbial communities act and vary are not well understood. In this study, we examined the denitrifying community within the beach aquifer at Huntington Beach, California, where high-nitrate groundwater is a persistent feature. Nitrite reductase-encoding gene fragments (nirKandnirS), responsible for the key step in the denitrification pathway, were PCR amplified, cloned, and sequenced from DNAs extracted from aquifer sediments collected along a cross-shore transect, where groundwater ranged in salinity from 8 to 34 practical salinity units and in nitrate concentration from 0.5 to 330 μM. We found taxonomically rich and novel communities, with allnirKclones exhibiting <85% identity andnirSclones exhibiting <92% identity at the amino acid level to those of cultivated denitrifiers and other environmental clones in the database. Unique communities were found at each site, despite being located within 40 m of each other, suggesting that the spatial scale at which denitrifier diversity and community composition vary is small. Statistical analyses ofnirsequences using the Monte Carlo-based program ∫-Libshuff confirmed that some populations were indeed distinct, although further sequencing would be required to fully characterize the highly diverse denitrifying communities at this site.
Discovery of a New Source of Rifamycin Antibiotics in Marine Sponge Actinobacteria by Phylogenetic Prediction
Authors: Kim Tae Kyung , Hewavitharana Amitha K. , Shaw P. Nicholas , Fuerst John A.
Published: March 2006

Phylogenetic analysis of the ketosynthase (KS) gene sequences of marine sponge-derivedSalinisporastrains of actinobacteria indicated that the polyketide synthase (PKS) gene sequence most closely related to that ofSalinisporawas the rifamycin B synthase ofAmycolatopsis mediterranei. This result was not expected from taxonomic species tree phylogenetics using 16S rRNA sequences. From the PKS sequence data generated from our sponge-derivedSalinisporastrains, we predicted that such strains might synthesize rifamycin-like compounds. Liquid chromatography-tandem mass spectrometry (LC/MS/MS) analysis was applied to one sponge-derivedSalinisporastrain to test the hypothesis of rifamycin synthesis. The analysis reported here demonstrates that thisSalinisporaisolate does produce compounds of the rifamycin class, including rifamycin B and rifamycin SV. A rifamycin-specific KS primer set was designed, and that primer set increased the number of rifamycin-positive strains detected by PCR screening relative to the number detectable using a conserved KS-specific set. Thus, theSalinisporagroup of actinobacteria represents a potential new source of rifamycins outside the genusAmycolatopsisand the first recorded source of rifamycins from marine bacteria.
Global Carbon Utilization Profiles of Wild-Type, Mutant, and Transformant Strains of
Hypocrea jecorina
Authors: Druzhinina Irina S. , Schmoll Monika , Seiboth Bernhard , Kubicek Christian P.
Published: March 2006

The ascomycete
Hypocrea jecorina
(
Trichoderma reesei
), an industrial producer of cellulases and hemicellulases, can efficiently degrade plant polysaccharides. However, the catabolic pathways for the resulting monomers and their relationship to enzyme induction are not well known. Here we used the Biolog Phenotype MicroArrays technique to evaluate the growth of
H. jecorina
on 95 carbon sources. For this purpose, we compared several wild-type isolates, mutants producing different amounts of cellulases, and strains transformed with a heterologous antibiotic resistance marker gene. The wild-type isolates and transformed strains had the highest variation in growth patterns on individual carbon sources. The cellulase mutants were relatively similar to their parental strains. Both in the mutant and in the transformed strains, the most significant changes occurred in utilization of xylitol, erythritol,
d
-sorbitol,
d
-ribose,
d
-galactose,
l
-arabinose,
N
-acetyl-
d
-glucosamine, maltotriose, and β-methyl-glucoside. Increased production of cellulases was negatively correlated with the ability to grow on γ-aminobutyrate, adonitol, and 2-ketogluconate; and positively correlated with that on
d
-sorbitol and saccharic acid. The reproducibility, relative simplicity, and high resolution (±10% of increase in mycelial density) of the phenotypic microarrays make them a useful tool for the characterization of mutant and transformed strains and for a global analysis of gene function.
Minimization of Glycerol Production during the High-Performance Fed-Batch Ethanolic Fermentation Process in
Saccharomyces cerevisiae
, Using a Metabolic Model as a Prediction Tool
Authors: Bideaux Carine , Alfenore Sandrine , Cameleyre Xavier , Molina-Jouve Carole , Uribelarrea Jean-Louis , Guillouet Stéphane E.
Published: March 2006

On the basis of knowledge of the biological role of glycerol in the redox balance of
Saccharomyces cerevisiae
, a fermentation strategy was defined to reduce the surplus formation of NADH, responsible for glycerol synthesis. A metabolic model was used to predict the operating conditions that would reduce glycerol production during ethanol fermentation. Experimental validation of the simulation results was done by monitoring the inlet substrate feeding during fed-batch
S. cerevisiae
cultivation in order to maintain the respiratory quotient (RQ) (defined as the CO
2
production to O
2
consumption ratio) value between 4 and 5. Compared to previous fermentations without glucose monitoring, the final glycerol concentration was successfully decreased. Although RQ-controlled fermentation led to a lower maximum specific ethanol production rate, it was possible to reach a high level of ethanol production: 85 g · liter
−1
with 1.7 g · liter
−1
glycerol in 30 h. We showed here that by using a metabolic model as a tool in prediction, it was possible to reduce glycerol production in a very high-performance ethanolic fermentation process.
Influence of Freeze-Thaw Stress on the Structure and Function of Microbial Communities and Denitrifying Populations in Soil
Authors: Sharma Shilpi , Szele Zsofia , Schilling Rolf , Munch Jean Charles , Schloter Michael
Published: March 2006

Microbial N
2
O release during the course of thawing of soil was investigated in model experiment focusing on denitrification, since freeze-thaw has been shown to cause significant physical and biological changes in soil, including a surge of N
2
O and CO
2
. The origin of these is still controversially discussed. The increase in denitrification after thawing may be attributed to the diffusion of organic substrates newly available to denitrifiers from disrupted soil aggregates, leading to an increase in microbial activity. Laboratory experiments with upper soil layer of a grassland were conducted in microcosms for real-time gas measurements during the entire phase of freeze and thaw. Shifts in microbial communities were evident on resolution of 16S and 18S rRNA genes and transcripts by denaturing gradient gel electrophoresis (DGGE). Microbial expression profiles were compared by RNA-arbitrarily primed PCR technique and subsequent resolution of amplified products on acrylamide gels. Differences in expression levels of periplasmic nitrate reductase gene (
napA
) and cytochrome
cd
1
nitrite reductase (
nirS
) were observed by most-probable-number-reverse transcription-PCR, with higher levels of expression occurring just after thawing began, followed by a decrease.
napA
and
nirS
DGGE profiles showed no change in banding patterns with fingerprints derived from DNA, whereas those derived from cDNA showed a clear succession of denitrifying bacteria, with the most complex pattern being observed at the end of the N
2
O surge. This study provides insight into the structural community changes and expression dynamics of denitrifiers as a result of freeze-thaw stress. Also, the results presented here support the belief that the gas fluxes observed during thawing is a result of freezing initiated high microbial activity.
Identification and Characterization of a Novel Intracellular Alkaline α-Amylase from the Hyperthermophilic Bacterium
Thermotoga maritima
MSB8
Authors: Ballschmiter Meike , Fütterer Ole , Liebl Wolfgang
Published: March 2006

The gene for a novel α-amylase, designated AmyC, from the hyperthermophilic bacterium
Thermotoga maritima
was cloned and heterologously overexpressed in
Escherichia coli
. The putative intracellular enzyme had no amino acid sequence similarity to glycoside hydrolase family (GHF) 13 α-amylases, yet the range of substrate hydrolysis and the product profile clearly define the protein as an α-amylase. Based on sequence similarity AmyC belongs to a subgroup within GHF 57. On the basis of amino acid sequence similarity, Glu185 and Asp349 could be identified as the catalytic residues of AmyC. Using a 60-min assay, the maximum hydrolytic activity of the purified enzyme, which was dithiothreitol dependent, was found to be at 90°C. AmyC displayed a remarkably high pH optimum of pH 8.5 and an unusual sensitivity towards both ATP and EDTA.
Application of Genotyping during an Extensive Outbreak of Waterborne Giardiasis in Bergen, Norway, during Autumn and Winter 2004
Authors: Robertson L. J. , Hermansen L. , Gjerde B. K. , Strand E. , Alvsvåg J. O. , Langeland N.
Published: March 2006

During the autumn and winter of 2004 and 2005, an extensive outbreak of waterborne giardiasis occurred in Bergen, Norway. Over 1,500 patients were diagnosed with giardiasis. Analysis of water from the implicated source revealed low numbers of
Giardia
cysts, but the initial contamination event probably occurred up to 10 weeks previously. While sewage leakage from a residential area is now considered to be the probable source of contamination, during the episode waste from one particular septic tank was thought to be a possible source. Genotyping of cysts from the septic tank demonstrated that they were assemblage A cysts, although the sequences were not identical to any previously published sequences. For the β-giardin gene, the closest published subgenotype was subgenotype A3; for the
gdh
gene, the closest published subgenotype was subgenotype A2. Genotyping of cysts from 21 patient samples revealed that they were assemblage B cysts; thus, the septic tank was unlikely to be the contamination source. Sequencing of the β-giardin and
gdh
genes from patient samples and a comparison of the sequences gave complex results. For the β-giardin gene, three isolates had sequences identical to subgenotype B3 sequences. However, other isolates had between one and four single-nucleotide polymorphisms (SNPs). For the
gdh
gene, none of the sequences were identical to the sequence published for subgenotype B3, and the sequences had between one and three SNPs. One isolate, which was identical to subgenotype B3 at the β-giardin gene, was more similar to subgenotype B2 at the
gdh
gene. Grouping the isolates on the basis of SNPs resulted in different groups for the two genes. The results are discussed in relation to giardiasis in Norway and to other
Giardia
genotyping studies.