bluuebunny/crossref_metadata_2025 huggingface dataset:
Abundant transcripts of malting barley identified by serial analysis of gene expression (SAGE)
Authors: White Jessica , Pacey‐Miller Toni , Crawford Allison , Cordeiro Giovanni , Barbary Daniel , Bundock Peter , Henry Robert
Published: January 2006

Serial analysis of gene expression (SAGE) was applied to the major cereal crop barley (Hordeum vulgare) to characterize the transcriptional profile of grain during the malting process. Seven SAGE libraries were generated from seed at different time points during malting, in addition to one library from dry mature seed. A total of 155 206 LongSAGE tags, representing 41 909 unique sequences, was generated. This study reports an in‐depth analysis of the most abundant transcripts from each of eight specific time points in a malting barley time course. The 100 most abundant tags from each library were analysed to identify the putative functional role of highly abundant transcripts. The largest functional groups included transcripts coding for stress response and cell defence, ribosomal proteins and storage proteins. The most abundant tag represented B22EL8, a barley metallothionein, which showed significant up‐regulation across the malting time course. Considerable changes in the abundance profiles of some of the highly abundant tags occurred at 24 h post‐steeping, indicating that it may be an important time point for gene expression changes associated with barley seed germination.
FISCAL POLICY, MACROECONOMIC STABILITY AND FINITE HORIZONS
Authors: Andrés Javier , Doménech Rafael , Leith Campbell
Published: January 2006

In this paper we analyse the stabilisation properties of distortionary taxes in a new‐Keynesian model with overlapping generations of finitely lived consumers. In this framework, government debt is part of net wealth and this adds a number of interesting channels through which fiscal policy could affect output and inflation. Output volatility, in the presence of technology shocks, is not substantially affected by the operation of automatic stabilisers, but we find interesting composition effects. While the presence of finitely lived households strengthens the stabilisation performance of distortionary taxes through the reduction of the volatility of consumption, it does so at the cost of more volatile investment and real money balances.
THE PRICE LEVEL, THE QUANTITY THEORY OF MONEY, AND THE FISCAL THEORY OF THE PRICE LEVEL
Authors: Gordon David B. , Leeper Eric M.
Published: January 2006

This paper examines price‐level determination from the perspective of portfolio choice. Arbitrages among money balances, bonds, and investment goods determine their relative demands. Returns to real balance holdings and after‐tax returns to investment goods determine the relative values of nominal and real assets. Because expectations of government policies ultimately determine the expected returns to both nominal and real assets, the price level depends on interactions among current and expected future monetary and fiscal policies. The quantity theory and the fiscal theory emerge as special cases produced by restricting both the margins and the policies considered.
SOME WELFARE IMPLICATIONS OF OPTIMAL STABILISATION POLICY IN AN ECONOMY WITH CAPITAL AND STICKY PRICES
Authors: Damjanovic Tatiana , Nolan Charles
Published: January 2006

In this paper we review and extend some of the key lessons that seem to be emerging from the Ramsey‐inspired theory of dynamic optimal monetary and fiscal policies. We construct measures of the key distortions in our economy; we label these ‘dynamic wedges’. Inflation, actual or anticipated, distorts these wedges in the present period, shrinks the tax base and increases the deadweight loss. We show that, if possible, labour as well as capital ought to be subsidised in steady state. We point to a number of extensions to the Ramsey literature that may help in the formulation of actual policy.
The epidemiology of onychomycoses in Crete, Greece, between 1992 and 2001
Authors: Ioannidou DJ , Maraki S , Krasagakis SK , Tosca A , Tselentis Y
Published: January 2006

Background  Onychomycosis represents the most frequently encountered nail disease, which is difficult to eradicate with drug treatment. Epidemiological studies concerning onychomycosis have been performed in many countries worldwide. Differences in the incidence of onychomycosis have been reported not only for various geographical areas, but also for different regions of the same country.

Objective  This survey was undertaken to determine the epidemiology of onychomycosis in the population of Crete, Greece.

Methods  In a prospective study, the fingernails and toenails of all new patients presenting to the outpatient Dermatology Department of the University Hospital of Crete were examined by certified dermatologists. If they appeared abnormal, nail material was obtained for mycologic examination.

Results  A total of 23 477 patients were examined during the study period (1992–2001). Of them, 19 556 (83.3%) participated in the investigation. Clinical abnormal nails were observed in 2098 (10.7%) patients, 36.7% males and 63.3% females. Mycologically confirmed onychomycosis was detected in 511 (24.3%). Toenail onychomycosis was found in 283 (55.4%) patients, fingernail onychomycosis in 210 (41%), and both toenail and fingernail onychomycosis in 18 (3.6%). In the toenail infections, dermatophytes were most frequently isolated (52%), followed by yeasts (24.7%) and moulds (15.5%); 7.8% of the infections were mixed. In the fingernail infections, yeasts were most often isolated (82.9%), followed by dermatophytes (10%), and moulds (1.9%); 5.2% of the infections were mixed.

Conclusion  Because the pattern of onychomycoses in a country is changing with time, epidemiological studies are necessary for determining the prevalence and the causative agents of the infection.
Three cases of severe Rhus dermatitis in an English primary school
Authors: Gach JE , Tucker W , Hill VA
Published: January 2006

We report three paediatric cases of severe allergic contact dermatitis caused by a Japanese lacquer tree (Rhus verniciflua), which is a rare specimen plant in the UK. The diagnosis of allergic contact dermatitis produced by plants that are not indigenous to a particular country is more likely to be delayed, as well as mistaken for cellulitis.
Eruptive vellus hair cysts: an effective extraction technique for treatment and diagnosis
Authors: Kaya TI , Tataroglu C , Tursen U , Ikizoglu G
Published: January 2006

Background  Eruptive vellus hair cysts are uncommon developmental anomalies of vellus hair follicles that are observed in young patients. Two patients were evaluated for asymptomatic flesh‐coloured papules appearing on the chest, abdomen and axillae. Lesions of both patients were diagnosed histologically as eruptive vellus hair cyst, which is a disorder with no standard treatment option.

Objective  To introduce an alternative treatment and diagnostic technique for eruptive vellus hair cysts and to evaluate this technique's effectiveness histopathologically.

Materials and methods  We treated the lesions by using cautery and a standard dissecting forceps under topical anaesthesia. After puncturing the overlying skin in the centre using the sharp‐tipped cautery point, we grasped the base of the cyst using a standard dissecting forceps and we extracted the cysts out. In patient 1, extracted cysts were sent for histopathological examination; in patient 2, microscopic examination of the extracted material was performed in potassium hydroxide preparations.

Results  Over 150 cysts were extracted with this technique in two patients. The technique was simple, quick and tolerated well. No lesion recurrence was observed after 4 months. On histopathological examination, extracted cysts were observed to have cyst walls. In patient 2, the diagnosis was made within minutes by microscopic examination of the potassium hydroxide preparations of the extracted material.

Conclusion  Our extraction technique is a simple and effective treatment option for these cysts. It can be regarded as curative since the cyst walls were also extracted. Microscopic examination of the potassium hydroxide preparation of the extracted cysts suggests that this technique can also be used as a simple diagnostic procedure.
2‐Deoxyglucose and NMDA inhibit protein synthesis in neurons and regulate phosphorylation of elongation factor‐2 by distinct mechanisms
Authors: Maus M. , Torrens Y. , Gauchy C. , Bretin S. , Nairn A. C. , Glowinski J. , Premont J.
Published: January 2006

Cerebral ischaemia is associated with brain damage and inhibition of neuronal protein synthesis. A deficit in neuronal metabolism and altered excitatory amino acid release may both contribute to those phenomena. In the present study, we demonstrate that both NMDA and metabolic impairment by 2‐deoxyglucose or inhibitors of mitochondrial respiration inhibit protein synthesis in cortical neurons through the phosphorylation of eukaryotic elongation factor (eEF‐2), without any change in phosphorylation of initiation factor eIF‐2α. eEF‐2 kinase may be activated both by Ca2+‐independent AMP kinase or by an increase in cytosolic Ca2+. Although NMDA decreases ATP levels in neurons, only the effects of 2‐deoxyglucose on protein synthesis and phosphorylation of elongation factor eEF‐2 were reversed by Na+ pyruvate. Protein synthesis inhibition by 2‐deoxyglucose was not as a result of a secondary release of glutamate from cortical neurons as it was not prevented by the NMDA receptor antagonist 5‐methyl‐10,11‐dihydro‐5H‐dibenzo‐(a,d)‐cyclohepten‐5,10‐imine hydrogen maleate (MK 801), nor to an increase in cytosolic‐free Ca2+. Conversely, 2‐deoxyglucose likely activates eEF‐2 kinase through a process involving phosphorylation by AMP kinase. In conclusion, we provide evidence that protein synthesis can be inhibited by NMDA and metabolic deprivation by two distinct mechanisms involving, respectively, Ca2+‐dependent and Ca2+‐independent eEF‐2 phosphorylation.
In vivo post‐transcriptional regulation of GAP‐43 mRNA by overexpression of the RNA‐binding protein HuD
Authors: Bolognani Federico , Tanner Daniel C. , Merhege Melissa , Deschênes‐Furry Julie , Jasmin Bernard , Perrone‐Bizzozero Nora I.
Published: January 2006

HuD is a neuronal‐specific RNA‐binding protein that binds to and stabilizes the mRNAs of growth‐associated protein‐43 (GAP‐43) and other neuronal proteins. HuD expression increases during brain development, nerve regeneration, and learning and memory, suggesting that this protein is important for controlling gene expression during developmental and adult plasticity. To examine the function of HuD in vivo, we generated transgenic mice overexpressing human HuD under the control of the calcium–calmodulin‐dependent protein kinase IIα promoter. The transgene was expressed at high levels throughout the forebrain, including the hippocampal formation, amygdala and cerebral cortex. Using quantitative in situ hybridization, we found that HuD overexpression led to selective increases in GAP‐43 mRNA in hippocampal dentate granule cells and neurons in the lateral amygdala and layer V of the neorcortex. In contrast, GAP‐43 pre‐mRNA levels were unchanged or decreased in the same neuronal populations. Comparison of the levels of mature GAP‐43 mRNA and pre‐mRNA in the same neurons of transgenic mice suggested that HuD increased the stability of the transcript. Confirming this, mRNA decay assays revealed that the GAP‐43 mRNA was more stable in brain extracts from HuD transgenic mice than non‐transgenic littermates. In conclusion, our results demonstrate that HuD overexpression is sufficient to increase GAP‐43 mRNA stability in vivo.
Sites in TM2 and 3 are critical for alcohol‐induced conformational changes in GABAA receptors
Authors: Jung Sangwook , Harris R. Adron
Published: January 2006

γ‐Aminobutyric acid type A (GABAA) receptors are molecular targets for alcohols. Previous work suggests that S270 and A291 residues in the transmembrane (TM) 2 and 3 domains of the GABAA receptor α subunit are components of an alcohol‐binding pocket, and S270I and A291W mutants abolished ethanol potentiation. Our results showed that A295C and F296C residues in the TM3 of the GABAA receptor α1 subunit are accessible to hexylmethanethiosulfonate (HMTS) in the alcohol‐bound state, but not in the resting state. Thus, the A295C and F296C sites become water‐accessible as a result of alcohol‐induced conformational changes. If S270 or A291 residues are sites of alcohol binding, then S270I or A291W mutations should prevent alcohol‐induced conformational movements within the TM3 domain. To investigate this question, the accessibility of HMTS reagent to double mutants (A291W/A295C, A291W/F296C, S270I/A295C or S270I/F296C) in the presence of ethanol or hexanol was tested. The A291W or S270I mutations markedly reduced the accessibility of HMTS to all the double mutants in the ethanol‐bound state, and to S270I/F296C, A291W/A295C and A291W/F296C double mutants in the hexanol‐bound state, suggesting that the A291 or S270 residues are critical sites for alcohol binding and alcohol‐induced conformational changes.
Avian Community Response to Lowland Tropical Rainforest Isolation: 40 Years of Change at La Selva Biological Station, Costa Rica
Authors: SIGEL BRYAN J. , SHERRY THOMAS W. , YOUNG BRUCE E.
Published: January 2006

Abstract:  Since 1960, most of the forest surrounding the La Selva Biological Station, an intensively studied tropical research facility in Costa Rica, has been converted to agricultural uses. We used quantitative censuses and analysis of previously published categorical abundances to assess changes in the bird community, and we evaluated potential causes of species‐specific changes by assessing their association with habitat, diet, participation in mixed‐species flocks, and nest type. Approximately the same percentage of species increased as decreased in abundance from 1960 to 1999 (10–20% of all species, depending on method of assessment). Diet was the single most important trait associated with declining species. At least 50% of the species that declined have insectivorous diets. Use of forest habitat and participation in mixed‐species flocks were also significant factors associated with declines, but nest type was unrelated to change in abundance. The species that increased in abundance tended to occur in open habitats and have omnivorous diets. These results reinforce the importance of several population risk factors associated with tropical understory insectivory and mixed‐species flocking: patchy spatial distribution, low population density, large home range, and dietary specialization. La Selva's protected area (1611 ha), despite a forested connection on one boundary with a higher elevation national park, is apparently too small to maintain at least one major guild (understory insectivores). This first quantitative assessment of bird community change at La Selva highlights the need to intensify study of the mechanisms and consequences of biological diversity change in tropical forest fragments.
Accidental Entrapment of an Endo‐Bronchial Blocker Tip by a Surgical Stapler During Selective Ventilation for Lung Lobectomy in a Dog
Authors: LEVIONNOIS OLIVIER L. , BERGADANO ALESSANDRA , SCHATZMANN URS
Published: January 2006

Objective— To describe the use of an endobronchial blocker (EBB) and to perform selective ventilation during pulmonary lobe resection via thoracotomy in a dog and report its accidental stapling in the resection site.

Study Design— Clinical case report.

Animal— One female dog with a suspected abscess or neoplasia of the right caudal pulmonary lobe.

Methods— One‐lung ventilation was performed using a wire‐guided EBB to seal the contaminated parenchyma and facilitate surgical access. The affected lung parenchyma was resected and the resection site was closed with staples.

Results— Lobar resection was performed successfully, but the loop of the EBB guide wire was inadvertently entrapped in the staple line of the lobectomy. Staples were removed to release the wire loop, and the resulting air leak caused loss of ventilation control until the parenchyma was re‐sealed.

Conclusions— We recommend removing the wire guide associate with the EBB after successful lung separation to avoid accidents that could have life‐threatening consequences if not recognized.

Clinical Relevance— One‐lung ventilation is useful to isolate healthy parenchyma from diseased parenchyma during lobectomy. Anesthesiologists and surgeons need to be aware of the potential complications associated with use of EBB.
Use of Scintigraphy for Diagnosis of Equine Paranasal Sinus Disorders
Authors: BARAKZAI SAFIA , TREMAINE HENRY , DIXON PADDY
Published: January 2006

Objectives— To describe and analyze subjective and objective scintigraphic findings in horses with paranasal sinus disorders.

Animals— Horses with suspected disorders of the paranasal sinuses or cheek teeth (n=48) and control horses (n=30).

Methods— 99Technetium‐methylenediphosphate (99Tc‐MDP; 7500 MBq) was administered for scintigraphic examination of the skull. Abnormal patterns of increased radionuclide uptake (IRU) were identified and subjectively described. Scintigrams and radiographs were blindly assessed by 2 clinicians and the accuracy of the imaging modalities was compared. Objective analysis was performed by comparing regions of interest (ROI) drawn over areas of abnormal IRU to the equivalent area on the unaffected side using the Mann–Whitney test. An ROI ratio (affected:non‐affected sides) was also calculated for each lesion.

Results— Twenty‐eight horses were diagnosed with primary (15) or secondary (13) sinusitis. Nine primary sinusitis cases had focal area(s) of moderate or marked IRU in addition to a more diffuse uptake in the affected sinus(es). κ was marginally higher for scintigraphic assessment of paranasal sinus disorders than for radiographic assessment. There were significant differences in mean counts/pixel in ROI drawn on the affected side compared with the non‐affected side. There was considerable overlap between the magnitudes of ROI ratios found with many different disorders.

Conclusions— Scintigraphy can be useful for differentiation of sinusitis of dental origin from other causes of sinusitis. Quantitative analysis of scintigrams is not reliable for identification of particular disorders.

Clinical Relevance— Focal areas of moderate or marked IRU may be observed with primary sinusitis. Careful 3‐dimensional localization of the lesion, along with consideration of other clinical and diagnostic findings should be performed to prevent false‐positive diagnoses of periapical infection in such instances.
Coronary to Pulmonary Artery Fistula—Diagnosis by Transesophageal Echocardiography
Authors: Okwuosa Tochi M. , Gundeck Ethan L. , Ward R. Parker
Published: January 2006

Coronary artery fistulas (CAFs) are rare coronary anomalies which generally require coronary angiography for definitive diagnosis. CAFs most commonly drain into the right atrium or right ventricle, but occasionally can drain into the pulmonary artery. We report the case of an asymptomatic coronary to pulmonary artery fistula, which could only be definitively diagnosed with transesophageal echocardiography.
Psychological impact on implant patients' oral health‐related quality of life
Authors: Ra'ed Omar Abu Hantash , Mahmoud Khalid AL‐Omiri , Ahed Mahmoud AL‐Wahadni
Published: January 2006

Objectives: The literature has shown that patients' satisfaction with dental prostheses is associated with the existence of certain personality profiles. It is important to study such relationships in dental implant patients.

Material and methods: Fifty patients (28 men and 22 women), aged between 22 and 71 years (mean age 43.22 years, SD 12.24 years), who were partially edentulous and were seeking dental implant therapy were entered into this study. The patients were requested to answer two reliable and valid questionnaires – the Dental Impact on Daily Living (DIDL) and the Neuroticism Extraversion Openness Five‐Factor Inventory (NEO‐FFI) – before implant treatment and 2–3 months after prosthodontic rehabilitation therapy.

Results: Certain personality traits were found to have a significant relationship with patients' satisfaction with dental implants both before and after implant therapy (P<0.05). Neuroticism score had valuable features in predicting patients' total satisfaction ratings (P=0), satisfaction with appearance dimension (P=0), satisfaction with oral comfort dimension (P=0.005) as well as satisfaction with general performance dimension (P=0).

Conclusion: Personality traits have an impact on patients' satisfaction with dental implant therapy. In addition, personality traits provide valuable information for the prediction of patients' satisfaction with their implant‐supported prostheses. Neuroticism, openness, agreeableness and consciousness are very helpful in this regard. Neuroticism was found the main predictor of the patients' oral health‐related quality of life following implant treatment.
Radiographic assessment of bone density around integrated titanium implants after ovariectomy in rats
Authors: Sakakura Celso Eduardo , Giro Gabriela , Gonçalves Daniela , Pereira Rosa Maria Rodrigues , Orrico Silvana Regina Perez , Marcantonio Jr Elcio
Published: January 2006

Objectives: This study evaluated the influence of ovariectomy 8 weeks after implant placement on bone integrated to titanium implants.

Material and methods: Thirty‐eight female rats were submitted to a titanium implant at the tibiae proximal methaphysis. After a healing period of 8 weeks the animals were randomly divided into three groups: control (CTL‐10 animals), sham‐operated (SHAM‐14 animals) and ovariectomy (OVX‐14 animals). The CTL group was then sacrificed in order to confirm integration of the implant. The SHAM group was submitted to fictitious surgery and the OVX group was submitted to bilateral ovariectomy. After 12 weeks post‐implant placement, the SHAM and OVX groups were sacrificed. In order to confirm the systemic osteopenia in rats, a dual‐energy X‐ray absorptiometry (DXA) was performed. For the evaluation of bone density, digital radiographs were taken. The grey level of the bone adjacent to implant was measured using image software and the bone density was calculated at six points on both sides of the implant.

Results: Densitometry measurements of the femur confirmed systemic bone mass loss in the OVX group. Individualized bone analyses of different regions surrounding the implant showed a significantly lower radiographic bone density (P<0.05) in the cancellous region of the OVX group (77.48±23.39 grey levels) when compared with the CTL and SHAM groups (91.61±32.10 and 102.57±32.50 grey levels, respectively).

Conclusions: The present study showed a decrease of the radiographic bone density in the cancellous region of bone around titanium implants placed 8 weeks before ovariectomy in rats.
Two Di‐Leucine Motifs Regulate Trafficking of Mucolipin‐1 to Lysosomes
Authors: Vergarajauregui Silvia , Puertollano Rosa
Published: January 2006

Mutations in the mucolipin‐1 gene have been linked to mucolipidosis type IV, a lysosomal storage disorder characterized by severe neurological and ophthalmologic abnormalities. Mucolipin‐1 is a membrane protein containing six putative transmembrane domains with both its N‐ and C‐termini localized facing the cytosol. To gain information on the sorting motifs that mediate the trafficking of this protein to lysosomes, we have generated chimeras in which the N‐ and C‐ terminal tail portions of mucolipin‐1 were fused to a reporter gene. In this article, we report the identification of two separate di‐leucine‐type motifs that co‐operate to regulate the transport of mucolipin‐1 to lysosomes. One di‐leucine motif is positioned at the N‐terminal cytosolic tail and mediates direct transport to lysosomes, whereas the other di‐leucine motif is found at the C‐terminal tail and functions as an adaptor protein 2‐dependent internalization motif. We have also found that the C‐terminal tail of mucolipin‐1 is palmitoylated and that this modification might regulate the efficiency of endocytosis. Finally, the mutagenesis of both di‐leucine motifs abrogated lysosomal accumulation and resulted in cell‐surface redistribution of mucolipin‐1. Taken together, these results reveal novel information regarding the motifs that regulate mucolipin‐1 trafficking and suggest a role for palmitoylation in protein sorting.
Composite Zirconium Silicides Through an In Situ Process
Authors: Canel Jérôme , Zaman Jérôme , Bettembourg Julie , Le Flem Marion , Poissonnet Sylvie
Published: January 2006

Zirconium silicides are being investigated for use as neutron reflector materials for the next generation of nuclear power devices. Hot isostatically pressed monolithic Zr3Si2 and reactive sintered ZrXSiY composite are currently under development. The composite is obtained in situ and contains a ZrSi matrix embedding shell‐like Zr, Zr2Si, and ZrSi2 domains with volume ratios depending on the initial Zr/Si ratio. Despite the lack of information on the mechanical properties of zirconium silicides, the composite structure is assumed to have enhanced fracture toughness; conditions to improve it further are discussed on the basis of microstructural observations of crack deflection.
Factors Limiting Post‐logging Seedling Regeneration by Big‐leaf Mahogany (Swietenia macrophylla) in Southeastern Amazonia, Brazil, and Implications for Sustainable Management1
Authors: Grogan James , Galvão Jurandir
Published: January 2006

Post‐logging seedling regeneration density by big‐leaf mahogany (Swietenia macrophylla), a nonpioneer light‐demanding timber species, is generally reported to be low to nonexistent. To investigate factors limiting seedling density following logging within the study region, we quantified seed production rates, germinability, dispersal patterns, and seed fates on the forest floor through germination and the first seedling growing season in southeastern Amazonia, Brazil. Fruit production rates were low by three logged and one unlogged populations compared to reports from other regions. Commercial‐sized trees (>60 cm diameter) were more fecund than noncommercial trees (30–60 cm diameter) at two sites, averaging 14.5 vs. 3.9 fruits/tree/year, respectively, at Marajoara, a logged site, over 8 yr. Fruit capsules contained an average of 60.3 seeds/fruit, 70 percent of which appeared viable by visual inspection. Sixty‐seven to 72 percent of apparently viable seeds germinated in nursery beds 2.5 mo after the dispersal period, when wet season rains began. Dry season winds blew most seeds west‐northwest of parent trees, with median dispersal distances of 28 and 9 m on west and east sides of parent trees, respectively. Nearly 100 percent of seeds fell within an area of 0.91 ha. On the forest floor beneath closed canopies, mammals, invertebrates, and fungal pathogens killed 40 percent of apparently viable seeds, while 36 percent germinated. Nine months after seedling establishment—midway through the first logging season following seed dispersal—14 percent of outplanted seeds survived as seedlings, representing 5.8 seeds/fruit. We conclude that seedlings are likely to survive in logging gaps at appreciable densities only in rare cases where previous year fruit production rates by logged trees were high (4–12.5% of commercial‐sized trees/year at Marajoara) and where tree crowns were felled in west or northwest directions.
Exact Solution for Two‐Dimensional Flow to a Well in an Anisotropic Domain
Authors: Fitts Charles R.
Published: January 2006

Although most current applications of the analytic element method are formulated for isotropic hydraulic conductivity, anisotropic domains can be modeled with analytic elements using the well‐known coordinate transformation where one coordinate axis is scaled by the square root of the anisotropy ratio. If the standard analytic solution for steady radial flow to a well is used with this coordinate transformation, the resulting solution correctly models the far field but it does not meet the constant head boundary condition at the well radius. This could be a significant shortcoming if you are interested in the flow field close to the well or want to estimate the head at the pumping well. A new solution for two‐dimensional steady flow to a well in an anisotropic domain is presented. This solution satisfies the governing equations exactly and meets the constant head boundary condition at the well radius exactly. It was derived using a conformal mapping.
Synergistic and Antagonistic Effects of Combined Subzero Temperature and High Pressure on Inactivation ofEscherichia coli
Authors: Moussa Marwen , Perrier-Cornet Jean-Marie , Gervais Patrick
Published: January 2006

The combined effects of subzero temperature and high pressure on the inactivation ofEscherichia coliK12TG1 were investigated. Cells of this bacterial strain were exposed to high pressure (50 to 450 MPa, 10-min holding time) at two temperatures (−20°C without freezing and 25°C) and three water activity levels (aw) (0.850, 0.992, and ca. 1.000) achieved with the addition of glycerol. There was a synergistic interaction between subzero temperature and high pressure in their effects on microbial inactivation. Indeed, to achieve the same inactivation rate, the pressures required at −20°C (in the liquid state) were more than 100 MPa less than those required at 25°C, at pressures in the range of 100 to 300 MPa with an awof 0.992. However, at pressures greater than 300 MPa, this trend was reversed, and subzero temperature counteracted the inactivation effect of pressure. When the amount of water in the bacterial suspension was increased, the synergistic effect was enhanced. Conversely, when the awwas decreased by the addition of solute to the bacterial suspension, the baroprotective effect of subzero temperature increased sharply. These results support the argument that water compression is involved in the antimicrobial effect of high pressure. From a thermodynamic point of view, the mechanical energy transferred to the cell during the pressure treatment can be characterized by the change in volume of the system. The amount of mechanical energy transferred to the cell system is strongly related to cell compressibility, which depends on the water quantity in the cytoplasm.
Production and Characterization of a Thermostable Alcohol Dehydrogenase That Belongs to the Aldo-Keto Reductase Superfamily
Authors: Machielsen Ronnie , Uria Agustinus R. , Kengen Servé W. M. , van der Oost John
Published: January 2006

The gene encoding a novel alcohol dehydrogenase that belongs to the aldo-keto reductase superfamily has been identified in the hyperthermophilic archaeon
Pyrococcus furiosus
. The gene, referred to as
adhD
, was functionally expressed in
Escherichia coli
and subsequently purified to homogeneity. The enzyme has a monomeric conformation with a molecular mass of 32 kDa. The catalytic activity of the enzyme increases up to 100°C, and a half-life value of 130 min at this temperature indicates its high thermostability. AdhD exhibits a broad substrate specificity with, in general, a preference for the reduction of ketones (pH optimum, 6.1) and the oxidation of secondary alcohols (pH optimum, 8.8). Maximal specific activities were detected with 2,3-butanediol (108.3 U/mg) and diacetyl-acetoin (22.5 U/mg) in the oxidative and reductive reactions, respectively. Gas chromatrography analysis indicated that AdhD produced mainly (S)-2-pentanol (enantiomeric excess, 89%) when 2-pentanone was used as substrate. The physiological role of AdhD is discussed.
Identification, Mutagenesis, and Transcriptional Analysis of the Methanesulfonate Transport Operon of
Methylosulfonomonas methylovora
Authors: Jamshad Mohammed , De Marco Paolo , Pacheco Catarina C. , Hanczar Timea , Murrell J. Colin
Published: January 2006

Recently identified genes located downstream (3′) of the
msmEF
(transport encoding) gene cluster,
msmGH
, and located 5′ of the structural genes for methanesulfonate monooxygenase (MSAMO) are described from
Methylosulfonomonas methylovora
. Sequence analysis of the derived polypeptide sequences encoded by these genes revealed a high degree of identity to ABC-type transporters. MsmE showed similarity to a putative periplasmic substrate binding protein, MsmF resembled an integral membrane-associated protein, and MsmG was a putative ATP-binding enzyme. MsmH was thought to be the cognate permease component of the sulfonate transport system. The close association of these putative transport genes to the MSAMO structural genes
msmABCD
suggested a role for these genes in transport of methanesulfonic acid (MSA) into
M. methylovora. msmEFGH
and
msmABCD
constituted two operons for the coordinated expression of MSAMO and the MSA transporter systems. Reverse-transcription-PCR analysis of
msmABCD
and
msmEFGH
revealed differential expression of these genes during growth on MSA and methanol. The
msmEFGH
operon was constitutively expressed, whereas MSA induced expression of
msmABCD
. A mutant defective in
msmE
had considerably slower growth rates than the wild type, thus supporting the proposed role of MsmE in the transport of MSA into
M. methylovora
.
Real-Time PCR for the Detection and Quantification of
Geodermatophilaceae
from Stone Samples and Identification of New Members of the Genus
Blastococcus
Authors: Salazar Oscar , Valverde Aranzazu , Genilloud Olga
Published: January 2006

Real-time PCR (RT-PCR) technology was used for the specific detection and quantification of members of the family
Geodermatophilaceae
in stone samples. Differences in the nucleotide sequences of the 16S rRNA gene region were used to design a pair of family-specific primers that were used to detect and quantify by RT-PCR DNA from members of this family in stone samples from different geographical origins in Spain. These primers were applied later to identify by PCR-specific amplification new members of the family
Geodermatophilaceae
isolated from the same stone samples. The diversity and taxonomic position of the wild-type strains identified from ribosomal sequence analysis suggest the presence of a new lineage within the genus
Blastococcus
.
A
Bacillus thuringiensis
S-Layer Protein Involved in Toxicity against
Epilachna varivestis
(Coleoptera: Coccinellidae)
Authors: Peña Guadalupe , Miranda-Rios Juan , de la Riva Gustavo , Pardo-López Liliana , Soberón Mario , Bravo Alejandra
Published: January 2006

The use of
Bacillus thuringiensis
as a biopesticide is a viable alternative for insect control since the insecticidal Cry proteins produced by these bacteria are highly specific; harmless to humans, vertebrates, and plants; and completely biodegradable. In addition to Cry proteins,
B. thuringiensis
produces a number of extracellular compounds, including S-layer proteins (SLP), that contribute to virulence. The S layer is an ordered structure representing a proteinaceous paracrystalline array which completely covers the surfaces of many pathogenic bacteria. In this work, we report the identification of an S-layer protein by the screening of
B. thuringiensis
strains for activity against the coleopteran pest
Epilachna varivestis
(Mexican bean beetle; Coleoptera: Coccinellidae). We screened two
B. thuringiensis
strain collections containing unidentified Cry proteins and also strains isolated from dead insects. Some of the
B. thuringiensis
strains assayed against
E. varivestis
showed moderate toxicity. However, a
B. thuringiensis
strain (GP1) that was isolated from a dead insect showed a remarkably high insecticidal activity. The parasporal crystal produced by the GP1 strain was purified and shown to have insecticidal activity against
E. varivestis
but not against the lepidopteran
Manduca sexta
or
Spodoptera frugiperda
or against the dipteran
Aedes aegypti
. The gene encoding this protein was cloned and sequenced. It corresponded to an S-layer protein highly similar to previously described SLP in
Bacillus anthracis
(EA1) and
Bacillus licheniformis
(OlpA). The phylogenetic relationships among SLP from different bacteria showed that these proteins from
Bacillus cereus
,
Bacillus sphaericus
,
B. anthracis
,
B. licheniformis
, and
B. thuringiensis
are arranged in the same main group, suggesting similar origins. This is the first report that demonstrates that an S-layer protein is directly involved in toxicity to a coleopteran pest.
Routes of Intraspecies Transmission ofMycobacterium aviumsubsp.paratuberculosisin Rabbits (Oryctolagus cuniculus): a Field Study
Authors: Judge Johanna , Kyriazakis Ilias , Greig Alastair , Davidson Ross S. , Hutchings Michael R.
Published: January 2006

Rabbits have been increasingly linked to the persistence of paratuberculosis (Johne's disease) in domestic ruminants in the United Kingdom. The aims of this study were to determine the routes of intraspecies transmission ofMycobacterium aviumsubspeciesparatuberculosis(MAP) in rabbits and to estimate the probability of transmission via each route, in order to gain understanding of the dynamics of MAP in this host. Rabbits were sampled from two sites where MAP had previously been isolated from the livestock and rabbit populations. No pathology was noted in any animals, but the overall prevalence of MAP in rabbits was high at both sites studied, 39.7% and 23.0%, respectively. MAP was isolated from the testes, uterus, placenta, fetuses, and milk. This is the first time that the bacterium has been isolated from any of these tissues in a nonruminant wildlife species. These results suggest that transmission may occur vertically, pseudovertically, and horizontally. Vertical, i.e., transplacental, and/or pseudo-vertical, i.e., through the ingestion of contaminated milk and/or feces, transmission occurred in 14% of offspring entering the population at 1 month of age. As infection via these routes is only possible from infected adult females, this equates to a probability of infection via this route of 0.326. Probability of infection via horizontal transmission (including interspecies transmission) occurred at up to 0.037 per month. The presence of these routes of transmission within natural rabbit populations will contribute to the maintenance of MAP infections within such populations and, therefore, the environment.
Expression of Transcription Activator ComK of
Bacillus subtilis
in the Heterologous Host
Lactococcus lactis
Leads to a Genome-Wide Repression Pattern: a Case Study of Horizontal Gene Transfer
Authors: Susanna Kim A. , den Hengst Chris D. , Hamoen Leendert W. , Kuipers Oscar P.
Published: January 2006

Horizontal gene transfer (HGT) is generally considered a possible mechanism by which bacteria acquire new genetic properties. Especially when pathogenicity genes are involved, HGT might have important consequences for humans. In this report we describe a case study of HGT in which a transcriptional activator, ComK of
Bacillus subtilis
, was introduced into a heterologous host species,
Lactococcus lactis
. ComK is the central regulator of competence development, activating transcription by binding to a ComK-binding site, a so-called K-box. Interestingly,
L. lactis
does not contain a
comK
gene, but it does contain almost 400 putatively functional K-boxes, as well as homologues of a number of competence genes. In this study, the effect of HGT of
B. subtilis comK
into
L. lactis
was investigated by determining the effects on the transcription profile using DNA microarray analyses. Production of wild-type ComK was shown to stimulate the transcription of 89 genes and decrease the expression of 114 genes. Notably, potential direct effects (i.e., genes preceded by a K-box) were found mainly among repressed genes, suggesting that ComK functions as a repressor in
L. lactis
. This is a remarkable difference between
L. lactis
and
B. subtilis
, in which ComK almost exclusively activates transcription. Additional DNA microarray analyses with a transcription activation-deficient but DNA-binding ComK variant, ComKΔC25, demonstrated that there were similar effects on gene regulation with this variant and with wild-type ComK, confirming that the direct effects of ComK result from interference with normal transcription through binding to available K-boxes. This study demonstrates that horizontal gene transfer can have dramatic effects that are very different than those that are expected on basis of the original functionality of a gene.
Growth of
Dehalobacter
and
Dehalococcoides
spp. during Degradation of Chlorinated Ethanes
Authors: Grostern Ariel , Edwards Elizabeth A.
Published: January 2006

Mixed anaerobic microbial subcultures enriched from a multilayered aquifer at a former chlorinated solvent disposal facility in West Louisiana were examined to determine the organism(s) involved in the dechlorination of the toxic compounds 1,2-dichloroethane (1,2-DCA) and 1,1,2-trichloroethane (1,1,2-TCA) to ethene. Sequences phylogenetically related to
Dehalobacter
and
Dehalococcoides
, two genera of anaerobic bacteria that are known to respire with chlorinated ethenes, were detected through cloning of bacterial 16S rRNA genes. Denaturing gradient gel electrophoresis analysis of 16S rRNA gene fragments after starvation and subsequent reamendment of culture with 1,2-DCA showed that the
Dehalobacter
sp. grew during the dichloroelimination of 1,2-DCA to ethene, implicating this organism in degradation of 1,2-DCA in these cultures. Species-specific real-time quantitative PCR was further used to monitor proliferation of
Dehalobacter
and
Dehalococcoides
during the degradation of chlorinated ethanes and showed that in fact both microorganisms grew simultaneously during the degradation of 1,2-DCA. Conversely,
Dehalobacter
grew during the dichloroelimination of 1,1,2-TCA to vinyl chloride (VC) but not during the subsequent reductive dechlorination of VC to ethene, whereas
Dehalococcoides
grew only during the reductive dechlorination of VC but not during the dichloroelimination of 1,1,2-TCA. This demonstrated that in mixed cultures containing multiple dechlorinating microorganisms, these organisms can have either competitive or complementary dechlorination activities, depending on the chloro-organic substrate.
Effects of Endogenous Substrates on Adaptation of Anaerobic Microbial Communities to 3-Chlorobenzoate
Authors: Becker Jennifer G. , Berardesco Gina , Rittmann Bruce E. , Stahl David A.
Published: January 2006

Lengthy adaptation periods in laboratory studies evaluating the potential for contaminant biodegradation in natural or engineered environments may indicate that the native microbial communities are not metabolizing the contaminants in situ. In this study, we characterized the adaptation period preceding the biodegradation of 3-chlorobenzoate in anaerobic communities derived from lake sediment and wastewater sludge digesters. The importance of alternative mechanisms of adaptation of the anaerobic communities to 3-chlorobenzoate was evaluated by monitoring the concentrations of metabolic substrates and products as well as the levels of total small subunit (SSU) rRNA and SSU rRNA from populations thought to be important in 3-chlorobenzoate mineralization. The anaerobic environments from which the 3-chlorobenzoate-degrading communities were derived contained different levels of endogenous substrates. Increasing methane levels in the digester and sediment communities and decreasing chemical oxygen demand concentrations in the sediment community during the adaptation periods revealed that endogenous substrates were preferentially utilized relative to 3-chlorobenzoate. Methane and chemical oxygen demand concentrations leveled off concomitantly with the onset of 3-chlorobenzoate biodegradation, suggesting that depletion of the preferentially degraded endogenous substrates stimulated 3-chlorobenzoate metabolism. Consistent with these observations, adaptation to 3-chlorobenzoate occurred more rapidly in digester samples that were depleted of endogenous substrates compared to samples that contained high levels of these biodegradable compounds. Other potential adaptation mechanisms, e.g., genetic change or selective population enrichment, appeared to be less important based on the reproducibility and relative lengths of the adaptation events, trends in the SSU rRNA levels, and/or amplification of SSU rRNA genes from key populations.
Involvement of Cyclopropane Fatty Acids in the Response of
Pseudomonas putida
KT2440 to Freeze-Drying
Authors: Muñoz-Rojas Jesús , Bernal Patricia , Duque Estrella , Godoy Patricia , Segura Ana , Ramos Juan-Luis
Published: January 2006

Pseudomonas putida
KT2440, a saprophytic soil bacterium that colonizes the plant root, is a suitable microorganism for the removal of pollutants and a stable host for foreign genes used in biotransformation processes. Because of its potential use in agriculture and industry, we investigated the conditions for the optimal preservation of the strain and its derivatives for long-term storage. The highest survival rates were achieved with cells that had reached the stationary phase and which had been subjected to freeze-drying in the presence of disaccharides (trehalose, maltose, and lactose) as lyoprotectants. Using fluorescence polarization techniques, we show that cell membranes of KT2440 were more rigid in the stationary phase than in the exponential phase of growth. This is consistent with the fact that cells grown in the stationary phase exhibited a higher proportion of C
17:cyclopropane
as a fatty acid than cells in the exponential phase. Mutants for the
cfaB
gene, which encodes the main C
17:cyclopropane
synthase, and for the
cfaA
gene, which encodes a minor C
17:cyclopropane
synthase, were constructed. These mutants were more sensitive to freeze-drying than wild-type cells, particularly the mutant with a knockout in the
cfaB
gene that produced less than 2% of the amount of C
17:cyclopropane
produced by the parental strain.
Real-Time Fluorogenic Reverse Transcription-PCR Assays for Detection of Bacteriophage MS2
Authors: O'Connell Kevin P. , Bucher Jennifer R. , Anderson Patricia E. , Cao Cheng J. , Khan Akbar S. , Gostomski Mark V. , Valdes James J.
Published: January 2006

Bacteriophage MS2 is used in place of pathogenic viruses in a wide variety of studies that range from testing of compounds for disinfecting surfaces to studying environmental transport and fate of pathogenic viruses in groundwater. MS2 is also used as a pathogen simulant in the research, development, and testing (including open air tests) of methods, systems, and devices for the detection of pathogens in both the battlefield and homeland defense settings. PCR is often used as either an integral part of such detection systems or as a reference method to assess the sensitivity and specificity of microbial detection. To facilitate the detection of MS2 by PCR, we describe here a set of real-time fluorogenic reverse transcription-PCR assays. The sensitivity of the assays (performed with primer pairs and corresponding dye-labeled probes) ranged from 0.4 to 40 fg of MS2 genomic RNA (200 to 20,000 genome equivalents). We also demonstrate the usefulness of the primer pairs in assays without dye-labeled probe that included the DNA-binding dye SYBR green. None of the assays gave false-positive results when tested against 400 pg of several non-MS2 nucleic acid targets.