bluuebunny/crossref_metadata_2025 huggingface dataset:
Effect of Telephone Counseling on Physical Activity for Low‐Active Older People in Primary Care: A Randomized, Controlled Trial
Authors: Kolt Gregory S. , Schofield Grant M. , Kerse Ngaire , Garrett Nick , Oliver Melody
Published: May 2007

OBJECTIVES: To assess the long‐term effectiveness of a telephone counseling intervention on physical activity and health‐related quality of life in low‐active older adults recruited through their primary care physician.

DESIGN: Randomized, controlled trial.

SETTING: Three primary care practices from different socioeconomic regions of Auckland, New Zealand.

PARTICIPANTS: One hundred and eighty‐six low‐active adults (aged 65) recruited from their primary care physicians' patient databases.

INTERVENTION: Eight telephone counseling sessions over 12 weeks based on increasing physical activity. Control patients received usual care.

MEASUREMENTS: Change in physical activity (as measured using the Auckland Heart Study Physical Activity Questionnaire) and quality of life (as measured using the Short Form‐36 Health Survey (SF‐36)) over a 12‐month period.

RESULTS: Moderate leisure physical activity increased by 86.8 min/wk more in the intervention group than in the control group (P=.007). More participants in the intervention group reached 2.5 hours of moderate or vigorous leisure physical activity per week after 12 months (42% vs 23%, odds ratio=2.9, 95% confidence interval=1.33–6.32, P=.007). No differences on SF‐36 measures were observed between the groups at 12 months.

CONCLUSION: Telephone‐based physical activity counseling is effective at increasing physical activity over 12 months in previously low‐active older adults.
An Electronic Medical Record Intervention Increased Nursing Home Advance Directive Orders and Documentation
Authors: Lindner Serge A. , Ben Davoren J. , Vollmer Andrew , Williams Brie , Seth Landefeld C.
Published: May 2007

OBJECTIVES: To develop an electronic medical record intervention to improve documentation of patient preferences about life‐sustaining care, detail of resuscitation and treatment‐limiting orders, and concordance between these orders and patient preferences.

DESIGN: Prospective before–after intervention trial.

SETTING: Veterans Affairs nursing home with an electronic medical record for all clinical information, including clinician orders.

PARTICIPANTS: All 224 nursing home admissions from May 1 to October 31, 2004.

MEASUREMENTS: Completion of an advance directive discussion note by the primary clinician, clinician orders about resuscitation and other life‐sustaining treatments, and concordance between these orders and documented patient preferences.

INTERVENTION: The electronic medical record was modified so that an admission order would specify resuscitation status. Additionally, the intervention alerted the primary clinician to complete a templated advance directive discussion note for documentation of life‐sustaining treatment preferences.

RESULTS: Primary clinicians completed an advance directive discussion note for five of 117 (4%) admissions pre‐intervention and 67 of 107 (63%) admissions post‐intervention (P<.001). In multivariate analysis, the intervention was independently associated with advance directive discussion note completion (odds ratio=42, 95% confidence interval=15–120). Of patients who preferred do‐not‐resuscitate (DNR) status, a DNR order was written for 86% pre‐intervention versus 98% post‐intervention (P=.07); orders to limit other life‐sustaining treatments were written for 16% and 40%, respectively (P=.01).

CONCLUSIONS: A targeted electronic medical record intervention increased completion of advance directive discussion notes in seriously ill patients. For patients who preferred DNR status, the intervention also increased the frequency of DNR orders and of orders to limit other life‐sustaining treatments.
A Judicial Abandonment of Blacks? Rethinking the “State Action” Cases of the Waite Court
Authors: Brandwein Pamela
Published: May 2007

This article reconsiders the conventional wisdom that the Supreme Court definitively abandoned the freedmen to their former masters through the “state action” decisions of the 1870s and 1880s. Arguing that anachronisms distort our understanding of this critical period, I offer an historical institutional analysis of state action doctrine by recovering the legal categories, assumptions, and distinctions that constituted judicial discourse about the state action rule. Showing that federal power to protect blacks was more intact than scholars realize, I also add a perspective from the sociology of knowledge. By examining a series of modern developments that erased the contexts of the state action decisions, I show how institutional practices gave rise to the anachronisms that this article seeks to correct.
Splitting the Difference: Modeling Appellate Court Decisions with Mixed Outcomes
Authors: Lindquist Stefanie A. , Martinek Wendy L. , Hettinger Virginia A.
Published: May 2007

In rendering a decision in a particular case, judges are not limited to finding simply for the appellant or for the respondent. Rather, in many cases, they have the option to find for the former on one or more issues and for the latter on one or more other issues. By thus “splitting the difference,” judges can render a judgment that favors both litigants to some degree. What accounts for such mixed outcomes? Several theoretical perspectives provide potential explanations for this phenomenon. First, Galanter (1974) suggests that litigants with greater resources will achieve more favorable outcomes in the courts. Where two high-resource, repeat-player litigants meet in the appeals courts, these more sophisticated and successful parties may be able to persuade the court to render decisions with mixed outcomes that at least partially favor each party. Second, split outcomes may result from strategic interactions among the appeals court judges on the decisionmaking panel. Where majority opinion writers seek to accommodate other judges on the panel, split outcomes have the potential to serve as an inducement for more ideologically extreme judges to join the majority opinion. Finally, Shapiro and Stone Sweet (Stone Sweet 2000; Shapiro & Stone Sweet 2002) propose that courts will sometimes split the difference in order to enhance their legitimacy (and ultimately enhance compliance by losing parties). For example, in highly salient cases, where noncompliance would more clearly threaten court legitimacy, judges may be more likely to split the difference in order to mollify even the losing party. We develop an empirical model of mixed outcomes to test these propositions using data available from the U. S. Courts of Appeals Database and find evidence supportive of all three theoretical perspectives.
Armed, and Dangerous (?): Motivating Rule Adherence Among Agents of Social Control
Authors: Tyler Tom R. , Callahan Patrick E. , Frost Jeffrey
Published: May 2007

A key concern within democracies is effectively regulating the behavior of societies' agents of social control, who have coercive power and considerable discretion over their use of that power. This can result in failures to adhere to the rules, policies, and laws dictating appropriate and lawful behavior. This article explores the effectiveness of motivating rule adherence among law enforcement officers and soldiers by focusing upon whether they believe that organizational authorities are legitimate or that rules and policies are morally right or wrong. The results suggest that both values have an important influence upon rule adherence. Further, aspects of organizational culture that encourage such values are identified and shown to be influential in this setting. Results show that the procedural justice of the organization is central to rule adherence. These findings support the argument that encouraging self-regulation via appeals to the values of law enforcement officers and soldiers is a viable strategy for minimizing misconduct, and they suggest how to effectively implement such approaches.
Urban Ethnography's “Saloon Problem” and Its Challenge to Public Sociology
Authors: Mattson Greggor
Published: May 2007

This essay assesses the legacy of urban ethnography's (UE) early engagement with the “saloon problem.” Early sociologists (1880–1915) intervened in the national debate on alcohol on the basis of their long–term, in–depth understanding of the urban poor. Ethnographers highlighted the role of the saloon as a haven for maintaining social ties while socializing immigrants to American norms. Instead of prohibition or temperance, sociologists advocated replacing the saloon's positive functions with more democratic institutions, especially an egalitarian domestic sphere. This position was shared by both academic and settlement house sociologists whose saloon investigations offer a coherent sociological research paradigm that antedates the Chicago School. The activism of early sociologists exemplifies the characteristics of Michael Burawoy's recent call for public sociology. Yet the early sociologists failed to redeem the saloon amongst Progressives, who increasingly rallied around the National Anti–Saloon League and constitutional Prohibition. By only investigating alcohol in its public manifestations, sociologists failed to challenge the way the social problem was framed and may even have contributed to the stigmatization of the saloon. This voyeuristic opportunism has plagued the American tradition of urban ethnography, the ineffective legacy of which poses a challenge to a contemporary revival of public sociology.
Women, Race, and Ethnicity: Exploring Earnings Differentials in Metropolitan America
Authors: Reid Lesley Williams , Adelman Robert M. , Jaret Charles
Published: May 2007

We draw on leading theories about the structural causes of inequality in the United States to explore inter–metropolitan differences in average earnings for white, black, Hispanic, and Asian women. Our analysis utilizes 2000 census data for a sample of 150 metropolitan areas to investigate the determinants of both women's median earnings and earnings’ inequality by race and ethnicity. We find substantial differences between the earnings of minority and white women across metropolitan areas, although the differences are not in the same direction for all groups. Among other findings, our results indicate: (1) The more retail trade and educational, health, and social service employment, the lower the earnings of most women; (2) the larger the immigrant population in an area, the higher the earnings of white and Asian, but not black or Hispanic women; and (3) residing in the South increases levels of inequality between black and white women. In summary, our results indicate that conventional predictors of aggregate earnings and earnings’ inequality operate differently for white, black, Hispanic, and Asian women at the metropolitan level. Structural characteristics of metropolitan areas all have some influence on women's economic outcomes; but those influences are consistent neither for the earnings of all groups of women nor for earnings’ inequality between groups of women.
Subprime Refinancing: Equity Extraction and Mortgage Termination
Authors: Pennington‐Cross Anthony , Chomsisengphet Souphala
Published: May 2007

This article examines the choice of borrowers to extract wealth from housing in the high‐cost (subprime) segment of the mortgage market and assesses the prepayment and default performance of these cash‐out refinance loans relative to the rate of refinance loans. Consistent with survey evidence, the propensity to extract equity is sensitive to the relative interest rates of other forms of consumer debt. After the loan is originated, our results indicate that cash‐out refinances perform differently from non–cash‐out refinances. For example, cash‐outs are less likely to default or prepay, and the termination of cash‐outs is more sensitive to changing interest rates and house prices.
Cardiac Resynchronization Therapy: Variations in Echo‐Guided Optimized Atrioventricular and Interventricular Delays During Follow‐Up
Authors: Valzania Cinzia , Biffi Mauro , Martignani Cristian , Diemberger Igor , Bertini Matteo , Ziacchi Matteo , Bacchi Letizia , Rocchi Guido , Rapezzi Claudio , Branzi Angelo , Boriani Giuseppe
Published: May 2007

Background: Relatively few data are available on long‐term echocardiographic optimization of atrioventricular (AV) and interventricular (VV) delay programming in cardiac resynchronization therapy (CRT). We assessed variations in optimized AV and VV delays during long‐term follow‐up. Methods: Thirty‐seven consecutive heart failure patients received Doppler echocardiographic optimization of AV and VV delay within 48 hours from CRT device implantation, at 6 months and at 12 months (the last for the first enrolled 14 patients). Results: After implantation, median optimized AV delay was 100 ms (range, 45 ms); VV optimization led to simultaneous biventricular activation in 4 patients, left ventricular preactivation in 17 patients and right ventricular preactivation in 16 patients. At 12 months median AV delay decreased to 85 ms (23 ms) (P < 0.05 vs. baseline). With respect to previous assessment, VV delay variations ≥40 ms were observed in 41% of the patients at 6 months and in 57% of the tested patients at 12 months. A nonconcordance (by Kappa test) of optimized VV delays was found between each new assessment and the previous one. VV delay optimization was associated with significant (P < 0.001) increases in aortic velocity time integral both at baseline and during follow‐up. Conclusions: Echocardiographic optimization of AV and VV delay is associated with broad intraindividual variability during follow‐up. A new assessment of optimized VV delays during long‐term follow‐up reveals a nonconcordance with previous values and provides increases in forward stroke volume.
An Allegory of Bildung
Authors: Kluge Sofie
Published: May 2007

This article proposes an interpretation of Friedrich Schlegel's critical essay ‘Über Goethes Meister’ (1798). Conspicuously subjecting Goethe's classical Bildungsroman to what we may term a ‘deconstructive’ reading ante terminem, Schlegel's famous review may be interpreted as an underplayed Romantic refutation of Goethe's Classicist aesthetics. Advancing a daring allegorical interpretation of the novel's central concept of Bildung– not just referring to ‘formation’ in the delimited social and educational sense, but also to ‘formation’ as the infinite principle of aesthetic creation, the inexhaustible source of art – Schlegel claims the literary text's deviation from the author's official aesthetic programme, the organic aesthetics of the symbol, and provocatively inserts it into a transcendental allegorical aesthetics. He thereby ultimately turns Wilhelm Meisters Lehrjahre, the emblem of the secular bourgeois novel, into a paradigm of the Romantic work of art as progressive Universalpoesie.
Periodontal tissue regeneration by recombinant human transforming growth factor‐β3 in Papio ursinus
Authors: Teare J. A. , Ramoshebi L. N. , Ripamonti U.
Published: May 2007

Background and Objective:  Osteogenic proteins of the transforming growth factor‐β superfamily induce periodontal tissue regeneration in animal models, including primates. To our knowledge, no studies have been performed in periodontal regeneration using the transforming growth factor‐β3 isoform. In the present study, recombinant human transforming growth factor‐β3 was examined for its ability to induce periodontal tissue regeneration in the nonhuman primate, Papio ursinus.

Material and Methods:  Class II furcation defects were surgically created bilaterally in the maxillary and mandibular molars of four adult baboons. Heterotopic ossicles, for transplantation to selected furcation defects, were induced within the rectus abdominis muscle by recombinant human transforming growth factor‐β3. Forty days later, the periodontal defects were implanted with recombinant human transforming growth factor‐β3 in Matrigel® as the delivery system, with recombinant human transforming growth factor‐β3 plus minced muscle tissue in Matrigel®, or with the harvested recombinant human transforming growth factor‐β3‐induced ossicles. Sixty days after periodontal implantation, the animals were killed and the specimens harvested. Histological analysis on undecalcified sections measured the area and volume of new alveolar bone and the coronal extension of newly formed alveolar bone and cementum.

Results:  Morphometric analyses showed pronounced periodontal regeneration in experimental defects compared with controls. Substantial regeneration was observed in defects implanted with fragments of heterotopically induced ossicles and with recombinant human transforming growth factor‐β3 plus minced muscle tissue.

Conclusion:  Recombinant human transforming growth factor‐β3 in Matrigel® significantly enhanced periodontal tissue regeneration in the nonhuman primate, P. ursinus.
Esthetic and Functional Rehabilitation of a Patient with Nonsyndromic Oligodontia: A Case Report from China
Authors: He Xiangyi , Shu Weina , Kang Yanli , Li Zhiqiang , Zhang Jing , Kari Kirsti , Meurman Jukka H.
Published: May 2007

Statement of the Problem:  Oligodontia (severe partial anodontia) is a developmental dental anomaly. It is either an isolated trait or part of a syndrome. Oligodontia is characterized by the congenital absence of more than six permanent teeth except the third molars. Treatment often calls for facilitated surgical techniques, but less severe cases can be treated conventionally in a normally equipped dental office.

Case Presentation:  A case of oligodontia treated with prosthetic rehabilitation is presented in this article to remind clinicians of the anomaly. A medical history was taken and clinical and radiographic examinations were made. It is emphasized that conventional prosthetic treatment can lead to a satisfactory result. The patient was treated with fully extending denture prostheses. An overdenture was designed in the maxillary arch, while a removable partial denture was made in the mandibular arch.

Results:  The patient’s speech and masticatory function improved greatly. He was also pleased with better facial esthetics. Observed temporomandibular joint dysfunction also ameliorated after the treatment.

Conclusion:  Dental clinicians should keep in mind that there are good possibilities with conventional prosthodontic techniques to help patients with dental anomalies. Treatment not only improves speech and masticatory function but also has psychological implications that may greatly help in regaining self‐confidence.

CLINICAL SIGNIFICANCE

Patients suffering from oligodontia may have severe psychological, esthetic, and functional problems. Thus, early diagnosis and treatment of these patients are necessary.
Predictive Medical Information and Underwriting
Authors: Dodge John H.
Published: May 2007

Predictive medical information is used by underwriters to assess the future risk of a claim in medically based insurance products such as health, life, and disability insurance. Medical underwriting involves the science of evaluating medical information to determine the risk for groups of individuals with various medical conditions. In disability insurance, this involves an evaluation of medical information to predict the risk of becoming disabled.

Before discussing medical underwriting, an understanding of certain terms used by disability insurance companies and the products that are available is required. The first is the definition of disability. The Americans with Disabilities Act (ADA) defines disability as the following:

A physical or mental impairment that substantially limits one or more of the major life activities; or has a record of such an impairment; or is regarded as having such an impairment even when no impairment exists, no substantial limitation results from the impairment, or the impairment is only substantially limiting because of the attitudes of others.
Genetic Screening and Disability Insurance: What Can We Learn from the Health Insurance Experience?
Authors: Kass Nancy , Medley Amy
Published: May 2007

The Human Genome Project has allowed researchers to gain new insights into the genetic causes of health and disease. With this knowledge comes the potential to develop new genetic tests that are capable of predicting the risk of disease or disability among presently healthy individuals. This information is potentially beneficial in that it may allow individuals to develop strategies to reduce their risk of illness and may allow health providers to recognize and treat the early stages of disease more effectively. As knowledge about genetic contributions to disease continues to grow and genetic testing technology becomes more widespread, there is a risk that personal genetic information could be used in ways potentially concerning to consumers. Genetic testing technology could potentially be used to discriminate against individuals in terms of their ability to obtain health, life, and disability insurance. Decreased access to insurance, in turn, significantly hinders access to health care and income for oneself and/or for one's dependent(s).
Workers' Compensation, Social Security Disability, SSI, and Genetic Testing
Authors: Sedo Kathryn J.
Published: May 2007

In addition to disability insurance purchased privately by individuals or employers, three other major types of disability insurance are available: Workers’ Compensation, Social Security Disability Insurance (SSDI), and Supplemental Security Insurance (SSI). The first two, Workers’ Compensation and SSDI, are available to individuals with work connections. The third, SSI, does not require a work connection.

Workers’ Compensation laws were initially passed to provide economic protection for workers and their families when a worker suffered an accident on the job resulting in an injury. The first laws passed were subject to court challenges based on their constitutionality. Ultimately courts approved the constitutionality of the Workers’ Compensation laws rationalizing that the laws created a compromise whereby an employee gave up her right to sue for a tort recovery in exchange for the employer's agreement to pay benefits without regard to the cause of the accident.
An Interval Analysis Algorithm for Automated Controller Synthesis in QFT Designs
Authors: Nataraj P. S. V. , Tharewal Sachin
Published: May 2007

In this paper, an interval analysis algorithm is proposed for the automatic synthesis of fixed structure controllers in quantitative feedback theory (QFT). The proposed algorithm is tested on several examples and compared with the controller designs given in the QFT literature. Compared to the existing methods for QFT controller synthesis, the proposed algorithm yields considerable improvement in the high frequency gain of the controller in all examples, and improvements in the cutoff frequency of the controller in all but one examples. Notation: R denotes the field of real numbers, Rn is the vector space of column vectors of length n with real entries. A real closed nonempty interval is a one-dimensional box, i.e., a pair x=[x̱,x¯] consisting of two real numbers x̱ and x¯ with x̱⩽x¯. The set of all intervals is IR. A box may be considered as an interval vector x=(x1,…,xn)T with components xk=[x̱k,x¯k]. A box x can also be identified as a pair x=[x̱,x¯] consisting of two real column vectors x̱ and x¯ of length n with x̱⩽x¯. A vector x∊Rn is contained in a box x, i.e., x∊x iff x̱⩽x⩽x¯. The set of all boxes of dimension n is IRn. The width of a box x is wid x=x¯−x̱. The range of a function f:Rn→R over a box x is range(f,x)={f(x)∣x∊x}. A natural interval extension of f on the box x is obtained by replacing in the expression for f, all occurrences of reals xi with intervals xi and all real operations with the corresponding interval operations. The natural interval evaluation of f on x is written as f(x). The interval function f(x) is said to be of convergent of order α if widf(x)−wid{range(f,x)}⩽c{widx}α. By the inclusion property of interval arithmetic, range (f,x)⊆f(x).
Two Transition Phase Control Methods for Hard Contact
Authors: Doh Nakju Lett , Chung Wan Kyun , Youm Youngil , Oh Yongwhan
Published: May 2007

A hard contact is an impact that occurs when the robot contacts a stiff environment with a high velocity. In the pretransition phase of the hard contact, it is important to maintain stability while reducing succeeding force peaks and the duration time. For these purposes, we propose two control methods. One is a suppression controller that suppresses position rebounds in-between the impact time. The other is a flexible-damped joint with a joint damping controller. The flexible-damped joint is a new joint type designed to increase the hardware damping. It is controlled by a novel controller, called the joint damping controller, to enhance damping. The major advantage of the proposed methods is a guaranteed stability. The performance of the proposed methods is validated via several comparative experiments.
Adaptive Synchronization Control of Multiple Spacecraft Formation Flying
Authors: Liu Hong-Tao , Shan Jinjun , Sun Dong
Published: May 2007

An adaptive nonlinear synchronization control approach is developed for multiple spacecraft formation flying with elliptical reference orbits. It can guarantee that both the tracking errors and the synchronization errors of the relative positions converge to zero globally, even in the presence of uncertain parameters. The generalized synchronization concept allows us to design various synchronization errors so that different synchronization performance can be obtained. Simulation results of a leader-follower spacecraft pair and the maneuvering of multiple spacecraft in formation flying are presented to verify the effectiveness of the proposed control technique.
Observer-Based Fault Diagnosis of Satellite Systems Subject to Time-Varying Thruster Faults
Authors: Chen Wen , Saif Mehrdad
Published: May 2007

This paper presents a novel fault diagnosis approach in satellite systems for identifying time-varying thruster faults. To overcome the difficulty in identifying time-varying thruster faults by adaptive observers, an iterative learning observer (ILO) is designed to achieve estimation of time-varying faults. The proposed ILO-based fault-identification strategy uses a learning mechanism to perform fault estimation instead of using integrators that are commonly used in classical adaptive observers. The stability of estimation-error dynamics is established and proved. An illustrative example clearly shows that time-varying thruster faults can be accurately identified.
An Approach for Model Updating of a Multiphysics MEMS Micromirror
Authors: Link Ryan J. , Zimmerman David
Published: May 2007

In this work, a general approach is formulated for updating the parameters of systems governed by multiphysics equations. The optimization technique is based on Genetic Algorithms (GAs). GAs represent a class of probabilistic optimization strategies loosely patterned after a simplified evolutionary scheme and Darwin’s “survival of the fittest” concepts. The GA is coupled to a commercially available multiphysics finite element program. In the context of this, issues of mode tracking, eigenvector comparisons, and approximate function evaluations are discussed. The approach is demonstrated on a micro-electro-mechanical (MEMS) micromirror which is governed by both structural and electrostatic physics. The MEMS mirror is characterized dynamically using a laser vibrometer. These experimental measurements are then used in the model updating of the finite element structural model and the electrostatic model. Several interesting observations that were encountered in this development will also be discussed.
The Shaft Torque of a Tandem Axial-Piston Pump
Authors: Manring Noah D. , Mehta Viral S. , Raab Frank J. , Graf Kevin J.
Published: May 2007

The objective of this study is to identify the best indexed position of two rotating groups within a tandem axial-piston pump for attenuating the torque ripple amplitude that is exerted on the shaft. By attenuating the torque ripple characteristics of the pump, other vibration aspects of the machine are also expected to be reduced. In particular, the objectives of this paper are aimed at reducing the noise that is generated by the pump. This paper begins by considering the theoretical torque ripple that is created by the discrete pumping elements of a single rotating group within an axial piston machine. From this analysis, an equation is produced that describes a single pulse for the torque ripple as a function of the average torque and the total number of pistons that are used within the rotating group. By superposing another rotating group on top of the first, and by indexing the angular position of one rotating group relative to the other, a second equation is produced for describing the theoretical torque ripple of a tandem pump design. This equation is also a function of the average shaft torque and the total number of pistons that are used within a single rotating group; however, an additional parameter known as the index angle also appears in this result. This index angle is shown to amplify or attenuate the amplitude of the torque ripple depending upon its value. From these results, it is shown that a proper selection of the index angle can reduce the torque ripple amplitude by as much as 75%.
Rapid Engineering of Bacterial Reporter Gene Fusions by Using Red Recombination
Authors: Gerlach Roman G. , Hölzer Stefanie U. , Jäckel Daniela , Hensel Michael
Published: May 2007

Reporter gene fusions are essential tools for the investigation of gene regulation. Such fusions are traditionally generated by transposon mutagenesis and identified by a suitable selection procedure. Alternatively, specific reporter fusions can be generated by cloning of DNA fragments containing promoters or other regulatory elements in reporter plasmids. Here, we describe a novel approach for the rapid generation of reporter gene fusions in single copies at defined positions in bacterial genomes. This technique utilizes the Red recombinase for the homologous recombination of PCR-generated cassettes containing various currently used reporter genes, such as those for β-galactosidase, luciferase, and green fluorescent protein. The approach allows the generation of transcriptional or translational reporter fusions in a single step without the requirement for recombinant DNA constructs and is applicable to various enterobacterial species. Generation of reporter fusions by Red recombination is rapid, overcomes the current limitations of transposon mutagenesis or reporter plasmids, and offers new options for the study of bacterial gene regulation.
Metabolic Characterization of
Escherichia coli
Strains Adapted to Growth on Lactate
Authors: Hua Qiang , Joyce Andrew R. , Palsson Bernhard Ø. , Fong Stephen S.
Published: May 2007

In comparison with intensive studies of genetic mechanisms related to biological evolutionary systems, much less analysis has been conducted on metabolic network responses to adaptive evolution that are directly associated with evolved metabolic phenotypes. Metabolic mechanisms involved in laboratory evolution of
Escherichia coli
on gluconeogenic carbon sources, such as lactate, were studied based on intracellular flux states determined from
13
C tracer experiments and
13
C-constrained flux analysis. At the end point of laboratory evolution, strains exhibited a more than doubling of the average growth rate and a 50% increase in the average biomass yield. Despite different evolutionary trajectories among parallel evolved populations, most improvements were obtained within the first 250 generations of evolution and were generally characterized by a significant increase in pathway capacity. Partitioning between gluconeogenic and pyruvate catabolic flux at the pyruvate node remained almost unchanged, while flux distributions around the key metabolites phosphoenolpyruvate, oxaloacetate, and acetyl-coenzyme A were relatively flexible over the course of evolution on lactate to meet energetic and anabolic demands during rapid growth on this gluconeogenic carbon substrate. There were no clear qualitative correlations between most transcriptional expression and metabolic flux changes, suggesting complex regulatory mechanisms at multiple levels of genetics and molecular biology. Moreover, higher fitness gains for cell growth on both evolutionary and alternative carbon sources were found for strains that adaptively evolved on gluconeogenic carbon sources compared to those that evolved on glucose. These results provide a novel systematic view of the mechanisms underlying microbial adaptation to growth on a gluconeogenic substrate.
Source Tracking Aerosols Released from Land-Applied Class B Biosolids during High-Wind Events
Authors: Baertsch Carolina , Paez-Rubio Tania , Viau Emily , Peccia Jordan
Published: May 2007

DNA-based microbial source tracking (MST) methods were developed and used to specifically and sensitively track the unintended aerosolization of land-applied, anaerobically digested sewage sludge (biosolids) during high-wind events. Culture and phylogenetic analyses of bulk biosolids provided a basis for the development of three different MST methods. They included (i) culture- and 16S rRNA gene-based identification of
Clostridium bifermentans
, (ii) direct PCR amplification and sequencing of the 16S rRNA gene for an uncultured bacterium of the class
Chloroflexi
that is commonly present in anaerobically digested biosolids, and (iii) direct PCR amplification of a 16S rRNA gene of the phylum
Euryarchaeota
coupled with terminal restriction fragment length polymorphism to distinguish terminal fragments that are unique to biosolid-specific microorganisms. Each method was first validated with a broad group of bulk biosolids and soil samples to confirm the target's exclusive presence in biosolids and absence in soils. Positive responses were observed in 100% of bulk biosolid samples and in less than 11% of the bulk soils tested. Next, a sampling campaign was conducted in which all three methods were applied to aerosol samples taken upwind and downwind of fields that had recently been land applied with biosolids. When average wind speeds were greater than 5 m/s, source tracking results confirmed the presence of biosolids in 56% of the downwind samples versus 3% of the upwind samples. During these high-wind events, the biosolid concentration in downwind aerosols was between 0.1 and 2 μg/m
3
. The application of DNA-based source tracking to aerosol samples has confirmed that wind is a possible mechanism for the aerosolization and off-site transport of land-applied biosolids.
Molecular Characterization ofhsp20, Encoding a Small Heat Shock Protein ofBifidobacterium breveUCC2003
Authors: Ventura Marco , Canchaya Carlos , Zhang Ziding , Fitzgerald Gerald F. , van Sinderen Douwe
Published: May 2007

Small heat shock proteins (sHSPs) are members of a diverse family of stress proteins that are important in cells to protect proteins under stressful conditions. Genome analysis ofBifidobacterium breveUCC2003 revealed a single sHSP-encoding gene, which was classified as ahsp20gene by comparative analyses. Genomic surveillance of available genome sequences indicated thathsp20homologs are not widely distributed in bacteria. In members of the genusBifidobacterium, this gene appears to be present in only 7 of the 30 currently described species. Moreover, phylogenetic analysis using all available bacterial and eukaryotic sHSP sequences revealed a close relationship between bifidobacterial HSP20 and the class B sHSPs found in members of the divisionFirmicutes. The results of this comparative analysis and variation in codon usage content suggest thathsp20was acquired by certain bifidobacteria through horizontal gene transfer. Analysis by slot blot, Northern blot, and primer extension experiments showed that transcription ofhsp20is strongly induced in response to severe heat shock regimens and by osmotic shock.
Postinoculation Protozoan Establishment and Association Patterns of Methanogenic Archaea in the Ovine Rumen
Authors: Ohene-Adjei Samuel , Teather Ronald M. , Ivan Michael , Forster Robert J.
Published: May 2007

Association patterns between archaea and rumen protozoa were evaluated by analyzing archaeal 16S rRNA gene clone libraries from ovine rumen inoculated with different protozoa. Five protozoan inoculation treatments, fauna free (negative control), holotrich and cellulolytic protozoa,
Isotricha
and
Dasytricha
spp.,
Entodinium
spp., and total fauna (type A) were tested. We used denaturing gradient gel electrophoresis, quantitative PCR, and phylogenetic analysis to evaluate the impact of the protozoan inoculants on the respective archaeal communities. Protozoan 18S ribosomal DNA clone libraries were also evaluated to monitor the protozoal population that was established by the inoculation. Phylogenetic analysis suggested that archaeal clones associated with the fauna-free, the
Entodinium
, and the type A inoculations clustered primarily with uncultured phylotypes.
Polyplastron multivesiculatum
was the predominant protozoan strain established by the holotrich and cellulolytic protozoan treatment, and this resulted predominantly in archaeal clones affiliated with uncultured and cultured methanogenic phylotypes (
Methanosphaera stadtmanae
,
Methanobrevibacter ruminantium
, and
Methanobacterium bryantii
). Furthermore, the
Isotricha
and
Dasytricha
inoculation treatment resulted primarily in archaeal clones affiliated with
Methanobrevibacter smithii
. This report provides the first assessment of the influence of protozoa on archaea within the rumen microbial community and provides evidence to suggest that different archaeal phylotypes associate with specific groups of protozoa. The observed patterns may be linked to the evolution of commensal and symbiotic relationships between archaea and protozoa in the ovine rumen environment. This report further underscores the prevalence and potential importance of a rather large group of uncultivated archaea in the ovine rumen, probably unrelated to known methanogens and undocumented in the bovine rumen.
Luciferase Detection during Stationary Phase in
Lactococcus lactis
Authors: Bachmann Herwig , Santos Filipe , Kleerebezem Michiel , van Hylckama Vlieg Johan E. T.
Published: May 2007

The luminescence signal of
luxAB
-encoded bacterial luciferase strongly depends on the metabolic state of the host cell, which restricts the use of this reporter system to metabolically active bacteria. Here we show that in stationary-phase cells of
Lactococcus lactis
, detection of luciferase is significantly improved by the addition of riboflavin or flavin mononucleotide to whole-cell assay systems.
Characterization of Three Classes of Membrane Proteins Involved in Fungal Azole Resistance by Functional Hyperexpression inSaccharomyces cerevisiae
Authors: Lamping Erwin , Monk Brian C. , Niimi Kyoko , Holmes Ann R. , Tsao Sarah , Tanabe Koichi , Niimi Masakazu , Uehara Yoshimasa , Cannon Richard D.
Published: May 2007

The study of eukaryotic membrane proteins has been hampered by a paucity of systems that achieve consistent high-level functional protein expression. We report the use of a modified membrane protein hyperexpression system to characterize three classes of fungal membrane proteins (ABC transporters Pdr5p, CaCdr1p, CaCdr2p, CgCdr1p, CgPdh1p, CkAbc1p, and CneMdr1p, the major facilitator superfamily transporter CaMdr1p, and the cytochrome P450 enzyme CaErg11p) that contribute to the drug resistance phenotypes of five pathogenic fungi and to express human P glycoprotein (HsAbcb1p). The hyperexpression system consists of a set of plasmids that direct the stable integration of a single copy of the expression cassette at the chromosomalPDR5locus of a modified hostSaccharomyces cerevisiaestrain, ADΔ. Overexpression of heterologous proteins at levels of up to 29% of plasma membrane protein was achieved. Membrane proteins were expressed with or without green fluorescent protein (GFP), monomeric red fluorescent protein, His, FLAG/His, Cys, or His/Cys tags. Most GFP-tagged proteins tested were correctly trafficked within the cell, and His-tagged proteins could be affinity purified. Kinetic analysis of ABC transporters indicated that the apparentKmvalue and theVmaxvalue of ATPase activities were not significantly affected by the addition of His tags. The efflux properties of seven fungal drug pumps were characterized by their substrate specificities and their unique patterns of inhibition by eight xenobiotics that chemosensitizedS. cerevisiaestrains overexpressing ABC drug pumps to fluconazole. The modified hyperexpression system has wide application for the study of eukaryotic membrane proteins and could also be used in the pharmaceutical industry for drug screening.
Use of RecombinantEntamoeba histolyticaCysteine Proteinase 1 To Identify a Potent Inhibitor of Amebic Invasion in a Human Colonic Model
Authors: Meléndez-López Samuel G. , Herdman Scott , Hirata Ken , Choi Min-Ho , Choe Youngchool , Craik Charles , Caffrey Conor R. , Hansell Elisabeth , Chávez-Munguía Bibiana , Chen Yen Ting , Roush William R. , McKerrow James , Eckmann Lars , Guo Jianhua , Stanley Samuel L. , Reed Sharon L.
Published: May 2007

Cysteine proteinases are key virulence factors of the protozoan parasiteEntamoeba histolytica. We have shown that cysteine proteinases play a central role in tissue invasion and disruption of host defenses by digesting components of the extracellular matrix, immunoglobulins, complement, and cytokines. Analysis of theE. histolyticagenome project has revealed more than 40 genes encoding cysteine proteinases. We have focused onE. histolyticacysteine proteinase 1 (EhCP1) because it is one of two cysteine proteinases unique to invasiveE. histolyticaand is highly expressed and released. Recombinant EhCP1 was expressed inEscherichia coliand refolded to an active enzyme with a pH optimum of 6.0. We used positional-scanning synthetic tetrapeptide combinatorial libraries to map the specificity of the P1 to P4 subsites of the active site cleft. Arginine was strongly preferred at P2, an unusual specificity among clan CA proteinases. A new vinyl sulfone inhibitor, WRR483, was synthesized based on this specificity to target EhCP1. Recombinant EhCP1 cleaved key components of the host immune system, C3, immunoglobulin G, and pro-interleukin-18, in a time- and dose-dependent manner. EhCP1 localized to large cytoplasmic vesicles, distinct from the sites of other proteinases. To gain insight into the role of secreted cysteine proteinases in amebic invasion, we tested the effect of the vinyl sulfone cysteine proteinase inhibitors K11777 and WRR483 on invasion of human colonic xenografts. The resultant dramatic inhibition of invasion by both inhibitors in this human colonic model of amebiasis strongly suggests a significant role of secreted amebic proteinases, such as EhCP1, in the pathogenesis of amebiasis.
Genomic and Population Analyses of the Mating Type Loci in
Coccidioides
Species Reveal Evidence for Sexual Reproduction and Gene Acquisition
Authors: Mandel M. Alejandra , Barker Bridget M. , Kroken Scott , Rounsley Steven D. , Orbach Marc J.
Published: May 2007

Coccidioides
species, the fungi responsible for the valley fever disease, are known to reproduce asexually through the production of arthroconidia that are the infectious propagules. The possible role of sexual reproduction in the survival and dispersal of these pathogens is unexplored. To determine the potential for mating of
Coccidioides
, we analyzed genome sequences and identified mating type loci characteristic of heterothallic ascomycetes.
Coccidioides
strains contain either a
MAT1-1
or a
MAT1-2
idiomorph, which is 8.1 or 9 kb in length, respectively, the longest reported for any ascomycete species. These idiomorphs contain four or five genes, respectively, more than are present in the
MAT
loci of most ascomycetes. Along with their cDNA structures, we determined that all genes in the
MAT
loci are transcribed. Two genes frequently found in common sequences flanking
MAT
idiomorphs,
APN2
and
COX13
, are within the
MAT
loci in
Coccidioides
, but the
MAT1-1
and
MAT1-2
copies have diverged dramatically from each other. Data indicate that the acquisition of these genes in the
MAT
loci occurred prior to the separation of
Coccidioides
from
Uncinocarpus reesii
. An analysis of 436
Coccidioides
isolates from patients and the environment indicates that in both
Coccidioides immitis
and
C. posadasii
, there is a 1:1 distribution of
MAT
loci, as would be expected for sexually reproducing species. In addition, an analysis of isolates obtained from 11 soil samples demonstrated that at three sampling sites, strains of both mating types were present, indicating that compatible strains were in close proximity in the environment.
Type IV Fimbrial Biogenesis Is Required for Protease Secretion and Natural Transformation inDichelobacter nodosus
Authors: Han Xiaoyan , Kennan Ruth M. , Parker Dane , Davies John K. , Rood Julian I.
Published: May 2007

The objective of this study was to develop an understanding of the molecular mechanisms by which type IV fimbrial biogenesis, natural transformation, and protease secretion are linked in the ovine foot rot pathogen,Dichelobacter nodosus. We have shown that like theD. nodosusfimbrial subunit FimA, the pilin-like protein PilE and the FimN, FimO, and FimP proteins, which are homologs of PilB, PilC, and PilD fromPseudomonas aeruginosa, are essential for fimbrial biogenesis and natural transformation, indicating that transformation requires an intact type IV fimbrial apparatus. The results also showed that extracellular protease secretion in thefimN,fimO,fimP, andpilEmutants was significantly reduced, which represents the first time that PilB, PilC, and PilE homologs have been shown to be required for the secretion of unrelated extracellular proteins in a type IV fimbriate bacterium. Quantitative real-time PCR analysis of the three extracellular protease genesaprV2,aprV5, andbprVshowed that the effects on protease secretion were not mediated at the transcriptional level. Bioinformatic analysis did not identify a classical type II secretion system, and the putative fimbrial biogenesis genepilQwas the only outer membrane secretin gene identified. Based on these results, it is postulated that inD. nodosus, protease secretion occurs by a type II secretion-related process that directly involves components of the type IV fimbrial biogenesis machinery, which represents the only type II secretion system encoded by the small genome of this highly evolved pathogen.
Genetic Interactions of DNA Repair Pathways in the PathogenNeisseria meningitidis
Authors: Davidsen Tonje , Tuven Hanne K. , Bjørås Magnar , Rødland Einar A. , Tønjum Tone
Published: May 2007

The current increase in the incidence and severity of infectious diseases mandates improved understanding of the basic biology and DNA repair profiles of virulent microbes. In our studies of the major pathogen and model organismNeisseria meningitidis, we constructed a panel of mutants inactivating genes involved in base excision repair, mismatch repair, nucleotide excision repair (NER), translesion synthesis, and recombinational repair pathways. The highest spontaneous mutation frequency among theN. meningitidissingle mutants was found in the MutY-deficient strain as opposed tomutSmutants inEscherichia coli, indicating a role for meningococcal MutY in antibiotic resistance development. Recombinational repair was recognized as a major pathway counteracting methyl methanesulfonate-induced alkylation damage in theN. meningitidis. In contrast to what has been shown in other species, meningococcal NER did not contribute significantly to repair of alkylation-induced DNA damage, and meningococcal recombinational repair may thus be one of the main pathways for removal of abasic (apurinic/apyrimidinic) sites and strand breaks in DNA. Conversely, NER was identified as the main meningococcal defense pathway against UV-induced DNA damage.N. meningitidisRecA single mutants exhibited only a moderate decrease in survival after UV exposure as opposed toE. coli recAstrains, which are extremely UV sensitive, possibly reflecting the lack of a meningococcal SOS response. In conclusion, distinct differences betweenN. meningitidisand established DNA repair characteristics inE. coliand other species were identified.