bluuebunny/crossref_metadata_2025 huggingface dataset:
Introducing Backstage – a digital backchannel for large class lectures
Authors: Pohl Alexander , Gehlen‐Baum Vera , Bry François
Published: September 2011

Purpose

This paper seeks to report on the conception of a novel digital backchannel, Backstage, dedicated to large classes, aiming at empowering not only the audience but also the speaker, at promoting the awareness of both audience and speaker, and at promoting an active participation of students in the lecture. In learning settings with a large and typically passive audience, the goal of Backstage is to foster active participation and facilitate collaboration akin to small‐learning groups.

Design/methodology/approach

The authors present the concept of a novel kind of digital backchannel that supports different forms of inter‐student communication via short microblog messages, social evaluation, and ranking of messages by the audience. The backchannel further supports immediate concise feedback to the lecturer of selected and aggregated students' opinions, making it possible to strengthen the lecturer's awareness of students' difficulties.

Findings

The concept is the outcome of a joint effort between computer scientists and educational scientists, and unifies technical and usability aspects with educational claims.

Originality/value

The concept combines computer‐mediated communication with elements of social media that not only foster active participation and collaboration among a large and otherwise passive audience, but also provide social means to self‐regulation and management of the backchannel discourse. The design of the backchannel is strongly influenced by findings of educational sciences.
Retesting the CCAPM Euler equations
Authors: Azar Samih
Published: September 2011

Purpose

This paper seeks to reconsider the Euler equation of the Consumption Capital Asset Pricing Model (CCAPM), to derive a regression‐based model to test it, and to present evidence that the model is consistent with reasonable values for the coefficient of relative risk aversion (CRRA). This runs contrary to the findings of the literature on the equity premium puzzle, but is in agreement with the literature that estimates the CRRA for the purpose of computing the social discount rate, and is in line with the research on labor supply. Tests based on General Method of Moments (GMM) for the same sample produce results that are extremely disparate and unstable. The paper aims to check and find support for the robustness of the regression‐based tests. Habit formation models are also to be evaluated with regression‐based and GMM tests. However, the validity of the regression‐based models depends critically on their functional forms.

Design/methodology/approach

The paper presents empirical evidence that the conventional use of GMM fails because of four pathological features of GMM that are referred to under the general caption of “weak identification”. In addition to GMM, the paper employs linear regression analysis to test the CCAPM, and it is found that the regression residuals follow well‐behaved distributional properties, making valid all statistical inferences, while GMM estimates are highly unstable.

Findings

Four unexpected findings are reported. The first is that the regression‐based models are consistent with reasonable values for the CRRA, i.e. estimates that are below 4. The second is that the regression‐based tests are robust, while the GMM‐based tests are not. The third is that regression‐based tests with habit formation depend crucially on the specification of the model. The fourth is that there is evidence that market stock returns are sensitive to both consumption and dividends. The author calls the latter “extra sensitivity of market stock returns”, and it is described as a new puzzle.

Originality/value

The regression‐based models of the CCAPM Euler equation are novel. The comparison between GMM and regression‐based models for the same sample is original. The regression‐based models with habit formation are new. The equity premium puzzle disappears because the estimates of the CRRA are reasonable. But another puzzle is documented, which is the “extra sensitivity of market stock returns” to consumption and dividends together.
Born global acquirers from Indian IT: an exploratory case study
Authors: Varma Sumati
Published: September 2011

Purpose

The purpose of this paper is to examine the phenomenon of the born global firm (BGF) in the Indian context. Specifically, the paper explores firm‐level characteristics (from the Indian IT industry) which act as initiating forces for firms which have made cross‐border acquisitions within five years of incorporation.

Design/methodology/approach

The paper uses inductive reasoning to derive a theoretical framework of analysis, using constructs from the resource‐based view and institutional theory.

Findings

The paper posits that firm resources, personal and network characteristics of its founders, age of the firm and institutional factors act as the initiating factors for a BGF.

Originality/value

This is the first study on the phenomenon of the BGF from India, which is an important emerging market, and has pioneering value, contributing to the growing literature on the topic.
Reducing opportunistic behaviour through a project alliance
Authors: Laan Albertus , Voordijk Hans , Dewulf Geert
Published: September 2011

Purpose

The purpose of this paper is to provide insights into how a project alliance contract is conducive to the development of cooperative relationships between client and contractor organizations involved in a complex project.

Design/methodology/approach

A longitudinal case study of a complex construction project was conducted in which the contract was changed at the end of the negotiation period from a design‐build into a project alliance form.

Findings

Data show that opportunistic behaviour is reduced when there is an incentive structure, as is to be found in project alliances, for client and contractor organizations to cooperatively realize the project. However, it is not sufficient for project partners to agree upon an appropriate incentive structure. For cooperative relationships to develop, they also have to put substantial efforts into reducing their remaining inclinations to make use of opportunities that arise to deviate from the alliance contract.

Practical implications

It is shown that both principals and contractors not only need to carefully select staff for such projects; they also have to work with the people employed such that appropriate attitudes are reinforced and rewarded. Developing cooperative relationships in project alliances needs the surrounding working methods to offer support.

Originality/value

The longitudinal character of the case study offers exceptional opportunities for studying the dynamics in preventing and overcoming the deteriorating patterns of opportunistic behaviour that organizations regularly face in many traditional and design‐build projects.
Study on entrepreneurial process model for SIFE student team based on Timmons model
Authors: Zeng Fan‐qi , Bu Xiang‐zhi , Su Li
Published: September 2011

Purpose

The purpose of this paper is to find the characteristic of entrepreneurial process for the student in free enterprise (SIFE) team in China, and to provide theoretical guidance for the entrepreneurial process of SIFE student team through a new Timmons model.

Design/methodology/approach

Taking an entrepreneurial project of Shantou University SIFE team as example, a Timmons model was proposed describing the SIFE student team entrepreneurial process based on the famous entrepreneurial process model established by Jeffry A. Timmons. The application value of the new Timmons model was proved by case analysis.

Findings

The paper summarized the main characteristics of the entrepreneurial model for the SIFE student team as: the nature of creation free enterprises, the excellent entrepreneurship networks, and the spirit of social entrepreneurship orientation. By case study, the paper proved the practical value of the new Timmons model by analysis of the key factors such as business opportunities, resources and entrepreneurial team, and their dynamic balance process.

Practical implications

The new model proposed in the paper will have theoretical value to provide a direction for student entrepreneurial practice, the analysis of the characteristics of the new model will also enrich the research on entrepreneurship theory.

Originality/value

This paper is the first research on SIFE student entrepreneurial process in China. In the past two years, the authors observed dozens of successful worldwide SIFE students' practical projects, but few of them appeared in the literature. It is hoped that this paper can offer some constructive advice to the entrepreneurial process of the SIFE student team and enrich the theory of general entrepreneurial education.
The major difficulties and countermeasures of current university graduates' entrepreneurship in China
Authors: Qunlian Hong
Published: September 2011

Purpose

College students' employment is a significant problem in the process of China's economic development. Owing to the comprehensive effect, entrepreneurship has become the important way to solve the problem of university students' employment. The purpose of this paper is to analyze systematically the main difficulties of university students' entrepreneurship in China and present corresponding policy recommendations.

Design/methodology/approach

This paper takes the conceptual analysis approach. Referring to the Nyawali and Fogel model of 1994, the paper constructs the key‐determinant model of China's university students' entrepreneurship and analyzes and explains the situation by adopting existing statistics and survey data about China's university students' entrepreneurship.

Findings

After the global financial crisis, encouraging university graduates' entrepreneurship to create more employment opportunities has become the key strategy to deal with employment problems for China's government. This paper finds that, although the university students have some advantages, there are still several difficulties for university graduates' entrepreneurship. Effective countermeasures must be taken to strengthen the entrepreneurship capacity by enhancing entrepreneurship education, increasing entrepreneurship opportunities by improving the entrepreneurship environment and raising the entrepreneurship desire by perfecting financing system and then encouraging university graduates to start up businesses actively, so that more college students could become entrepreneurs.

Originality/value

This paper systematically constructs the key‐determinant model of China's university students' entrepreneurship. The policy recommendations for college students' entrepreneurship presented in this paper might be referred to by government to make decisions to improve university students' entrepreneurship support policy and enhance university students' entrepreneurship ability.
Creating a global network of shared service centres for accounting
Authors: Lindvall Jan , Iveroth Einar
Published: September 2011

Purpose

The purpose of this paper is to contribute to the understanding of the practice of IT‐enabled management control change, in particular how IT‐driven change is made possible from a practical perspective in a global context. It does so by investigating the redesign of the telecommunications company Ericsson's global finance and accounting function from an independent structure of numerous national chief financial officer units into one interdependent global network of shared service centres.

Design/methodology/approach

Ericsson's transformation was followed by drawing mainly on interviews and documents. The data were analysed using narrative and temporal bracketing strategies for theorising from process data.

Findings

The paper illustrates how IT‐enabled management control change unfolds as a continuous interaction between a dynamic organisational structure (social dimension) and a less, but still, dynamic IT (material dimension) across time. The study also highlights how such a process is metaphorically similar to the form of a hermeneutic spiral rather than the common perspective of an arrow from the present to the future.

Research limitations/implications

The focus of the paper is on positive organisational change and how transformation is possible from a strategic and managerial point of view. Hence, less focus is placed on the employee perspective.

Practical implications

This paper stresses the importance of pre‐understanding, an openness to trials and learning, and a dynami stance towards the moving targets of IT and organisation.

Originality/value

The paper provides rich empirical material. The analysis includes contemporary issues, and the practice of IT‐enabled management control change.
Corporate social responsibility implementation
Authors: Smith Alan D.
Published: September 2011

Purpose

Corporate social responsibility (CSR)‐based strategies have become important concepts in dealing with firms' stakeholders. The purpose of this paper is to focus on the processes of stakeholder legitimacy and interest detection, namely ethical considerations and community obligations, to promote CSR as an intangible strategic asset.

Design/methodology/approach

The two relatively large service‐based firms (contrasting not‐for‐profit with for‐profit) that were selected for study were Pittsburgh‐based, namely the largest single employer of the metropolitan area of Pittsburgh (The University of Pittsburgh Medical Center (UPMC)) and WESCO, a distributor of electrical construction products and electrical and industrial maintenance products and the largest domestic provider of integrated supply services.

Findings

It was found that the management teams at UPMC and WESCO approach CSR‐based strategies and its initiative from completely different perspectives. UPMC functions a not‐for‐profit organization while WESCO is a for‐profit entity. They also approach CSR differently as a result from its geographically based service and product offerings, UPMC being more local/regional and WESCO going for global markets. These differences promote differences in the groups and types of CSR that each company is currently engaged in promoting.

Originality/value

The paper demonstrates that both not‐for‐profit and for‐profit entities have a reason to be socially responsible, whether they are local or global firms. The overarching fact is that consumers expect firms to be conscience of the social concerns of the community in which they operate and socially responsible to the various stakeholder groups they serve.
Is saving really prior to investment? Time series and panel evidence in the WAEMU
Authors: Roche Seka Pierre
Published: September 2011

Purpose

This study aims to re‐examine the direction of causality between investment and saving in the West African Economic and Monetary Union (WAEMU).

Design/methodology/approach

The study is empirical, testing for Granger causality between investment and saving as well as for other pertinent variables in the determination of the two variables of interest. It uses two methods: co‐integration and decomposition of variances on the one hand, and dynamic panel on the other.

Findings

The use of recent developments in the treatment and analysis of time series data and the inclusion of relevant variables omitted in prior studies help to shed more light on the contradictory results that exist so far. The empirical result is a proof that saving is a real constraint on investment in the financially moderate economies of the WAEMU.

Practical implications

The paper encourages own resource mobilisation for economic growth and development. Ideas generated in the study suggest that financial liberalization per se will not work unless enough flow of domestic savings exists in a country.

Originality/value

It is one of the recent attempts to investigate this issue within a group of African countries operating in an economic and monetary union. The strength of the paper is the use of various econometric methods to address the issue.
Limited attention and stock price drift following earnings announcements and 10‐K filings
Authors: You Haifeng , Zhang Xiao‐Jun
Published: September 2011

Purpose

This study aims to examine whether limited attention leads to the market underreaction to earnings announcement and 10‐K filings.

Design/methodology/approach

This is an empirical study involving statistical analysis of a large sample of data, obtained from Compustat, CRSP and Xignite Inc. Both portfolio analysis and multivariate regressions are used in hypotheses testing.

Findings

The following key findings are presented in the paper. First, we show that among large firms, investors under‐react more to the information contained in 10‐K filings than earnings announcements. Second, underreaction to earnings announcements tends to be stronger for small firms than large firms. Third, we find that companies report their earnings and 10‐Ks earlier when there is a higher demand for such information, and document a negative relationship between the degree of underreaction and the timeliness of such information release. Finally, we show that the recent ruling by SEC to accelerate 10‐K filing has little impact on the degree of investors' underreaction to 10‐K information.

Research limitations/implications

The findings of this study suggest that investors' failure to devote enough attention to an economic event leads to underreaction, and the degree of underreaction is negatively correlated with the amount of investor attention.

Practical implications

Investors need to periodically reassess the informational contents of economic events, and allocate their attention accordingly, in order to avoid underreaction.

Originality/value

This study analyzes and the roles of limited attention in determining the degree of investor underreaction to earnings announcement and 10‐K filings. The comparison of the two related but distinct financial reporting events yields interesting insights.
Parity and disparity between two Chama oysters: the reproductive biology of the Indo‐Pacific C. pacifica Broderip, invasive to the Mediterranean Sea; and C. savignyi Lamy, indigenous to the Red Sea
Authors: Zurel Dror , Gophna Uri , Benayahu Yehuda
Published: September 2011

The Indo‐Pacific oyster Chama pacifica Broderip, 1835 (Mollusca, Bivalvia, Chamidae) is rarely found in the Northern Red Sea reefs of Eilat (Gulf of Aqaba), where it is outnumbered by its indigenous congener, Chama savignyi Lamy, 1921. The influx of Eritrean biota from the Red Sea into the Mediterranean Sea via the Suez Canal has led to the formation of massive Chama oyster beds along the Eastern Mediterranean shore. However, unlike the Northern Red Sea, the Eastern Mediterranean Chama beds are dominated by C. pacifica oysters, whereas C. savignyi is absent from this region. In an attempt to understand this difference in their respective distribution, the reproductive biology of both species was compared. Histological analysis of the male and female gonads, monitored monthly from March 2009 to August 2010 in both regions, revealed a similar reproductive cycle, comprising six stages: onset of gametogenesis, advanced gametogenesis, ripe, ready to spawn, spent and sexual rest. Female gonads demonstrated an additional, seventh stage – restoration, coinciding with inferred spawning of ripe gametes. Both species were found to be dioecious spawners, with a single, annual, temperature‐dependent inferred spawning period. Chama pacifica was found to reproduce efficiently in maximal Mediterranean seawater temperatures not experienced by the Northern Red Sea Chama populations. This study demonstrates the high invasive potential of an oyster species despite its rarity in its source region.
Characteristics and clinical significance of prostate cancers missed by initial transrectal 12‐core biopsy
Authors: Numao Noboru , Kawakami Satoru , Sakura Mizuaki , Yoshida Soichiro , Koga Fumitaka , Saito Kazutaka , Masuda Hitoshi , Fujii Yasuhisa , Yamamoto Shinya , Yonese Junji , Ishikawa Yuichi , Fukui Iwao , Kihara Kazunori
Published: September 2011

Study Type – Diagnostic (exploratory cohort)

Level of Evidence 3a

What's known on the subject? and What does the study add?

Initial transrectal 12‐core biopsy has a small but definite risk of missing anterior significant prostate cancers irrespective of age, PSA, prostate volume and DRE findings.

Our study yields valuable information for diagnosis and treatment decision of prostate cancer based on transrectal 12‐core biopsy.

OBJECTIVE

To characterize prostate cancers missed by initial transrectal 12‐core biopsy.

PATIENTS AND METHODS

Between 2002 and 2008, 715 men with prostate‐specific antigen levels in the range 2.5–20 ng/mL or abnormal digital rectal examination underwent three‐dimensional 26‐core prostate biopsy (i.e. a combination of transrectal 12‐core biopsy and transperineal 14‐core biopsy) on initial examination.

Of the 257 patients diagnosed with cancer, 120 patients subsequently underwent radical prostatectomy.

Cancers were grouped into TR12‐negative cancers (i.e. not detected through transrectal 12‐core biopsy but detected through transperineal 14‐core biopsy) and TR12‐positive (i.e. detected through transrectal 12‐core biopsy) cancers.

Clinicopathological characteristics of the TR12‐negative and TR12‐positive cancers were evaluated.

RESULTS

TR12‐negative cancers comprised 21% of the three‐dimensional 26‐core biopsy‐detected cancers.

The frequency of cancers with a biopsy Gleason score ≤6 and that of cancers with a biopsy primary Gleason grade ≤3 was higher in TR12‐negative cancers, at 58% and 83%, respectively, than in TR12‐positive cancers, at 25% (P < 0.001) and 53% (P < 0.001), respectively.

The median number of positive cores in TR12‐negative cancers was two out of 26.

TR12‐negative cancers were more frequently located anteriorly than posteriorly.

The incidence of the TR12‐negative cancers was not associated significantly with any clinical variable.

CONCLUSION

Many of the cancers missed by initial transrectal 12‐core biopsy are probably low‐grade and low‐volume diseases, although initial transrectal 12‐core biopsy has a small but definite risk of missing anterior significant cancers.
The Unbearable Humanness of Drone Warfare in FATA, Pakistan
Authors: Ronald Shaw Ian Graham , Akhter Majed
Published: September 2011

Abstract: This paper provides a critical analysis of how and why US‐led drone warfare is conducted in the Federally Administered Tribal Areas (FATA) of Pakistan. First, we provide detailed statistics on the scale and funding of US drone operations, noting a rapid acceleration of its adoption by the military. This is then situated within an overarching narrative of the logic of “targeting”. Second, we study a legal document called the “Frontier Crimes Regulation” of 1901 that defines the relationship of FATA to the rest of Pakistan as an “exceptional” place. In the third section, we argue that the drone is a political actor with a fetishized existence, and this enables it to violate sovereign Pakistani territory. In this sense, the continued violence waged by robots in Pakistan's tribal areas is a result of the deadly interaction between law and technology. The paper concludes by noting the proliferation of drones in everyday life.
Functional Analysis of Interaction Sites on the N‐Terminal Domain of Clathrin Heavy Chain
Authors: Willox Anna K. , Royle Stephen J.
Published: September 2011

In clathrin‐mediated membrane traffic, clathrin does not bind directly to cargo and instead binds to adaptors that mediate this function. For endocytosis, the main adaptor is the adaptor protein (AP)‐2 complex, but it is uncertain how clathrin contacts AP‐2. Here we tested in human cells the importance of the three binding sites that have been identified so far on the N‐terminal domain (NTD) of clathrin. We find that mutation of each of the three sites on the NTD, alone or in combination, does not block clathrin/AP‐2‐mediated endocytosis in the same way as deletion of the NTD. We report here the fourth and final site on the NTD that is required for clathrin/AP‐2‐mediated endocytic function. Each of the four interaction sites can operate alone to mediate endocytosis. The observed functional redundancy between interaction sites on the NTD explains how productivity of clathrin‐coated vesicle formation is ensured.
Preliminary evidence of abnormal white matter related to the fusiform gyrus in Williams syndrome: a diffusion tensor imaging tractography study
Authors: Haas B. W. , Hoeft F. , Barnea‐Goraly N. , Golarai G. , Bellugi U. , Reiss A. L.
Published: September 2011

Williams syndrome (WS) is a genetic condition caused by a hemizygous microdeletion on chromosome 7q11.23. WS is characterized by a distinctive social phenotype composed of increased drive toward social engagement and attention toward faces. In addition, individuals with WS exhibit abnormal structure and function of brain regions important for the processing of faces such as the fusiform gyrus. This study was designed to investigate if white matter tracts related to the fusiform gyrus in WS exhibit abnormal structural integrity as compared to typically developing (TD; age matched) and developmentally delayed (DD; intelligence quotient matched) controls. Using diffusion tensor imaging data collected from 40 (20 WS, 10 TD and 10 DD) participants, white matter fibers were reconstructed that project through the fusiform gyrus and two control regions (caudate and the genu of the corpus callosum). Macro‐structural integrity was assessed by calculating the total volume of reconstructed fibers and micro‐structural integrity was assessed by calculating fractional anisotropy (FA) and fiber density index (FDi) of reconstructed fibers. WS participants, as compared to controls, exhibited an increase in the volume of reconstructed fibers and an increase in FA and FDi for fibers projecting through the fusiform gyrus. No between‐group differences were observed in the fibers that project through the control regions. Although preliminary, these results provide further evidence that the brain anatomy important for processing faces is abnormal in WS.
Analysis of enquiries to the National Pharmacy Association following major changes to controlled drug legislation in the UK
Authors: Gallagher Cathal T , Hickman Adrienne C , Hannbeck Leyla , Flynn Robert W
Published: September 2011

Objectives

The National Pharmacy Association (NPA) provides an advice service to community pharmacists in the UK, and keeps a database of the enquiries it receives. The aim of this research was to analyse the database for the period of October 2007 to March 2008 to gain an insight into how well pharmacists coped with legislative changes directly affecting pharmacy by identifying which changes generated the most enquiries during these 6 months and ascertaining in which months these queries were at their highest levels.

Methods

Anonymised telephone enquiries regarding controlled drugs (CDs) received by the NPA from pharmacists during a 6-month period were reviewed and categorised according to the legislative change or other CD issue to which they related. A Poisson model was applied to determine whether there was a significant difference in the total number of CD queries generated each month.

Key findings

Altogether 6082 queries regarding CDs were received, of which 57% related to legislative changes. The three legislative changes that took place during the 6-month period all generated a significant increase in numbers of queries around the time of the change. Queries regarding the new form of CD register comprised the largest single category.

Conclusions

Community pharmacists seek information regarding legislative changes when such changes come into force to a greater degree than when the legislation is drafted, consulted upon or enacted. The high number of queries received by the NPA does reflect that there was a difficulty in applying the various legislative changes in practice.
Centenary of the Anglo‐American high‐altitude expedition to Pikes Peak
Authors: West John B.
Published: September 2011

This year (2011) marks the centenary of one of the most important early research expeditions to high altitude. The principal participants were J. S. Haldane and C. G. Douglas from Oxford, Y. Henderson from Yale and E. C. Schneider from Colorado College. Pikes Peak just outside Colorado Springs proved to be an excellent venue because of its substantial altitude of 4300 m, convenient access via a cog railway and comfortable living accommodation. The expedition had a classical design, with measurements made first at sea level, then on the summit for 5 weeks and then at sea level again. The extensive scientific programme included descriptions of acute mountain sickness, many measurements of partial pressures in alveolar gas and arterial blood, changes in ventilation, including periodic breathing, exercise measurements and a large number of blood studies. One error was the conclusion that the arterial 
 could considerably exceed the alveolar value, implying oxygen secretion by the lung, but this should not detract from the other important advances. Mabel FitzGerald was invited to be a member of the expedition but did not join the men on the summit. Instead, she visited various mining camps in Colorado at somewhat lower altitudes and carried out classical studies of alveolar gas partial pressures and haemoglobin values. Indeed, her results of alveolar 
 and 
 are frequently cited today. The Pikes Peak expedition was a model and it had an extensive influence on later studies.
Computational analyses of CO‐rebreathing methods for estimating haemoglobin mass in humans
Authors: Chada Kinnera E. , Bruce Eugene N.
Published: September 2011

Measurement of haemoglobin mass (MHb) is used to quantify alterations in oxygen delivery during exercise training or acclimatization to altitude. Uptake of carbon monoxide by haemoglobin is the basis of the common non‐radioactive methods to determine MHb in humans. This study used a validated mathematical model to simulate CO uptake during rebreathing protocols and to determine sources of errors in estimation of MHb. Our previously published model was validated using experimentally measured carboxyhaemoglobin levels (%HbCO) from arterial, capillary and venous blood sites of human subjects during CO‐rebreathing protocols. This model was then used to simulate various CO‐rebreathing protocols in 24 human subjects with known MHb. Using variables generated by the model, MHb was estimated on the basis of assumptions typically made for calculating the volume of CO bound to myoglobin, the volume of CO exhaled and the volume of CO in the rebreathing system. It was found that inaccurate estimation of the volume of CO bound to myoglobin was the major source of error in determination of MHb. Additionally, the size of the error was found to depend on the site of blood sampling because of differences in %HbCO. Regression equations were developed to improve the estimation of volume of CO bound to myoglobin, and a new protocol that is less dependent on the site of blood sampling is proposed.
Salmonella enterica in Swine Production: Assessing the Association between Amplified Fragment Length Polymorphism and Epidemiological Units of Concern
Authors: Wang Bing , Wang Chong , McKean James D. , Logue Catherine M. , Gebreyes Wondwossen A. , Tivendale Kelly A. , O'Connor Annette M.
Published: September 2011

The aims of this study were to determine the ability of amplified fragment length polymorphism (AFLP) to differentiate
Salmonella
isolates from different units of swine production and to demonstrate the relatedness of
Salmonella
between farms and abattoirs by AFLP. Twenty-four farms in the midwestern United States were visited four times from 2006 to 2009. At each farm or abattoir visit, 30 fecal samples or 30 mesenteric lymph nodes were collected, respectively. A total of 220
Salmonella
isolates were obtained, serotyped, and genotyped by multilocus sequence typing (MLST) and AFLP. These 220 isolates clustered into 21 serotypes, 18 MLST types, and 14 predominant AFLP clusters based on a genetic similarity threshold level of 60%. To assess genetic differentiation between farms, harvest cohorts, and pigs, analysis of molecular variance was conducted using AFLP data. The results showed 65.62% of overall genetic variation was attributed to variance among pigs, 27.21% to farms, and 7.17% to harvest cohorts. Variance components at the farm (
P
= 0.003) and pig (
P
= 0.001) levels were significant, but not at the harvest cohort level (
P
= 0.079). A second analysis, a permutation test using AFLP data, indicated that on-farm and at-abattoir
Salmonella
from pigs of the same farms were more related than from different farms. Therefore, among the three subtyping methods, serotyping, MLST, and AFLP, AFLP was the method that was able to differentiate among
Salmonella
isolates from different farms and link contamination at the abattoir to the farm of origin.
Phenotypic Diversity Caused by Differential RpoS Activity among Environmental Escherichia coli Isolates
Authors: Chiang Sarah M. , Dong Tao , Edge Thomas A. , Schellhorn Herb E.
Published: September 2011

Enteric bacteria deposited into the environment by animal hosts are subject to diverse selective pressures. These pressures may act on phenotypic differences in bacterial populations and select adaptive mutations for survival in stress. As a model to study phenotypic diversity in environmental bacteria, we examined mutations of the stress response sigma factor, RpoS, in environmental
Escherichia coli
isolates. A total of 2,040 isolates from urban beaches and nearby fecal pollution sources on Lake Ontario (Canada) were screened for RpoS function by examining growth on succinate and catalase activity, two RpoS-dependent phenotypes. The
rpoS
sequence was determined for 45 isolates, including all candidate RpoS mutants, and of these, six isolates were confirmed as mutants with the complete loss of RpoS function. Similarly to laboratory strains, the RpoS expression of these environmental isolates was stationary phase dependent. However, the expression of RpoS regulon members KatE and AppA had differing levels of expression in several environmental isolates compared to those in laboratory strains. Furthermore, after plating
rpoS
+
isolates on succinate, RpoS mutants could be readily selected from environmental
E. coli
. Naturally isolated and succinate-selected RpoS mutants had lower generation times on poor carbon sources and lower stress resistance than their
rpoS
+
isogenic parental strains. These results show that RpoS mutants are present in the environment (with a frequency of 0.003 among isolates) and that, similarly to laboratory and pathogenic strains, growth on poor carbon sources selects for
rpoS
mutations in environmental
E. coli
. RpoS selection may be an important determinant of phenotypic diversification and, hence, the survival of
E. coli
in the environment.
Membrane Stresses Induced by Overproduction of Free Fatty Acids in Escherichia coli
Authors: Lennen Rebecca M. , Kruziki Max A. , Kumar Kritika , Zinkel Robert A. , Burnum Kristin E. , Lipton Mary S. , Hoover Spencer W. , Ranatunga Don R. , Wittkopp Tyler M. , Marner Wesley D. , Pfleger Brian F.
Published: September 2011

Microbially produced fatty acids are potential precursors to high-energy-density biofuels, including alkanes and alkyl ethyl esters, by either catalytic conversion of free fatty acids (FFAs) or enzymatic conversion of acyl-acyl carrier protein or acyl-coenzyme A intermediates. Metabolic engineering efforts aimed at overproducing FFAs inEscherichia colihave achieved less than 30% of the maximum theoretical yield on the supplied carbon source. In this work, the viability, morphology, transcript levels, and protein levels of a strain ofE. colithat overproduces medium-chain-length FFAs was compared to an engineered control strain. By early stationary phase, an 85% reduction in viable cell counts and exacerbated loss of inner membrane integrity were observed in the FFA-overproducing strain. These effects were enhanced in strains endogenously producing FFAs compared to strains exposed to exogenously fed FFAs. Under two sets of cultivation conditions, long-chain unsaturated fatty acid content greatly increased, and the expression of genes and proteins required for unsaturated fatty acid biosynthesis were significantly decreased. Membrane stresses were further implicated by increased expression of genes and proteins of the phage shock response, the MarA/Rob/SoxS regulon, and thenuoandcyooperons of aerobic respiration. Gene deletion studies confirmed the importance of the phage shock proteins and Rob for maintaining cell viability; however, little to no change in FFA titer was observed after 24 h of cultivation. The results of this study serve as a baseline for future targeted attempts to improve FFA yields and titers inE. coli.
One-Pot Synthesis of Diverse
dl
-Configuration Dipeptides by a Streptomyces
d
-Stereospecific Amidohydrolase
Authors: Arima Jiro , Usuki Hirokazu , Hatanaka Tadashi , Mori Nobuhiro
Published: September 2011

The synthesis of diverse
dl
-configuration dipeptides in a one-pot reaction was demonstrated by using a function of the aminolysis reaction of a
d
-stereospecific amidohydrolase from
Streptomyces
sp., a clan SE, S12 family peptidase categorized as a peptidase with
d
-stereospecificity. The enzyme was able to use various aminoacyl derivatives, including
l
-aminoacyl derivatives, as acyl donors and acceptors. Investigations of the specificity of the peptide synthetic activity revealed that the enzyme preferentially used
d
-aminoacyl derivatives as acyl donors. In contrast,
l
-amino acids and their derivatives were preferentially used as acyl acceptors. Consequently, the synthesized dipeptides had a
dl
-configuration when
d
- and
l
-aminoacyl derivatives were mixed in a one-pot reaction. This report also describes that the enzyme produced cyclo(
d
–Pro-
l
-Arg), a specific inhibitor of family 18 chitinase, with a conversion rate for
d
-Pro benzyl ester and
l
-Arg methyl ester to cyclo(
d
-Pro–
l
-Arg) of greater than 65%. Furthermore, based on results of cyclo(
d
-Pro–
l
-Arg) synthesis, we propose a reaction mechanism for cyclo(
d
-Pro–
l
-Arg) production.
Use of Stable Isotopes To Measure the Metabolic Activity of the Human Intestinal Microbiota
Authors: Reichardt Nicole , Barclay Andrew R. , Weaver Lawrence T. , Morrison Douglas J.
Published: September 2011

The human intestinal microbiota is a complex biological system comprising a vast repertoire of microbes with considerable metabolic activity relevant to both bacterial growth and host health. Greater strides have been made in the analysis of microbial diversity than in the measurement of functional activity, particularlyin vivo. Stable isotope probing offers a new approach by coupling measurements of metabolic activity with microbial identification. Using a low-enrichment labeling strategyin vitro, this study has identified metabolically active bacterial groups via magnetic-bead capture methodology and stable isotope ratio analysis. Using five probes (EUB338, Bac303, Bif164, EREC482, and Clep866), changes in the activities of key intestinal microbial groups were successfully measured by exploiting tracers ofde novoRNA synthesis. Perturbation of the nutrient source with oligofructose generated changes in the activity of bifidobacteria as expected, but also in theBacteroides-Prevotellagroup, theEubacterium rectale-Clostridium coccoidesgroup, and theClostridium leptumsubgroup. Changes in activity were also observed in response to the medium type. This study suggests that changes in the functional activity of the gut microbiota can be assessed using tracers ofde novonucleic acid synthesis combined with measurement of low isotopic enrichment in 16S rRNA. Such tracers potentially limit substrate bias because they are universally available to bacteria. This low-enrichment labeling approach does not depend on the commercial availability of specific labeled substrates and can be easily translated toin vivoprobing experiments of the functional activity of the microbiota in the human gut.
Leaching of Cryptosporidium parvum Oocysts, Escherichia coli, and a Salmonella enterica Serovar Typhimurium Bacteriophage through Intact Soil Cores following Surface Application and Injection of Slurry
Authors: Forslund Anita , Markussen Bo , Toenner-Klank Lise , Bech Tina B. , Jacobsen Ole Stig , Dalsgaard Anders
Published: September 2011

Increasing amounts of livestock manure are being applied to agricultural soil, but it is unknown to what extent this may be associated with contamination of aquatic recipients and groundwater if microorganisms are transported through the soil under natural weather conditions. The objective of this study was therefore to evaluate how injection and surface application of pig slurry on intact sandy clay loam soil cores influenced the leaching ofSalmonella entericaserovar Typhimurium bacteriophage 28B,Escherichia coli, andCryptosporidium parvumoocysts. All three microbial tracers were detected in the leachate on day 1, and the highest relative concentration was detected on the fourth day (0.1 pore volume). Although the concentration of the phage 28B declined over time, the phage was still found in leachate at day 148.C. parvumoocysts and chloride had an additional rise in the relative concentration at a 0.5 pore volume, corresponding to the exchange of the total pore volume. The leaching ofE. coliwas delayed compared with that of the added microbial tracers, indicating a stronger attachment to slurry particles, butE. colicould be detected up to 3 months. Significantly enhanced leaching of phage 28B and oocysts by the injection method was seen, whereas leaching of the indigenousE. coliwas not affected by the application method. Preferential flow was the primary transport vehicle, and the diameter of the fractures in the intact soil cores facilitated transport of all sizes of microbial tracers under natural weather conditions.
Influence of Host Phylogeographic Patterns and Incomplete Lineage Sorting on Within-Species Genetic Variability in Wigglesworthia Species, Obligate Symbionts of Tsetse Flies
Authors: Symula Rebecca E. , Marpuri Ian , Bjornson Robert D. , Okedi Loyce , Beadell Jon , Alam Uzma , Aksoy Serap , Caccone Adalgisa
Published: September 2011

Vertical transmission of obligate symbionts generates a predictable evolutionary history of symbionts that reflects that of their hosts. In insects, evolutionary associations between symbionts and their hosts have been investigated primarily among species, leaving population-level processes largely unknown. In this study, we investigated the tsetse (Diptera: Glossinidae) bacterial symbiont,
Wigglesworthia glossinidia
, to determine whether observed codiversification of symbiont and tsetse host species extends to a single host species (
Glossina fuscipes fuscipes
) in Uganda. To explore symbiont genetic variation in
G. f. fuscipes
populations, we screened two variable loci (
lon
and
lepA
) from the
Wigglesworthia glossinidia
bacterium in the host species
Glossina fuscipes fuscipes
(
W. g. fuscipes
) and examined phylogeographic and demographic characteristics in multiple host populations. Symbiont genetic variation was apparent within and among populations. We identified two distinct symbiont lineages, in northern and southern Uganda. Incongruence length difference (ILD) tests indicated that the two lineages corresponded exactly to northern and southern
G. f. fuscipes
mitochondrial DNA (mtDNA) haplogroups (
P
= 1.0). Analysis of molecular variance (AMOVA) confirmed that most variation was partitioned between the northern and southern lineages defined by host mtDNA (85.44%). However, ILD tests rejected finer-scale congruence within the northern and southern populations (
P
= 0.009). This incongruence was potentially due to incomplete lineage sorting that resulted in novel combinations of symbiont genetic variants and host background. Identifying these novel combinations may have public health significance, since tsetse is the sole vector of sleeping sickness and
Wigglesworthia
is known to influence host vector competence. Thus, understanding the adaptive value of these host-symbiont combinations may afford opportunities to develop vector control methods.
Localization Pattern of Conjugation Machinery in a Gram-Positive Bacterium
Authors: Bauer Theresa , Rösch Thomas , Itaya Mitsuhiro , Graumann Peter L.
Published: September 2011

Conjugation is an efficient way for transfer of genetic information between bacteria, even between highly diverged species, and a major cause for the spreading of resistance genes. We have investigated the subcellular localization of several conserved conjugation proteins carried on plasmid pLS20 found in
Bacillus subtilis
. We show that VirB1, VirB4, VirB11, VirD2, and VirD4 homologs assemble at a single cell pole, but also at other sites along the cell membrane, in cells during the lag phase of growth. Bimolecular fluorescence complementation analyses showed that VirB4 and VirD4 interact at the cell pole and, less frequently, at other sites along the membrane. VirB1 and VirB11 also colocalized at the cell pole. Total internal reflection fluorescence microscopy showed that pLS20 is largely membrane associated and is frequently found at the cell pole, indicating that transfer takes place at the pole, which is a preferred site for the assembly of the active conjugation apparatus, but not the sole site. VirD2, VirB4, and VirD4 started to localize to the pole or the membrane in stationary-phase cells, and VirB1 and VirB11 were observed as foci in cells resuspended in fresh medium but no longer in cells that had entered exponential growth, although at least VirB4 was still expressed. These data reveal an unusual assembly/disassembly timing for the pLS20 conjugation machinery and suggest that specific localization of conjugation proteins in lag-phase cells and delocalization during growth are the reasons why pLS20 conjugation occurs only during early exponential phase.
Probing the Impact of Ligand Binding on the Acyl-Homoserine Lactone-Hindered Transcription Factor EsaR of Pantoea stewartii subsp. stewartii
Authors: Schu Daniel J. , Ramachandran Revathy , Geissinger Jared S. , Stevens Ann M.
Published: September 2011

The quorum-sensing regulator EsaR from
Pantoea stewartii
subsp.
stewartii
is a LuxR homologue that is inactivated by acyl-homoserine lactone (AHL). In the corn pathogen
P. stewartii
, production of exopolysaccharide (EPS) is repressed by EsaR at low cell densities. However, at high cell densities when high concentrations of its cognate AHL signal are present, EsaR is inactivated and derepression of EPS production occurs. Thus, EsaR responds to AHL in a manner opposite to that of most LuxR family members. Depending on the position of its binding site within target promoters, EsaR serves as either a repressor or activator in the absence rather than in the presence of its AHL ligand. The effect of AHL on LuxR homologues has been difficult to study
in vitro
because AHL is required for purification and stability. EsaR, however, can be purified without AHL enabling an
in vitro
analysis of the response of the protein to ligand. Western immunoblots and pulse-chase experiments demonstrated that EsaR is stable
in vivo
in the absence or presence of AHL. Limited
in vitro
proteolytic digestions of a biologically active His-MBP tagged version of EsaR highlighted intradomain and interdomain conformational changes that occur in the protein in response to AHL. Gel filtration chromatography of the full-length fusion protein and cross-linking of the N-terminal domain both suggest that this conformational change does not impact the multimeric state of the protein. These findings provide greater insight into the diverse mechanisms for AHL responsiveness found within the LuxR family.