bluuebunny/crossref_metadata_2025 huggingface dataset:
Working through the Problems of Study Abroad Using the Methodologies of Religious Studies
Authors: Siegler Elijah
Published: January 2015

After illustrating the joys of teaching religious studies abroad with an anecdote from my trip to China, I warn of some of its inherent pedagogical and ethical challenges. I argue that teaching some of the “new directions” in religious studies scholarship might address these challenges. These include a turning away from the abstract (texts, beliefs, theologies) and towards the concrete (bodies, places, rituals); moving away from teaching religions as unchanging, ancient verities and instead emphasizing the impact that colonialism, modernization, and secularism have had; moving from searching for authenticity to questioning it; and emphasizing methodological self‐consciousness. Keeping these new directions in mind will help ensure the study abroad experience is educationally successful. This essay serves as an introduction to a series of six additional essays comprising a special section of the journal (see Teaching Theology and Religion 18:1, January 2015).
What Do We Compare When We Compare Religions? Philosophical Remarks on the Psychology of Studying Comparative Religion Abroad
Authors: Irvine Andrew
Published: January 2015

The issue of comparison is a vexing one in religious and theological studies, not least for teachers of comparative religion in study abroad settings. We try to make familiar ideas fresh and strange, in settings where students may find it hard not to take “fresh” and “strange” as signs of existential threat. The author explores this delicate pedagogical situation, drawing on several years' experience directing a study abroad program and on the thought of figures from the Western existentialist tradition and Chinese Confucian philosophy. The article focuses particularly on “oh events” – defined as moments when one learns one has something to learn and something to unlearn. The author argues that the experience of shame that is typical of oh events can become a valuable resource for cross‐cultural learning and personal transformation, if teachers assist students to reflect on the experience as a sign of differing, but potentially harmonizable, cultural expectations. This essay is published alongside of six other essays, including a response from John Barbour, comprising a special section of the journal (see Teaching Theology and Religion 18:1, January 2015).
The Immersion Experience: Lessons from Study Abroad in Religion
Authors: Mitchell Kerry
Published: January 2015

This paper discusses strategies I employed during seven years of teaching within a study abroad program focusing on religion. This year‐long program traveled to four Asian countries and included immersion experiences in monasteries, ashrams, and other religious institutions. I identify four principles and discuss accompanying exercises that guided my teaching: (1) Accept and observe anxiety. Inability to understand is a sign that direct and deep contact is taking place. (2) Educate about education. Help students to see the aims, assumptions, and context of the teaching strategies religious practitioners employ. (3) Make it practical. Devise exercises that students can do and do well and that do not demand synthetic, systematic comprehension even as a goal. (4) Stop making sense. Build pauses and breaks into the train of reflection on the meaning of experience. These spaces give room for the shifts in the ways of learning that study abroad demands. This essay is published alongside of six other essays, including a response from John Barbour, comprising a special section of the journal (see Teaching Theology and Religion 18:1, January 2015).
Inverting the Object of Study: Recalibrating the Frame of Reference in Study Abroad Experiences
Authors: Palmer Norris W.
Published: January 2015

This essay is concerned with study abroad experiences as opportunities for student cognitive development, using the interpretive lens of educational psychologist William G. Perry. A standard and often valuable assignment in courses on world religions is a site visit to a religious institution in one's local area. This may concretize otherwise abstract materials and help students reflect on ways in which the lived experience of religion differs from its presentation in course texts and other academic materials. Increasingly, study abroad trips are being offered as extended and more intensive ways of bringing this material to life, offering students opportunity to see lived religion within another cultural framework. At the heart of this paper is the contention that such study abroad experiences function not simply as longer, more intense versions of site visits but, rather, as experiences that invert the subject and object of study. The worldview of the student becomes a primary object of study, which is examined, as it were, by the particulars of the religion(s) under investigation and the cultures of which said religion(s) are a part. Where site visits offer students an opportunity to visit the strange amidst the familiar, study abroad trips provide opportunities for students to become the strange within a recalibrated familiar. The subject becomes the object and is interrogated by the context of study. While local, stateside site visits can offer a degree of such dislocation, their brevity, together with some degree of assimilation to the larger culture flows on the part of the local religious institution being visited, most often mitigates any significant inversion. Students generally see such institutions as either mildly or wildly exotic, but always within their frame of reference, which constitutes the norm. When abroad, the normative experience of students is often subverted in ways that lay bare the assumptions behind such views and makes possible another world in which to live. Simply put, the subject and object of study change places. If this inversion is carefully attended to, it can provide rich insight into not only the topics nominally being studied but also occasion opportunity for real cognitive development on the part of the student. This essay is published alongside of six other essays, including a response from John Barbour, comprising a special section of the journal (see Teaching Theology and Religion 18:1, January 2015).
The Politics of Teaching of Indigenous Traditions in Aotearoa/New Zealand
Authors: Wiseman Wendy A.
Published: January 2015

Reflecting on two study abroad trips to New Zealand in 2005 and 2007, I suggest in this essay that it is possible to mitigate the risk of (American or European) students recapitulating imperial attitudes through development of a rigorous curriculum focusing on the legacies of colonialism, institutional racism, and the somewhat dubious phenomenon of “post‐colonialism.” Readings, I argue, should be in continual play during cultural and social activities, operating in a dialectal move toward an “ethics of respect.” Such an ethics remains aporetic, or uncertain, insofar as no code of behavior can render us immune to the political and polemical effects of past and present forms of imperialism. However, a cultivated respect for distance and difference, including regarding questions of “authenticity,” can help to actualize the transformative promise of studying (indigenous) religion abroad. This essay is published alongside of six other essays, including a response from John Barbour, comprising a special section of the journal (see Teaching Theology and Religion 18:1, January 2015).
Finding Freedom Abroad: Working with Conservative Christian Students in Study Abroad Programs
Authors: Mercer Calvin
Published: January 2015

Conservative (fundamentalist, evangelical) Christian students present a general theological worldview that often correlates with significant anxiety. In a foreign setting, the anxiety of conservative students, removed from their supportive infrastructure, can be considerably heightened. This structure of thinking and emotion presents distinctive challenges and opportunities. Drawing upon my work as a clinician and as a religion professor who conducted study abroad programs, I make suggestions for working effectively with conservative Christian students in study abroad contexts. Suggestions include predeparture, in‐country, and post‐trip strategies. Specific examples of conversations with students are provided to illustrate the challenges and strategies. This essay is published alongside of seven other essays, including a response from John Barbour, comprising a special section of the journal (seeTeaching Theology and Religion18:1, January 2015).
Heart failure in sub‐Saharan Africa: review of the aetiology of heart failure and the role of point‐of‐care biomarker diagnostics
Authors: Glezeva Nadezhda , Gallagher Joe , Ledwidge Mark , O'Donoghue John , McDonald Kenneth , Chipolombwe John , Watson Chris
Published: January 2015

Within Africa, the burden of heart failure is significant. This arises from the increase in cardiovascular disease and associated risk factors such as hypertension and diabetes, as well as causes of heart failure which are particular to sub‐Saharan Africa, such as endomyocardial fibrosis. The lack of access to echocardiography and other imaging modalities, from a cost and technical perspective, combined with the predominantly rural nature of many countries with poor transport links, means that the vast majority of people never obtain an appropriate diagnosis. Similarly, research has been limited on the causes and treatment of heart failure in Africa and in particular endemic causes such as EMF and rheumatic heart disease. This review outlines the burden of heart failure in Africa and highlights the opportunity to expand diagnosis through the use of biomarkers, in particular natriuretic peptides. This builds on the success of point‐of‐care testing in human immunodeficiency virus and tuberculosis which have been extensively deployed in community settings in Africa.
The Curse of Being Landlocked: Institutions Rather than Trade
Authors: Carmignani Fabrizio
Published: January 2015

This article revisits the long‐term economic effects of being landlocked. The conventional wisdom, which also prevails in policy circles, is that landlockedness hurts development by reducing trade. Gravity models of bilateral trade seem to confirm this view. However, there is no evidence in cross‐country data of a systematic relationship between landlockedness and country's trade to GDP ratio. Drawing on this stylised fact, the paper explores the possibility that landlockedness might affect GDP independently from its effect on trade. Theoretical considerations suggest that institutional quality could be a relevant transmission mechanism. The estimation of a system of three equations confirms that landlockedness has a negative effect on GDP and that this negative effect is transmitted through institutions rather than trade. Moreover, after controlling for the transmission via institutions and trade, landlockedness has a further negative effect on GDP. These findings call for a review of the policy approach to the development of landlocked countries.
Biogeography of Heterotrophic Flagellate Populations Indicates the Presence of Generalist and Specialist Taxa in the Arctic Ocean
Authors: Thaler Mary , Lovejoy Connie
Published: January 2015

Heterotrophic marine flagellates (HF) are ubiquitous in the world's oceans and represented in nearly all branches of the domain Eukaryota. However, the factors determining distributions of major taxonomic groups are poorly known. The Arctic Ocean is a good model environment for examining the distribution of functionally similar but phylogenetically diverse HF because the physical oceanography and annual ice cycles result in distinct environments that could select for microbial communities or favor specific taxa. We reanalyzed new and previously published high-throughput sequencing data from multiple studies in the Arctic Ocean to identify broad patterns in the distribution of individual taxa. HF accounted for fewer than 2% to over one-half of the reads from the water column and for up to 60% of reads from ice, which was dominated by
Cryothecomonas
. In the water column, many HF phylotypes belonging to Telonemia and Picozoa, uncultured marine stramenopiles (MAST), and choanoflagellates were geographically widely distributed. However, for two groups in particular, Telonemia and
Cryothecomonas
, some species level taxa showed more restricted distributions. For example, several phylotypes of Telonemia favored open waters with lower nutrients such as the Canada Basin and offshore of the Mackenzie Shelf. In summary, we found that while some Arctic HF were successful over a range of conditions, others could be specialists that occur under particular conditions. We conclude that tracking species level diversity in HF not only is feasible but also provides a potential tool for understanding the responses of marine microbial ecosystems to rapidly changing ice regimes.
Counting Viruses and Bacteria in Photosynthetic Microbial Mats
Authors: Carreira Cátia , Staal Marc , Middelboe Mathias , Brussaard Corina P. D.
Published: January 2015

Viral abundances in benthic environments are the highest found in aquatic systems. Photosynthetic microbial mats represent benthic environments with high microbial activity and possibly high viral densities, yet viral abundances have not been examined in such systems. Existing extraction procedures typically used in benthic viral ecology were applied to the complex matrix of microbial mats but were found to inefficiently extract viruses. Here, we present a method for extraction and quantification of viruses from photosynthetic microbial mats using epifluorescence microscopy (EFM) and flow cytometry (FCM). A combination of EDTA addition, probe sonication, and enzyme treatment applied to a glutaraldehyde-fixed sample resulted in a substantially higher viral (5- to 33-fold) extraction efficiency and reduced background noise compared to previously published methods. Using this method, it was found that in general, intertidal photosynthetic microbial mats harbor very high viral abundances (2.8 × 10
10
± 0.3 × 10
10
g
−1
) compared with benthic habitats (10
7
to 10
9
g
−1
). This procedure also showed 4.5- and 4-fold-increased efficacies of extraction of viruses and bacteria, respectively, from intertidal sediments, allowing a single method to be used for the microbial mat and underlying sediment.
Xylan Utilization Regulon in Xanthomonas citri pv. citri Strain 306: Gene Expression and Utilization of Oligoxylosides
Authors: Chow V. , Shantharaj D. , Guo Y. , Nong G. , Minsavage G. V. , Jones J. B. , Preston J. F.
Published: January 2015

Xanthomonas citri
pv. citri strain 306 (
Xcc
306), a causative agent of citrus canker, produces endoxylanases that catalyze the depolymerization of cell wall-associated xylans. In the sequenced genomes of all plant-pathogenic xanthomonads, genes encoding xylanolytic enzymes are clustered in three adjacent operons. In
Xcc
306, these consecutive operons contain genes encoding the glycoside hydrolase family 10 (GH10) endoxylanases Xyn10A and Xyn10C, the
agu67
gene, encoding a GH67 α-glucuronidase (Agu67), the
xyn43E
gene, encoding a putative GH43 α-
l
-arabinofuranosidase, and the
xyn43F
gene, encoding a putative β-xylosidase. Recombinant Xyn10A and Xyn10C convert polymeric 4-
O
-methylglucuronoxylan (MeGX

n

) to oligoxylosides methylglucuronoxylotriose (MeGX
3
), xylotriose (X
3
), and xylobiose (X
2
).
Xcc
306 completely utilizes MeGX

n

predigested with Xyn10A or Xyn10C but shows little utilization of MeGX

n

.
Xcc
306 with a deletion in the gene encoding α-glucuronidase (
Xcc
306 Δ
agu67
) will not utilize MeGX
3
for growth, demonstrating the role of Agu67 in the complete utilization of GH10-digested MeGX

n

. Preferential growth on oligoxylosides compared to growth on polymeric MeGX

n

indicates that GH10 xylanases, either secreted by
Xcc
306
in planta
or produced by the plant host, generate oligoxylosides that are processed by Xyn10 xylanases and Agu67 residing in the periplasm. Coordinate induction by oligoxylosides of
xyn10
,
agu67
,
cirA
, the
tonB
receptor, and other genes within these three operons indicates that they constitute a regulon that is responsive to the oligoxylosides generated by the action of
Xcc
306 GH10 xylanases on MeGX

n

. The combined expression of genes in this regulon may allow scavenging of oligoxylosides derived from cell wall deconstruction, thereby contributing to the tissue colonization and/or survival of
Xcc
306 and, ultimately, to plant disease.
Metabolic Engineering of Corynebacterium glutamicum for Methanol Metabolism
Authors: Witthoff Sabrina , Schmitz Katja , Niedenführ Sebastian , Nöh Katharina , Noack Stephan , Bott Michael , Marienhagen Jan
Published: January 2015

Methanol is already an important carbon feedstock in the chemical industry, but it has found only limited application in biotechnological production processes. This can be mostly attributed to the inability of most microbial platform organisms to utilize methanol as a carbon and energy source. With the aim to turn methanol into a suitable feedstock for microbial production processes, we engineered the industrially important but nonmethylotrophic bacterium
Corynebacterium glutamicum
toward the utilization of methanol as an auxiliary carbon source in a sugar-based medium. Initial oxidation of methanol to formaldehyde was achieved by heterologous expression of a methanol dehydrogenase from
Bacillus methanolicus
, whereas assimilation of formaldehyde was realized by implementing the two key enzymes of the ribulose monophosphate pathway of
Bacillus subtilis
: 3-hexulose-6-phosphate synthase and 6-phospho-3-hexuloisomerase. The recombinant
C. glutamicum
strain showed an average methanol consumption rate of 1.7 ± 0.3 mM/h (mean ± standard deviation) in a glucose-methanol medium, and the culture grew to a higher cell density than in medium without methanol. In addition, [
13
C]methanol-labeling experiments revealed labeling fractions of 3 to 10% in the m + 1 mass isotopomers of various intracellular metabolites. In the background of a
C. glutamicum
Δ
ald
Δ
adhE
mutant being strongly impaired in its ability to oxidize formaldehyde to CO
2
, the m + 1 labeling of these intermediates was increased (8 to 25%), pointing toward higher formaldehyde assimilation capabilities of this strain. The engineered
C. glutamicum
strains represent a promising starting point for the development of sugar-based biotechnological production processes using methanol as an auxiliary substrate.
Listeria Phage and Phage Tail Induction Triggered by Components of Bacterial Growth Media (Phosphate, LiCl, Nalidixic Acid, and Acriflavine)
Authors: Lemaître Jean-Paul , Duroux Amandine , Pimpie Romain , Duez Jean-Marie , Milat Marie-Louise
Published: January 2015

The detection of
Listeria monocytogenes
from food is currently carried out using a double enrichment. For the ISO methodology, this double enrichment is performed using half-Fraser and Fraser broths, in which the overgrowth of
L. innocua
can occur in samples where both species are present. In this study, we analyzed the induction of phages and phage tails of
Listeria
spp. in these media and in two brain heart infusion (BHI) broths (BHIM [bioMérieux] and BHIK [Biokar]) to identify putative effectors. It appears that Na
2
HPO
4
at concentrations ranging from 1 to 40 g/liter with an initial pH of 7.5 can induce phage or phage tail production of
Listeria
spp., especially with 10 g/liter of Na
2
HPO
4
and a pH of 7.5, conditions present in half-Fraser and Fraser broths. Exposure to LiCl in BHIM (18 to 21 g/liter) can also induce phage and phage tail release, but in half-Fraser and Fraser broths, the concentration of LiCl is much lower (3 g/liter). Low phage titers were induced by acriflavine and/or nalidixic acid. We also show that the production of phages and phage tails can occur in half-Fraser and Fraser broths. This study points out that induction of phages and phage tails could be triggered by compounds present in enrichment media. This could lead to a false-negative result for the detection of
L. monocytogenes
in food products.
Identification and Characterization of Carboxyl Esterases of Gill Chamber-Associated Microbiota in the Deep-Sea Shrimp Rimicaris exoculata by Using Functional Metagenomics
Authors: Alcaide María , Tchigvintsev Anatoli , Martínez-Martínez Mónica , Popovic Ana , Reva Oleg N. , Lafraya Álvaro , Bargiela Rafael , Nechitaylo Taras Y. , Matesanz Ruth , Cambon-Bonavita Marie-Anne , Jebbar Mohamed , Yakimov Michail M. , Savchenko Alexei , Golyshina Olga V. , Yakunin Alexander F. , Golyshin Peter N. , Ferrer Manuel
Published: January 2015

The shrimpRimicaris exoculatadominates the fauna in deep-sea hydrothermal vent sites along the Mid-Atlantic Ridge (depth, 2,320 m). Here, we identified and biochemically characterized three carboxyl esterases from microbial communities inhabiting theR. exoculatagill that were isolated by naive screens of a gill chamber metagenomic library. These proteins exhibit low to moderate identity to known esterase sequences (≤52%) and to each other (11.9 to 63.7%) and appear to have originated from unknown species or from genera ofProteobacteriarelated toThiothrix/Leucothrix(MGS-RG1/RG2) and to theRhodobacteraceaegroup (MGS-RG3). A library of 131 esters and 31 additional esterase/lipase preparations was used to evaluate the activity profiles of these enzymes. All 3 of these enzymes had greater esterase than lipase activity and exhibited specific activities with ester substrates (≤356 U mg−1) in the range of similar enzymes. MGS-RG3 was inhibited by salts and pressure and had a low optimal temperature (30°C), and its substrate profile clustered within a group of low-activity and substrate-restricted marine enzymes. In contrast, MGS-RG1 and MGS-RG2 were most active at 45 to 50°C and were salt activated and barotolerant. They also exhibited wider substrate profiles that were close to those of highly active promiscuous enzymes from a marine hydrothermal vent (MGS-RG2) and from a cold brackish lake (MGS-RG1). The data presented are discussed in the context of promoting the examination of enzyme activities of taxa found in habitats that have been neglected for enzyme prospecting; the enzymes found in these taxa may reflect distinct habitat-specific adaptations and may constitute new sources of rare reaction specificities.
Bacteriophage PBC1 and Its Endolysin as an Antimicrobial Agent against Bacillus cereus
Authors: Kong Minsuk , Ryu Sangryeol
Published: January 2015

Bacillus cereus
is an opportunistic human pathogen responsible for food poisoning and other, nongastrointestinal infections. Due to the emergence of multidrug-resistant
B. cereus
strains, the demand for alternative therapeutic options is increasing. To address these problems, we isolated and characterized a
Siphoviridae
virulent phage, PBC1, and its lytic enzymes. PBC1 showed a very narrow host range, infecting only 1 of 22
B. cereus
strains. Phylogenetic analysis based on the major capsid protein revealed that PBC1 is more closely related to the
Bacillus clarkii
phage BCJA1c and phages of lactic acid bacteria than to the phages infecting
B. cereus
. Whole-genome comparison showed that the late-gene region, including the terminase gene, structural genes, and holin gene of PBC1, is similar to that from
B. cereus
temperate phage 250, whereas their endolysins are different. Compared to the extreme host specificity of PBC1, its endolysin, LysPBC1, showed a much broader lytic spectrum, albeit limited to the genus
Bacillus
. The catalytic domain of LysPBC1 when expressed alone also showed
Bacillus
-specific lytic activity, which was lower against the
B. cereus
group but higher against the
Bacillus subtilis
group than the full-length protein. Taken together, these results suggest that the virulent phage PBC1 is a useful component of a phage cocktail to control
B. cereus
, even with its exceptionally narrow host range, as it can kill a strain of
B. cereus
that is not killed by other phages, and that LysPBC1 is an alternative biocontrol agent against
B. cereus
.
Effects of Norspermidine and Spermidine on Biofilm Formation by Potentially Pathogenic Escherichia coli and Salmonella enterica Wild-Type Strains
Authors: Nesse Live L. , Berg Kristin , Vestby Lene K.
Published: January 2015

Polyamines are present in all living cells. In bacteria, polyamines are involved in a variety of functions, including biofilm formation, thus indicating that polyamines may have potential in the control of unwanted biofilm. In the present study, the effects of the polyamines norspermidine and spermidine on biofilms of 10 potentially pathogenic wild-type strains of
Escherichia coli
serotype O103:H2,
Salmonella enterica
subsp.
enterica
serovar Typhimurium, and
S. enterica
serovar Agona were investigated. We found that exogenously supplied norspermidine and spermidine did not mediate disassembly of preformed biofilm of any of the
E. coli
and
S. enterica
strains. However, the polyamines did affect biofilm production. Interestingly, the two species reacted differently to the polyamines. Both polyamines reduced the amount of biofilm formed by
E. coli
but tended to increase biofilm formation by
S. enterica
. Whether the effects observed were due to the polyamines specifically targeting biofilm formation, being toxic for the cells, or maybe a combination of the two, is not known. However, there were no indications that the effect was mediated through binding to exopolysaccharides, as earlier suggested for
E. coli
. Our results indicate that norspermidine and spermidine do not have potential as inhibitors of
S. enterica
biofilm. Furthermore, we found that the commercial polyamines used contributed to the higher pH of the test medium. Failure to acknowledge and control this important phenomenon may lead to misinterpretation of the results.
Retention in Treated Wastewater Affects Survival and Deposition of Staphylococcus aureus and Escherichia coli in Sand Columns
Authors: Li Jiuyi , Zhao Xiaokang , Tian Xiujun , Li Jin , Sjollema Jelmer , Wang Aimin
Published: January 2015

The fate and transport of pathogenic bacteria from wastewater treatment facilities in the Earth's subsurface have attracted extensive concern over recent decades, while the impact of treated-wastewater chemistry on bacterial viability and transport behavior remains unclear. The influence of retention time in effluent from a full-scale municipal wastewater treatment plant on the survival and deposition of
Staphylococcus aureus
and
Escherichia coli
strains in sand columns was investigated in this paper. In comparison to the bacteria cultivated in nutrient-rich growth media, retention in treated wastewater significantly reduced the viability of all strains. Bacterial surface properties, e.g., zeta potential, hydrophobicity, and surface charges, varied dramatically in treated wastewater, though no universal trend was found for different strains. Retention in treated wastewater effluent resulted in changes in bacterial deposition in sand columns. Longer retention periods in treated wastewater decreased bacterial deposition rates for the strains evaluated and elevated the transport potential in sand columns. We suggest that the wastewater quality should be taken into account in estimating the fate of pathogenic bacteria discharged from wastewater treatment facilities and the risks they pose in the aquatic environment.
Involvement of the Cytochrome P450 System EthBAD in the
N
-Deethoxymethylation of Acetochlor by Rhodococcus sp. Strain T3-1
Authors: Wang Fei , Zhou Jie , Li Zhoukun , Dong Weiliang , Hou Ying , Huang Yan , Cui Zhongli
Published: January 2015

Acetochlor [2-chloro-
N
-(ethoxymethyl)-
N
-(2-ethyl-6-methylphenyl)-acetamide] is a widely applied herbicide with potential carcinogenic properties.
N
-Deethoxymethylation is the key step in acetochlor biodegradation.
N
-Deethoxymethylase is a multicomponent enzyme that catalyzes the conversion of acetochlor to 2′-methyl-6′-ethyl-2-chloroacetanilide (CMEPA). Fast detection of CMEPA by a two-enzyme (
N
-deethoxymethylase–amide hydrolase) system was established in this research. Based on the fast detection method, a three-component enzyme was purified from
Rhodococcus
sp. strain T3-1 using ammonium sulfate precipitation and hydrophobic interaction chromatography. The molecular masses of the components of the purified enzyme were estimated to be 45, 43, and 11 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Based on the results of peptide mass fingerprint analysis, acetochlor
N
-deethoxymethylase was identified as a cytochrome P450 system, composed of a cytochrome P450 oxygenase (43-kDa component; EthB), a ferredoxin (45 kDa; EthA), and a reductase (11 kDa; EthD), that is involved in the degradation of methyl
tert
-butyl ether. The gene cluster
ethABCD
was cloned by PCR amplification and expressed in
Escherichia coli
BL21(DE3). Resting cells of a recombinant
E. coli
strain showed deethoxymethylation activity against acetochlor. Subcloning of
ethABCD
showed that
ethABD
expressed in
E. coli
BL21(DE3) has the activity of acetochlor
N
-deethoxymethylase and is capable of converting acetochlor to CMEPA.
Thermal and Solvent Stress Cross-Tolerance Conferred to Corynebacterium glutamicum by Adaptive Laboratory Evolution
Authors: Oide Shinichi , Gunji Wataru , Moteki Yasuhiro , Yamamoto Shogo , Suda Masako , Jojima Toru , Yukawa Hideaki , Inui Masayuki
Published: January 2015

Reinforcing microbial thermotolerance is a strategy to enable fermentation with flexible temperature settings and thereby to save cooling costs. Here, we report on adaptive laboratory evolution (ALE) of the amino acid-producing bacteriumCorynebacterium glutamicumunder thermal stress. After 65 days of serial passage of the transgenic strain GLY3, in which the glycolytic pathway is optimized for alanine production under oxygen deprivation, three strains adapted to supraoptimal temperatures were isolated, and all the mutations they acquired were identified by whole-genome resequencing. Of the 21 mutations common to the three strains, one large deletion and two missense mutations were found to promote growth of the parental strain under thermal stress. Additive effects on thermotolerance were observed among these mutations, and the combination of the deletion with the missense mutation onotsA, encoding a trehalose-6-phosphate synthase, allowed the parental strain to overcome the upper limit of growth temperature. Surprisingly, the three evolved strains acquired cross-tolerance for isobutanol, which turned out to be partly attributable to the genomic deletion associated with the enhanced thermotolerance. The deletion involved loss of two transgenes,pfkandpyk, encoding the glycolytic enzymes, in addition to six native genes, and elimination of the transgenes, but not the native genes, was shown to account for the positive effects on thermal and solvent stress tolerance, implying a link between energy-producing metabolism and bacterial stress tolerance. Overall, the present study provides evidence that ALE can be a powerful tool to refine the phenotype ofC. glutamicumand to investigate the molecular bases of stress tolerance.
Novel Hypovirulence-Associated RNA Mycovirus in the Plant-Pathogenic Fungus Botrytis cinerea: Molecular and Biological Characterization
Authors: Yu Lin , Sang Wen , Wu Ming-De , Zhang Jing , Yang Long , Zhou Ying-Jun , Chen Wei-Dong , Li Guo-Qing
Published: January 2015

Botrytis cinerea
is a pathogenic fungus causing gray mold on numerous economically important crops and ornamental plants. This study was conducted to characterize the biological and molecular features of a novel RNA mycovirus, Botrytis cinerea RNA virus 1 (BcRV1), in the hypovirulent strain BerBc-1 of
B. cinerea
. The genome of BcRV1 is 8,952 bp long with two putative overlapped open reading frames (ORFs), ORF1 and ORF2, coding for a hypothetical polypeptide (P1) and RNA-dependent RNA polymerase (RdRp), respectively. A −1 frameshifting region (designated the KNOT element) containing a shifty heptamer, a heptanucleotide spacer, and an H-type pseudoknot was predicted in the junction region of ORF1 and ORF2. The −1 frameshifting role of the KNOT element was experimentally confirmed through determination of the production of the fusion protein red fluorescent protein (RFP)-green fluorescent protein (GFP) by the plasmid containing the construct
dsRed-
KNOT-
eGFP
in
Escherichia coli
. BcRV1 belongs to a taxonomically unassigned double-stranded RNA (dsRNA) mycovirus group. It is closely related to grapevine-associated totivirus 2 and Sclerotinia sclerotiorum nonsegmented virus L. BcRV1 in strain BerBc-1 was found capable of being transmitted vertically through macroconidia and horizontally to other
B. cinerea
strains through hyphal contact. The presence of BcRV1 was found to be positively correlated with hypovirulence in
B. cinerea
, with the attenuation effects of BcRV1 on mycelial growth and pathogenicity being greatly affected by the accumulation level of BcRV1.
Identification and Characterization of LFD-2, a Predicted Fringe Protein Required for Membrane Integrity during Cell Fusion in Neurospora crassa
Authors: Palma-Guerrero Javier , Zhao Jiuhai , Gonçalves A. Pedro , Starr Trevor L. , Glass N. Louise
Published: January 2015

The molecular mechanisms of membrane merger during somatic cell fusion in eukaryotic species are poorly understood. In the filamentous fungus
Neurospora crassa
, somatic cell fusion occurs between genetically identical germinated asexual spores (germlings) and between hyphae to form the interconnected network characteristic of a filamentous fungal colony. In
N. crassa
, two proteins have been identified to function at the step of membrane fusion during somatic cell fusion: PRM1 and LFD-1. The absence of either one of these two proteins results in an increase of germling pairs arrested during cell fusion with tightly appressed plasma membranes and an increase in the frequency of cell lysis of adhered germlings. The level of cell lysis in Δ
Prm1
or Δ
lfd-1
germlings is dependent on the extracellular calcium concentration. An available transcriptional profile data set was used to identify genes encoding predicted transmembrane proteins that showed reduced expression levels in germlings cultured in the absence of extracellular calcium. From these analyses, we identified a mutant (
lfd-2
, for

l

ate

f

usion

d

efect-2
) that showed a calcium-dependent cell lysis phenotype.
lfd-2
encodes a protein with a Fringe domain and showed endoplasmic reticulum and Golgi membrane localization. The deletion of an additional gene predicted to encode a low-affinity calcium transporter,
fig1
, also resulted in a strain that showed a calcium-dependent cell lysis phenotype. Genetic analyses showed that LFD-2 and FIG1 likely function in separate pathways to regulate aspects of membrane merger and repair during cell fusion.
Molecular Characterization of Hap Complex Components Responsible for Methanol-Inducible Gene Expression in the Methylotrophic Yeast Candida boidinii
Authors: Oda Saori , Yurimoto Hiroya , Nitta Nobuhisa , Sasano Yu , Sakai Yasuyoshi
Published: January 2015

We identified genes encoding components of the Hap complex,
CbHAP2
,
CbHAP3
, and
CbHAP5
, as transcription factors regulating methanol-inducible gene expression in the methylotrophic yeast
Candida boidinii
. We found that the
Cbhap2
Δ,
Cbhap3
Δ, and
Cbhap5
Δ gene-disrupted strains showed severe growth defects on methanol but not on glucose and nonfermentable carbon sources such as ethanol and glycerol. In these disruptants, the transcriptional activities of methanol-inducible promoters were significantly decreased compared to those of the wild-type strain, indicating that CbHap2p, CbHap3p, and CbHap5p play indispensable roles in methanol-inducible gene expression. Further molecular and biochemical analyses demonstrated that CbHap2p, CbHap3p, and CbHap5p localized to the nucleus and bound to the promoter regions of methanol-inducible genes regardless of the carbon source, and heterotrimer formation was suggested to be necessary for binding to DNA. Unexpectedly, distinct from
Saccharomyces cerevisiae
, the Hap complex functioned in methanol-specific induction rather than glucose derepression in
C. boidinii
. Our results shed light on a novel function of the Hap complex in methanol-inducible gene expression in methylotrophic yeasts.
A FACS-Optimized Screen Identifies Regulators of Genome Stability in Candida albicans
Authors: Loll-Krippleber Raphaël , Feri Adeline , Nguyen Marie , Maufrais Corinne , Yansouni Jennifer , d'Enfert Christophe , Legrand Mélanie
Published: January 2015

Loss of heterozygosity (LOH) plays important roles in genome dynamics, notably, during tumorigenesis. In the fungal pathogen
Candida albicans
, LOH contributes to the acquisition of antifungal resistance. In order to investigate the mechanisms that regulate LOH in
C. albicans
, we have established a novel method combining an artificial heterozygous locus harboring the blue fluorescent protein and green fluorescent protein markers and flow cytometry to detect LOH events at the single-cell level. Using this fluorescence-based method, we have confirmed that elevated temperature, treatment with methyl methanesulfonate, and inactivation of the Mec1 DNA damage checkpoint kinase triggered an increase in the frequency of LOH. Taking advantage of this system, we have searched for
C. albicans
genes whose overexpression triggered an increase in LOH and identified four candidates, some of which are known regulators of genome dynamics with human homologues contributing to cancer progression. Hence, the approach presented here will allow the implementation of new screens to identify genes that are important for genome stability in
C. albicans
and more generally in eukaryotic cells.
Electroencephalography as a clinical tool for diagnosing and monitoring attention deficit hyperactivity disorder: a cross-sectional study
Authors: Helgadóttir Halla , Gudmundsson Ólafur Ó , Baldursson Gísli , Magnússon Páll , Blin Nicolas , Brynjólfsdóttir Berglind , Emilsdóttir Ásdís , Gudmundsdóttir Gudrún B , Lorange Málfrídur , Newman Paula K , Jóhannesson Gísli H , Johnsen Kristinn
Published: January 2015

Objectives

The aim of this study was to develop and test, for the first time, a multivariate diagnostic classifier of attention deficit hyperactivity disorder (ADHD) based on EEG coherence measures and chronological age.

Setting

The participants were recruited in two specialised centres and three schools in Reykjavik.

Participants

The data are from a large cross-sectional cohort of 310 patients with ADHD and 351 controls, covering an age range from 5.8 to 14 years. ADHD was diagnosed according to the Diagnostic and Statistical Manual of Mental Disorders fourth edition (DSM-IV) criteria using the K-SADS-PL semistructured interview. Participants in the control group were reported to be free of any mental or developmental disorders by their parents and had a score of less than 1.5 SDs above the age-appropriate norm on the ADHD Rating Scale-IV. Other than moderate or severe intellectual disability, no additional exclusion criteria were applied in order that the cohort reflected the typical cross section of patients with ADHD.

Results

Diagnostic classifiers were developed using statistical pattern recognition for the entire age range and for specific age ranges and were tested using cross-validation and by application to a separate cohort of recordings not used in the development process. The age-specific classification approach was more accurate (76% accuracy in the independent test cohort; 81% cross-validation accuracy) than the age-independent version (76%; 73%). Chronological age was found to be an important classification feature.

Conclusions

The novel application of EEG-based classification methods presented here can offer significant benefit to the clinician by improving both the accuracy of initial diagnosis and ongoing monitoring of children and adolescents with ADHD. The most accurate possible diagnosis at a single point in time can be obtained by the age-specific classifiers, but the age-independent classifiers are also useful as they enable longitudinal monitoring of brain function.
Establishment of a prospective cohort of mechanically ventilated patients in five intensive care units in Lima, Peru: protocol and organisational characteristics of participating centres
Authors: Denney Joshua A , Capanni Francesca , Herrera Phabiola , Dulanto Augusto , Roldan Rollin , Paz Enrique , Jaymez Amador A , Chirinos Eduardo E , Portugal Jose , Quispe Rocio , Brower Roy G , Checkley William ,   INTENSIVOS Cohort Study
Published: January 2015

Introduction

Mechanical ventilation is a cornerstone in the management of critically ill patients worldwide; however, less is known about the clinical management of mechanically ventilated patients in low and middle income countries where limitation of resources including equipment, staff and access to medical information may play an important role in defining patient-centred outcomes. We present the design of a prospective, longitudinal study of mechanically ventilated patients in Peru that aims to describe a large cohort of mechanically ventilated patients and identify practices that, if modified, could result in improved patient-centred outcomes and lower costs.

Methods and analysis

Five Peruvian intensive care units (ICUs) and the Medical ICU at the Johns Hopkins Hospital were selected for this study. Eligible patients were those who underwent at least 24 h of invasive mechanical ventilation within the first 48 h of admission into the ICU. Information on ventilator settings, clinical management and treatment were collected daily for up to 28 days or until the patient was discharged from the unit. Vital status was assessed at 90 days post enrolment. A subset of participants who survived until hospital discharge were asked to participate in an ancillary study to assess vital status, and physical and mental health at 6, 12, 24 and 60 months after hospitalisation, Primary outcomes include 90-day mortality, time on mechanical ventilation, hospital and ICU lengths of stay, and prevalence of acute respiratory distress syndrome. In subsequent analyses, we aim to identify interventions and standardised care strategies that can be tailored to resource-limited settings and that result in improved patient-centred outcomes and lower costs.

Ethics and dissemination

We obtained ethics approval from each of the four participating hospitals in Lima, Peru, and at the Johns Hopkins School of Medicine, Baltimore, USA. Results will be disseminated as several separate publications in different international journals.
Detection for pathway effect contributing to disease in systems epidemiology with a case–control design
Authors: Ji Jiadong , Yuan Zhongshang , Zhang Xiaoshuai , Li Fangyu , Xu Jing , Liu Ying , Li Hongkai , Wang Jia , Xue Fuzhong
Published: January 2015

Objectives

Identification of pathway effects responsible for specific diseases has been one of the essential tasks in systems epidemiology. Despite some advance in procedures for distinguishing specific pathway (or network) topology between different disease status, statistical inference at a population level remains unsolved and further development is still needed. To identify the specific pathways contributing to diseases, we attempt to develop powerful statistics which can capture the complex relationship among risk factors.

Setting and participants

Acute myeloid leukaemia (AML) data obtained from 133 adults (98 patients and 35 controls; 47% female).

Results

Simulation studies indicated that the proposed Pathway Effect Measures (PEM) were stable; bootstrap-based methods outperformed the others, with bias-corrected bootstrap CI method having the highest power. Application to real data of AML successfully identified the specific pathway (Treg→TGFβ→Th17) effect contributing to AML with p values less than 0.05 under various methods and the bias-corrected bootstrap CI (−0.214 to −0.020). It demonstrated that Th17–Treg correlation balance was impaired in patients with AML, suggesting that Th17–Treg imbalance potentially plays a role in the pathogenesis of AML.

Conclusions

The proposed bootstrap-based PEM are valid and powerful for detecting the specific pathway effect contributing to disease, thus potentially providing new insight into the underlying mechanisms and ways to study the disease effects of specific pathways more comprehensively.
Differentiation between mycoplasma and viral community-acquired pneumonia in children with lobe or multi foci infiltration: a retrospective case study
Authors: Guo Wan-liang , Wang Jian , Zhu Li-yuan , Hao Chuang-li
Published: January 2015

Objectives

To analyse the clinical features, inflammatory markers and radiographs of community-acquired pneumonia (CAP) cases with lobe or multi foci infiltration; with a special focus on factors which allow the differential diagnosis of viral and mycoplasma pneumonia.

Setting

Retrospective chart review of CAP cases in a large university teaching hospital.

Participants

126 paediatric CAP cases, with lobe or multi foci infiltration, presenting between May 2012 and April 2013. Demographic data, clinical presentation on admission or referral, laboratory tests, prior history, and radiography were collected for each case if available.

Primary and secondary outcome measures

We used univariate and multivariate logistic regression to determine the significant factors which allow the differential diagnosis of viral and mycoplasma CAP with lobe or multi foci infiltration.

Results

There were 71 (56%) male and 55 (44%) female CAP cases with lobar or multi foci infiltration. 70 pneumonia cases were caused byMycoplasma pneumoniaeand 18 by viruses. Univariate analysis of the mycoplasma and viral causes of the CAP revealed that increased respiratory rate, wheeze, male gender and lymphocyte percentage were the factors associated with the differentiation of mycoplasma and viral aetiologies of pneumonia (p<0.05). A stepwise logistic regression analysis was performed to assess independent factors which allow the differential diagnosis of viral and mycoplasma pneumonia. Increased respiratory rate, wheeze, and lymphocyte percentage were reliable independent factors which allow the differential diagnosis of viral and mycoplasma CAP with lobar or multi foci infiltration.

Conclusions

Whether the CAP with lobar or multi foci infiltration was caused by mycoplasma species or viruses could not be inferred from the radiological patterns. Wheeze, lymphocyte percentage and respiratory rate were independent factors which allowed the differential diagnosis of viral and mycoplasma CAP with lobar or multi foci infiltration.
The impact of a streamlined funding application process on application time: two cross-sectional surveys of Australian researchers
Authors: Barnett Adrian G , Graves Nicholas , Clarke Philip , Herbert Danielle
Published: January 2015

Objective

To examine if streamlining a medical research funding application process saved time for applicants.

Design

Cross-sectional surveys before and after the streamlining.

Setting

The National Health and Medical Research Council (NHMRC) of Australia.

Participants

Researchers who submitted one or more NHMRC Project Grant applications in 2012 or 2014.

Main outcome measures

Average researcher time spent preparing an application and the total time for all applications in working days.

Results

The average time per application increased from 34 working days before streamlining (95% CI 33 to 35) to 38 working days after streamlining (95% CI 37 to 39; mean difference 4 days, bootstrap p value <0.001). The estimated total time spent by all researchers on applications after streamlining was 614 working years, a 67-year increase from before streamlining.

Conclusions

Streamlined applications were shorter but took longer to prepare on average. Researchers may be allocating a fixed amount of time to preparing funding applications based on their expected return, or may be increasing their time in response to increased competition. Many potentially productive years of researcher time are still being lost to preparing failed applications.