bluuebunny/crossref_metadata_2025 huggingface dataset:
Applying Molecular Dynamics Simulations to Identify Rarely Sampled Ligand‐bound Conformational States of Undecaprenyl Pyrophosphate Synthase, an Antibacterial Target
Authors: Sinko William , de Oliveira César , Williams Sarah , Van Wynsberghe Adam , Durrant Jacob D. , Cao Rong , Oldfield Eric , McCammon J. Andrew
Published: February 2011

Undecaprenyl pyrophosphate synthase is a cis‐prenyltransferase enzyme, which is required for cell wall biosynthesis in bacteria. Undecaprenyl pyrophosphate synthase is an attractive target for antimicrobial therapy. We performed long molecular dynamics simulations and docking studies on undecaprenyl pyrophosphate synthase to investigate its dynamic behavior and the influence of protein flexibility on the design of undecaprenyl pyrophosphate synthase inhibitors. We also describe the first X‐ray crystallographic structure of Escherichia coli apo‐undecaprenyl pyrophosphate synthase. The molecular dynamics simulations indicate that undecaprenyl pyrophosphate synthase is a highly flexible protein, with mobile binding pockets in the active site. By carrying out docking studies with experimentally validated undecaprenyl pyrophosphate synthase inhibitors using high‐ and low‐populated conformational states extracted from the molecular dynamics simulations, we show that structurally dissimilar compounds can bind preferentially to different and rarely sampled conformational states. By performing structural analyses on the newly obtained apo‐undecaprenyl pyrophosphate synthase and other crystal structures previously published, we show that the changes observed during the molecular dynamics simulation are very similar to those seen in the crystal structures obtained in the presence or absence of ligands. We believe that this is the first time that a rare ‘expanded pocket’ state, key to drug design and verified by crystallography, has been extracted from a molecular dynamics simulation.
Rib metastases from a non‐tonsillar squamous cell carcinoma in a dog
Authors: Clarke B. S. , Mannion P. A. , White R. A. S.
Published: February 2011

Metastatic spread to the ribs in a 15‐year‐old, male, neutered, Irish setter is reported occurring secondary to an oral squamous cell carcinoma (SCC). The dog presented with a history of a rapidly growing SCC of the right upper incisive region, which was confirmed by histopathology as a SCC. Thoracic radiographs showed bony lesions associated with the body of the right third rib, and the fifth and seventh costal cartilages. A rostral partial maxillectomy was performed as palliative treatment for the oral mass and a core biopsy of the lesion on the third rib was performed. The rib lesion was identified histopathologically as a metastatic SCC. A review of the literature of oropharyngeal SCC and the metastatic potential of non‐tonsillar SCC is presented, in particular metastatic bone disease. This case report suggests possible implications of metastatic bone disease for treatment and prognosis for future cases of non‐tonsillar SCC.
Postoperative haemorrhage associated with active suction drains in two dogs
Authors: Lynch A. M. , Bound N. J. , Halfacree Z. J. , Baines S.
Published: February 2011

This article describes two dogs in which an active suction drain was placed to manage dead space at the surgical site and acute haemorrhage and hypovolaemia occurred postoperatively. In both instances, fluid resuscitation and temporary discontinuation of drainage resulted in resolution of clinical signs. Although the underlying cause of haemorrhage was not definitively identified, the use of low‐pressure drainage systems and avoidance of interference with local blood vessels should be considered. This is a previously undocumented complication of active suction drain use in veterinary patients.
Japan's Bubble, the USA's Bubble and China's Bubble
Authors: Ueda Kazuo
Published: February 2011

This paper compares the three recent episodes of boom and bust cycles in asset prices: Japan in the late 1980s to the 1990s; the USA since the mid‐1990s; and China during the past decade. Although we have not yet seen a collapse of Chinese property prices, their increases so far are comparable to those in the other two episodes and a careful comparative study is warranted. The present paper first examines the behavior of asset prices, of property prices in particular, in the three cases, and highlights some similarities. The paper emphasizes the role played by extremely easy monetary policy in generating bubble‐like asset price behavior in the three cases. The reason for easy monetary policies is investigated. In the US case, the monetary authority was concerned about the risk of deflation in the early to mid‐2000s. The experiences of Japan and China are quite similar in that the monetary authorities of both countries were seriously concerned about the possible deflationary effects of exchange rate appreciation on the economy. The implications of such a finding for the future of Chinese macroeconomic policy are discussed.
Solvent Dependence of the Photophysical Properties of 2‐Chlorothioxanthone, the Principal Photoproduct of Chlorprothixene
Authors: Piñero Santiago Luis E. , García Carmelo , Lhiaubet‐Vallet Virginie , Miranda Miguel A. , Oyola Rolando
Published: February 2011

2‐chlorothioxanthone (CTX) is used as photoinitiator for the reticulation of synthetic resins and for the preparation of pharmaceuticals. It was previously determined that CTX is the primary photoproduct of z‐chlorprothixene (CPTX) when irradiated at 313 nm and is formed in an autocatalyzed reaction through an energy‐transfer mechanism (Piñeroet al.[2009]Photochem. Photobiol.,85, 895–900). In this work, the photophysical properties of CTX were measured in acetonitrile/water solutions to determine if their magnitude can affect the side effects of CPTX. The results show that CTX has higher absorption coefficients in the visible region (400–420 nm) and higher triplet quantum yields than its parent compound. Similar to TX, both properties strongly depend on the solvent polarity/hydroxylicity. The quantum yield of the triplet intermediate is very close to the value of the phenothiazine triplets. The phenothiazines are the most phototoxic antidepressants. Therefore, given the appropriate microenvironment, the photosensitization side effects of CPTX can be intensified on the production of CTX.
Two highly validated multiplexes (12‐plex and 8‐plex) for species delimitation and parentage analysis in oaks (Quercus spp.)
Authors: GUICHOUX E. , LAGACHE L. , WAGNER S. , LÉGER P. , PETIT R.J.
Published: February 2011

Multiplex PCR is a fast and cost‐effective technique allowing increased genotyping throughput of microsatellites. We developed two multiplexes for Quercus petraea and Q. robur, a 12‐plex of EST‐SSRs (eSSRs) and an 8‐plex of genomic SSRs (gSSRs). We studied the origin of allele calling errors at the human reader and software levels. We showed that the robustness of allele identification can be improved by binning on raw peak sizes prior to genetic data analysis. We checked through simulation the power of these markers for species delimitation and hybrid detection. The resolution achieved with all 20 markers was greatly improved compared to that of previous studies based on a subset of the markers. Preliminary PCR tests suggest that these multiplexes might be useful to study other oak species as well. The strategy used for multiplex microsatellite development (from PCR conditions to the definition of allele calling rules) should be broadly applicable.
Crocus ilvensis sp. nov. (sect. Crocus, Iridaceae), endemic to Elba Island (Tuscan Archipelago, Italy)
Authors: Peruzzi Lorenzo , Carta Angelino
Published: February 2011

A new species of Crocus sect. Crocus is described as an endemic of Elba Island (Tuscan Archipelago, Italy): C. ilvensis Peruzzi & Carta sp. nov. (2n=8). The new species was wrongly referred to C. corsicus Vanucchi (2n=18), C. etruscus Parl. (2n=8) or C. vernus Hill. subsp. vernus (2n=8, 16) by previous authors. Its karyotype structure, asymmetry and chromosome dimensions show affinity with C. etruscus, and no clear relations with other related taxa. From a morphological point of view, C. ilvensis appears intermediate between C. etruscus and C. vernus subsp. vernus, showing however a peculiar combination of character‐states. The new species is completely allopatric with other Crocus sect. Crocus taxa and its possible role in the origin of tetraploid races of C. vernus is discussed.
A mathematical model for evaluating tuberculosis screening strategies
Authors: Jia Zhongwei , Cheng Shiming , Jia Xiaowei
Published: February 2011

In order to better predict epidemic of tuberculosis (TB) and evaluate the effect of TB control strategies, we added the effect of transmission from permanent residents to migrants to our previous TB model. We simulated the behavior of TB transmission by the extended model. The numerical simulation indicated that the basic reproductive numbers must be less than one in both permanent residents and migrants in order to eliminate the disease from total population. We also evaluated the Canada's TB screening strategy and observe that TB is sensitive to the strategy. Our study suggests that immigrants have a considerable influence on the overall transmission dynamics behavior of tuberculosis. The effective TB control measures should incorporate migrant as well the permanent residents.
The Predictive Ability of Business Survey Indices*
Authors: Chindamo Phillip
Published: February 2011

This article assesses the predictive ability of the survey‐based business performance indices produced by the Australian Industry Group with respect to “official” Australian economic data. The index related to manufacturing sector performance is a useful predictor of official economic data for the manufacturing sector. This usefulness is both in terms of providing early information on current period economic conditions and providing information regarding future period economic conditions. The results for the services and construction sector indices are mixed, with limited evidence of Granger causality from the services sector index to the corresponding official economic data for the services sector. There is some value in the construction sector index as both a leading indicator as well as a timelier contemporary indicator of construction sector economic activity.
Monitoring unfractionated heparin therapy in dogs by measuring thrombin generation
Authors: Allegret Virginie , Dunn Marilyn , Bédard Christian
Published: February 2011

Background: The calibrated automated thrombogram (CAT), an assay that permits measurement of thrombin generation in plasma, may be useful in studying hemostatic disorders and anticoagulant therapy in animals.

Objectives: The aims of the study were to measure thrombin generation in healthy Beagle dogs and to evaluate the potential use of the CAT assay for monitoring therapy with unfractionated heparin (UFH).

Methods: Individual platelet‐poor plasma samples and a plasma pool from 20 healthy adult Beagles were prepared. Serial UFH plasma dilutions were used to establish an in vitro heparin‐sensitivity curve. The pharmacodynamic effects of heparin in vivo were evaluated in Beagles using the CAT assay to measure thrombin generation with tissue factor at a concentration of 5 pM for initiation.

Results: In healthy Beagles, the range of endogenous thrombin potential (ETP) was 238.7–414.0 nM/min (mean ± SD, 340.4 ± 63.1 nM/min). ETP intra‐assay and interassay variations were 7.1% and 12.9%, respectively. In vitro, a UFH concentration ≥0.4 U/mL resulted in total inhibition of thrombin generation. In vivo, the maximal effect of UFH on ETP was observed at 170 ± 36 minutes (range, 120–210 minutes) and resulted in a decrease in ETP of 38.5 ± 7.8% (range, 26.5–50.3%). In 210–420 minutes, ETP returned to baseline in 5 dogs.

Conclusion: Our study demonstrates that thrombin generation can be measured in canine plasma and may be useful in assessing the degree of anticoagulation provided by UFH.
Moderately and slightly siderophile element constraints on the depth and extent of melting in early Mars
Authors: RIGHTER Kevin , CHABOT Nancy L.
Published: February 2011

Abstract–

The thermal history of Mars during accretion and differentiation is important for understanding some fundamental aspects of its evolution such as crust formation, mantle geochemistry, chronology, volatile loss and interior degassing, and atmospheric development. In light of data from new Martian meteorites and exploration rovers, we have made a new estimate of Martian mantle siderophile element depletions. New high pressure and temperature metal–silicate experimental partitioning data and expressions are also available. Using these new constraints, we consider the conditions under which the Martian mantle may have equilibrated with metallic liquid. The resulting conditions that best satisfy six siderophile elements—Ni, Co, W, Mo, P, and Ga—and are consistent with the solidus and liquidus of the Martian mantle phase diagram are a pressure of 14 ± 3 GPa and temperature of 2100 ± 200 K. The Martian mantle depletions of Cr and V are also consistent with metal–silicate equilibration in this pressure and temperature range if deep mantle silicate phases are also taken into account. The results are not consistent with either metal–silicate equilibrium at the surface or at the current‐day Martian core–mantle boundary. Recent measurements and modeling have concluded that deep (∼17 GPa or 1350 km) mantle melting is required to explain isotopic data for Martian meteorites and the nature of differentiation into core, mantle, and crust. This is in general agreement with our estimates of the conditions of Martian core formation based on siderophile elements that result in an intermediate depth magma ocean scenario for metal–silicate equilibrium.
How can we enhance girls’ interest in scientific topics?
Authors: Kerger Sylvie , Martin Romain , Brunner Martin
Published: February 2011

Background. Girls are considerably less interested in scientific subjects than boys. One reason may be that scientific subjects are considered to be genuinely masculine. Thus, being interested in science may threaten the self‐perception of girls as well as the femininity of their self‐image.

Aims. If scientific topics that are considered to be stereotypically feminine were chosen, however, this potential threat might be overcome which, in turn, might lead to an increase in girls’ interest in science. This hypothesis was empirically tested by means of two studies.

Sample. Participants were 294 (Study 1) and 190 (Study 2) Grade 8 to Grade 9 students.

Method. Gender differences in students’ interest in masculine and feminine topics were investigated for a range of scientific concepts (Study 1) as well as for a given scientific concept (Study 2) for four scientific subjects (i.e., biology, physics, information technology, and statistics), respectively.

Results. Both studies indicated that the mean level of girls’ scientific interest was higher when scientific concepts were presented in the context of feminine topics and boys’ level of scientific interests was higher when scientific concepts were presented in the context of masculine topics.

Conclusion. Girls’ interest in science could be substantially increased by presenting scientific concepts in the context of feminine topics. Gender differences as well as individual differences in the level of interest in scientific topics may be taken into account by creating learning environments in which students could select the context in which a certain scientific concept is embedded.
Achievement goals as mediators of the relationship between competence beliefs and test anxiety
Authors: Putwain David W. , Symes Wendy
Published: February 2011

Background. Previous work suggests that the expectation of failure is related to higher test anxiety and achievement goals grounded in a fear of failure.

Aim. To test the hypothesis, based on the work of Elliot and Pekrun (2007), that the relationship between perceived competence and test anxiety is mediated by achievement goal orientations.

Sample. Self‐report data were collected from 275 students in post‐compulsory education following courses in A Level Psychology.

Results. Competence beliefs were inversely related to the worry and tension components of test anxiety, both directly and indirectly through a performance‐avoidance goal orientation. A mastery‐avoidance goal orientation offered an indirect route from competence beliefs to worry only.

Conclusion. These findings provide partial support for Elliot and Pekrun's (2007) model. Although significant mediating effects were found for mastery‐avoidance and performance‐avoidance goals, they were small and there may be other mechanisms to account for the relations between competence beliefs and test anxiety.
Personal reputation in organizations: Two‐study constructive replication and extension of antecedents and consequences
Authors: Zinko Robert , Ferris Gerald R. , Humphrey Stephen E. , Meyer Christopher J. , Aime Federico
Published: February 2011

Personal reputation has been argued to demonstrate important influences on work outcomes. However, substantive research on personal reputation is relatively scarce. This two‐study investigation empirically supports and extends existing theory regarding the temporal development of personal reputation (i.e., antecedents and consequences), and thus contributes to a more informed understanding of both the construct and criterion‐related validity of this important construct. Study 1 is conducted longitudinally, in order to assess the development of personal reputation over time, which is undertaken to demonstrate the effects of human capital and social effectiveness as antecedents of reputation. Study 2 complements and extends the first study by conducting a field investigation examining the effects of time, human capital, and social effectiveness as antecedents of personal reputation, while also exploring the reputation consequences of autonomy, power, and career success. Our findings suggest that human capital, time, and social effectiveness play a part in the development of a reputation. Furthermore, career success, power, and autonomy were shown to be outcomes of the reputation construct. Contributions and strengths of this investigation, limitations, and directions for future research are discussed.
Peer socialization of masculinity and femininity: Differential effects of overt and relational forms of peer victimization
Authors: Ewing Lee Elizabeth A. , Troop‐Gordon Wendy
Published: February 2011

Although peer influence has been implicated in recent theories of gender socialization, few investigations have tested whether children's gendered behaviours change over time as a function of peer experiences and whether some peer experiences may exacerbate, rather than dampen, gender non‐conformity. Accordingly, the current study examined prospective links between specific forms of peer victimization and children's adherence to traditional gender roles. Peer reports of victimization and self‐reports of engagement in stereotypically masculine and feminine activities were collected from 199 children (104 girls; 95 boys) in the Fall and Spring of their fifth‐grade year. Multi‐group path analysis was used to explore the relations between forms of victimization and masculinity and femininity for girls and boys. For girls, peer victimization predicted withdrawal from both feminine and masculine behaviours. For boys, physical, verbal, and general victimization predicted lower levels of feminine behaviours, but social exclusion forecast heightened engagement in traditionally feminine activities. These findings underscore how social experiences can amplify, as well as reduce, gender non‐conformity.
Surface Chemistry of ZnO Nanowires for Nanosensor Applications
Authors: Gupta Anurag , Kim Bruce C. , Watkins Clifton C. , Street Shane C. , Edwards Eugene , Brantley Christina , Ruffin Paul
Published: February 2011

In this work, surface modification of ZnO nanowires (NWs) with oleic acid is reported. ZnO NWs were synthesized in a customized chemical vapor deposition furnace and were characterized with scanning electron microscopy, X-ray diffraction, and energy dispersive spectroscopy. Raman and infrared spectroscopies were used to determine that oleic acid modifies the ZnO NW surface with the carboxylate group attached to the surface. The study of the surface chemistry of ZnO NWs provides information about the class of compounds that modify ZnO NWs. This information will assist in tailoring the interface of NWs with appropriate compounds for fabricating nanosensors with high selectivity.
Measurement of a Bluff Body Aerodynamic Yaw Moment Magnification and Damping Using a Dynamic Wind Tunnel Facility
Authors: Mansor Shuhaimi , Passmore Martin
Published: February 2011

This paper describes a technique for determining the dynamic aerodynamic yaw moment derivative based on the time response data using an oscillating model rig. The aerodynamic yaw moment derivatives are initially estimated using oscillation frequency and amplitude decay. The results from the dynamic measurement are compared with conventional static test and presented in the form of aerodynamic magnification. The yaw moment derivative exceeds that determined statically across reduced frequency range measured. The yaw damping derivative was found to be a function of freestream speed; at low velocities it is negative but progressively increases to a positive value. With further increases in speed, a self-sustained oscillation is observed with almost constant frequency and amplitude. This result is attributed to coupling between the model wake and the model stability; however, the exact behavior of the interaction is not fully understood; this phenomenon is under further investigation.
Multiple Approaches To Enhance the Cultivability of Bacteria Associated with the Marine Sponge
Haliclona
(
gellius
) sp
Authors: Sipkema Detmer , Schippers Klaske , Maalcke Wouter J. , Yang Yu , Salim Sina , Blanch Harvey W.
Published: February 2011

Three methods were examined to cultivate bacteria associated with the marine sponge
Haliclona
(
gellius
) sp.: agar plate cultures, liquid cultures, and floating filter cultures. A variety of oligotrophic media were employed, including media with aqueous and organic sponge extracts, bacterial signal molecules, and siderophores. More than 3,900 isolates were analyzed, and 205 operational taxonomic units (OTUs) were identified. Media containing low concentrations of mucin or a mixture of peptone and starch were most successful for the isolation of diversity, while the commonly used marine broth did not result in a high diversity among isolates. The addition of antibiotics generally led to a reduced diversity on plates but yielded different bacteria than other media. In addition, diversity patterns of isolates from agar plates, liquid cultures, and floating filters were significantly different. Almost 89% of all isolates were
Alphaproteobacteria
; however, members of phyla that are less commonly encountered in cultivation studies, such as
Planctomycetes
,
Verrucomicrobia
, and
Deltaproteobacteria
, were isolated as well. The sponge-associated bacteria were categorized into three different groups. The first group represented OTUs that were also obtained in a clone library from previously analyzed sponge tissue (group 1). Furthermore, we distinguished OTUs that were obtained from sponge tissue (in a previous study) but not from sponge isolates (group 2), and there were also OTUs that were not obtained from sponge tissue but were obtained from sponge isolates (group 3). The 17 OTUs categorized into group 1 represented 10 to 14% of all bacterial OTUs that were present in a large clone library previously generated from
Haliclona
(
gellius
) sp. sponge tissue, which is higher than previously reported cultivability scores for sponge-associated bacteria. Six of these 17 OTUs were not obtained from agar plates, which underlines that the use of multiple cultivation methods is worthwhile to increase the diversity of the cultivable microorganisms from sponges.
Parameters Affecting Spore Recovery from Wipes Used in Biological Surface Sampling
Authors: Da Silva Sandra M. , Filliben James J. , Morrow Jayne B.
Published: February 2011

The need for the precise and reliable collection of potential biothreat contaminants has motivated research in developing a better understanding of the variability in biological surface sampling methods. In this context, the objective of this work was to determine parameters affecting the efficiency of extracting
Bacillus anthracis
Sterne spores from commonly used wipe sampling materials and to describe performance using the interfacial energy concept. In addition, surface thermodynamics was applied to understand and predict surface sampling performance. Wipe materials were directly inoculated with known concentrations of
B. anthracis
spores and placed into extraction solutions, followed by sonication or vortexing. Experimental factors investigated included wipe material (polyester, cotton, and polyester-rayon), extraction solution (sterile deionized water [H
2
O], deionized water with 0.04% Tween 80 [H
2
O-T], phosphate-buffered saline [PBS], and PBS with 0.04% Tween 80 [PBST]), and physical dissociation method (vortexing or sonication). The most efficient extraction from wipes was observed for solutions containing the nonionic surfactant Tween 80. The increase in extraction efficiency due to surfactant addition was attributed to an attractive interfacial energy between Tween 80 and the centrifuge tube wall, which prevented spore adhesion. Extraction solution significantly impacted the extraction efficiency, as determined by statistical analysis (
P
< 0.05). Moreover, the extraction solution was the most important factor in extraction performance, followed by the wipe material. Polyester-rayon was the most efficient wipe material for releasing spores into solution by rank; however, no statistically significant difference between polyester-rayon and cotton was observed (
P
> 0.05). Vortexing provided higher spore recovery in H
2
O and H
2
O-T than sonication, when all three wipe materials and the reference control were considered (
P
< 0.05).
Validation of a
Clostridium
Endospore Viability Assay and Analysis of Greenland Ices and Atacama Desert Soils
Authors: Yang Wan-Wan , Ponce Adrian
Published: February 2011

A microscopy-based endospore viability assay (micro-EVA) capable of enumerating germinable
Clostridium
endospores (GCEs) in less than 30 min has been validated and employed to determine GCE concentrations in Greenland ices and Atacama Desert soils. Inoculation onto agarose doped with Tb
3+
and
d
-alanine triggers
Clostridium
spore germination and the concomitant release of ∼10
8
molecules of dipicolinic acid (DPA) per endospore, which, under pulsed UV excitation, enables enumeration of resultant green Tb
3+
-DPA luminescent spots as GCEs with time-gated luminescence microscopy. The intensity time courses of the luminescent spots were characteristic of stage I
Clostridium
spore germination dynamics. Micro-EVA was validated against traditional CFU cultivation from 0 to 1,000 total endospores/ml (i.e., phase-bright bodies/ml), yielding 56.4% ± 1.5% GCEs and 43.0% ± 1.0% CFU. We also show that
d
-alanine serves as a
Clostridium
-specific germinant (three species tested) that inhibits
Bacillus
germination of spores (five species tested) in that endospore concentration regime. Finally, GCE concentrations in Greenland ice cores and Atacama Desert soils were determined with micro-EVA, yielding 1 to 2 GCEs/ml of Greenland ice (versus <1 CFU/ml after 6 months of incubation) and 66 to 157 GCEs/g of Atacama Desert soil (versus 40 CFU/g soil).
Characterization of
Actinoplanes missouriensis
Spore Flagella
Authors: Uchida Kaoru , Jang Moon-Sun , Ohnishi Yasuo , Horinouchi Sueharu , Hayakawa Masayuki , Fujita Nobuyuki , Aizawa Shin-Ichi
Published: February 2011

Actinoplanes missouriensis
spores swim with a tuft of flagella. Flagella of newborn spores are wrapped with a membranous sheath. When the sheath is unwrapped, spores start swimming. Flagellar length is kept short, at around 1.9 μm, which covers half the circumference of the spore.
Antibacterial Activity and Mechanism of Action of Zinc Oxide Nanoparticles against
Campylobacter jejuni
Authors: Xie Yanping , He Yiping , Irwin Peter L. , Jin Tony , Shi Xianming
Published: February 2011

The antibacterial effect of zinc oxide (ZnO) nanoparticles on
Campylobacter jejuni
was investigated for inhibition and inactivation of cell growth. The results showed that
C. jejuni
was extremely sensitive to treatment with ZnO nanoparticles. The MIC of ZnO nanoparticles for
C. jejuni
was determined to be 0.05 to 0.025 mg/ml, which is 8- to 16-fold lower than that for
Salmonella enterica
serovar Enteritidis and
Escherichia coli
O157:H7 (0.4 mg/ml). The action of ZnO nanoparticles against
C. jejuni
was determined to be bactericidal, not bacteriostatic. Scanning electron microscopy examination revealed that the majority of the cells transformed from spiral shapes into coccoid forms after exposure to 0.5 mg/ml of ZnO nanoparticles for 16 h, which is consistent with the morphological changes of
C. jejuni
under other stress conditions. These coccoid cells were found by ethidium monoazide-quantitative PCR (EMA-qPCR) to have a certain level of membrane leakage. To address the molecular basis of ZnO nanoparticle action, a large set of genes involved in cell stress response, motility, pathogenesis, and toxin production were selected for a gene expression study. Reverse transcription-quantitative PCR (RT-qPCR) showed that in response to treatment with ZnO nanoparticles, the expression levels of two oxidative stress genes (
katA
and
ahpC
) and a general stress response gene (
dnaK
) were increased 52-, 7-, and 17-fold, respectively. These results suggest that the antibacterial mechanism of ZnO nanoparticles is most likely due to disruption of the cell membrane and oxidative stress in
Campylobacter
.
Development of an O-Antigen Serotyping Scheme for
Cronobacter sakazakii
Authors: Sun Yamin , Wang Min , Liu Hongbo , Wang Jingjing , He Xin , Zeng Jing , Guo Xi , Li Kun , Cao Boyang , Wang Lei
Published: February 2011

Cronobacter sakazakii
is an opportunistic pathogen that can cause severe infections. Serotyping provides a basis for the categorization of bacterial strains and is an important tool for epidemiological and surveillance purposes. In this study, of the 135
Cronobacter
strains tested initially, 119 were identified as
C. sakazakii
and used. A serotyping scheme for
C. sakazakii
that classifies strains based on their different O antigens was developed. Seven antisera that exhibited high agglutinin titers (>640) were produced. O2 and O6 antisera were specific for their homologous strains, O4 and O7 antisera gave heterologous titers with O1 and O6 antigens, respectively, and O1, O3, and O5 antisera cross-reacted with each other and require preabsorption with the other two antigens. All of these 119
C. sakazakii
strains were clearly assigned to these seven serotypes. O1 and O2 are the dominant serotypes, comprising 69.7% of the isolates. We also characterized the O-antigen gene clusters using restriction fragment length polymorphism (RFLP). The grouping of
C. sakazakii
strains based on their RFLP banding patterns correlated well with the grouping of strains based on our serotyping scheme. The serotype scheme presented here could prove to be a useful tool for serotyping
C. sakazakii
isolates.
Diversity of and Selection Acting on Cylindrospermopsin
cyrB
Gene Adenylation Domain Sequences in Florida
Authors: Yilmaz Mete , Phlips Edward J.
Published: February 2011

Aphanizomenon ovalisporum
is the only confirmed cylindrospermopsin producer identified in the United States to date. On the other hand,
Cylindrospermopsis raciborskii
is a prominent feature of many lakes in Florida and other regions of the United States. To see the variation in cylindrospermopsin
cyrB
gene adenylation domain sequences and possibly discover new cylindrospermopsin producers, we collected water samples for a 3-year period from 17 different systems in Florida. Positive amplicons were cloned and sequenced, revealing that approximately 92% of sequences were
A. ovalisporum
-like (>99% identity). Interestingly, 6% of sequences were very similar (>99% identity) to
cyrB
sequences of
C. raciborskii
from Australia and of
Aphanizomenon
sp. from Germany. Neutrality tests suggest that
A. ovalisporum
-like
cyrB
adenylation domain sequences are under purifying selection, with abundant low-frequency polymorphisms within the population. On the other hand, when compared between species by codon-based methods, amino acids of CyrB also seem to be under purifying selection, in accordance with the one proposed amino acid thought to be activated by the CyrB adenylation domain.
Mutacins from
Streptococcus mutans
UA159 Are Active against Multiple Streptococcal Species
Authors: Hossain Mohammad Shahnoor , Biswas Indranil
Published: February 2011

Streptococcus mutans
UA159, whose genome is completely sequenced, produces two nonlantibiotic mutacins, mutacin IV (encoded by
nlmAB
) and mutacin V (encoded by
nlmC
). In this study, we investigated the contribution of
nlmA
and
nlmB
to mutacin IV activity and demonstrated by performing genetic studies as well as by using semipurified molecules that, in contrast to a previous report, both of these genes are required for optimum mutacin IV activity. We also showed that mutacin IV is active against multiple
Streptococcus
species. In contrast, mutacin V displayed a narrower inhibitory range than mutacin IV. Our results suggest that mutacin IV and mutacin V may act synergistically to inhibit various organisms.
The Centromere Site of the Segregation Cassette of Broad-Host-Range Plasmid RA3 Is Located at the Border of the Maintenance and Conjugative Transfer Modules
Authors: Kulińska Anna , Cao Yunhong , Macioszek Małgorzata , Hayes Finbarr , Jagura-Burdzy Grażyna
Published: February 2011

RA3 is a low-copy-number, broad-host-range (BHR) conjugative plasmid of the IncU incompatibility group isolated originally fromAeromonasspp. A 4.9-kb fragment of RA3 is sufficient to stabilize an otherwise unstable replicon inEscherichia coli. This fragment specifies thekorA-incC-korB-orf11operon coding for an active partition system related to the central control operon of IncP-1 plasmids and found also in BHR environmental plasmids recently classified as the PromA group. All four genes in the cassette are necessary for segregation. IncC and KorB of RA3 belong to the ParA and ParB families of partitioning proteins, respectively. In contrast with IncP-1 plasmids, neither KorB nor IncC are involved in transcriptional autoregulation. Instead, KorA exerts transcriptional control of the operon by binding to a palindromic sequence that overlaps the putative −35 promoter motif of the cassette. The Orf11 protein is not required for regulation, but its absence decreases the stabilization potential of the segregation module. A region discontiguous from the cassette harbors a set of unrelated repeat motifs distributed over ∼300 bp. Dissection of this region identified the centromere sequence that is vital for partitioning. The ∼300-bp fragment also encompasses the origin of conjugative transfer,oriT, and the promoter that drives transcription of the conjugative transfer operon. A similar set ofcis-acting motifs are evident in the PromA group of environmental plasmids, highlighting a common evolutionary origin of segregation and conjugative transfer modules in these plasmids and members of the IncU group.
Enhancing Isoprene Production by Genetic Modification of the 1-Deoxy-
d
-Xylulose-5-Phosphate Pathway in
Bacillus subtilis
Authors: Xue Junfeng , Ahring Birgitte K.
Published: February 2011

To enhance the production of isoprene, a volatile 5-carbon hydrocarbon, in the Gram-positive spore-forming rod-shaped bacterium
Bacillus subtilis
, 1-deoxy-
d
-xylulose-5-phosphate synthase (Dxs) and 1-deoxy-
d
-xylulose-5-phosphate reductoisomerase (Dxr) were overexpressed in
B. subtilis
DSM 10. For the strain that overexpresses Dxs, the yield of isoprene was increased 40% over that by the wild-type strain. In the Dxr overexpression strain, the level of isoprene production was unchanged. Overexpression of Dxr together with Dxs showed an isoprene production level similar to that of the Dxs overproduction strain. The effects of external factors, such as stress factors including heat (48°C), salt (0.3 M NaCl), ethanol (1%), and oxidative (0.005% H
2
O
2
) stress, on isoprene production were further examined. Heat, salt, and H
2
O
2
induced isoprene production; ethanol inhibited isoprene production. In addition, induction and repression effects are independent of SigB, which is the general stress-responsive alternative sigma factor of Gram-positive bacteria.
Effect of Spatial Differences in Microbial Activity, pH, and Substrate Levels on Methanogenesis Initiation in Refuse
Authors: Staley Bryan F. , de los Reyes Francis L. , Barlaz Morton A.
Published: February 2011

The initiation of methanogenesis in refuse occurs under high volatile fatty acid (VFA) concentration and low pH (5.5 to 6.25), which generally are reported to inhibit methanogenic
Archaea
. One hypothesized mechanism for the initiation of methanogenesis in refuse decomposition is the presence of pH-neutral niches within the refuse that act as methanogenesis initiation centers. To provide experimental support for this mechanism, laboratory-scale landfill reactors were operated and destructively sampled when methanogenesis initiation was observed. The active bacterial and archaeal populations were evaluated using RNA clone libraries, RNA terminal restriction fragment length polymorphism (T-RFLP), and reverse transcription-quantitative PCR (RT-qPCR). Measurements from 81 core samples from vertical and horizontal sections of each reactor showed large spatial differences in refuse pH, moisture content, and VFA concentrations. No pH-neutral niches were observed prior to methanogenesis. RNA clone library results showed that active bacterial populations belonged mostly to
Clostridiales
, and that methanogenic
Archaea
activity at low pH was attributable to
Methanosarcina barkeri
. After methanogenesis began, pH-neutral conditions developed in high-moisture-content areas containing substantial populations of
M. barkeri
. These areas expanded with increasing methane production, forming a reaction front that advanced to low-pH areas. Despite low-pH conditions in >50% of the samples within the reactors, the leachate pH was neutral, indicating that it is not an accurate indicator of landfill microbial conditions. In the absence of pH-neutral niches, this study suggests that methanogens tolerant to low pH, such as
M. barkeri
, are required to overcome the low-pH, high-VFA conditions present during the anaerobic acid phase of refuse decomposition.
Versatile Metabolic Adaptations ofRalstonia eutrophaH16 to a Loss of PdhL, the E3 Component of the Pyruvate Dehydrogenase Complex
Authors: Raberg Matthias , Bechmann Jan , Brandt Ulrike , Schlüter Jonas , Uischner Bianca , Voigt Birgit , Hecker Michael , Steinbüchel Alexander
Published: February 2011

A previous study reported that the Tn5-induced poly(3-hydroxybutyric acid) (PHB)-leaky mutantRalstonia eutrophaH1482 showed a reduced PHB synthesis rate and significantly lower dihydrolipoamide dehydrogenase (DHLDH) activity than the wild-typeR. eutrophaH16 but similar growth behavior. Insertion of Tn5was localized in thepdhLgene encoding the DHLDH (E3 component) of the pyruvate dehydrogenase complex (PDHC). Taking advantage of the available genome sequence ofR. eutrophaH16, observations were verified and further detailed analyses and experiments were done.In silicogenome analysis revealed thatR. eutrophapossesses all five known types of 2-oxoacid multienzyme complexes and five DHLDH-coding genes. Of these DHLDHs, only PdhL harbors an amino-terminal lipoyl domain. Furthermore, insertion of Tn5inpdhLof mutant H1482 disrupted the carboxy-terminal dimerization domain, thereby causing synthesis of a truncated PdhL lacking this essential region, obviously leading to an inactive enzyme. The defined ΔpdhLdeletion mutant ofR. eutrophaexhibited the same phenotype as the Tn5mutant H1482; this excludes polar effects as the cause of the phenotype of the Tn5mutant H1482. However, insertion of Tn5or deletion ofpdhLdecreases DHLDH activity, probably negatively affecting PDHC activity, causing the mutant phenotype. Moreover, complementation experiments showed that different plasmid-encoded E3 components ofR. eutrophaH16 or of other bacteria, likeBurkholderia cepacia, were able to restore the wild-type phenotype at least partially. Interestingly, the E3 component ofB. cepaciapossesses an amino-terminal lipoyl domain, like the wild-type H16. A comparison of the proteomes of the wild-type H16 and of the mutant H1482 revealed striking differences and allowed us to reconstruct at least partially the impressive adaptations ofR. eutrophaH1482 to the loss of PdhL on the cellular level.
Altering the Composition of Caseicins A and B as a Means of Determining the Contribution of Specific Residues to Antimicrobial Activity
Authors: Norberg Sarah , O'Connor Paula M. , Stanton Catherine , Ross R. Paul , Hill Colin , Fitzgerald Gerald F. , Cotter Paul D.
Published: February 2011

Caseicin A (IKHQGLPQE) and caseicin B (VLNENLLR) are antimicrobial peptides generated through the bacterial fermentation of sodium caseinate, and on the basis of this and previous studies, they are active against many Gram-negative pathogens (
Cronobacter sakazakii
,
Cronobacter muytjensii
,
Salmonella enterica
serovar Typhimurium,
Escherichia coli
,
Klebsiella pneumoniae
, and
Pseudomonas fluorescens
) as well as the Gram-positive organism
Staphylococcus aureus
. Here we describe further studies with the aim of establishing the importance of specific (charged and nonpolar aliphatic) residues within the caseicin peptides and the effects that they have on the bacteria listed above. In order to achieve our objective, we created four derivatives of each caseicin (A1 to A4 and B1 to B4) in which specific residues were altered, and results obtained with these derivatives were compared to wild-type caseicin activity. Although conversion of cationic residues to alanine in caseicins B1 (R8A change), A1 (K2A), A2 (H3A), and A3 (K2A-H3A) generally resulted in their activity against microbial targets being reduced or unaltered,
C. sakazakii
DPC6440 was unusual in that it displayed enhanced sensitivity to three peptides (caseicins A1, A3, and B2) in which positively charged residues had been eliminated. While the replacement of leucine with alanine in selected variants (B3 and B4) resulted in reduced activity against a number of strains of
Cronobacter
and, in some cases,
S.
Typhimurium, these changes enhanced the activities of these peptides against DPC6440 and a number of
S. aureus
strains. It is thus apparent that the importance of specific residues within the caseicin peptides is dependent on the strain being targeted.
Significant Shift in Median Guinea Pig Infectious Dose Shown by an Outbreak-AssociatedListeria monocytogenesEpidemic Clone Strain and a Strain Carrying a Premature Stop Codon Mutation ininlA
Authors: Van Stelten A. , Simpson J. M. , Chen Y. , Scott V. N. , Whiting R. C. , Ross W. H. , Nightingale K. K.
Published: February 2011

Listeria monocytogenescontains (i) epidemic clone (EC) strains, which have been linked to the majority of listeriosis outbreaks worldwide and are overrepresented among sporadic cases in the United States, and (ii) strains commonly isolated from ready-to-eat foods that carry a mutation leading to a premature stop codon (PMSC) ininlA, which encodes the key virulence factor internalin A (InlA). Internalin A binds certain isoforms of the cellular receptor E-cadherin to facilitate crossing the intestinal barrier during the initial stages of anL. monocytogenesinfection. Juvenile guinea pigs, which express the human isoform of E-cadherin that binds InlA, were intragastrically challenged with a range of doses of (i) an EC strain associated with a listeriosis outbreak or (ii) a strain carrying a PMSC mutation ininlA. Recovery ofL. monocytogenesfrom tissues (i.e., liver, spleen, mesenteric lymph nodes, and ileum) was used to develop strain-specific dose-response curves on the basis of individual and combined organ data. Modeling of individual and combined organ data revealed an approximate 1.2 to 1.3 log10increase in the median infectious dose for the strain carrying a PMSC ininlArelative to that for the EC strain. Inclusion of the strain parameter significantly improved the goodness of fit for individual and combined organ models, indicating a significant shift in median infectious dose for guinea pigs challenged with aninlAPMSC strain compared to that for guinea pigs challenged with an EC strain. Results from this work provide evidence that theL. monocytogenesdose-response relationship is strain specific and will provide critical data for enhancement of current risk assessments and development of future risk assessments.
Phylogeny of Replication Initiator Protein TrfA Reveals a Highly Divergent Clade of Incompatibility Group P1 Plasmids
Authors: Stenger Drake C. , Lee Min Woo
Published: February 2011

Incompatibility group P1 (IncP-1) plasmid diversity was evaluated based on replication initiator protein (TrfA) phylogeny. A new and highly divergent clade was identified. Replication assays indicated that TrfA of recently discovered IncP-1 plasmids from
Xylella fastidiosa
and
Verminephrobacter eiseniae
initiated plasmid replication using cognate or heterologous origins of replication.
Efficient Generation of Unmarked Deletions in
Legionella pneumophila
Authors: Bryan Andrew , Harada Kaoru , Swanson Michele S.
Published: February 2011

Unmarked gene deletions facilitate studies of
Legionella pneumophila
multicomponent processes, such as motility and exonuclease activity. For this purpose,
FRT
-flanked alleles constructed in
Escherichia coli
using λ-Red recombinase were transferred to
L. pneumophila
by natural transformation. Resistance cassettes were then efficiently excised using the Flp site-specific recombinase encoded on a plasmid that is readily lost.
Shuttle Vector System for
Methanococcus maripaludis
with Improved Transformation Efficiency
Authors: Walters Alison D. , Smith Sarah E. , Chong James P. J.
Published: February 2011

We have identified an open reading frame and DNA element that are sufficient to maintain shuttle vectors in
Methanococcus maripaludis
. Strain S0001, containing ORF1 from pURB500 integrated into the
M. maripaludis
genome, supports a significantly smaller shuttle vector, pAW42, and a 7,000-fold increase in transformation efficiency for pURB500-based vectors.