bluuebunny/crossref_metadata_2025 huggingface dataset:
Impact of digital dermoscopy analysis on the decision to follow up or to excise a pigmented skin lesion: a multicentre study
Authors: Burroni Marco , Wollina Uwe , Torricelli Rocco , Gilardi Stefano , Dell'Eva Giordana , Helm Cathrine , Bardey Wolfgang , Nami Niccolò , Nobile Franco , Ceccarini Massimo , Pomponi Adriano , Alessandro Biondi , Rubegni Pietro
Published: March 2011

Background: The quality of early malignant melanoma (MM) diagnosis is dependent on the experience of dermatologists, tools like dermoscopy and histopathology, and awareness and education of the studied population. Does a higher rate of excision of pigmented skin lesions (PSL) increase the rate of detected melanomas?

Material and Methods: The DB‐MIPS objective tool, able to evaluate mathematical defined variables, has been used to verify the variability of measurements among PSL stored by five different centres located in Italy, Switzerland, and Germany.

Results: The objective analysis showed low differences in terms of moles' features among the different groups, arguing for robustness of the dermatological patient's PSL inspection. Differences in terms of false positives and predictive positive values have been detected. The tendency to follow up a lesion was proportional to the percentage of thin MM (<0.75 mm tumour thickness), while the interventism was proportional to the percentage of dysplastic moles. Similar percentage of thin melanoma has been observed in all the centres, indicating a standardization in early diagnosing among experienced dermatologists. The main difference among the centres was their mode of action, i.e. to follow up or remove suspicious PSL.

Conclusion: Interventism depends neither on the geographic site nor on the features of the observed moles. Higher removal rates do not correspond to higher MM detections: this means that an in‐depth knowledge of melanoma patterns is required and follow‐up of suspicious moles is highly suggested.
VIS2FIX: A High‐Speed Fixation Method for Immuno‐Electron Microscopy
Authors: Karreman Matthia A. , van Donselaar Elly G. , Gerritsen Hans C. , Verrips C. Theo , Verkleij Arie J.
Published: March 2011

Immuno‐transmission electron microscopy (TEM) is the technique of choice for high‐resolution localization of proteins in fixed specimen. Here we introduce 2 novel methods for the fixation of sections from cryo‐immobilized samples that result in excellent ultrastructural preservation. These high‐speed fixation techniques, both called VIS2FIX, allow for a reduction in sample preparation time from at least 1 week to only 8 h. The methods were validated in immuno‐TEM experiments on THP‐1 monocytes, human umbilical vein endothelial cells (HUVECs) and Madin–Darby canine kidney (MDCK‐II) cells. The fixation and retention of neutral lipids is demonstrated, offering unique prospects for the application of immuno‐TEM in the lipidomics field. Furthermore, the VIS2FIX methods were successfully employed in correlative fluorescence and electron microscopy.
Pseudo‐Bartter’s syndrome revealing cystic fibrosis in an infant caused by 3849 + 1G>A and 4382delA compound heterozygosity
Authors: Nahida El‐Rifai , Mohammed Hourani , Guy Lalau
Published: March 2011

Pseudo‐Bartter’s (PB) syndrome characterized by hypokalemic metabolic alkalosis and persistent failure to thrive constitutes a rare typical presentation of cystic fibrosis (CF) with prevalence of 16.8%. We present a case of CF presenting with failure to thrive, dehydration, PB syndrome associated with chest infection and primo‐colonization with Pseudomonas aeruginosa. Sweat chloride test was 102 mmol/L. DNA analysis identified 2 mutations 3849 + 1G>A (intron 19) and 4382delA (exon 24) present in heterozygous status. To the best of our knowledge, our case is the first reported case in the literature of CF manifested by PB syndrome associated with chest infection and primo‐colonization with Pseudomonas aeruginosa.

Conclusion:  The genotype 3849 + 1G>A/4382delA found in our patient is described for the first time in the literature. It explains the lung involvement with the dehydration and electrolyte disturbances. The role of the mutation in exon 24 in cases of CF with PB syndrome remains to be determined.
Anthropometric measurements and obesity diagnosis in schoolchildren
Authors: Cândido Ana PC , Freitas Sílvia N , Machado‐Coelho George LL
Published: March 2011

Aim:  To evaluate whether anthropometric methods such as arm muscle area, arm fat area, Rohrer index, conicity index, and waist‐to‐height ratio and the commonly used body mass index method are suitable for obesity screening in a multiethnic paediatric population.

Method:  A cross‐sectional population study was carried out in the city of Ouro Preto, Brazil, in 2006. Schoolchildren (n = 788) aged 6–15 years were selected by the stratified random sampling procedure by the proportion of students according to age and gender in all schools. Nonparametric receiver‐operating characteristic analysis was used to define the cut‐off for obesity for each method studied, using the tetrapolar bioelectrical impedance technique as the reference method.

Results:  Arm fat area measurements and Rohrer index were most suitable for adiposity screening in prepubertal and pubertal boys and in pubertal and prepubertal girls, respectively. In postpubertal period, the Rohrer index was the most suitable body fat assessment method in both genders.

Conclusion:  Body mass index presented the best balance between sensitivity and specificity only in the prepubertal period and postpubertal boys. Anthropometric measurements, especially arm fat area measurements and Rohrer index, are useful for screening of adiposity in a multiethnic paediatric population.
The complexity of amphibian population declines: understanding the role of cofactors in driving amphibian losses
Authors: Blaustein Andrew R. , Han Barbara A. , Relyea Rick A. , Johnson Pieter T.J. , Buck Julia C. , Gervasi Stephanie S. , Kats Lee B.
Published: March 2011

Population losses and extinctions of species are occurring at unprecedented rates, as exemplified by declines and extinctions of amphibians worldwide. However, studies of amphibian population declines generally do not address the complexity of the phenomenon or its implications for ecological communities, focusing instead on single factors affecting particular amphibian species. We argue that the causes for amphibian population declines are complex; may differ among species, populations, and life stages within a population; and are context dependent with multiple stressors interacting to drive declines. Because amphibians are key components of communities, we emphasize the importance of investigating amphibian declines at the community level. Selection pressures over evolutionary time have molded amphibian life history characteristics, such that they may remain static even in the face of strong, recent human‐induced selection pressures.
Anthropogenic impacts on marine ecosystems in Antarctica
Authors: Aronson Richard B. , Thatje Sven , McClintock James B. , Hughes Kevin A.
Published: March 2011

Antarctica is the most isolated continent on Earth, but it has not escaped the negative impacts of human activity. The unique marine ecosystems of Antarctica and their endemic faunas are affected on local and regional scales by overharvesting, pollution, and the introduction of alien species. Global climate change is also having deleterious impacts: rising sea temperatures and ocean acidification already threaten benthic and pelagic food webs. The Antarctic Treaty System can address local‐ to regional‐scale impacts, but it does not have purview over the global problems that impinge on Antarctica, such as emissions of greenhouse gases. Failure to address human impacts simultaneously at all scales will lead to the degradation of Antarctic marine ecosystems and the homogenization of their composition, structure, and processes with marine ecosystems elsewhere.
UPTAKE KINETICS AND INTRACELLULAR LOCALIZATION OF HYPOCRELLIN PHOTOSENSITIZERS FOR PHOTODYNAMIC THERAPY: A CONFOCAL MICROSCOPY STUDY
Authors: Miller Gerald G. , Brown Kevin , Moore Ronald B. , McPhee Malcolm S. , Diwu Zhenjun J. , Liu Jixiang , Huang Liren , Lown J. William , Begg David A. , Chlumecky Vera , Tulip John
Published: March 2011

Hypocrellins are naturally occurring compounds with photosensitizing properties in biological systems. We have prepared synthetic derivatives of hypocrellin B, which have promise as photosensitizers in the clinical application of photodynamic therapy. The intracellular localization and uptake kinetics of hypocrellin B and several selected hypocrellin congeners were determined semiquantitatively by fluorescence confocal microscopy in monolayer cultures of EMT6/Ed murine tumor cells. Each compound had unique uptake kinetics. Although no compound tested to date has demonstrated nuclear labeling, most could be detected in lysosomes, Golgi, endoplasmic reticulum and, to a minor extent, in cellular membranes. No two compounds gave identical labeling distributions. The differences are assumed to originate in physicochemical properties characteristic of each compound, which may ultimately impact upon the primary modality of phototoxicity.
INTERACTIONS BETWEEN N‐ASPARTYL CHLORIN E6, DETERGENT MICELLES AND PLASMA LIPOPROTEINS
Authors: Kessel David
Published: March 2011

Addition of plasma to the photosensitizing agent N‐aspartyl chlorin e6 (NPe6) in Triton X‐100 micelles results in a blue‐shift of fluorescence emission, corresponding to a change in the average environmental dielectric constant from 8 to 32. This effect was not observed with anionic or cationic detergents or with several other sensitizers including a close analog, chlorin e6. The apolipoprotein fraction of high‐density lipoproteins (HDL) was found responsible for this blue‐shift. These results indicate a highly selective interaction between HDL apolipoproteins and NPe6 molecules. Addition of the HDL apolipoprotein to NPe6 incorporated into Triton micelles decreased the fraction of NPe6 molecules accessible to Cd2+ ‐induced fluorescence quenching but resulted in an increase in the Stern‐Volmer quenching constant (an index of quenching efficiency).
The Second Order Statistics of High Waves in Wind Sea and Swell
Authors: Tromans Peter , Vanderschuren Luc , Ewans Kevin
Published: March 2011

The statistics of extreme wave crest elevation and wave height have been calculated for realistic, directionally spread sea and swell using a probabilistic method tested and described previously. The nonlinearity of steep waves is modeled to the second order using Sharma and Dean kinematics, and a response surface (reliability type) method is used to deduce the crest elevation or wave height corresponding to a given probability of exceedance. The effects of various combinations of sea and swell are evaluated. As expected, in all cases, nonlinearity makes extreme crests higher than the corresponding linear ones. The nonlinear effects on the wave height are relatively small.
Ultimate Strength
Analysis of Ship Hull Girder Under Random Material and Geometric
Properties
Authors: Vhanmane Suhas , Bhattacharya Baidurya
Published: March 2011

The ultimate strength of a ship’s hull depends on its material and geometric properties, some or all of which may be random in nature. In addition, initial imperfections in the form of initial deflection and residual welding stresses in plating between stiffeners can significantly affect the hull ultimate strength. In this paper, the effect of randomness in yield strength and in the initial imperfections on ultimate hull girder strength is determined. Different levels of statistical dependence between yield strength and initial imperfection of stiffeners and plating between stiffeners have been considered. The methodology is applied on a bulk carrier and a VLCC tanker.
Prevention and Intensification of Melt-Water Explosive Interactions
Authors: Zielinski Stephen M. , Sansone Anthony A. , Ziolkowski Matthew , Taleyarkhan Rusi P.
Published: March 2011

The combination of a hot fluid (e.g., molten metals) and a cold vaporizing fluid (e.g., water) can undergo spontaneous or externally assisted explosive interactions. Such explosions are a well-established contributor to the risk for nuclear reactors exemplified by the infamous Chernobyl accident. Once fundamentals are understood, it may be possible to not only prevent but also, more importantly, control the intensity for useful applications in the areas covering variable thrust propulsion with tailored pressure profiles, for enhancing rapid heat transfer, and also for powder metallurgy (i.e., supercooled powder production, wherein materials turn superplastic with enhanced ductility). This paper discusses results of experiments conducted with various molten metals, specifically, tin, gallium, galinstan, and aluminum interacting with water (with and without salt), and with and without noncondensable gases such as hydrogen or air. It is found that under the appropriate conditions, spontaneous and energetic phase changes can be initiated within milliseconds if the hot metal is tin or galinstan, including the timed feedback of shocks leading to chain-type reactions. Using 3–10 g of tin or galinstan, shock pressures up to 25 bars (350 psig) and mechanical power over ∼2-4 kW were monitored about 4 cm from the explosion zone. The interaction could be intensified more than ten folds by dropping the melt through an argon atmosphere. A slow metal quenching interaction occurring over tens of seconds could be turned explosive to transpire within milliseconds if the thermal states are within the so-called thermal interaction zone. Such explosive interactions did not transpire with gallium or aluminum due to tough oxide coatings. However, by adding ∼10 w/o of salt in water, molten Gallium readily exploded. It was also conclusively revealed that, for an otherwise spontaneously explosive interaction of tin-water or galinstan-water, the inclusion of trace (0.3 w/o) quantities of aluminum has a radical influence on stabilizing the system and ensuring conclusive prevention of explosion triggering. This paper compares and presents the results obtained in this study and draws analogies with industrial scale aluminum casthouse safety involving thousands of kilograms of melt. Insights are provided for enabling physics-based prevention, or, alternately, the intentional initiation of explosions.
When the Heart Forgets to Beat
Authors: Damani Samir
Published: March 2011

Whole-genome sequencing combined with a genome-wide association study yields important genetic predictors of sinus node dysfunction.
PEG Gets to the Heart of the Matter
Authors: Skelly Christopher L.
Published: March 2011

Polyethylene glycol improves cardiac function by inhibiting the apoptotic pathway induced by ischemia-reperfusion injury.
Surface Localization Determinants of Borrelia OspC/Vsp Family Lipoproteins
Authors: Kumru Ozan S. , Schulze Ryan J. , Rodnin Mykola V. , Ladokhin Alexey S. , Zückert Wolfram R.
Published: March 2011

The dimeric OspC/Vsp family surface lipoproteins of
Borrelia
spirochetes are crucial to the transmission and persistence of Lyme borreliosis and tick-borne relapsing fever. However, the requirements for their proper surface display remained undefined. In previous studies, we showed that localization of
Borrelia burgdorferi
monomeric surface lipoprotein OspA was dependent on residues in the N-terminal “tether” peptide. Here, site-directed mutagenesis of the
B. burgdorferi
OspC tether revealed two distinct regions affecting either release from the inner membrane or translocation through the outer membrane. Determinants of both of these steps appear consolidated within a single region of the
Borrelia turicatae
Vsp1 tether. Periplasmic OspC mutants still were able to form dimers. Their localization defect could be rescued by the addition of an apparently structure-destabilizing C-terminal epitope tag but not by coexpression with wild-type OspC. Furthermore, disruption of intermolecular Vsp1 salt bridges blocked dimerization but not surface localization of the resulting Vsp1 monomers. Together, these results suggest that
Borrelia
OspC/Vsp1 surface lipoproteins traverse the periplasm and the outer membrane as unfolded monomeric intermediates and assemble into their functional multimeric folds only upon reaching the spirochetal surface.
Evidence that WapB Is a 1,2-Glucosyltransferase of Pseudomonas aeruginosa Involved in Lipopolysaccharide Outer Core Biosynthesis
Authors: Kocíncová Dana , Hao Youai , Vinogradov Evgeny , Lam Joseph S.
Published: March 2011

Pseudomonas aeruginosa
is an important opportunistic pathogen infecting debilitated individuals. One of the major virulence factors expressed by
P. aeruginosa
is lipopolysaccharide (LPS), which is composed of lipid A, core oligosaccharide (OS), and O-antigen polysaccharide. The core OS is divided into inner and outer regions. Although the structure of the outer core OS has been elucidated, the functions and mechanisms of the glycosyltransferases involved in core OS biogenesis are currently unknown. Here, we show that a previously uncharacterized gene,
pa1014
, is involved in outer core biosynthesis, and we propose to rename this gene
wapB
. We constructed a chromosomal mutant,
wapB
::Gm, in a PAO1 (O5 serotype) strain background. Characterization of the LPS from the mutant by Western immunoblotting showed a lack of reactivity to PAO1 outer core-specific monoclonal antibody (MAb) 5c-101. The chemical structure of the core OS of the
wapB
mutant was elucidated using nuclear magnetic resonance spectroscopy and mass spectrometry techniques and revealed that the core OS of the
wapB
mutant lacked the terminal β-1,2-linked-
d
-glucose residue. Complementation of the mutant with
wapB in trans
restored the core structure to one that is identical to that of the wild type. Eleven of the 20
P. aeruginosa
International Antigenic Typing Scheme (IATS) serotypes produce LPSs that lack the terminal
d
-glucose residue (Glc
IV
). Interestingly, expressing
wapB
in each of these 11 serotypes modifies each of their outer core OS structures, which became reactive to MAb 5c-101 in Western immunoblotting, suggesting the presence of a terminal
d
-glucose in these core OS structures. Our results strongly suggested that
wapB
encodes a 1,2-glucosyltransferase.
The Phytopathogen Pseudomonas syringae pv. tomato DC3000 Has Three High-Affinity Iron-Scavenging Systems Functional under Iron Limitation Conditions but Dispensable for Pathogenesis
Authors: Jones Alexander M. , Wildermuth Mary C.
Published: March 2011

High-affinity iron scavenging through the use of siderophores is a well-established virulence determinant in mammalian pathogenesis. However, few examples have been reported for plant pathogens. Here, we use a genetic approach to investigate the role of siderophores inPseudomonas syringaepv. tomato DC3000 (DC3000) virulence in tomato. DC3000, an agronomically important pathogen, has two known siderophores for high-affinity iron scavenging, yersiniabactin and pyoverdin, and we uncover a third siderophore, citrate, required for growth when iron is limiting. Though growth of a DC3000 triple mutant unable to either synthesize or import these siderophores is severely restricted in iron-limited culture, it is fully pathogenic. One explanation for this phenotype is that the DC3000 triple mutant is able to directly pirate plant iron compounds such as heme/hemin or iron-nicotianamine, and our data indicate that DC3000 can import iron-nicotianamine with high affinity. However, an alternative explanation, supported by data from others, is that the pathogenic environment of DC3000 (i.e., leaf apoplast) is not iron limited but is iron replete, with available iron of >1 μM. Growth of the triple mutant in culture is restored to wild-type levels by supplementation with a variety of iron chelates at >1 μM, including iron(III) dicitrate, a dominant chelate of the leaf apoplast. This suggests that lower-affinity iron import would be sufficient for DC3000 iron nutritionin plantaand is in sharp contrast to the high-affinity iron-scavenging mechanisms required in mammalian pathogenesis.
RecA-Dependent Replication in the
nrdA101
(Ts) Mutant of Escherichia coli under Restrictive Conditions
Authors: Salguero Israel , Guarino Estrella , Guzmán Elena C.
Published: March 2011

Cells carrying the thermosensitive
nrdA101
allele are able to replicate entire chromosomes at 42°C when new DNA initiation events are inhibited. We investigated the role of the recombination enzymes on the progression of the DNA replication forks in the
nrdA101
mutant at 42°C in the presence of rifampin. Using pulsed-field gel electrophoresis (PFGE), we demonstrated that the replication forks stalled and reversed during the replication progression under this restrictive condition. DNA labeling and flow cytometry experiments supported this finding as the deleterious effects found in the RecB-deficient background were suppressed specifically by the absence of RuvABC; however, this did not occur in a RecG-deficient background. Furthermore, we show that the RecA protein is absolutely required for DNA replication in the
nrdA101
mutant at restrictive temperature when the replication forks are reversed. The detrimental effect of the
recA
deletion is not related to the chromosomal degradation caused by the absence of RecA. The inhibition of DNA replication observed in the
nrdA101 recA
mutant at 42°C in the presence of rifampin was reverted by the presence of the wild-type RecA protein expressed ectopically but only partially suppressed by the RecA protein with an S25P mutation [RecA(S25P)], deficient in the rescue of the stalled replication forks. We propose that RecA is required to maintain the integrity of the reversed forks in the
nrdA101
mutant under certain restrictive conditions, supporting the relationship between DNA replication and recombination enzymes through the stabilization and repair of the stalled replication forks.
Role of the Nfo and ExoA Apurinic/Apyrimidinic Endonucleases in Radiation Resistance and Radiation-Induced Mutagenesis of Bacillus subtilis Spores
Authors: Moeller Ralf , Setlow Peter , Pedraza-Reyes Mario , Okayasu Ryuichi , Reitz Günther , Nicholson Wayne L.
Published: March 2011

The roles of DNA repair by apurinic/apyrimidinic (AP) endonucleases alone, and together with DNA protection by α/β-type small acid-soluble spore proteins (SASP), in
Bacillus subtilis
spore resistance to different types of radiation have been studied. Spores lacking both AP endonucleases (Nfo and ExoA) and major SASP were significantly more sensitive to 254-nm UV-C, environmental UV (>280 nm), X-ray exposure, and high-energy charged (HZE)-particle bombardment and had elevated mutation frequencies compared to those of wild-type spores and spores lacking only one or both AP endonucleases or major SASP. These findings further implicate AP endonucleases and α/β-type SASP in repair and protection, respectively, of spore DNA against effects of UV and ionizing radiation.
The Moraxella catarrhalis Nitric Oxide Reductase Is Essential for Nitric Oxide Detoxification
Authors: Wang Wei , Kinkel Traci , Martens-Habbena Willm , Stahl David A. , Fang Ferric C. , Hansen Eric J.
Published: March 2011

Moraxella catarrhalis
is a Gram-negative obligate aerobe that is an important cause of human respiratory tract infections. The
M. catarrhalis
genome encodes a predicted truncated denitrification pathway that reduces nitrate to nitrous oxide. We have previously shown that expression of both the
M. catarrhalis aniA
(encoding a nitrite reductase) and
norB
(encoding a putative nitric oxide reductase) genes is repressed by the transcriptional regulator NsrR under aerobic conditions and that
M. catarrhalis
O35E
nsrR
mutants are unable to grow in the presence of low concentrations of nitrite (W. Wang, et al., J. Bacteriol. 190:7762–7772, 2008). In this study, we constructed an
M. catarrhalis norB
mutant and showed that planktonic growth of this mutant is inhibited by low levels of nitrite, whether or not an
nsrR
mutation is present. To determine the importance of NorB in this truncated denitrification pathway, we analyzed the metabolism of nitrogen oxides by
norB
,
aniA norB
, and
nsrR norB
mutants. We found that
norB
mutants are unable to reduce nitric oxide and produce little or no nitrous oxide from nitrite. Furthermore, nitric oxide produced from nitrite by the AniA protein is bactericidal for a
Moraxella catarrhalis
O35E
norB
mutant but not for wild-type O35E bacteria under aerobic growth conditions
in vitro
, suggesting that nitric oxide catabolism in
M. catarrhalis
is accomplished primarily by the
norB
gene product. Measurement of bacterial protein
S
-nitrosylation directly implicates nitrosative stress resulting from AniA-dependent nitric oxide formation as a cause of the growth inhibition of
norB
and
nsrR
mutants by nitrite.
An Antirepressor, SrpR, Is Involved in Transcriptional Regulation of the SrpABC Solvent Tolerance Efflux Pump of Pseudomonas putida S12
Authors: Sun Xu , Zahir Zileena , Lynch Karlene H. , Dennis Jonathan J.
Published: March 2011

Organic compounds exhibit various levels of toxicity toward living organisms based upon their ability to insert into biological membranes and disrupt normal membrane function. The primary mechanism responsible for organic solvent tolerance in many bacteria is energy-dependent extrusion via efflux pumps. One such bacterial strain,
Pseudomonas putida
S12, is known for its high tolerance to organic solvents as provided through the SrpABC resistance-nodulation-cell division (RND) family efflux pump. To determine how two putative regulatory proteins (SrpR and SrpS, encoded directly upstream of the SrpABC structural genes) influence SrpABC efflux pump expression, we conducted transcriptional analysis, β-galactosidase fusion experiments, electrophoretic mobility shift assays, and pulldown analysis. Together, the results of these experiments suggest that expression of the
srpABC
operon can be derepressed by two distinct but complementary mechanisms: direct inhibition of the SrpS repressor by organic solvents and binding of SrpS by its antirepressor SrpR.
A Novel Gene Inactivation System Reveals Altered Periplasmic Flagellar Orientation in aBorrelia burgdorferi fliLMutant
Authors: Motaleb M. A. , Pitzer Joshua E. , Sultan Syed Z. , Liu Jun
Published: March 2011

Motility and chemotaxis are essential components of pathogenesis for many infectious bacteria, includingBorrelia burgdorferi, the causative agent of Lyme disease. Motility and chemotaxis genes comprise 5 to 6% of the genome ofB. burgdorferi, yet the functions of most of those genes remain uncharacterized, mainly due to the paucity of a nonpolar gene inactivation system. In this communication, we describe the development of a novel gene inactivation methodology to targetB. burgdorferi fliL, a putative periplasmic flagellar gene located in a large motility operon and transcribed by RNA polymerase containing σ70. Although the morphology of nonpolarfliLmutant cells was indistinguishable from that of wild-type cells, the mutant exhibited a defective-motility phenotype. Cryo-electron tomography (cryo-ET) of intact organisms revealed that the periplasmic flagella in thefliLmutant were frequently tilted toward the cell pole instead of their normal orientation toward the cell body. These defects were corrected when the mutant was complemented incis. Moreover, a comparative analysis of flagellar motors from the wild type and the mutant provides the first structural evidence that FliL is localized between the stator and rotor. Our results suggest that FliL is likely involved in coordinating or regulating the orientation of periplasmic flagella inB. burgdorferi.
Combinatorial Regulation by MrpC2 and FruA Involves Three Sites in the
fmgE
Promoter Region during Myxococcus xanthus Development
Authors: Son Bongjun , Liu Yu , Kroos Lee
Published: March 2011

Starvation causes cells in a dense population of
Myxococcus xanthus
to change their gliding movements and construct mounds. Short-range C-signaling between rod-shaped cells within mounds induces gene expression that promotes differentiation into spherical spores. Several C-signal-dependent genes have been shown to be regulated by cooperative binding of two transcription factors to the promoter region. These
F
ruA- and
M
rpC2-regulated
g
enes (designated
fmg
) each exhibit a different arrangement of binding sites. Here, we describe
fmgE
, which appears to be regulated by three sites for cooperative binding of FruA and MrpC2. Chromatin immunoprecipitation analysis showed that association of MrpC2 and/or its longer form, MrpC with the
fmgE
promoter region, depends on FruA, consistent with cooperative binding of the two proteins
in vivo
. Electrophoretic mobility shift assays with purified His
10
-MrpC2 and FruA-His
6
indicated cooperative binding
in vitro
to three sites in the
fmgE
promoter region. The effects of mutations on binding
in vitro
and on expression of
fmgE-lacZ
fusions correlated site 1 (at about position −100 relative to the transcriptional start site) with negative regulation and site 2 (just upstream of the promoter) and site 3 (at about position +100) with positive regulation. Site 3 was bound by His
10
-MrpC2 alone, or the combination of His
10
-MrpC2 and FruA-His
6
, with the highest affinity, followed by site 1 and then site 2, supporting a model in which site 3 recruits MrpC2 and FruA to the
fmgE
promoter region, site 1 competes with site 2 for transcription factor binding, and site 2 occupancy is required to activate the promoter but only occurs when C-signaling produces a high concentration of active FruA.
Transcription Antitermination by a Phosphorylated Response Regulator and Cobalamin-Dependent Termination at a B
12
Riboswitch Contribute to Ethanolamine Utilization in Enterococcus faecalis
Authors: Baker Kris Ann , Perego Marta
Published: March 2011

The ability of bacteria to utilize ethanolamine (EA) as a carbon and nitrogen source may confer an advantage for survival, colonization, and pathogenicity in the human intestinal tract.
Enterococcus faecalis
, a Gram-positive human commensal organism, depends on a two-component signaling system (TCS-17) for sensing EA and regulating the expression of the ethanolamine utilization genes. Multiple promoters participate in
eut
gene expression in the presence of EA as the sole carbon source and cobalamin (CoB12), an essential cofactor in the enzymatic degradation process. By means of
in vivo
and
in vitro
approaches, this study characterized the transcriptional activity identified in the
eutT-eutG
intergenic region of the
E. faecalis eut
cluster. Two novel promoters in this region were shown to be active
in vivo
. The distal P2-1 promoter was associated with a B12 riboswitch that terminated transcription in the presence of CoB12. Transcription elongation from the proximal P2-2 promoter was regulated by antitermination mediated by the phosphorylated form of the response regulator of TCS-17 (RR17). 3′-Rapid amplification of cDNA ends (RACE) analyses of the terminated RNA products allowed precise identification of the hairpin loop structures involved in termination/antitermination. The results uncovered the role of the B12 riboswitch and RR17 in
eut
gene expression, adding to the complexity of this regulatory pathway and extending the knowledge of possible means of transcription regulation in Gram-positive organisms.
Full-Length
De Novo
Sequence of the Chlamydophila psittaci Type Strain, 6BC
Authors: Voigt Anja , Schöfl Gerhard , Heidrich Alexander , Sachse Konrad , Saluz Hans Peter
Published: March 2011

Chlamydophila psittaci
is an obligate intracellular zoonotic pathogen, mainly of birds. It is the causative agent of psittacosis in birds and humans. Here we report the full-length
de novo
genome sequence of the avian isolate 6BC, the type strain of the species
C. psittaci
.
Genome Sequence of Acinetobacter calcoaceticus PHEA-2, Isolated from Industry Wastewater
Authors: Zhan Yuhua , Yan Yongliang , Zhang Wei , Yu Haiying , Chen Ming , Lu Wei , Ping Shuzhen , Peng Zixin , Yuan Menglong , Zhou Zhengfu , Elmerich Claudine , Lin Min
Published: March 2011

Genome analysis of
Acinetobacter calcoaceticus
PHEA-2 was undertaken because of the importance of this bacterium for bioremediation of phenol-polluted water and because of the close phylogenetic relationship of this species with the human pathogen
Acinetobacter baumannii
. To our knowledge, this is the first strain of
A. calcoaceticus
whose genome has been sequenced.
Genome Sequence of the Methanotrophic Alphaproteobacterium Methylocystis sp. Strain Rockwell (ATCC 49242)
Authors: Stein Lisa Y. , Bringel Françoise , DiSpirito Alan A. , Han Sukkyun , Jetten Mike S. M. , Kalyuzhnaya Marina G. , Kits K. Dimitri , Klotz Martin G. , Op den Camp Huub J. M. , Semrau Jeremy D. , Vuilleumier Stéphane , Bruce David C. , Cheng Jan-Fang , Davenport Karen W. , Goodwin Lynne , Han Shunsheng , Hauser Loren , Lajus Aurélie , Land Miriam L. , Lapidus Alla , Lucas Susan , Médigue Claudine , Pitluck Sam , Woyke Tanja
Published: March 2011

Methylocystis
sp. strain Rockwell (ATCC 49242) is an aerobic methane-oxidizing alphaproteobacterium isolated from an aquifer in southern California. Unlike most methanotrophs in the
Methylocystaceae
family, this strain has a single
pmo
operon encoding particulate methane monooxygenase but no evidence of the genes encoding soluble methane monooxygenase. This is the first reported genome sequence of a member of the
Methylocystis
species of the
Methylocystaceae
family in the order
Rhizobiales
.
Complete Genome Sequence of the Pathogenic Bacterium Riemerella anatipestifer Strain RA-GD
Authors: Yuan Jianfeng , Liu Wanfei , Sun Mingfei , Song Shuhui , Cai Jianping , Hu Songnian
Published: March 2011

Riemerella anatipestifer
is a well-described pathogen of waterfowl and other avian species which can cause a great loss to the poultry industry. Here we obtained the complete genome sequence of
R. anatipestifer
strain RA-GD, which was isolated from an infected duck in Guangzhou, China, and was cultivated in our laboratory.
Prophage-Stimulated Toxin Production in Clostridium difficile NAP1/027 Lysogens
Authors: Sekulovic Ognjen , Meessen-Pinard Mathieu , Fortier Louis-Charles
Published: March 2011

TcdA and TcdB exotoxins are the main virulence factors of
Clostridium difficile
, one of the most deadly nosocomial pathogens. Recent data suggest that prophages can influence the regulation of toxin expression. Here we present the characterization of φCD38-2, a
pac
-type temperate
Siphoviridae
phage that stimulates toxin expression when introduced as a prophage into
C. difficile
. Host range analysis showed that φCD38-2 was able to infect 99/207 isolates of
C. difficile
representing 11 different PCR ribotypes. Of 89 isolates corresponding to the NAP1/027 hypervirulent strain, which recently caused several outbreaks in North America and Europe, 79 (89%) were sensitive to φCD38-2. The complete double-stranded DNA (dsDNA) genome was determined, and a putative function could be assigned to 24 of the 55 open reading frames. No toxins or virulence factors could be identified based on bioinformatics analyses. Our data also suggest that φCD38-2 replicates as a circular plasmid in
C. difficile
lysogens. Upon introduction of φCD38-2 into a NAP1/027 representative isolate, up to 1.6- and 2.1-fold more TcdA and TcdB, respectively, were detected by immunodot blotting in culture supernatants of the lysogen than in the wild-type strain. In addition, real-time quantitative reverse transcriptase PCR (qRT-PCR) analyses showed that the mRNA levels of all five pathogenicity locus (PaLoc) genes were higher in the CD274 lysogen. Our study provides the first genomic sequence of a new
pac
-type
Siphoviridae
phage family member infecting
C. difficile
and brings further evidence supporting the role of prophages in toxin production in this important nosocomial pathogen.
Identification of ArgP and Lrp as Transcriptional Regulators of lysP, the Gene Encoding the Specific Lysine Permease of Escherichia coli
Authors: Ruiz Jimena , Haneburger Ina , Jung Kirsten
Published: March 2011

Expression of
lysP
, which encodes the lysine-specific transporter LysP in
Escherichia coli
, is regulated by the concentration of exogenous available lysine. In this study, the LysR-type transcriptional regulator ArgP was identified as the activator of
lysP
expression. At lysine concentrations higher than 25 μM,
lysP
expression was shut off and phenocopied an
argP
deletion mutant. Purified ArgP-His
6
bound to the
lysP
promoter/control region at a sequence containing a conserved T-N
11
-A motif. Its affinity increased in the presence of lysine but not in the presence of the other known coeffector, arginine.
In vivo
data suggest that lysine-loaded ArgP and arginine-loaded ArgP compete at the
lysP
promoter. We propose that lysine-loaded ArgP prevents
lysP
transcription at the promoter clearance step, as described for the lysine-dependent regulation of
argO
(R. S. Laishram and J. Gowrishankar, Genes Dev. 21:1258-1272, 2007). The global regulator Lrp also bound to the
lysP
promoter/control region. An
lrp
mutant exhibited reduced
lysP
expression in the absence of external lysine. These results indicate that ArgP is a major regulator of
lysP
expression but that Lrp modulates
lysP
transcription under lysine-limiting conditions.
Flagellin Redundancy in Caulobacter crescentus and Its Implications for Flagellar Filament Assembly
Authors: Faulds-Pain Alexandra , Birchall Christopher , Aldridge Christine , Smith Wendy D. , Grimaldi Giulia , Nakamura Shuichi , Miyata Tomoko , Gray Joe , Li Guanglai , Tang Jay X. , Namba Keiichi , Minamino Tohru , Aldridge Phillip D.
Published: March 2011

Bacterial flagella play key roles in surface attachment and host-bacterial interactions as well as driving motility. Here, we have investigated the ability of
Caulobacter crescentus
to assemble its flagellar filament from six flagellins: FljJ, FljK, FljL, FljM, FljN, and FljO. Flagellin gene deletion combinations exhibited a range of phenotypes from no motility or impaired motility to full motility. Characterization of the mutant collection showed the following: (i) that there is no strict requirement for any one of the six flagellins to assemble a filament; (ii) that there is a correlation between slower swimming speeds and shorter filament lengths in Δ
fljK
Δ
fljM
mutants; (iii) that the flagellins FljM to FljO are less stable than FljJ to FljL; and (iv) that the flagellins FljK, FljL, FljM, FljN, and FljO alone are able to assemble a filament.
Unexpected Diversity of Chlorite Dismutases: a Catalytically Efficient Dimeric Enzyme from Nitrobacter winogradskyi
Authors: Mlynek Georg , Sjöblom Björn , Kostan Julius , Füreder Stephanie , Maixner Frank , Gysel Kira , Furtmüller Paul Georg , Obinger Christian , Wagner Michael , Daims Holger , Djinović-Carugo Kristina
Published: March 2011

Chlorite dismutase (Cld) is a unique heme enzyme catalyzing the conversion of ClO
2
−
to Cl
−
and O
2
. Cld is usually found in perchlorate- or chlorate-reducing bacteria but was also recently identified in a nitrite-oxidizing bacterium of the genus
Nitrospira
. Here we characterized a novel Cld-like protein from the chemolithoautotrophic nitrite oxidizer
Nitrobacter winogradskyi
which is significantly smaller than all previously known chlorite dismutases. Its three-dimensional (3D) crystal structure revealed a dimer of two identical subunits, which sharply contrasts with the penta- or hexameric structures of other chlorite dismutases. Despite a truncated N-terminal domain in each subunit, this novel enzyme turned out to be a highly efficient chlorite dismutase (

K
m

= 90 μM;
k
cat
= 190 s
−1
;
k
cat
/

K
m

= 2.1 × 10
6
M
−1
s
−1
), demonstrating a greater structural and phylogenetic diversity of these enzymes than was previously known. Based on comparative analyses of Cld sequences and 3D structures, signature amino acid residues that can be employed to assess whether uncharacterized Cld-like proteins may have a high chlorite-dismutating activity were identified. Interestingly, proteins that contain all these signatures and are phylogenetically closely related to the novel-type Cld of
N. winogradskyi
exist in a large number of other microbes, including other nitrite oxidizers.
Chlamydia trachomatis Secretion of an Immunodominant Hypothetical Protein (CT795) into Host Cell Cytoplasm
Authors: Qi Manli , Lei Lei , Gong Siqi , Liu Quanzhong , DeLisa Matthew P. , Zhong Guangming
Published: March 2011

The
Chlamydia
-specific hypothetical protein CT795 was dominantly recognized by human antisera produced during
C. trachomatis
infection but not by animal antisera raised against dead chlamydia organisms. The immundominant region recognized by the human antibodies was mapped to the N-terminal fragment T22-S69. The endogenous CT795 was detected in the cytoplasm of host cells during
C. trachomatis
infection and was highly enriched in the host cytosolic fraction but absent in the purified chlamydia organisms, suggesting that CT795 is synthesized and secreted into host cell cytoplasm without incorporation into the organisms. All
C. trachomatis
serovars tested secreted CT795. A predicted signal peptide of CT795 directed the mature PhoA to cross
Escherichia coli
inner membranes. The secretion of CT795 in
Chlamydia
-infected cells was inhibited by a C
16
compound targeting signal peptidase I, but not by a C
1
compound known to block the type III secretion pathway. These results suggest that CT795, like CPAF (a
Chlamydia
-secreted virulence factor), is secreted into the host cell cytoplasm via a sec-dependent mechanism and not by a type III secretion pathway. The above characterizations of CT795 have provided important information for further understanding the potential roles of CT795 in
C. trachomatis
pathogenesis.
Heme Utilization by Nontypeable Haemophilus influenzae Is Essential and Dependent on Sap Transporter Function
Authors: Mason Kevin M. , Raffel Forrest K. , Ray William C. , Bakaletz Lauren O.
Published: March 2011

Bacterial strategies of innate immune evasion and essential metabolic functions are critical for commensal-host homeostasis. Previously, we showed that Sap translocator function is necessary for nontypeable
Haemophilus influenzae
(NTHI) behaviors that mediate diseases of the human airway. Antimicrobial peptide (AP) lethality is limited by binding mediated by the Sap complex. SapA shares homology with the dipeptide-binding protein (DppA) and the heme-binding lipoprotein (HbpA), both of which have previously been shown to bind the iron-containing compound heme, whose acquisition is essential for
Haemophilus
survival. Computational modeling revealed conserved SapA residues, similarly modeled to mediate heme binding in HbpA. Here, we directly demonstrate that SapA bound heme and was essential for heme utilization by iron-starved NTHI. Further, the Sap translocator permease mediated heme transport into the bacterial cytoplasm, thus defining a heretofore unknown mechanism of intracytoplasmic membrane heme transport in
Haemophilus
. Since we demonstrate multiple ligand specificity for the SapA-binding protein, we tested whether APs would compete with heme for SapA binding. We showed that human β-defensins 2 and 3, human cathelicidin LL-37, human neutrophil protein 1, and melittin displaced heme bound to SapA, thus supporting a hierarchy wherein immune evasion supercedes even the needed iron acquisition functions of the Sap system.
The Maturation Factors HoxR and HoxT Contribute to Oxygen Tolerance of Membrane-Bound [NiFe] Hydrogenase in Ralstonia eutropha H16
Authors: Fritsch Johannes , Lenz Oliver , Friedrich Bärbel
Published: March 2011

The membrane-bound [NiFe] hydrogenase (MBH) of
Ralstonia eutropha
H16 undergoes a complex maturation process comprising cofactor assembly and incorporation, subunit oligomerization, and finally twin-arginine-dependent membrane translocation. Due to its outstanding O
2
and CO tolerance, the MBH is of biotechnological interest and serves as a molecular model for a robust hydrogen catalyst. Adaptation of the enzyme to oxygen exposure has to take into account not only the catalytic reaction but also biosynthesis of the intricate redox cofactors. Here, we report on the role of the MBH-specific accessory proteins HoxR and HoxT, which are key components in MBH maturation at ambient O
2
levels. MBH-driven growth on H
2
is inhibited or retarded at high O
2
partial pressure (pO
2
) in mutants inactivated in the
hoxR
and
hoxT
genes. The ratio of mature and nonmature forms of the MBH small subunit is shifted toward the precursor form in extracts derived from the mutant cells grown at high pO
2
. Lack of
hoxR
and
hoxT
can phenotypically be restored by providing O
2
-limited growth conditions. Analysis of copurified maturation intermediates leads to the conclusion that the HoxR protein is a constituent of a large transient protein complex, whereas the HoxT protein appears to function at a final stage of MBH maturation. UV-visible spectroscopy of heterodimeric MBH purified from
hoxR
mutant cells points to alterations of the Fe-S cluster composition. Thus, HoxR may play a role in establishing a specific Fe-S cluster profile, whereas the HoxT protein seems to be beneficial for cofactor stability under aerobic conditions.
Monofunctional Transglycosylases Are Not Essential for Staphylococcus aureus Cell Wall Synthesis
Authors: Reed Patricia , Veiga Helena , Jorge Ana M. , Terrak Mohammed , Pinho Mariana G.
Published: March 2011

The polymerization of peptidoglycan is the result of two types of enzymatic activities: transglycosylation, the formation of linear glycan chains, and transpeptidation, the formation of peptide cross-bridges between the glycan strands.
Staphylococcus aureus
has four penicillin binding proteins (PBP1 to PBP4) with transpeptidation activity, one of which, PBP2, is a bifunctional enzyme that is also capable of catalyzing transglycosylation reactions. Additionally, two monofunctional transglycosylases have been reported in
S. aureus
: MGT, which has been shown to have
in vitro
transglycosylase activity, and a second putative transglycosylase, SgtA, identified only by sequence analysis. We have now shown that purified SgtA has
in vitro
transglycosylase activity and that both MGT and SgtA are not essential in
S. aureus.
However, in the absence of PBP2 transglycosylase activity, MGT but not SgtA becomes essential for cell viability. This indicates that
S. aureus
cells require one transglycosylase for survival, either PBP2 or MGT, both of which can act as the sole synthetic transglycosylase for cell wall synthesis. We have also shown that both MGT and SgtA interact with PBP2 and other enzymes involved in cell wall synthesis in a bacterial two-hybrid assay, suggesting that these enzymes may work in collaboration as part of a larger, as-yet-uncharacterized cell wall-synthetic complex.