bluuebunny/crossref_metadata_2025 huggingface dataset:
A modular versatile chip carrier for high-throughput screening of cell–biomaterial interactions
Authors: Unadkat H. V. , Rewagad R. R. , Hulsman M. , Hulshof G. F. B. , Truckenmüller R. K. , Stamatialis D. F. , Reinders M. J. T. , Eijkel J. C. T. , van den Berg A. , van Blitterswijk C. A. , de Boer J.
Published: November 2012

The field of biomaterials research is witnessing a steady rise in high-throughput screening approaches, comprising arrays of materials of different physico-chemical composition in a chip format. Even though the cell arrays provide many benefits in terms of throughput, they also bring new challenges. One of them is the establishment of robust homogeneous cell seeding techniques and strong control over cell culture, especially for long time periods. To meet these demands, seeding cells with low variation per tester area is required, in addition to robust cell culture parameters. In this study, we describe the development of a modular chip carrier which represents an important step in standardizing cell seeding and cell culture conditions in array formats. Our carrier allows flexible and controlled cell seeding and subsequent cell culture using dynamic perfusion. To demonstrate the application of our device, we successfully cultured and evaluated C2C12 premyoblast cell viability under dynamic conditions for a period of 5 days using an automated pipeline for image acquisition and analysis. In addition, using computational fluid dynamics, lactate and BMP-2 as model molecules, we estimated that there is good exchange of nutrients and metabolites with the flowing medium, whereas no cross-talk between adjacent TestUnits should be expected. Moreover, the shear stresses to the cells can be tailored uniformly over the entire chip area. Based on these findings, we believe our chip carrier may be a versatile tool for high-throughput cell experiments in biomaterials sciences.
A Fast and Efficient Adaptive Threshold Rate Control Scheme for Remote Sensing Images
Authors: Chen Xiao , Xu Xiaoqing
Published: November 2012

The JPEG2000 image compression standard is ideal for processing remote sensing images. However, its algorithm is complex and it requires large amounts of memory, making it difficult to adapt to the limited transmission and storage resources necessary for remote sensing images. In the present study, an improved rate control algorithm for remote sensing images is proposed. The required coded blocks are sorted downward according to their numbers of bit planes prior to entropy coding. An adaptive threshold computed from the combination of the minimum number of bit planes, along with the minimum rate-distortion slope and the compression ratio, is used to truncate passes of each code block during Tier-1 encoding. This routine avoids the encoding of all code passes and improves the coding efficiency. The simulation results show that the computational cost and working buffer memory size of the proposed algorithm reach only 18.13 and 7.81%, respectively, of the same parameters in the postcompression rate distortion algorithm, while the peak signal-to-noise ratio across the images remains almost the same. The proposed algorithm not only greatly reduces the code complexity and buffer requirements but also maintains the image quality.
Lateral Transpsoas Fusion: Indications and Outcomes
Authors: Patel Vishal C. , Park Daniel K. , Herkowitz Harry N.
Published: November 2012

Spinal fusion historically has been used extensively, and, recently, the lateral transpsoas approach to the thoracic and lumbar spine has become an increasingly common method to achieve fusion. Recent literature on this approach has elucidated its advantage over more traditional anterior and posterior approaches, which include a smaller tissue dissection, potentially lower blood loss, no need for an access surgeon, and a shorter hospital stay. Indications for the procedure have now expanded to include degenerative disc disease, spinal stenosis, degenerative scoliosis, nonunion, trauma, infection, and low-grade spondylolisthesis. Lateral interbody fusion has a similar if not lower rate of complications compared to traditional anterior and posterior approaches to interbody fusion. However, lateral interbody fusion has unique complications that include transient neurologic symptoms, motor deficits, and neural injuries that range from 1 to 60% in the literature. Additional studies are required to further evaluate and monitor the short- and long-term safety, efficacy, outcomes, and complications of lateral transpsoas procedures.
Loss of Function of Arabidopsis C-Terminal Domain Phosphatase-Like1 Activates Iron Deficiency Responses at the Transcriptional Level
Authors: Aksoy Emre , Jeong In Sil , Koiwa Hisashi
Published: November 2012

The expression of genes that control iron (Fe) uptake and distribution (i.e. Fe utilization-related genes) is tightly regulated. Fe deficiency strongly induces Fe utilization-related gene expression; however, little is known about the mechanisms that regulate this response in plants. Transcriptome analysis of an Arabidopsis (Arabidopsis thaliana) mutant defective in RNA polymerase II C-terminal domain-phosphatase-like1 (CPL1) revealed significant up-regulation of Fe utilization-related genes (e.g. IRON-REGULATED TRANSPORTER1), suggesting the importance of RNA metabolism in Fe signaling. An analysis using multiple cpl1 alleles established that cpl1 mutations enhanced specific transcriptional responses to low Fe availability. Changes in protein level were less prominent than those in transcript level, indicating that cpl1-2 mainly affects the Fe deficiency response at the transcriptional level. However, Fe content was significantly increased in the roots and decreased in the shoots of cpl1-2 plants, indicating that the cpl1 mutations do indeed affect Fe homeostasis. Furthermore, root growth of cpl1-2 showed improved tolerance to Fe deficiency and cadmium (Cd) toxicity. cpl1-2 plants accumulated more Cd in the shoots, suggesting that Cd toxicity in the roots of this mutant is averted by the transport of excess Cd to the shoots. Genetic data indicate that cpl1-2 likely activates Fe deficiency responses upstream of both FE-DEFICIENCY-INDUCED TRANSCRIPTION FACTOR-dependent and -independent signaling pathways. Interestingly, various osmotic stress/abscisic acid (ABA)-inducible genes were up-regulated in cpl1-2, and the expression of some ABA-inducible genes was controlled by Fe availability. We propose that the cpl1 mutations enhance Fe deficiency signaling and promote cross talk with a branch of the osmotic stress/ABA signaling pathway.
PRODUCT DIFFERENTIATION UNDER CONGESTION: HOTELLING WAS RIGHT
Authors: AHLIN CHRISTIAN , AHLIN PETER D.
Published: November 2012

We introduce negative network externalities—“congestion costs”—into H. Hotelling's (1929) model of spatial competition with linear transportation costs. For any firm locations on opposite sides of the midpoint, a pure strategy price equilibrium exists and is unique if congestion costs are strong enough relative to transportation costs. We analyze product differentiation and find that Hotelling's Principle of Minimum Differentiation comes closer to holding in the presence of congestion costs. The greater are congestion costs, the less differentiated products can be in (locationally symmetric) equilibrium. In fact, minimum differentiation comes arbitrarily close to holding depending on the magnitude of these costs relative to transportation costs. Intuitively, greater congestion effects stabilize competition at closer quarters, eliminating aggressive pricing equilibria. Thus, negative network externalities can play a significant role in product differentiation. (JEL D21, L15, R12)
Six new records of IdaeaTreitschke (Lepidoptera: Geometridae, Sterrhinae) from Korea
Authors: Choi Sei‐Woong , Kim Sung‐Soo
Published: November 2012

Six species of sterrhine genus Idaea Treitschke are reported for the first time from Korea: Idaea roseomarginaria (Inoue), I. paraula (Prout), I. sugillata (Bastelberger), I. imbecilla (Inoue), I. kyushuensis Sato, and I. nudaria (Christoph). To date, 23 species of Idaea have been reported from Korea. Diagnosis and descriptions of adults and photos of male and female genitalia are provided.
Habitat use and demography of Mus musculus in a rural landscape of Argentina
Authors: LEÓN Vanina A. , FRASCHINA Jimena , GUIDOBONO Juan S. , BUSCH Maria
Published: November 2012

The main goal of the paper was to determine the habitat distribution of the house mouse (Mus musculus) within a rural landscape of Buenos Aires province, Argentina. We also studied the seasonal variation in abundance and reproductive activity. The habitats studied were poultry farms, human houses in a small village, cropfields, pastures, cropfield and pasture edges, riparian habitats (streams), railway embankments and woodlots. We captured 817 M. musculus and 690 individuals of 5 native rodent species. M. musculus was captured in poultry farms, houses, riparian habitats, cropfield and borders, but it showed a significantly higher abundance in poultry farms compared to the other habitats. Its presence outside poultry farms was significantly related to the distance to streams and poultry farms. The mean trapping success index of M. musculus did not show significant variations between periods, but the proportion of active males was significantly higher in the spring–summer period than in the autumn–winter period. All captures of M. musculus in cropfields, borders and riparian habitats occurred in the spring–summer period. The capture of M. musculus in many types of habitats suggests that it can disperse outside poultry farms, and streams may be used as corridors.
Body size variation of four latitudinally‐separated populations of a toad species: age and growth rate as the proximate determinants
Authors: YU Tonglei , LU Xin
Published: November 2012

Age and growth rate are critical parameters determining the body size of adult anurans, which grow indeterminately. Therefore, evaluating their relative importance and selective pressures acting on them may help to explain variation in body size of anurans across geographical populations. Based on the data on body size and skeletochronological age of 4 Asiatic toad (Bufo gargarizans) populations that are widely separated in latitude, the present study showed that toads at the northern extremity of the range were the oldest, on average, and grew relatively fast, toads at the southern end of the range were relatively old but grew fastest, whereas mid‐latitude toads were intermediate in both traits. These results indicated that relative contributions of age and growth rate to body size differed among populations. From a life history perspective, greater age attained by toads at higher latitudes should result from delayed maturation, and faster growth from an adaptive response to strong seasonal time constraints; in contrast, selective pressures on the 2 parameters at lower latitudes might be associated, respectively, with low adult death rates and longer favorable growing seasons.
What do patients find helpful in psychotherapy? Implications for the therapeutic relationship in mental health nursing
Authors: Cahill J. , Paley G. , Hardy G.
Published: November 2012

Accessible summary

A group of four mental health nurses were trained to deliver and evaluate psychodynamic interpersonal therapy, an evidence‐based model of therapy, in a University research setting.

We were able to compare data with a group of cognitive behavioural therapists in the same University setting, enabling us to replicate a controlled research study which had compared psychodynamic interpersonal therapy and cognitive behavioural therapy.

Patients reported minimal significant differences between the therapies in terms of what they found helpful and from the patient's point of view, the relationship is the most important factor across both therapies.

Regardless of the model in which they have been trained, mental health nurses need to keep a primary focus on the therapeutic relationship.

This study examined client perception of the therapeutic impact of two models of therapy delivered by mental health nurses and clinical psychologists respectively – psychodynamic interpersonal therapy (PIT) and cognitive behavioural therapy (CBT). A non‐equivalent groups design was used in order to benchmark results against Llewelyn et al.: one group received PIT and the other received CBT. This design was utilized principally because the research was conducted across two practice settings where randomization was not feasible. We used two intact groups in practice research settings that received the therapies as reported in Llewelyn et al. Sixty‐one clients receiving CBT or PIT in practice research settings completed a Helpful Aspects of Therapy form after each session in order to measure client perceptions of helpful and hindering events in therapy. Only two out of the 13 impacts were rated as significantly different. PIT clients reported higher levels of ‘awareness’ than CBT clients, whereas CBT clients reported higher levels of problem solution than PIT clients. The results replicate Llewelyn et al.'s findings in that clients experienced theoretically different models of therapy as broadly similar in their therapeutic impact. We argue that this provides some support for the influence of ‘common’ rather than ‘specific’ factors in psychotherapy effectiveness in mental health nursing.
γ‐Aminobutyric acid transaminase deficiency impairs central carbon metabolism and leads to cell wall defects during salt stress in Arabidopsis roots
Authors: RENAULT HUGUES , EL AMRANI ABDELHAK , BERGER ADELINE , MOUILLE GRÉGORY , SOUBIGOU‐TACONNAT LUDIVINE , BOUCHEREAU ALAIN , DELEU CAROLE
Published: November 2012

GABA (γ‐aminobutyric acid) is a ubiquitous non‐protein amino acid that commonly accumulates in plants in response to stressful conditions. However, functions of GABA in plants remain still unclear. In this study, we provide metabolic and transcriptional evidences that GABA metabolism is involved in central carbon metabolism regulation upon salt stress in roots. Furthermore, we show that GABA transaminase deficiency results in deleterious consequences for root development and cell wall elaboration during salt stress. Our work thus highlights the critical function of GABA metabolism in root metabolic acclimation to saline conditions.
Engineering Complex Tissues
Authors: Atala Anthony , Kasper F. Kurtis , Mikos Antonios G.
Published: November 2012

Advances in tissue engineering technologies will enable regeneration of complex tissues and organs.
Calling All Satellite Cells!
Authors: Baker Aaron B.
Published: November 2012

A signaling pathway crucial for satellite cell expansion may hold the key to enhancing cell-based therapies for muscle regeneration.
Osmoprotection of Bacillus subtilis through Import and Proteolysis of Proline-Containing Peptides
Authors: Zaprasis Adrienne , Brill Jeanette , Thüring Marietta , Wünsche Guido , Heun Magnus , Barzantny Helena , Hoffmann Tamara , Bremer Erhard
Published: November 2012

Bacillus subtilis
can attain cellular protection against the detrimental effects of high osmolarity through osmotically induced
de novo
synthesis and uptake of the compatible solute
l
-proline. We have now found that
B. subtilis
can also exploit exogenously provided proline-containing peptides of various lengths and compositions as osmoprotectants. Osmoprotection by these types of peptides is generally dependent on their import via the peptide transport systems (Dpp, Opp, App, and DtpT) operating in
B. subtilis
and relies on their hydrolysis to liberate proline. The effectiveness with which proline-containing peptides confer osmoprotection varies considerably, and this can be correlated with the amount of the liberated and subsequently accumulated free proline by the osmotically stressed cell. Through gene disruption experiments, growth studies, and the quantification of the intracellular proline pool, we have identified the PapA (YqhT) and PapB (YkvY) peptidases as responsible for the hydrolysis of various types of Xaa-Pro dipeptides and Xaa-Pro-Xaa tripeptides. The PapA and PapB peptidases possess overlapping substrate specificities. In contrast, osmoprotection by peptides of various lengths and compositions with a proline residue positioned at their N terminus was not affected by defects in the PapA and PapB peptidases. Taken together, our data provide new insight into the physiology of the osmotic stress response of
B. subtilis
. They illustrate the flexibility of this ubiquitously distributed microorganism to effectively exploit environmental resources in its acclimatization to sustained high-osmolarity surroundings through the accumulation of compatible solutes.
Combinatorial Mutagenesis and Selection of Improved Signal Sequences and Their Application for High-Level Production of Translocated Heterologous Proteins in Escherichia coli
Authors: Heggeset Tonje Marita Bjerkan , Kucharova Veronika , Nærdal Ingemar , Valla Svein , Sletta Håvard , Ellingsen Trond Erling , Brautaset Trygve
Published: November 2012

We previously designed the consensus signal peptide (CSP) and demonstrated that it can be used to strongly stimulate heterologous protein production in
Escherichia coli
. A comparative study using CSP and two bacterial signal sequences, pelB and ompA, showed that the effect of signal sequences on both expression level and translocation efficiency can be highly protein specific. We report here the generation of CSP mutant libraries by a combinatorial mutagenesis approach. Degenerated CSP oligonucleotides were cloned in frame with the 5′ end of the
bla
gene, encoding the mature periplasmic β-lactamase released from its native signal sequence. This novel design allows for a direct selection of improved signal sequences that positively affect the expression level and/or translocation efficiency of β-lactamase, based on the ampicillin tolerance level of the
E. coli
host cells. By using this strategy, 61 different CSP mutants with up to 8-fold-increased ampicillin tolerance level and up to 5.5-fold-increased β-lactamase expression level were isolated and characterized genetically. A subset of the CSP mutants was then tested with the alternative reporter gene
phoA
, encoding periplasmic alkaline phosphatase (AP), resulting in an up to 8-fold-increased production level of active AP protein in
E. coli
. Moreover, it was demonstrated that the CSP mutants can improve the production of the medically important human interferon α2b under high-cell-density cultivations. Our results show that there is a clear potential for improving bacterial signal sequences by using combinatorial mutagenesis, and bioinformatics analyses indicated that the beneficial mutations could not be rationally predicted.
Landscape and Meteorological Factors Affecting Prevalence of Three Food-Borne Pathogens in Fruit and Vegetable Farms
Authors: Strawn Laura K. , Fortes Esther D. , Bihn Elizabeth A. , Nightingale Kendra K. , Gröhn Yrjö T. , Worobo Randy W. , Wiedmann Martin , Bergholz Peter W.
Published: November 2012

Produce-related outbreaks have been traced back to the preharvest environment. A longitudinal study was conducted on five farms in New York State to characterize the prevalence, persistence, and diversity of food-borne pathogens in fresh produce fields and to determine landscape and meteorological factors that predict their presence. Produce fields were sampled four times per year for 2 years. A total of 588 samples were analyzed for
Listeria monocytogenes
,
Salmonella
, and Shiga toxin-producing
Escherichia coli
(STEC). The prevalence measures of
L. monocytogenes
,
Salmonella
, and STEC were 15.0, 4.6, and 2.7%, respectively.
L. monocytogenes
and
Salmonella
were detected more frequently in water samples, while STEC was detected with equal frequency across all sample types (soil, water, feces, and drag swabs).
L. monocytogenes sigB
gene allelic types 57, 58, and 61 and
Salmonella enterica
serovar Cerro were repeatedly isolated from water samples. Soil available water storage (AWS), temperature, and proximity to three land cover classes (water, roads and urban development, and pasture/hay grass) influenced the likelihood of detecting
L. monocytogenes
. Drainage class, AWS, and precipitation were identified as important factors in
Salmonella
detection. This information was used in a geographic information system framework to hypothesize locations of environmental reservoirs where the prevalence of food-borne pathogens may be elevated. The map indicated that not all croplands are equally likely to contain environmental reservoirs of
L. monocytogenes
. These findings advance recommendations to minimize the risk of preharvest contamination by enhancing models of the environmental constraints on the survival and persistence of food-borne pathogens in fields.
Taking the Starch out of Oral Biofilm Formation: Molecular Basis and Functional Significance of Salivary α-Amylase Binding to Oral Streptococci
Authors: Nikitkova Anna E. , Haase Elaine M. , Scannapieco Frank A.
Published: November 2012

α-Amylase-binding streptococci (ABS) are a heterogeneous group of commensal oral bacterial species that comprise a significant proportion of dental plaque microfloras. Salivary α-amylase, one of the most abundant proteins in human saliva, binds to the surface of these bacteria via specific surface-exposed α-amylase-binding proteins. The functional significance of α-amylase-binding proteins in oral colonization by streptococci is important for understanding how salivary components influence oral biofilm formation by these important dental plaque species. This review summarizes the results of an extensive series of studies that have sought to define the molecular basis for α-amylase binding to the surface of the bacterium as well as the biological significance of this phenomenon in dental plaque biofilm formation.
Influence of Salmonella enterica Serovar Enteritidis Infection on the Development of the Cecum Microbiota in Newly Hatched Chicks
Authors: Juricova H. , Videnska P. , Lukac M. , Faldynova M. , Babak V. , Havlickova H. , Sisak F. , Rychlik I.
Published: November 2012

Terminal restriction fragment length polymorphism and quantitative PCR showed that the cecal microbiota of chicks up to the age of 21 days was dominated by representatives of the orders
Enterobacteriales
,
Clostridiales
, and
Lactobacillales
.
Salmonella enterica
serovar Enteritidis infection caused the greatest changes in the gut microbiota when 1-day-old chicks were infected, compared with the infection of 4- and 16-day-old chicks.
A New, Sensitive Marine Microalgal Recombinant Biosensor Using Luminescence Monitoring for Toxicity Testing of Antifouling Biocides
Authors: Sanchez-Ferandin Sophie , Leroy Fanny , Bouget François-Yves , Joux Fabien
Published: November 2012

In this study, we propose the use of the marine green alga
Ostreococcus tauri
, the smallest free-living eukaryotic cell known to date, as a new luminescent biosensor for toxicity testing in the environment. Diuron and Irgarol 1051, two antifouling biocides commonly encountered in coastal waters, were chosen to test this new biosensor along with two degradation products of diuron. The effects of various concentrations of the antifoulants on four genetic constructs of
O. tauri
(based on genes involved in photosynthesis, cell cycle, and circadian clock) were compared using 96-well culture microplates and a luminometer to automatically measure luminescence over 3 days. This was compared to growth inhibition of
O. tauri
wild type under the same conditions. Luminescence appeared to be more sensitive than growth inhibition as an indicator of toxicity. Cyclin-dependent kinase (CDKA), a protein involved in the cell cycle, fused to luciferase (CDKA-Luc) was found to be the most sensitive of the biosensors, allowing an accurate determination of the 50% effective concentration (EC
50
) after only 2 days (diuron, 5.65 ± 0.44 μg/liter; Irgarol 1015, 0.76 ± 0.10 μg/liter). The effects of the antifoulants on the CDKA-Luc biosensor were then compared to growth inhibition in natural marine phytoplankton. The effective concentrations of diuron and Irgarol 1051 were found to be similar, indicating that this biosensor would be suitable as a reliable ecotoxicological test. The advantage of this biosensor over cell growth inhibition testing is that the process can be easily automated and could provide a high-throughput laboratory approach to perform short-term toxicity tests. The ability to genetically transform and culture recombinant
O. tauri
gives it huge potential for screening many other toxic compounds.
Comparative Analysis of Leptospira Strains Isolated from Environmental Soil and Water in the Philippines and Japan
Authors: Saito Mitsumasa , Villanueva Sharon Y. A. M. , Chakraborty Antara , Miyahara Satoshi , Segawa Takaya , Asoh Tatsuma , Ozuru Ryo , Gloriani Nina G. , Yanagihara Yasutake , Yoshida Shin-ichi
Published: November 2012

There have been few reports on the epidemiological analysis of environmental
Leptospira
isolates. This is probably because the isolation of leptospires from the environment was usually unsuccessful due to the overgrowth of contaminants and the slow growth of
Leptospira
. In this study, we collected a total of 88 samples of soil and water from three sites: Metro Manila and Nueva Ecija, Philippines (an area where
Leptospira
is now endemic), and Fukuoka, Japan (an area where
Leptospira
was once endemic). We succeeded in isolating
Leptospira
from 37 samples by using the novel combination of five antimicrobial agents reported in 2011. The frequencies of positive isolation of
Leptospira
in the Philippines and Japan were 40 and 46%, respectively. For
Leptospira
-positive samples, five colonies from each sample were isolated and analyzed by pulsed-field gel electrophoresis (PFGE). The isolates from each area showed their respective characteristics in phylogenetic trees based on the PFGE patterns. Some isolates were closely related to each other across borders. Based on 16S rRNA gene-based phylogenetic analysis, four isolates in Fukuoka were identified as a pathogenic species,
L. alstonii
; however, its virulence had been lost. One isolate from Nueva Ecija was identified as the intermediate pathogenic species
Leptospira licerasiae
. Most of the isolates from the environment belonged to nonpathogenic
Leptospira
species. We also investigated the strain variation among the isolates in a puddle over 5 months. We demonstrated, using PFGE analysis, that
Leptospira
survived in the wet soil on dry days and appeared in the surface water on rainy days. These results showed that the soil could be a reservoir of leptospires in the environment.
Impact of Long-Term Diesel Contamination on Soil Microbial Community Structure
Authors: Sutton Nora B. , Maphosa Farai , Morillo Jose A. , Abu Al-Soud Waleed , Langenhoff Alette A. M. , Grotenhuis Tim , Rijnaarts Huub H. M. , Smidt Hauke
Published: November 2012

Microbial community composition and diversity at a diesel-contaminated railway site were investigated by pyrosequencing of bacterial and archaeal 16S rRNA gene fragments to understand the interrelationships among microbial community composition, pollution level, and soil geochemical and physical properties. To this end, 26 soil samples from four matrix types with various geochemical characteristics and contaminant concentrations were investigated. The presence of diesel contamination significantly impacted microbial community composition and diversity, regardless of the soil matrix type. Clean samples showed higher diversity than contaminated samples (
P
< 0.001). Bacterial phyla with high relative abundances in all samples included
Proteobacteria
,
Firmicutes
,
Actinobacteria
,
Acidobacteria
, and
Chloroflexi
. High relative abundances of
Archaea
, specifically of the phylum
Euryarchaeota
, were observed in contaminated samples. Redundancy analysis indicated that increased relative abundances of the phyla
Chloroflexi
,
Firmicutes
, and
Euryarchaeota
correlated with the presence of contamination. Shifts in the chemical composition of diesel constituents across the site and the abundance of specific operational taxonomic units (OTUs; defined using a 97% sequence identity threshold) in contaminated samples together suggest that natural attenuation of contamination has occurred. OTUs with sequence similarity to strictly anaerobic
Anaerolineae
within the
Chloroflexi
, as well as to
Methanosaeta
of the phylum
Euryarchaeota
, were detected.
Anaerolineae
and
Methanosaeta
are known to be associated with anaerobic degradation of oil-related compounds; therefore, their presence suggests that natural attenuation has occurred under anoxic conditions. This research underscores the usefulness of next-generation sequencing techniques both to understand the ecological impact of contamination and to identify potential molecular proxies for detection of natural attenuation.
The Mangotoxin Biosynthetic Operon (
mbo
) Is Specifically Distributed within Pseudomonas syringae Genomospecies 1 and Was Acquired Only Once during Evolution
Authors: Carrión Víctor J. , Gutiérrez-Barranquero José A. , Arrebola Eva , Bardaji Leire , Codina Juan C. , de Vicente Antonio , Cazorla Francisco M. , Murillo Jesús
Published: November 2012

Mangotoxin production was first described in
Pseudomonas syringae
pv. syringae strains. A phenotypic characterization of 94
P. syringae
strains was carried out to determine the genetic evolution of the mangotoxin biosynthetic operon (
mbo
). We designed a PCR primer pair specific for the
mbo
operon to examine its distribution within the
P. syringae
complex. These primers amplified a 692-bp DNA fragment from 52 mangotoxin-producing strains and from 7 non-mangotoxin-producing strains that harbor the
mbo
operon, whereas 35 non-mangotoxin-producing strains did not yield any amplification. This, together with the analysis of draft genomes, allowed the identification of the
mbo
operon in five pathovars (pathovars aptata, avellanae, japonica, pisi, and syringae), all of which belong to genomospecies 1, suggesting a limited distribution of the
mbo
genes in the
P. syringae
complex. Phylogenetic analyses using partial sequences from housekeeping genes differentiated three groups within genomospecies 1. All of the strains containing the
mbo
operon clustered in groups I and II, whereas those lacking the operon clustered in group III; however, the relative branching order of these three groups is dependent on the genes used to construct the phylogeny. The
mbo
operon maintains synteny and is inserted in the same genomic location, with high sequence conservation around the insertion point, for all the strains in groups I and II. These data support the idea that the
mbo
operon was acquired horizontally and only once by the ancestor of groups I and II from genomospecies 1 within the
P. syringae
complex.
Characterization of a Newly Discovered Symbiont of the Whitefly Bemisia tabaci (Hemiptera: Aleyrodidae)
Authors: Bing Xiao-Li , Yang Jiao , Zchori-Fein Einat , Wang Xiao-Wei , Liu Shu-Sheng
Published: November 2012

Bemisia tabaci
(Hemiptera: Aleyrodidae) is a species complex containing >28 cryptic species, some of which are important crop pests worldwide. Like many other sap-sucking insects, whiteflies harbor an obligatory symbiont, “
Candidatus
Portiera aleyrodidarum,” and a number of secondary symbionts. So far, six genera of secondary symbionts have been identified in
B. tabaci
. In this study, we report and describe the finding of an additional bacterium in the indigenous
B. tabaci
cryptic species China 1 (formerly known as
B. tabaci
biotype ZHJ3). Phylogenetic analysis based on the 16S rRNA and
gltA
genes showed that the bacterium belongs to the
Alphaproteobacteria
subdivision of the
Proteobacteria
and has a close relationship with human pathogens of the genus
Orientia
. Consequently, we temporarily named it
Orientia
-like organism (OLO). OLO was found in six of eight wild populations of
B. tabaci
China 1, with the infection rate ranging from 46.2% to 76.8%. Fluorescence
in situ
hybridization (FISH) of
B. tabaci
China 1 in nymphs and adults revealed that OLOs are confined to the bacteriome and co-occur with “
Ca
. Portiera aleyrodidarum.” The vertical transmission of OLO was demonstrated by detection of OLO at the anterior pole end of the oocytes through FISH. Quantitative PCR analysis of population dynamics suggested a complex interaction between “
Ca
. Portiera aleyrodidarum” and OLO. Based on these results, we propose “
Candidatus
Hemipteriphilus asiaticus” for the classification of this symbiont from
B. tabaci
.
Improved Gene Ontology Annotation for Biofilm Formation, Filamentous Growth, and Phenotypic Switching in Candida albicans
Authors: Inglis Diane O. , Skrzypek Marek S. , Arnaud Martha B. , Binkley Jonathan , Shah Prachi , Wymore Farrell , Sherlock Gavin
Published: November 2012

The opportunistic fungal pathogen
Candida albicans
is a significant medical threat, especially for immunocompromised patients. Experimental research has focused on specific areas of
C. albicans
biology, with the goal of understanding the multiple factors that contribute to its pathogenic potential. Some of these factors include cell adhesion, invasive or filamentous growth, and the formation of drug-resistant biofilms. The Gene Ontology (GO) (
www.geneontology.org
) is a standardized vocabulary that the
Candida
Genome Database (CGD) (
www.candidagenome.org
) and other groups use to describe the functions of gene products. To improve the breadth and accuracy of pathogenicity-related gene product descriptions and to facilitate the description of as yet uncharacterized but potentially pathogenicity-related genes in
Candida
species, CGD undertook a three-part project: first, the addition of terms to the biological process branch of the GO to improve the description of fungus-related processes; second, manual recuration of gene product annotations in CGD to use the improved GO vocabulary; and third, computational ortholog-based transfer of GO annotations from experimentally characterized gene products, using these new terms, to uncharacterized orthologs in other
Candida
species. Through genome annotation and analysis, we identified candidate pathogenicity genes in seven non-
C. albicans Candida
species and in one additional
C. albicans
strain, WO-1. We also defined a set of
C. albicans
genes at the intersection of biofilm formation, filamentous growth, pathogenesis, and phenotypic switching of this opportunistic fungal pathogen, which provides a compelling list of candidates for further experimentation.
Identification of the Mating-Type (MAT) Locus That Controls Sexual Reproduction of Blastomyces dermatitidis
Authors: Li Wenjun , Sullivan Thomas D. , Walton Eric , Averette Anna Floyd , Sakthikumar Sharadha , Cuomo Christina A. , Klein Bruce S. , Heitman Joseph
Published: November 2012

Blastomyces dermatitidisis a dimorphic fungal pathogen that primarily causes blastomycosis in the midwestern and northern United States and Canada. While the genes controlling sexual development have been known for a long time, the genes controlling sexual reproduction ofB. dermatitidis(teleomorph,Ajellomyces dermatitidis) are unknown. We identified the mating-type (MAT) locus in theB. dermatitidisgenome by comparative genomic approaches. TheB. dermatitidis MATlocus resembles those of other dimorphic fungi, containing either an alpha-box (MAT1-1) or an HMG domain (MAT1-2) gene linked to theAPN2,SLA2, andCOX13genes. However, in some strains ofB. dermatitidis, theMATlocus harbors transposable elements (TEs) that make it unusually large compared to theMATlocus of other dimorphic fungi. Based on theMATlocus sequences ofB. dermatitidis, we designed specific primers for PCR determination of the mating type. TwoB. dermatitidisisolates of opposite mating types were cocultured on mating medium. Immature sexual structures were observed starting at 3 weeks of coculture, with coiled-hyphae-containing cleistothecia developing over the next 3 to 6 weeks. Genetic recombination was detected in potential progeny by mating-type determination, PCR-restriction fragment length polymorphism (PCR-RFLP), and random amplification of polymorphic DNA (RAPD) analyses, suggesting that a meiotic sexual cycle might have been completed. The F1 progeny were sexually fertile when tested with strains of the opposite mating type. Our studies provide a model for the evolution of theMATlocus in the dimorphic and closely related fungi and open the door to classic genetic analysis and studies on the possible roles of mating and mating type in infection and virulence.
Candida albicans Induces Arginine Biosynthetic Genes in Response to Host-Derived Reactive Oxygen Species
Authors: Jiménez-López Claudia , Collette John R. , Brothers Kimberly M. , Shepardson Kelly M. , Cramer Robert A. , Wheeler Robert T. , Lorenz Michael C.
Published: November 2012

The interaction of
Candida albicans
with phagocytes of the host's innate immune system is highly dynamic, and its outcome directly impacts the progression of infection. While the switch to hyphal growth within the macrophage is the most obvious physiological response, much of the genetic response reflects nutrient starvation: translational repression and induction of alternative carbon metabolism. Changes in amino acid metabolism are not seen, with the striking exception of arginine biosynthesis, which is upregulated in its entirety during coculture with macrophages. Using single-cell reporters, we showed here that arginine biosynthetic genes are induced specifically in phagocytosed cells. This induction is lower in magnitude than during arginine starvation
in vitro
and is driven not by an arginine deficiency within the phagocyte but instead by exposure to reactive oxygen species (ROS). Curiously, these genes are induced in a narrow window of sublethal ROS concentrations.
C. albicans
cells phagocytosed by primary macrophages deficient in the
gp91

phox

subunit of the phagocyte oxidase do not express the
ARG
pathway, indicating that the induction is dependent on the phagocyte oxidative burst.
C. albicans arg
pathway mutants are retarded in germ tube and hypha formation within macrophages but are not notably more sensitive to ROS. We also find that the
ARG
pathway is regulated not by the general amino acid control response but by transcriptional regulators similar to the
Saccharomyces cerevisiae
ArgR complex. In summary, phagocytosis induces this single amino acid biosynthetic pathway in an ROS-dependent manner.
Structural Comparison of ColH and ColG Collagen-Binding Domains from Clostridium histolyticum
Authors: Bauer Ryan , Wilson Jeffrey J. , Philominathan Sagaya Theresa Leena , Davis Dan , Matsushita Osamu , Sakon Joshua
Published: November 2012

Clostridium histolyticum
secretes collagenases, ColG and ColH, that cause extensive tissue destruction in myonecrosis. The C-terminal collagen-binding domain (CBD) of collagenase is required for insoluble collagen fibril binding and subsequent collagenolysis. The high-resolution crystal structures of ColG-CBD (s3b) and ColH-CBD (s3) are reported in this paper. The new X-ray structure of s3 was solved at 2.0-Å resolution (
R
= 17.4%;
R
free
= 23.3%), while the resolution of the previously determined s3b was extended to 1.4 Å (
R
= 17.9%;
R
free
= 21.0%). Despite sharing only 30% sequence identity, the molecules resemble one another closely (root mean square deviation [RMSD] C
α
= 1.5 Å). All but one residue, whose side chain chelates with Ca
2+
, are conserved. The dual Ca
2+
binding site in s3 is completed by an unconserved aspartate. Differential scanning calorimetric measurements showed that s3 gains thermal stability, comparable to s3b, by binding to Ca
2+
(
holo

T
m

= 94.1°C;
apo

T
m

= 70.2°C).
holo
s3 is also stabilized against chemical denaturants urea and guanidine HCl. The three most critical residues for collagen interaction in s3b are conserved in s3. The general shape of the binding pocket is retained by altered loop structures and side chain positions. Small-angle X-ray scattering data revealed that s3 also binds asymmetrically to minicollagen. Besides the calcium-binding sites and the collagen-binding pocket, architecturally important hydrophobic residues and the hydrogen-bonding network around the
cis
-peptide bond are well conserved within the metallopeptidase subfamily M9B. CBDs were previously shown to bind to the extracellular matrix of various tissues. Compactness and extreme stability in physiological Ca
2+
concentration possibly make both CBDs suitable for targeted growth factor delivery.
Genetic Characterization of a Novel Rhizobial Plasmid Conjugation System in Rhizobium leguminosarum bv. viciae Strain VF39SM
Authors: Ding Hao , Yip Cynthia B. , Hynes Michael F.
Published: November 2012

Rhizobium leguminosarum
strain VF39SM contains two plasmids that have previously been shown to be self-transmissible by conjugation. One of these plasmids, pRleVF39b, is shown in this study to carry a set of plasmid transfer genes that differs significantly from conjugation systems previously studied in the rhizobia but is similar to an uncharacterized set of genes found in
R. leguminosarum
bv. trifolii strain WSM2304. The entire sequence of the transfer region on pRleVF39b was determined as part of a genome sequencing project, and the roles of the various genes were examined by mutagenesis. The transfer region contains a complete set of mating pair formation (Mpf) genes, a
traG
gene, and a relaxase gene,
traA
, all of which appear to be necessary for plasmid transfer. Experimental evidence suggested the presence of two putative origins of transfer within the gene cluster. A regulatory gene,
trbR
, was identified in the region between
traA
and
traG
and was mutated. TrbR was shown to function as a repressor of both
trb
gene expression and plasmid transfer.
Pseudomonas syringae pv. tomato DC3000 CmaL (PSPTO4723), a DUF1330 Family Member, Is Needed To Produce
l
-
allo
-Isoleucine, a Precursor for the Phytotoxin Coronatine
Authors: Worley Jay N. , Russell Alistair B. , Wexler Aaron G. , Bronstein Philip A. , Kvitko Brian H. , Krasnoff Stuart B. , Munkvold Kathy R. , Swingle Bryan , Gibson Donna M. , Collmer Alan
Published: November 2012

Pseudomonas syringae
pv. tomato DC3000 produces the phytotoxin coronatine, a major determinant of the leaf chlorosis associated with DC3000 pathogenesis. The DC3000 PSPTO4723 (
cmaL
) gene is located in a genomic region encoding type III effectors; however, it promotes chlorosis in the model plant
Nicotiana benthamiana
in a manner independent of type III secretion. Coronatine is produced by the ligation of two moieties, coronafacic acid (CFA) and coronamic acid (CMA), which are produced by biosynthetic pathways encoded in separate operons. Cross-feeding experiments, performed in
N. benthamiana
with
cfa
,
cma
, and
cmaL
mutants, implicate CmaL in CMA production. Furthermore, analysis of bacterial supernatants under coronatine-inducing conditions revealed that mutants lacking either the
cma
operon or
cmaL
accumulate CFA rather than coronatine, supporting a role for CmaL in the regulation or biosynthesis of CMA. CmaL does not appear to regulate CMA production, since the expression of proteins with known roles in CMA production is unaltered in
cmaL
mutants. Rather, CmaL is needed for the first step in CMA synthesis, as evidenced by the fact that wild-type levels of coronatine production are restored to a Δ
cmaL
mutant when it is supplemented with 50 μg/ml
l
-
allo
-isoleucine, the starting unit for CMA production.
cmaL
is found in all other sequenced
P. syringae
strains with coronatine biosynthesis genes. This characterization of CmaL identifies a critical missing factor in coronatine production and provides a foundation for further investigation of a member of the widespread DUF1330 protein family.
Detoxification of 7-Dehydrocholesterol Fatal to Helicobacter pylori Is a Novel Role of Cholesterol Glucosylation
Authors: Shimomura Hirofumi , Hosoda Kouichi , McGee David J. , Hayashi Shunji , Yokota Kenji , Hirai Yoshikazu
Published: November 2012

The glucosylation of free cholesterol (FC) by
Helicobacter pylori
cells has various biological significances for the survival of this bacterium.
H. pylori
cells with glucosylated FC are capable of evading host immune systems, such as phagocytosis by macrophages and activation of antigen-specific T cells, and surviving in the gastric mucosal tissues for long periods. An additional role of cholesterol glucosylation in the survival of
H. pylori
which is distinct from the role of escaping the host immune system, however, has yet to be identified. This study demonstrated that 7-dehydrocholesterol (7dFC), an FC precursor, is a toxic compound fatal to
H. pylori
cells, but the cell membrane of
H. pylori
is capable of absorbing this toxic sterol via glucosylation. In contrast to the case with 7dFC, no toxicity to
H. pylori
cells was detected from the glucosylated 7dFC. In addition,
cgt
gene mutant
H. pylori
cells that cannot glucosylate cholesterols had higher susceptibility to the toxic action of 7dFC than wild-type
H. pylori
cells. These results indicate that the
cgt
gene product of
H. pylori
serves to detoxify the sterol fatal to this bacterium and to permit this toxic sterol as a cell membrane lipid component. In summary, this study defined a novel role of cholesterol glucosylation in
H. pylori
.
Characterization of BreR Interaction with the Bile Response Promoters
breAB
and
breR
in Vibrio cholerae
Authors: Cerda-Maira Francisca A. , Kovacikova Gabriela , Jude Brooke A. , Skorupski Karen , Taylor Ronald K.
Published: November 2012

The
Vibrio cholerae
BreR protein is a transcriptional repressor of the
breAB
efflux system operon, which encodes proteins involved in bile resistance. In a previous study (F. A. Cerda-Maira, C. S. Ringelberg, and R. K. Taylor, J. Bacteriol.
190:
7441–7452, 2008), we used gel mobility shift assays to determine that BreR binds at two independent binding sites at the
breAB
promoter and a single site at its own promoter. Here it is shown, by DNase I footprinting and site-directed mutagenesis, that BreR is able to bind at a distal and a proximal site in the
breAB
promoter. However, only one of these sites, the proximal 29-bp site, is necessary for BreR-mediated transcriptional repression of
breAB
expression. In addition, it was determined that BreR represses its own expression by recognizing a 28-bp site at the
breR
promoter. These sites comprise regions of dyad symmetry within which residues critical for BreR function could be identified. The BreR consensus sequence AANGTANAC-N
6
-GTNTACNTT overlaps the −35 region at both promoters, implying that the repression of gene expression is achieved by interfering with RNA polymerase binding at these promoters.
Bezafibrate to prevent relapsing pancreatitis in WAGR syndrome
Authors: Diacono Daniela , Fagbemi Andrew , Puleston Joanne , Banerjee Indraneel
Published: November 2012

Recurrent episodes of acute, severe pancreatitis are a feature of the rare WAGR congenital anomaly syndrome. Although pancreatitis is well recognised as a life-threatening association of WAGR syndrome, treatment to prevent pancreatitis is not recommended. We report the successful prevention of hypertriglyceridaemia-associated relapsing pancreatitis in a child with WAGR syndrome with Bezafibrate. In addition to dietary fat restriction, Bezafibrate may be a useful therapeutic adjunct in WAGR related relapsing pancreatitis.